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1 Introduction TB is a disease that is caused by the bacterium Mycobacterium tuberculosis. The bacteria usually attack the lungs, but TB bacteria can attack any part of the body, such as the kidney, spine, or brain. If not treated properly, TB can be fatal. TB was once the leading cause of death. 2 Case presentation A 7-year-old Saudi girl presented to the emergency department at this tertiary care hospital with a worsening course of fever, cough, weight loss, and abdominal pain over 3 weeks. She was admitted to a local hospital earlier and was suspected of having lymphoma. There was a history of recent contact with an aunt who was diagnosed as having pulmonary TB 6 months earlier and who was started on treatment with 4 drugs. The results of culture and susceptibility testing are not available. The child's place of residence in AL-Jouf was crowded, with many extended family members, and was poorly ventilated. Her father was 45 years old, and her mother was 35 years old. Both were healthy, but of poor socioeconomic status. On examination, the child looked acutely ill, pale, tachypneic, and tachycardic, with a temperature of 39.6 °C, WT 11.7 < 5th centile, and HT 106 cm > 50th centile. There were multiple cautery marks over the chest wall and upper abdomen, and on chest auscultation, there was a marked decrease of air entry on the right side, with bronchial breathing and basal crepitations.

achypneic, and tachycardic, with a temperature of 39.6 °C, WT 11.7 < 5th centile, and HT 106 cm > 50th centile. There were multiple cautery marks over the chest wall and upper abdomen, and on chest auscultation, there was a marked decrease of air entry on the right side, with bronchial breathing and basal crepitations. 2.1 Laboratory data (Table 1) The chest X-ray (Fig. 1) demonstrated an airspace disease involving the right lower lobe and a cystic change involving the right upper lobe with blunted cardiopulmonary angles. A CT scan of the chest (Fig. 2) demonstrated large airspace consolidation involving the right middle and lower lobes, with a large cavity in the right upper lobe, and bilateral miliary nodules that had a tree-in-bud appearance. There were multiple mediastinal necrotic lymph nodes suggestive of acute on top of chronic TB. The abdominal CT showed small hypo-dense splenic lesions.Table 1 Clinical investigation upon diagnosis and during treatment of DOT.

ity in the right upper lobe, and bilateral miliary nodules that had a tree-in-bud appearance. There were multiple mediastinal necrotic lymph nodes suggestive of acute on top of chronic TB. The abdominal CT showed small hypo-dense splenic lesions.Table 1 Clinical investigation upon diagnosis and during treatment of DOT. Laboratory data On diagnosis On DOT Normal range CBC and differential WBC, 109/L 19.17 5.86 4.30–11.30 RBC, 1012/L 3.49 4.34 4.30–5.50 hemoglobin, g/L 67 113 110–150 Hematocrit, L/L 0.294 0.339 0.350–0.450 MCVfL 73.7 78.1 75–95 MCHCpg 22.5 26 24–30 RDW, % 19 12.9 11–15 Platelet, 109/L 25 231 155–435 Neutrophil absolute, 109/L 6.20 1.71 1.35–7.50 Lymphocyte absolute, 109/L 1.21 2.88 1.90–4.90 ESR, mm/h 140 9 0–15 CRP, mg/L 103 0.4 ≤3 mg/L Albumin, g/L 17 42.9 32–48 AST, U/L 373 33.5 10–45 ALT, U/L 114 24.9 10–35 Bilirubin, total, umol/L 6.3 4.9 0–21 Alkaline phosphate, u/L 269.5 246 100–300 HIV 1–2 antibody screening Non reactive Figure 1 Airspace disease involving the right middle and lower lobe with a cystic change involving the right upper lobe. Figure 2 Improvement of the airspace disease in the right middle and lower lobe after starting treatment. Bronchial wall thickening with peribronchial infiltrates in the perihilar region.

Laboratory data On diagnosis On DOT Normal range CBC and differential WBC, 109/L 19.17 5.86 4.30–11.30 RBC, 1012/L 3.49 4.34 4.30–5.50 hemoglobin, g/L 67 113 110–150 Hematocrit, L/L 0.294 0.339 0.350–0.450 MCVfL 73.7 78.1 75–95 MCHCpg 22.5 26 24–30 RDW, % 19 12.9 11–15 Platelet, 109/L 25 231 155–435 Neutrophil absolute, 109/L 6.20 1.71 1.35–7.50 Lymphocyte absolute, 109/L 1.21 2.88 1.90–4.90 ESR, mm/h 140 9 0–15 CRP, mg/L 103 0.4 ≤3 mg/L Albumin, g/L 17 42.9 32–48 AST, U/L 373 33.5 10–45 ALT, U/L 114 24.9 10–35 Bilirubin, total, umol/L 6.3 4.9 0–21 Alkaline phosphate, u/L 269.5 246 100–300 HIV 1–2 antibody screening Non reactive Figure 1 Airspace disease involving the right middle and lower lobe with a cystic change involving the right upper lobe. Figure 2 Improvement of the airspace disease in the right middle and lower lobe after starting treatment. Bronchial wall thickening with peribronchial infiltrates in the perihilar region. Based on these findings, the patient was diagnosed as having pulmonary TB, iron deficiency anemia, and failure to thrive. Air borne isolation was initiated, and she was admitted to a negative air pressure room and started on INH, Rifampicin, pyrazinamide streptomycin, pyridoxine, and iron therapy. Four days after treatment, her platelet count decreased to 25 x 109/L. Rifampicin-induced thrombocytopenia was suspected, and the drug was therefore discontinued and replaced with ethambutol. Subsequently, the platelet count rebounded to a normal level.

Based on these findings, the patient was diagnosed as having pulmonary TB, iron deficiency anemia, and failure to thrive. Air borne isolation was initiated, and she was admitted to a negative air pressure room and started on INH, Rifampicin, pyrazinamide streptomycin, pyridoxine, and iron therapy. Four days after treatment, her platelet count decreased to 25 x 109/L. Rifampicin-induced thrombocytopenia was suspected, and the drug was therefore discontinued and replaced with ethambutol. Subsequently, the platelet count rebounded to a normal level. The Mantoux test was positive. Sputum for the AFB stain was positive, and the rapid DNA amplification test was positive for the M. tuberculosis complex. The culture was later reported to be positive for mycobacterium TB. The isolate was susceptible to streptomycin, isoniazid, rifampin, ethambutol, and pyrazinamide. The general condition of the patient improved on treatment, with a normalizing respiratory rate and other vital signs, but she experienced persistent episodes of low-grade fever. Her appetite improved, with good oral intake and increased weight. The baseline eye examination was normal, and follow ups were arranged. After 18 days of inpatient treatment, the patient was discharged, due in part to the insistence of the parents for an early discharge, on ethambutol, INH, Pyrazinamide, a Streptomycin IM injection once daily for 8 weeks (to be given at a local hospital), iron therapy, and pyridoxine with a follow-up OPD appointment.

d. After 18 days of inpatient treatment, the patient was discharged, due in part to the insistence of the parents for an early discharge, on ethambutol, INH, Pyrazinamide, a Streptomycin IM injection once daily for 8 weeks (to be given at a local hospital), iron therapy, and pyridoxine with a follow-up OPD appointment. On frequent repeated outpatient visits over the ensuing months, the response to treatment was suboptimal, as evidenced clinically and according to the investigation results. Poor compliance was suspected, and her antimicrobial serum levels were found to be undetectable. The local treating physician was contacted, and arrangements were made to initiate direct observation therapy (DOT); however, the family did not agree to have health care personnel in their home. This ultimately resulted in her clinical deterioration and a second admission. At this admission, there was a history of fever with chills for one month associated with decreased oral intake and activity. The parents noticed that she developed cervical and sternal ulcers, which were progressively worsening and discharging yellow, milky material.

On frequent repeated outpatient visits over the ensuing months, the response to treatment was suboptimal, as evidenced clinically and according to the investigation results. Poor compliance was suspected, and her antimicrobial serum levels were found to be undetectable. The local treating physician was contacted, and arrangements were made to initiate direct observation therapy (DOT); however, the family did not agree to have health care personnel in their home. This ultimately resulted in her clinical deterioration and a second admission. At this admission, there was a history of fever with chills for one month associated with decreased oral intake and activity. The parents noticed that she developed cervical and sternal ulcers, which were progressively worsening and discharging yellow, milky material. On examination, she was conscious, listless, unwell in appearance, pale, and dehydrated. Her vital signs were as follows: temperature: 38.4 °C, heart rate: 147 beats per minute, respiratory rate: 28 per minute, O2 saturation: 100% in room air, and blood pressure: 87/48 mmHg. A chest exam showed clear vesicular breathing, with no added sounds and decreased air entry, mainly on the right side. A cardiovascular exam showed normal S1 and S2, with no murmur. The abdomen was mildly distended with no organomegaly. She had multiple ulcerative skin lesions on the right side of the neck, over the sternum, and chest as well as bilateral axillary lymph nodes, which were ulcerating and discharging milky serous material. Her hemoglobin was 5 g/dl, indicating poor nutrition and poor compliance with all medications, including iron therapy.

e had multiple ulcerative skin lesions on the right side of the neck, over the sternum, and chest as well as bilateral axillary lymph nodes, which were ulcerating and discharging milky serous material. Her hemoglobin was 5 g/dl, indicating poor nutrition and poor compliance with all medications, including iron therapy. She was admitted with the impression of pulmonary and extra pulmonary (scrofuloderma) TB secondary to poor compliance with medical therapy in addition to social issues. The patient received packed RBCs. A review of the cultures and susceptibility results of the specimens from the sputum and lesions taken earlier showed that the organism had become first resistant to INH, then to ethionamide and streptomycin. The patient was therefore started on ethambutol, pyrazinamide, cycloserine, moxifloxacin, and pyridoxine (vitamin B6) to reduce the risk of the neurotoxicity of cycloserine and iron therapy. A specimen was submitted to a specialized center abroad, and PZA was tested by broth dilution using critical concentrations and CLSI interpretive criteria. All other agents were tested by the microbroth dilution with the (minimal inhibitory concentration) MIC reported and using laboratory developed interpretive criteria. The blood culture was negative. The wound culture was positive for Streptococcus pneumonia and for MRSA, which were treated accordingly. C-reactive protein was 239. A CT chest scan with contrast (Fig. 3) showed an interval increase in the number, size, and necrosis of the previously seen enlarged cervical and axillary lymph nodes, with abscess formation in addition to patchy pneumonic consolidations of the upper, middle, and right lower lobes. A bone scan showed mild increased activity in the left clavicle, likely related to a chronic infectious process. However, according to Pediatric Surgery, there was no role for surgical intervention.Figure 3 Improvement in the miliary nodules as well as the endobronchial tree-in-bud involvement of the right upper lobe and middle lobe after starting treatment.

ty in the left clavicle, likely related to a chronic infectious process. However, according to Pediatric Surgery, there was no role for surgical intervention.Figure 3 Improvement in the miliary nodules as well as the endobronchial tree-in-bud involvement of the right upper lobe and middle lobe after starting treatment. Figure 4 Chest CT: Large airspace consolidation involving the right middle and lower lobes with cavitary areas, a large cavity in the right upper lobe, bilateral miliary nodules and tree-in-bud appearance, with multiple mediastinal necrotic lymph nodes. The Child Advocacy Committee (CAC) was involved because of obvious parental neglect in the form of poor compliance with medications and refusal to cooperate with local as well as this hospital's medical staff. After several sessions with the parents and in coordination with the local Ministry of Health (MOH) authorities, an arrangement was made for admission to a local hospital where the child would receive her medications as an inpatient and then be followed up later with DOT therapy along with appointments at our hospital for a total of at least 9 months. On outpatient follow-up, the patient continued on ethambutol, pyrazinamide, cycloserine, moxifloxacin, pyridoxine, and iron therapy. She has markedly improved, with an absence of fever, weight gain, normal inflammatory markers, and clearing CXR (Fig. 4), Table 1.

The Child Advocacy Committee (CAC) was involved because of obvious parental neglect in the form of poor compliance with medications and refusal to cooperate with local as well as this hospital's medical staff. After several sessions with the parents and in coordination with the local Ministry of Health (MOH) authorities, an arrangement was made for admission to a local hospital where the child would receive her medications as an inpatient and then be followed up later with DOT therapy along with appointments at our hospital for a total of at least 9 months. On outpatient follow-up, the patient continued on ethambutol, pyrazinamide, cycloserine, moxifloxacin, pyridoxine, and iron therapy. She has markedly improved, with an absence of fever, weight gain, normal inflammatory markers, and clearing CXR (Fig. 4), Table 1. 3 Discussion TB remains a significant global health problem, with nearly 9 million new cases and 1.5 million deaths estimated annually [1], [2]. The estimated incidence of TB in Saudi Arabia is 14/100,000 [3]. Because of increased resistance to commonly used anti-TB drugs, management has become more complex and difficult, requiring the use of second-line drugs and, in some cases, surgical resection. Drug-resistant TB is defined as an infection caused by a strain of M. tuberculosis that is resistant to one of the first-line anti-TB drugs: isoniazid, rifampin, pyrazinamide, ethambutol, or streptomycin. Multidrug-resistant TB (MDR-TB) is defined as an infection caused by a strain of M. tuberculosis that is resistant to at least isoniazid and rifampin. Extensively drug-resistant TB (XDR-TB) is defined as an infection caused by a strain of M. tuberculosis that is resistant to at least isoniazid, rifampin, and fluoroquinolones, as well as either aminoglycosides (amikacin and kanamycin), capreomycin or both [4]. Totally drug-resistant TB (TDR-TB) is defined as an infection caused by a strain of M. tuberculosis that is resistant to all locally tested medications [5], [6]. Primary drug resistance is said to occur in a patient who has never received anti-TB therapy, and secondary drug resistance refers to the development of resistance during or following chemotherapy in patients who had previously had drug-susceptible TB.

losis that is resistant to all locally tested medications [5], [6]. Primary drug resistance is said to occur in a patient who has never received anti-TB therapy, and secondary drug resistance refers to the development of resistance during or following chemotherapy in patients who had previously had drug-susceptible TB. The first representative national survey conducted in Saudi Arabia showed that of 1904 patients, 79.3% had pulmonary and 20% had extra pulmonary TB, with lymph nodes being the most common site of infection (56.2%) [7]. Of the 1609 isolates from new cases of TB, 16.4% were resistant to at least one first-line drug and 1.8% were MDR. Of the 295 isolates from previously treated TB cases, 63.4% were resistant to at least one first-line drug and 15.9% were MDR. The prevalence of resistance to anti-tuberculosis agents was highest for INH followed by streptomycin and rifampicin. Regarding the geographical distribution of MDR-TB in the country, the highest prevalence was detected in the northern and southern provinces, followed by the western and central provinces, and the lowest proportion was reported in the eastern province. The treatment of patients with INH monoresistant TB has not been evaluated rigorously in randomized trials, and therefore, approaches are generally based on expert opinion derived from retrospective or single arm studies. In general, most patients with INH monoresistance should be treated with a rifamycin (rifampin or rifabutin), pyrazinamide, and ethambutol.

sistant TB has not been evaluated rigorously in randomized trials, and therefore, approaches are generally based on expert opinion derived from retrospective or single arm studies. In general, most patients with INH monoresistance should be treated with a rifamycin (rifampin or rifabutin), pyrazinamide, and ethambutol. The duration of therapy should be six to nine months (or four months after culture conversion) [8], [9], based on trials conducted by the Hong Kong Chest Service/British Medical Research Council, which demonstrated success rates of 95–98% among 107 patients with INH-resistant disease [10]. This finding was also supported by a retrospective study of patients with INH-monoresistant TB treated for six months, which showed comparable rates of failure or relapse among patients with drug-susceptible TB [9]. Data supporting the addition of a fluoroquinolone for the treatment of INH-monoresistant TB is inconsistent with one study of 328 patients with smear-positive TB who were randomized to receive either INH or moxifloxacin (in addition to rifampin, pyrazinamide, and ethambutol), in which culture negativity after eight weeks was comparable between the two groups (55 and 60%, respectively) [11]. On the other hand, a retrospective Danish study suggested that the inclusion of a fluoroquinolone in the treatment regimen was associated with a slightly higher success rate [12].

amide, and ethambutol), in which culture negativity after eight weeks was comparable between the two groups (55 and 60%, respectively) [11]. On the other hand, a retrospective Danish study suggested that the inclusion of a fluoroquinolone in the treatment regimen was associated with a slightly higher success rate [12]. As per the World Health Organization (WHO) recommendation, the treatment of patients for whom drug susceptibility testing is not available but are known to reside in a region with a background level of INH resistance > 7%, an acceptable approach consists of a standard initial phase (e.g., INH, rifampin, pyrazinamide, and ethambutol), followed by a continuation phase with INH and rifampin as well as the addition of ethambutol (rather than only INH and rifampin). Overall, effective therapy for INH-monoresistant TB is associated with very high bacteriologic, clinical response rates (>95%), and low relapse rates (<5%). National TB treatment guidelines strongly recommend using a patient-cantered case management approach, including directly observed therapy (“DOT”), when treating persons with active TB. DOT is especially critical for patients with drug-resistant TB, HIV-infected patients, and those on intermittent treatment regimens (i.e., 2 or 3 times weekly). DOT means that a trained health care worker or other designated individual (excluding a family member) provides the prescribed TB drugs and watches the patient swallow every dose to ensure that the medications are administered as directed. Because TB treatment entails several drugs taken regularly for several months, people from all social classes, educational backgrounds, ages, genders, and ethnicities may find it difficult to correctly adhere to the instructions. Studies show that 86–90% of patients receiving DOT complete therapy, compared to 61% for those on self-administered therapy. DOT helps patients finish TB therapy as quickly as possible, without unnecessary gaps, prevents TB from spreading to others, decreases the risk of drug-resistance resulting from erratic or incomplete treatment and decreases the chances of treatment failure and relapse [13], [14], [15], [16].

self-administered therapy. DOT helps patients finish TB therapy as quickly as possible, without unnecessary gaps, prevents TB from spreading to others, decreases the risk of drug-resistance resulting from erratic or incomplete treatment and decreases the chances of treatment failure and relapse [13], [14], [15], [16]. Serious reactions to antituberculosis drugs are uncommon. Rifampicin-induced thrombocytopenia is an uncommon but potentially life-threatening complication of antituberculosis treatment [17]. Rifampicin-induced thrombocytopenia was first reported by Blajchman and co-colleagues [18] in 1970. Most of the described cases were observed with high dose intermittent therapy (1200 mg twice weekly) [19]. Only a few cases of thrombocytopenia have occurred during daily treatment or after the administration of rifampicin following an interruption of therapy [18], [20]. The Tuberculosis Research Centre of Chennai reported only a single case of rifampicin-induced thrombocytopenia among more than over 8000 patients treated for tuberculosis over 30 years [21]. It has been observed that rifampicin-induced thrombocytopenia is caused by the presence of anti-rifampicin antibodies [20]. These antibodies fix a complement on the platelets in the presence of rifampicin, resulting in platelet destruction [21]. It has been found that antibodies against rifampicin are significant in number among patients who have stopped therapy; yet, rifampicin-induced thrombocytopenia is still relatively rare [18], [21]. The low incidence of thrombocytopenia due to daily doses of rifampicin has been attributed to the possible development of neutralizing antibodies formed during continuous treatment or may occur because the antigen–antibody complex is continuously removed without causing an allergic reaction [22]. Thus, daily doses of rifampicin may result in immunologic tolerance, whereas intermittent dosing favors sensitization [23]. It has been recommended that rifampicin-induced thrombocytopenia be an absolute contraindication to further therapy with rifampicin [19]. However, Bhasin and co-colleagues [24] suggested that re-challenges should be performed before withdrawing rifampicin. When it is necessary to re-start rifampicin, one should check the platelets counts frequently, under supervision and with the use of steroids.

contraindication to further therapy with rifampicin [19]. However, Bhasin and co-colleagues [24] suggested that re-challenges should be performed before withdrawing rifampicin. When it is necessary to re-start rifampicin, one should check the platelets counts frequently, under supervision and with the use of steroids. We present this case as an example of a child with complicated TB as illustrated by the initial poor clinical response to therapy caused by the emergence of resistance. On presentation, the organism cultured was sensitive to all first line agents, but because of poor compliance as documented by undetectable drug levels, the patient developed polydrug resistance. The improvement in the intake of medications and the resulting treatment success in this case was undoubtedly achieved by following the DOT method. Moreover, collaboration with the local physician and health care authorities was key in the implementation process. Confirmation of rifampicin-induced thrombocytopenia at the time of initial presentation is not often possible because tests for drug-dependent anti-platelet antibodies are not available in most laboratories. Discontinuation of the suspected drug leading to the resolution of thrombocytopenia provides strong evidence of rifampicin-induced thrombocytopenia. TB is nearly always curable if patients are treated with effective and uninterrupted antituberculous therapy. In children, parental support and an understanding of the importance of adherence to treatment as instructed is critical in bringing about cure, controlling the spread of infection, and minimizing the development of drug resistance. In the U.S.A., a Task Force composed of representatives of many federal agencies has developed a National Action Plan for addressing MDR-TB. The Task Force identified a number of objectives that must be met if MDR-TB is to be successfully combatted.

pread of infection, and minimizing the development of drug resistance. In the U.S.A., a Task Force composed of representatives of many federal agencies has developed a National Action Plan for addressing MDR-TB. The Task Force identified a number of objectives that must be met if MDR-TB is to be successfully combatted. These objectives fall under the categories of a) surveillance and epidemiology to determine the magnitude and nature of the problem; b) laboratory diagnostics to improve the rapidity, sensitivity, and reliability of diagnostic methods for MDR-TB; c) patient management to effectively managing patients who have MDR-TB and to prevent patients with drug-susceptible TB from developing drug-resistant disease; d) screening and preventive therapy to identify persons who are infected with or at risk of developing MDR-TB and to prevent them from developing clinically active TB; e) infection control to minimize the risk of transmission of MDR-TB to patients, workers, and others in institutional settings; f) outbreak control; g) program evaluation to ensure that TB programs are effective in managing patients and preventing MDR-TB; h) information dissemination/training and education; and i) research to provide new more effective tools with which to combat MDR-TB [25]. Conflict of interest Authors declare no competing interests. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction The term “genetic counseling” was first introduced in 1947 by Sheldon Reed [1]. The American Society of Human Genetics proposed a definition of genetic counseling in 1975, which was redefined after the professional society of genetic counselors known as “The National Society of Genetic Counselors” was incorporated in 1979. Genetic counseling is a “process of helping people understand and adapt to the medical, psychological and familial implications of genetic contributions to disease” [2]. The first genetic counseling program was established by Professor Melissa Richter in 1969 at Sarah Lawrence College, which is located in New York, in the United States [3]. At present, there are several genetic counseling Master's programs in the United States, Canada, Europe and South Africa.

ibutions to disease” [2]. The first genetic counseling program was established by Professor Melissa Richter in 1969 at Sarah Lawrence College, which is located in New York, in the United States [3]. At present, there are several genetic counseling Master's programs in the United States, Canada, Europe and South Africa. Training in genetic counseling was not available in the Middle Eastern countries until 2003, when the late Professor Ahmed Teebi was hired by King Faisal Specialist Hospital and Research Center (KFSHRC) as Head of the Department of Genetics. He championed the development of a training program in the kingdom and hired a Canadian genetic counselor, Shelley Kennedy, as supervisor of the program to develop its curriculum. One year later, with Professor Moeenaldeen Al-Sayed, as Medical Director, a diploma in genetic counseling was established at KFSHRC in Riyadh, Saudi Arabia [4]. Nine years later, Saudi Arabia witnessed the birth of a Master's program in genetic counseling. As an expansion of the successful development of this field in Saudi Arabia, the Master's program in genetic counseling was recognized and accredited by the Saudi Commission for Health Specialties in 2015. The inclusion of this Master's program in the Ministry of Higher Education in Saudi Arabia will provide genetic counselors throughout the Kingdom, which will alleviate the burden placed on non-geneticist health care providers who have little training in medical genetics. The availability of genetic counselors in many of the governmental hospitals in the Kingdom of Saudi Arabia will provide tremendous advantages to patients and to other health care providers. This article reviews the genetic counselor's scope of practice at KFSHRC, placing particular emphasis on the challenges encountered in our genetic counseling service.

s in many of the governmental hospitals in the Kingdom of Saudi Arabia will provide tremendous advantages to patients and to other health care providers. This article reviews the genetic counselor's scope of practice at KFSHRC, placing particular emphasis on the challenges encountered in our genetic counseling service. 1.1 Genetic counselors' scope of practice Genetic counselors are Master's trained health care professionals who combine their knowledge of basic science, medical genetics, epidemiological principles, counseling theory with their skills in genetic risk assessment, education, interpersonal communication and counseling to provide services to patients and their families for a diverse set of genetic or genomic indications. Some genetic counselors offer general genetic counseling, while others sub-specialize in a particular area of interest, such as cancer or prenatal care, assisted reproduction, cardiovascular health, research, public health and education.

o patients and their families for a diverse set of genetic or genomic indications. Some genetic counselors offer general genetic counseling, while others sub-specialize in a particular area of interest, such as cancer or prenatal care, assisted reproduction, cardiovascular health, research, public health and education. Genetic counselors see patients and their families for several reasons, including but not limited to, a family history of an inherited condition, a previous child with intellectual disability, multiple congenital anomalies or birth defects, repeated pregnancy loss or infertility, a positive newborn screening test, a newly diagnosed abnormality or genetic condition and to enroll patients in research studies. Moreover, they provide carrier testing, premarital genetic testing and counseling regarding preventative reproductive options, which include prenatal diagnosis (PND) and pre-implantation genetic diagnosis (PGD) for various underlying genetic defects and chromosomal abnormalities. 1.2 Genetic counseling clinics: the KFSHRC experience Our genetic counseling clinics serve patients from different provinces of Saudi Arabia, the gulf and Arab countries. We have six genetic counseling clinics per week. These clinics are separate from the genetic and metabolic clinics, which are operated by geneticists. The time allocated for each patient ranges from 25 min for a follow-up to 75 min for a new case. Some cases require multiple counseling sessions, whereas other cases require only one or two visits to the genetic counseling clinic.

s are separate from the genetic and metabolic clinics, which are operated by geneticists. The time allocated for each patient ranges from 25 min for a follow-up to 75 min for a new case. Some cases require multiple counseling sessions, whereas other cases require only one or two visits to the genetic counseling clinic. Cases are referred to our clinics from different specialties both within and outside of KFSHRC, such as medical genetics, high risk obstetric gynecology, cancer, dermatology, neurology, hematology, pediatrics and in vitro fertilization clinics. Referrals are primarily for extensive genetic counseling, molecular and cytogenetic testing, premarital screening, carrier testing and preventative reproductive options. Many families seek genetic counseling to understand the nature and consequences of genetic conditions, the risk of recurrence and preventative reproductive options and to address their uncertainty regarding genetics and inheritance. Our teams provide support for these families by identifying their concerns, addressing their needs, providing psychosocial counseling and promoting their decision-making process regarding testing or methods of prevention. 2 Challenges in genetic counseling services: the KFSHRC experience Genetic counseling often raises ethical and professional challenges. The most frequently encountered challenges among physicians in Western countries such as in Austria have been informed consent, organizational constraints, withholding information, and attaining/maintaining proficiency. [5].

ces: the KFSHRC experience Genetic counseling often raises ethical and professional challenges. The most frequently encountered challenges among physicians in Western countries such as in Austria have been informed consent, organizational constraints, withholding information, and attaining/maintaining proficiency. [5]. The major ethical principles that govern the attitudes of genetic counselors include respect for the patients' autonomy and right to make their own decisions, beneficence, i.e., taking actions to help others, non-maleficence, i.e., to do no harm, and justice, or administering services fairly between others [6]. There are several unique and difficult issues that are faced routinely in genetic counseling clinics in Saudi Arabia. Below is an overview of these issues based on several experiences at KFSHRC. 2.1 Consanguinity Consanguineous marriage is a common practice in Saudi Arabia. Consanguinity refers to the marriage of parents with a recent common ancestor [7]. The overall prevalence of consanguineous marriage in Saudi Arabia is 56% and is 33.6% between first-degree cousins [8]. When families with a positive family history for an autosomal recessive condition are seen for genetic counseling, the role of consanguinity in increasing the chances of having children with genetic disease is always discussed; however, most of the time these families continue practicing consanguineous marriage and have more affected children. In addition, it is not uncommon to see more than one genetic disease segregate with a family.

role of consanguinity in increasing the chances of having children with genetic disease is always discussed; however, most of the time these families continue practicing consanguineous marriage and have more affected children. In addition, it is not uncommon to see more than one genetic disease segregate with a family. In Saudi Arabia, the high rate of consanguinity may be attributed to different social and traditional factors, in addition to the desire to keep property within families. Main factors that inspire consanguinity include social and economic benefits and the stability of marriage between cousins. When the man and woman are raised in the same or a similar family environment, it is believed that they will adjust more easily to the marriage. In addition, marriage between relatives is considered to be beneficial because it maintains family fortunes within the same family structure. Anthropologists have long agreed that the primary achievement of consanguineous marriages is the inheritance of family structure and property [9], [10], [11]. Another cultural belief is in polygamy, which the husbands in some Saudi families believe is the practical solution to this situation.

the same family structure. Anthropologists have long agreed that the primary achievement of consanguineous marriages is the inheritance of family structure and property [9], [10], [11]. Another cultural belief is in polygamy, which the husbands in some Saudi families believe is the practical solution to this situation. 2.2 The evil eye The evil eye is a “folk belief elicited by the good luck of fortunate people whether in the form of material possessions including livestock, or possessing beauty, health, or offspring, may result in their misfortune” (Evil eye Encyclopedia). Some families relate their children's inherited disease to belief in the evil eye. The evil eye belief is widespread, according to which people can cause harm by merely an envious glance at desired objects or their owners [12]. The belief in the evil eye is ancient and present in every culture. The oldest reference appears in the cuneiform texts of the Sumerians, Babylonians and Assyrians around 3000 BC (Ancient History Encyclopedia). At that time, it was called “drishti” or “nazar” in India, and it was also called “nazar” in Turkey. Some people believe that the evil eye can place a curse on victims through a malevolent gaze of their magical eye. Others believe that envying others through the use of the evil eye can happen unintentionally. Furthermore, some cultures believe that the evil eye can suddenly bring bad luck by looking unintentionally at people who are unlucky enough to be cursed with the power. The evil eye was mentioned in the Quran in Sorat Al-Nisa verse: “or envy people for what God has given them of His bounty gave Ibrahim the Book and Wisdom and gave him the great king”. In Islam, Prophet Muhammad states: “The influence of an evil eye is a fact.” All Muslims believe in the evil eye because it is mentioned in the Quran. Some of the families at the genetic counseling clinics explained that they expected the disease to be caused by the evil eye. According to some health care providers, others held this belief but they did not share it.

e of an evil eye is a fact.” All Muslims believe in the evil eye because it is mentioned in the Quran. Some of the families at the genetic counseling clinics explained that they expected the disease to be caused by the evil eye. According to some health care providers, others held this belief but they did not share it. 2.3 Preventative reproductive options and termination of pregnancy Preventative reproductive strategies are urgently needed to combat the medical, financial and social burden of genetic diseases in Saudi Arabia. Preventative reproductive options are governed by religious rulings and/or political influence worldwide. While PGD and PND diagnoses are permitted in majority of countries worldwide, the practice of PGD is prohibited in Germany to protect embryos [13], [14].

ncial and social burden of genetic diseases in Saudi Arabia. Preventative reproductive options are governed by religious rulings and/or political influence worldwide. While PGD and PND diagnoses are permitted in majority of countries worldwide, the practice of PGD is prohibited in Germany to protect embryos [13], [14]. In the past few years, several studies have been conducted in Saudi Arabia to compare the Saudi preference for PGD versus PND. In a study conducted in 2006, 30 couples who had been treated at KFSHRC were surveyed regarding their attitudes towards preventative reproductive options. It was found that 27% of the couples preferred PGD compared to 13% who accepted PND. In addition, 10% were reluctant to choose any preventative options [15]. Similar findings were observed in a study conducted with a Lebanese population that examined their attitudes towards PGD as an alternative option to PND. The results indicated that 68% of the participating women accepted PGD [16]. The factors that contributed to the avoidance of PGD included receiving an IVF procedure, long waiting lists, fear of mixing embryos, the possibility of misdiagnosis and/or failure to conceive. However, the aforementioned studies showed an overall preference for PGD compared with PND primarily due to religious rulings that govern the termination of pregnancy in Arab countries.

ed receiving an IVF procedure, long waiting lists, fear of mixing embryos, the possibility of misdiagnosis and/or failure to conceive. However, the aforementioned studies showed an overall preference for PGD compared with PND primarily due to religious rulings that govern the termination of pregnancy in Arab countries. The potential benefits of PGD are not confined to preventing the occurrence of genetic disease; they expand to include advantages that are strongly valued by families. One advantage is the use of unaffected IVF-embryos for affected siblings who would benefit from hematopoietic stem cell transplantation for different genetic diseases, such as Fanconi Anemia and hemoglobinopathies [17]. Another benefit is the use of sex selection for medical reasons, e.g., in case of an affected child with X-linked conditions. This practice is permitted in Islam [18]. However, some families request sex selection in genetic counseling session because of social preferences, which is not accepted in Saudi Arabia. Similarly, sex selection for social reasons is prohibited in other countries, such as in Turkey [19].

child with X-linked conditions. This practice is permitted in Islam [18]. However, some families request sex selection in genetic counseling session because of social preferences, which is not accepted in Saudi Arabia. Similarly, sex selection for social reasons is prohibited in other countries, such as in Turkey [19]. Prenatal diagnoses are accepted in Islam before 120 days of conception if the fetus is grossly malformed with an untreatable severe condition and are based on the parent's request [20]. Cases that receive consensus by three geneticists can be terminated before 120 days; other patients that do not fulfill the criteria for termination of pregnancy are provided with a letter describing the medical aspects of the disease to seek support from scholars by getting a Fatwa. PND and termination of an affected pregnancy present a real challenge to genetic counselors, geneticists and maternal fetal medicine consultants, especially when the pregnant woman reports to the clinic late in her first trimester and/or with a lack of identifiable pathogenic mutations underlying the cause of the index case.

d termination of an affected pregnancy present a real challenge to genetic counselors, geneticists and maternal fetal medicine consultants, especially when the pregnant woman reports to the clinic late in her first trimester and/or with a lack of identifiable pathogenic mutations underlying the cause of the index case. A study conducted with 32 families with hemoglobinopathies to explore their attitudes towards PND and termination of pregnancy found that 81.3% of the families accepted PND. However, termination of pregnancy was viewed differently because of religious differences, and no other factors were identified to contribute to the families' decisions [21]. Another study included a survey of 200 parents of affected Saudi children and explored their views on termination of pregnancy for 30 genetic conditions. The results indicated that mothers were more interested in termination of pregnancy than fathers of the patients [22]. Overall, PND is widely accepted by the Saudi population, but termination of pregnancy remains a dilemma for some families. Pregnancy at the time of mutation analysis is also common in Saudi Arabia. At the genetic counseling clinic, we routinely meet couples and review the preventative options with them. However, couples often indicate that an unplanned pregnancy has occurred. A similar problem arose when families were referred to the PGD clinic; this indicates lack of proper use of contraception, either intentionally or unintentionally.

g clinic, we routinely meet couples and review the preventative options with them. However, couples often indicate that an unplanned pregnancy has occurred. A similar problem arose when families were referred to the PGD clinic; this indicates lack of proper use of contraception, either intentionally or unintentionally. Mitochondrial diseases due to mtDNA defects are a challenging category of genetic disease that present to genetic counseling for preventative reproductive options. The reasons these cases are difficult include social stigma for carrier mothers and technical limitations of PGD application. Tissue specificity, heteroplasmy, mutant load and bottleneck effect are barriers to performing PGD with no residual risk of recurrence for a concerned family [23]. The situation is different in Western countries such as in the UK, where parents of an affected child with an mtDNA defect have alternatives to PGD. These alternatives include oocyte donation and embryo manipulation to replace the defective mtDNA with unaffected mtDNA through different mitochondrial gene replacement techniques that have been approved ethically and legally (http://www.nuffieldbioethics.org/mitochondrial-dna-disorders) [24], [25]. Both approaches are prohibited in Islam [26], [27]. Accordingly, these families suffer physically and psychosocially due to their limited reproductive options in Saudi Arabia.

replacement techniques that have been approved ethically and legally (http://www.nuffieldbioethics.org/mitochondrial-dna-disorders) [24], [25]. Both approaches are prohibited in Islam [26], [27]. Accordingly, these families suffer physically and psychosocially due to their limited reproductive options in Saudi Arabia. 2.4 Manifesting carriers Our society is highly inbred, favoring first cousin marriages [28]. Therefore, most of our patients are affected with autosomal recessive conditions, in particular inborn errors of metabolism [29]. The carriers for autosomal recessive or X-linked conditions are typically asymptomatic. However, it is known that heterozygous individuals may manifest some signs of disease based on biochemical, radiological and clinical findings. Examples include hyperammonemia in case of carriers for X-linked conditions, ornithine transcarbamylase deficiency, calvarial thickening in heterozygote individuals with autosomal recessive disorders, Marshall syndrome reported by Khalifa et al [30] and phenotypic spectrums seen in heterozygotes for familial Mediterranean fever. The question that becomes apparent is whether we should provide management and treatment for the manifesting heterozygotes. This demonstrates the crucial role of genetic counselors in providing pre-test and post-test counseling to enable these individuals to understand the implications of genetic testing and to adapt physically and cope psychosocially to carrier test results. Without proper education and counseling, this could lead to alterations in parenting, increased anxiety, negative self-concept and stigmatization. In Western countries, additional burdens on manifesting carriers include health insurance and/or employment discrimination [31].

cope psychosocially to carrier test results. Without proper education and counseling, this could lead to alterations in parenting, increased anxiety, negative self-concept and stigmatization. In Western countries, additional burdens on manifesting carriers include health insurance and/or employment discrimination [31]. Carrier testing identifies heterozygote individuals with autosomal recessive and X-linked conditions; these findings indicate additional family members who are at-risk for manifesting signs of the disease and/or having affected children, which elicits a complex counseling session. Ataxia telangiectasia (AT) is an autosomal recessive condition affecting 1:40,000–1:100,000 live births in the United States. Carriers of AT are at a four-fold increased risk of developing cancer and heart disease in comparison to general population [32]. AT carriers are often hypersensitive to radiation and radiomimetic drugs. Genotype–phenotype correlation may provide insight into at-risk carriers' predisposition to cancer. However, the lack of reported correlation between a novel genetic alteration and the risks mentioned above complicates genetic counseling. This ultimately affects individual's ability to make an informed decision regarding management. Urgent issues such as parental surveillance, referral to other specialties and carrier testing for unaffected offspring emerge and challenge the entire genetic counseling process.

tioned above complicates genetic counseling. This ultimately affects individual's ability to make an informed decision regarding management. Urgent issues such as parental surveillance, referral to other specialties and carrier testing for unaffected offspring emerge and challenge the entire genetic counseling process. 2.5 Emotional and social impact on carrier women The emotional and social impact on parents with X-linked conditions differs from that of parents with recessive diseases. In a study conducted by James et al [33], they found that carrier mothers of X-linked conditions experienced more guilt, anger and self-blame while parents of children with autosomal recessive conditions worried more about reproductive risk. The social burden on female carriers in form of stigmatization is no longer a phenomenon that is uniquely related to X-linked conditions or mitochondrial disease due to maternal inheritance, as previously believed. Our clinical experience expands that list to include carrier mothers for autosomal recessive conditions. We believe that the stigmatization of carrier mothers of autosomal recessive conditions in our community is determined by the lack of adequate knowledge regarding the inheritance and etiology of the disease, the education level of the parents and social influence.

ude carrier mothers for autosomal recessive conditions. We believe that the stigmatization of carrier mothers of autosomal recessive conditions in our community is determined by the lack of adequate knowledge regarding the inheritance and etiology of the disease, the education level of the parents and social influence. The diagnosis of carrier status in mothers of affected boys with X-linked conditions has been found to affect their relationship with their daughters. Some mothers have reported a feeling of closeness, which enabled them to talk openly with their daughters, while others found that the results of the carrier test drove them apart [34]. In our population, the stigmatization that occurs when mothers receive a positive carrier test result may, in their opinion, justify mothers' choice to not inform at-risk daughters about the availability of genetic testing. Mothers who experience stigma from their husbands and relatives show social isolation and tend to hide genetic test information from their daughters. These mothers clearly indicate that the underlying reason for their behavior is fear for their daughter's future (Personal communications).

ailability of genetic testing. Mothers who experience stigma from their husbands and relatives show social isolation and tend to hide genetic test information from their daughters. These mothers clearly indicate that the underlying reason for their behavior is fear for their daughter's future (Personal communications). One of the dilemmas we face in genetic counseling clinic is requesting carrier testing for the fiancé of a carrier female. In our clinical practice, the father of a carrier female brings the fiancé and requests us to perform carrier testing on him without providing information about the genetic disease in question. Nevertheless, some fiancés attend genetic counseling clinics unaware about the genetic condition in the female's family. This imposes a great challenge on the genetic counselors, who must value the ethical principles in practice for all of the involved parties while considering the burden placed on the woman's family, the confidentiality of the woman's test result and the right of the fiancé to be fully counseled before having the genetic test performed.

eat challenge on the genetic counselors, who must value the ethical principles in practice for all of the involved parties while considering the burden placed on the woman's family, the confidentiality of the woman's test result and the right of the fiancé to be fully counseled before having the genetic test performed. 2.6 Complex molecular test findings Advances in molecular testing have benefited families because it resolves the ambiguity associated with undiagnosed cases. It has simultaneously provided opportunities to extended family members to pursue premarital screening and to parents to pursue preventative reproductive options. In spite of these perceived benefits, advances in genetic testing will continue to become more complex [35]. In particular, exome sequencing presents challenges to genetic counselors. Examples of issues encountered in our routine clinical practice include the identification of variants of unknown clinical significance, which limits our ability to offer PND and termination of pregnancy, and the identification of pathogenic mutations in multiple genes causing a number of genetic conditions in a family. This certainly warrants careful and thorough genetic counseling. The major dilemma in these findings is related to PGD, as only two mutations are tested using the PGD approach. This places a large burden on geneticist to carefully review each case with the family and choose two candidate diseases for PGD based on several parameters, such as a disease severity and prognostic outcome. In such cases, PND would be offered to detect other causative mutations that would not be excluded by PGD, in the event that the pregnancy continued after PGD. The decision-making process for the parents becomes increasingly difficult as other issues arise, e.g., identifying at-risk family members and cascade genetic testing. Another issue encountered in the clinic was receiving negative results of exome sequencing, which required further consideration of reanalyzing the patient's sample with a platform characterized by in-depth coverage, homozygosity mapping if multiple affected family members are available or the use of another alternative molecular approach. Finally, incidental findings and ethical challenges of whether to report such findings were another challenge faced.

yzing the patient's sample with a platform characterized by in-depth coverage, homozygosity mapping if multiple affected family members are available or the use of another alternative molecular approach. Finally, incidental findings and ethical challenges of whether to report such findings were another challenge faced. The application of guidelines developed by The American College of Medical Genetics and Genomics (ACMG) helps address this dilemma and emphasizes the importance of pre-test genetic counseling in the era of exome sequencing [36]. It becomes apparent that the results from advanced molecular technologies in the field of medical genetics provide additional complexity to the already complex family situation and reproductive history. 3 Conclusion Marriage customs in Saudi Arabia present challenges to genetic counselors, as multiple diseases can segregate within a single family. This practice warrants cascade genetic testing for extended family members and raises ethical challenges. Genetic counseling becomes a very complex process in view of the available advanced molecular techniques and a lack of clinical significance of the reported variants. The profession of genetic counseling in the kingdom will continue to evolve to incorporate growing complexity due to molecular advances while addressing the unique social and cultural norms of the region. Conflict of interest None declared. Ethical approval There is no need for ethical approval for this review. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Breast lumps are uncommon in children. The most common type of breast mass found in the adolescent population is a fibroadenoma. Phyllodes tumors are rare fibroepithelial tumors that account for 0.3–0.5% of all breast tumors in females. They are rarely observed in adolescents, with only 20 cases reported [1]. Here, we report a case of an adolescent with a lump in her left breast which the histopathology revealed to be a benign phyllodes tumor. 2 Case summary An 11-year-old pre-menarcheal girl presented with a lump in her left breast that was gradually increasing in size. Local examination revealed a 15 × 15 cm, well-circumscribed, firm, and freely mobile lump occupying the entire left breast (Fig. 1). The right breast was normal, and there was no axillary lymphadenopathy. Ultrasound revealed a 9 × 5 × 6 cm homogenous, hypoechoic, and encapsulated lesion with minimal vascularity and no calcification (Fig. 2). Cytology was suggestive of a fibroadenoma. The patient underwent an excision biopsy using an inframammary incision. A 9 × 9 cm encapsulated lump weighing 250 gm with smooth surfaces was excised. The histopathologic analysis revealed that it was a benign phyllodes tumor with margins reaching the inked surface (Fig. 3).Figure 1 Clinical photograph of the patient showing the left breast lump. Figure 2 Ultrasonographic imaging of the left breast lump. Figure 3 Histopathological analysis of the specimen –Hematoxylin and Eosin staining showing the biphasic tumor and a magnified histopathological image showing nuclear pleomorphism and mitotic figures.

2 Case summary An 11-year-old pre-menarcheal girl presented with a lump in her left breast that was gradually increasing in size. Local examination revealed a 15 × 15 cm, well-circumscribed, firm, and freely mobile lump occupying the entire left breast (Fig. 1). The right breast was normal, and there was no axillary lymphadenopathy. Ultrasound revealed a 9 × 5 × 6 cm homogenous, hypoechoic, and encapsulated lesion with minimal vascularity and no calcification (Fig. 2). Cytology was suggestive of a fibroadenoma. The patient underwent an excision biopsy using an inframammary incision. A 9 × 9 cm encapsulated lump weighing 250 gm with smooth surfaces was excised. The histopathologic analysis revealed that it was a benign phyllodes tumor with margins reaching the inked surface (Fig. 3).Figure 1 Clinical photograph of the patient showing the left breast lump. Figure 2 Ultrasonographic imaging of the left breast lump. Figure 3 Histopathological analysis of the specimen –Hematoxylin and Eosin staining showing the biphasic tumor and a magnified histopathological image showing nuclear pleomorphism and mitotic figures. 3 Discussion The evaluation of an adolescent presenting with a breast mass differs substantially from that of an adult because of marked differences in breast cancer risk and breast architecture. There is less emphasis on exclusion of malignancy, as pediatric breast masses are typically benign (95% benign fibroadenomas) [2].

ussion The evaluation of an adolescent presenting with a breast mass differs substantially from that of an adult because of marked differences in breast cancer risk and breast architecture. There is less emphasis on exclusion of malignancy, as pediatric breast masses are typically benign (95% benign fibroadenomas) [2]. Management of pediatric breast masses is primarily conservative. Clinical observation over two to four months is appropriate. Masses that increase by more than 1 cm and those larger than 2 cm warrant ultrasonographic percutaneous biopsies to confirm the benign nature [2]. According to Stanford University, the following criteria for juvenile fibroadenoma are used: 1] circumscribed and rarely multiple; 2] biphasic stromal and epithelial process lacking a leaf-like growth pattern in a uniformly hypercellular stroma; 3] a lack of atypical features and a stroma-like periductal increase in cellularity, stromal overgrowth, and cytologic atypia, as well as a mitotic rate <3/hpf; 4] frequent epithelial and myoepithelial hyperplasia; and 5] an age of between 10 and 20 years [3]. Giant fibroadenomas are defined as tumors >500 g. Phyllodes tumors are rare fibroepithelial tumors that account for 0.3–0.5% of female breast tumors, the peak of which occurs in women between the ages of 45 and 49 years [4]. This type of tumor is rarely found in adolescents. Only about 20 cases have been reported in children [1].

adenomas are defined as tumors >500 g. Phyllodes tumors are rare fibroepithelial tumors that account for 0.3–0.5% of female breast tumors, the peak of which occurs in women between the ages of 45 and 49 years [4]. This type of tumor is rarely found in adolescents. Only about 20 cases have been reported in children [1]. A large breast lump with history of rapidly increasing size and ultrasound features suggestive of a fibroadenoma (except a size > 2 cm) should arouse high suspicion of a phyllodes tumor [5]. Axillary node involvement is rare. Another characteristic feature of these tumors is a high rate of local recurrences (5–20%) [6]. Fibroadenomas and phyllodes tumors share many common features. Clinically, both present as rounded, circumscribed, and moveable masses. Histologically, both can be grouped as “fibroepithelial lesions”. Preoperative diagnosis poses a diagnostic difficulty, as fine needle aspiration cytology and core needle biopsy may not be able to distinguish a phyllodes tumor from a fibroadenoma [5].

ically, both present as rounded, circumscribed, and moveable masses. Histologically, both can be grouped as “fibroepithelial lesions”. Preoperative diagnosis poses a diagnostic difficulty, as fine needle aspiration cytology and core needle biopsy may not be able to distinguish a phyllodes tumor from a fibroadenoma [5]. Microscopically, phyllodes tumors are characterized by a double-layered epithelial component arrayed in clefts and surrounded by a hypercellular stromal mesenchymal component. The stroma often protrudes into the epithelial lining spaces, forming a slit-like space or a leaf-like pattern; hence, the name phyllodes, which means “leaf-like” in Greek. The morphologic features that have to be accounted for are the following: 1) the degree of stromal hypercellularity, 2) stromal overgrowth, 3) nuclear atypia, 4) number of mitoses, 5) amount of stroma relative to epithelium, and 6) infiltrative tumor borders. However, the natural history of these tumors is often different, and the differences are tabulated in Table 1. Phyllodes tumors tend to grow more rapidly and may recur if incompletely excised. Moreover, phyllodes tumors may metastasize. In contrast, fibroadenomas usually do not need to be removed, and even when surgery is needed, enucleation is sufficient.Table 1 Differences between fibroadenoma and Phyllodes tumors.

n Table 1. Phyllodes tumors tend to grow more rapidly and may recur if incompletely excised. Moreover, phyllodes tumors may metastasize. In contrast, fibroadenomas usually do not need to be removed, and even when surgery is needed, enucleation is sufficient.Table 1 Differences between fibroadenoma and Phyllodes tumors. Fibroadenoma Phyllodes tumor History Long duration and slow growing Short duration and rapidly growing Histopathologic characteristics Duct-like spaces surrounded by fibrous stroma Characteristic ‘leaf-like pattern’ Mitotic figures Not observed Present Stromal overgrowth and stromal infiltration Not observed Present Surgical management Enucleation Wide local excision Local recurrence Not known Common Metastasis Not known May be observed in malignant phyllodes Excisional biopsy is required in the majority of cases of suspected phyllodes tumors. The World Health Organization (WHO) classifies them as either benign, borderline, or malignant based on histopathological features [7] (Table 2).Table 2 WHO classification of phyllodes tumors. Margin Stromal atypia Mitoses/10 HPF Stromal overgrowth Benign Pushing Minimal <5 Absent Borderline Pushing/infiltrating Moderate <10 Present Malignant Infiltrating Severe >/=10 Present The benign variant is most common, with only 10–25% of cases being malignant. The rate of distant metastasis for the malignant tumors is 15–25% [6].

Margin Stromal atypia Mitoses/10 HPF Stromal overgrowth Benign Pushing Minimal <5 Absent Borderline Pushing/infiltrating Moderate <10 Present Malignant Infiltrating Severe >/=10 Present The benign variant is most common, with only 10–25% of cases being malignant. The rate of distant metastasis for the malignant tumors is 15–25% [6]. Phyllodes tumors are managed by wide local excision. In cases of large lumps, a mastectomy may be necessary. A recent study by Yom et al concluded that a clear margin of 0.1 mm is equivalent to a margin of 1 cm [8]. Due to the rarity of the condition in younger age groups, an individualized, case-based approach, and regular follow up are advisable. As per the National Comprehensive Cancer Network (NCCN) guidelines, in cases with local recurrence, resection with wide, tumor-free surgical margins should be performed. Adjuvant therapy has no proven effect. In cases of systemic metastasis, treatment is based on the soft tissue sarcoma protocol [9]. Patients may experience postoperative cosmetic deformity or secondary asymmetry. However, reconstructive surgery is usually not considered until at least one year after the procedure and after the patient has reached skeletal maturity. Additionally, the breast parenchyma may expand to fill the resulting defect and resolve any deformities over the course of development. Surgeons should discuss all potential outcomes, expectations of aesthetic results, and the need for reconstructive surgery with patients prior to surgery and at subsequent follow-up appointments [10].

e breast parenchyma may expand to fill the resulting defect and resolve any deformities over the course of development. Surgeons should discuss all potential outcomes, expectations of aesthetic results, and the need for reconstructive surgery with patients prior to surgery and at subsequent follow-up appointments [10]. 4 Conclusion When an adolescent presents with a large breast lump, the possibility of a phyllodes tumor, though a rare differential, should be considered. A case-based, individualized approach is recommended, as there are no set protocols. Due to the extreme rarity of this tumor and because of the frequently benign nature of tumors in this age group, a more conservative approach with regular follow up is advisable. This prevents cosmetic and psychological distress in young adolescent girls. Conflict of interest The authors have no conflict of interest to report. Source of funding No funding was obtained to conduct the study. Acknowledgment Department of Histopathology, Tata Memorial Hospital, Mumbai, Maharashtra, in India. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

A 10-year-old boy was diagnosed with Down syndrome, and he had a duodenal atresia repaired when he was 10 days old. He presented with a history of jaundice and pale stools for the past 2 months. Laboratory studies yielded the following results: Alanine Aminotransferase 71.8 U/L, Aspartate Aminotransferase 81.3 U/L, Alkaline Phosphatase 1367 U/L, Gamma-Glutamyl transpeptidase 588 IU/L, Amylase 64 U/L, and Lipase 24 IU/L. Ultrasound of the abdomen exhibited a shadowing calculus in the distal Common bile duct (CBD) in the region of the head of the pancreas measuring 1 cm and causing intra- and extrahepatic biliary duct dilatation. The maximum CBD diameter was 1 cm with the intrahepatic duct measuring 0.3 cm (see Fig. 1).Figure 1 Abdominal US showing a distal CBD stone. The patient underwent Endoscopic Retrograde Cholangiopancreatography (ERCP) under general anesthesia. It was highly difficult to locate the ampulla of Vater endoscopically. However, after multiple attempts, a stone measuring approximately 1 cm was extracted via endoscopic sphincterotomy. No stent was inserted. Post-ERCP, the patient started to complain of mild abdominal pain that was not aggravated by food intake. His serum amylase and lipase levels remained normal. An abdominal X-ray is shown in Fig. 2 (Fig. 3).Figure 2 Abdominal X-rays: A: Standing position, B: Left lateral decubitus position. Figure 3 Retroperitoneal air. ↑: sub-diaphragmatic air did not move between standing and the lateral decubitus positions, A & B. →: retroperitoneal air outlining the right kidney. For the answer: visit: www.elsevier.com/locate/ijpam.

Post-ERCP, the patient started to complain of mild abdominal pain that was not aggravated by food intake. His serum amylase and lipase levels remained normal. An abdominal X-ray is shown in Fig. 2 (Fig. 3).Figure 2 Abdominal X-rays: A: Standing position, B: Left lateral decubitus position. Figure 3 Retroperitoneal air. ↑: sub-diaphragmatic air did not move between standing and the lateral decubitus positions, A & B. →: retroperitoneal air outlining the right kidney. For the answer: visit: www.elsevier.com/locate/ijpam. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Motor vehicle accidents are the leading cause of death in adolescents and young adults worldwide. According to the Centers for Disease Control and Prevention [1], unintentional injuries related to motor vehicle accidents are consistently the leading cause of mortality from ages 5 through 24 in the United States. Road traffic accidents (RTAs) are a major public health concern worldwide that have resulted in nearly 1.24 million deaths and 20–50 million non-fatal injuries. RTAs are also ranked as the ninth-highest cause of disability-adjusted life years (DALY) lost, and they are estimated to rise by one-third by 2020 [2]. According to the 2013 World Health Organization (WHO) report, there are only 19 countries in the world with higher per capita road traffic death rates than the Kingdom of Saudi Arabia (KSA), where nearly one person is killed and four are injured every hour on average [2]. Excessive speed and disobeying traffic signals cause over 65% of traffic accidents, which result in considerable economic losses to victims, their families, and to nations as a whole [3]. RTAs are the second-greatest public health concern in the Kingdom, as they contribute to emergency and outpatient hospital admissions and are responsible for a yearly estimated loss of 500 million U.S. dollars [4]. In a recent review article, Mansuri et al [5] reported that over the past 25 years, RTAs accounted for 83.4% of all trauma admissions. Nearly 80% of total accidents are primarily due to speeding and non-compliance with right-of-way rules by drivers and pedestrians [6].

ly estimated loss of 500 million U.S. dollars [4]. In a recent review article, Mansuri et al [5] reported that over the past 25 years, RTAs accounted for 83.4% of all trauma admissions. Nearly 80% of total accidents are primarily due to speeding and non-compliance with right-of-way rules by drivers and pedestrians [6]. According to WHO [7], Saudi Arabia has the world's highest number of deaths from RTAs per capita. Road traffic accidents are now the primary cause of death, injury, and disability in adult males aged 16–36 years in the Kingdom and result in significant health care costs. For example, the cost of treating people injured due to road traffic crashes during 2002 was estimated at 652.5 million Saudi riyals (US$ 174 million). According to recent estimates, the traffic accident death rate (age adjusted) in Saudi Arabia is 23.2 per 100,000 [8]. According to the Director of the General Directorate of Traffic, there were more than 544,000 accidents in Saudi Arabia during 2011, resulting in more than 39,000 injuries and 7153 deaths. Three-fourths of these accidents involved young people. Moreover, RTA losses amounted to more than 80% of the deaths and 7 billion in economic losses every year [9]. On average, 17 Saudis, primarily male, die on the roads each day. Furthermore, road traffic victims occupy one-fifth of hospital beds, with the majority of victims requiring long-term rehabilitation [10].

ople. Moreover, RTA losses amounted to more than 80% of the deaths and 7 billion in economic losses every year [9]. On average, 17 Saudis, primarily male, die on the roads each day. Furthermore, road traffic victims occupy one-fifth of hospital beds, with the majority of victims requiring long-term rehabilitation [10]. The number of road deaths per year has risen during recent years to a rate equal to an average of 19 deaths per day [5]. The most common human factors contributing towards traffic accidents include speeding (in 65% of accidents), driver error (in 80% of accidents), violation of traffic signals at intersections (in 50% of accidents), and illegal U-turns. Other causes are related to vehicles, the road, and the environment (e.g., road layout, which contributes to 20% of accidents) [11]. Excessive speeding was the most common cause reported in all recent and past studies. Driver error was identified as the main contributing factor in about two-thirds of all RTAs, mainly characterized as reckless driving and excess speeding [5], [12].

ment (e.g., road layout, which contributes to 20% of accidents) [11]. Excessive speeding was the most common cause reported in all recent and past studies. Driver error was identified as the main contributing factor in about two-thirds of all RTAs, mainly characterized as reckless driving and excess speeding [5], [12]. Attitudes and behaviors in general are founded very early in life and are crucial for safe driving behavior. Adult role models and certain cultural factors may promote risky behaviors that contribute to unintentional injuries that result in premature death or lifelong impairment. Many young men in the Arab world treat driving like a hobby or a sport. It is not unusual to see teenage boys participate in “Saudi drifting” with a crowd of spectators, for example. The major objective of the paper is to examine attitudes and behavior that encourage risky driving called “drifting” among Saudi youth. The capital, and the actual birthplace of the “Arab Drift” (called “Tafheet” or “Hajwalah” in Arabic) is Riyadh. The combination of good roadways and fine desert sand give the drifter the sensation of driving on water rather than asphalt. Such thrill-seeking behavior is very alluring to young Saudi males. Many of them adopt “Tafheet” as their leisure-time activity, usually during weekends and late nights. Tafheet is an illegal street racing phenomenon that emerged in the late 1970s that involves trying to “drift” a motor vehicle at speeds of up to 260 km/h (160 mph) across wide highways. Racers often drive dangerously close to traffic at high speed or slide around on a wide flat straight road section at high speed, drifting sideways and ignoring road barriers. This activity often attracts spectators, who watch from the roadside without any protection. Tafheet practices and events occur with little to no concern for the safety of vehicle occupants, other drivers or spectators. Tafheet events are commonly seen on the wide-sectioned highways of Riyadh, Al-Qassim Province, and less notably in other parts of Saudi Arabia [12]. Because they occur in a secretive manner, there seems to be no police records or documentation by highway authorities regarding these events.

other drivers or spectators. Tafheet events are commonly seen on the wide-sectioned highways of Riyadh, Al-Qassim Province, and less notably in other parts of Saudi Arabia [12]. Because they occur in a secretive manner, there seems to be no police records or documentation by highway authorities regarding these events. To date, no research studies have investigated youth attitudes favoring risky driving behaviors (pro-attitudes), including “Tafheet”, that are undoubtedly associated with unintentional injuries. Our study is the first of its kind to address this problem among school children using general population school samples. The main objective of the paper is to investigate youth risk behaviors related to driving, perceptions, and attitudes concerning driving behavior, including “Tafheet”, in Saudi Arabia. We expect that behaviors and/or attitudes favoring thrill seeking and risk taking are associated with participation in this dangerous sport.

ain objective of the paper is to investigate youth risk behaviors related to driving, perceptions, and attitudes concerning driving behavior, including “Tafheet”, in Saudi Arabia. We expect that behaviors and/or attitudes favoring thrill seeking and risk taking are associated with participation in this dangerous sport. 2 Subjects and Methods The study was conducted in high schools in Riyadh, the capital and the largest city in Saudi Arabia. The study employed a multistage probability sampling scheme. Schools were selected based on geographical location and type of institution. Riyadh was divided into three geographical areas, North, South, and Middle for sampling purposes. In each geographical area, the two largest public schools, one private school, and one international school were selected. Public schools were single-gender institutions, while private and international schools were coeducational. Approximately 60 classes with a free period were selected at random. Each class had 25–40 students per classroom on average. All students in the selected classrooms were eligible to participate in the study.

ted. Public schools were single-gender institutions, while private and international schools were coeducational. Approximately 60 classes with a free period were selected at random. Each class had 25–40 students per classroom on average. All students in the selected classrooms were eligible to participate in the study. Prior to administering the survey, a committee visited each school to explain the purpose of the study to students, informational letters, and consent forms were sent home to parents. Students were informed that participation in the study was completely voluntary. Four research teams composed of health professionals (e.g., nurses, health coordinators) collected the data from November 2010 to February 2012. Ethical approval for the study was obtained from King Fahad Medical City Institutional Review Board, along with permission from the Ministry of Education in Riyadh to administer the survey in schools. 2.1 Survey development and validity A comprehensive adolescent health survey was developed for Saudi youth based on an extensive literature review of adolescent health, along with questions adopted from the Youth Risk Behavior Survey used by the United States Centers for Disease Control and Prevention (CDC). The Youth Risk Behavior Survey has been validated over the past several years in the USA. Using these standard questions helps confer high face validity and credibility to our survey. Questions regarding family and school connectedness were also developed by consulting experts in adolescent health to achieve further face validity.

sk Behavior Survey has been validated over the past several years in the USA. Using these standard questions helps confer high face validity and credibility to our survey. Questions regarding family and school connectedness were also developed by consulting experts in adolescent health to achieve further face validity. The survey inquired about adolescents' behaviors and attitudes and included questions on smoking behavior, perceptions of school and family, use of technology, school activities, leisure time activities, and driving. The survey items were translated to Arabic and then were back-translated to English by bilingual professionals. The survey was administered in the language preferred by the student. A total of 1668 students participated in the survey. The current analysis included only boys, (N = 799) as females are prohibited from driving. Accordingly, questions pertaining to driving are not relevant for female students. 2.2 Measures Outcome: In this study, we measured Tafheet as a proxy for risky driving behavior by asking “Have you ever engaged in “Tafheet”? with response choices “yes” and “no”. 2.2.1 Independent predictors Personality factor: Those who responded either Strongly Agree or Agree to the question, “even if dangerous, I like to do exciting things”, were grouped in the “yes” category for this predictor. Those who responded “Disagree” or “Strongly Disagree” were grouped as “no”.

2.2 Measures Outcome: In this study, we measured Tafheet as a proxy for risky driving behavior by asking “Have you ever engaged in “Tafheet”? with response choices “yes” and “no”. 2.2.1 Independent predictors Personality factor: Those who responded either Strongly Agree or Agree to the question, “even if dangerous, I like to do exciting things”, were grouped in the “yes” category for this predictor. Those who responded “Disagree” or “Strongly Disagree” were grouped as “no”. Thoughts about “Tafheet”: Two questions were used to assess students' thoughts:(1) You think that “Tafheet” is a sort of a) Talent, b) Sport, c) Violence, d) Delinquency, or e) Cool activity (2) You think that “Tafheet” should be a) Prohibited, b) Supported by doing special clubs, c) Supported by making yearly tournament, or d) Supported by training and by providing the needed safety equipment.

Thoughts about “Tafheet”: Two questions were used to assess students' thoughts:(1) You think that “Tafheet” is a sort of a) Talent, b) Sport, c) Violence, d) Delinquency, or e) Cool activity (2) You think that “Tafheet” should be a) Prohibited, b) Supported by doing special clubs, c) Supported by making yearly tournament, or d) Supported by training and by providing the needed safety equipment. Attitudes toward unintentional injury behaviors: Six questions were asked to measure students' attitudes. These questions assessed whether the teen thinks there is the potential for harm to himself or others (physically or otherwise) across different driving scenarios. The available responses are Always, Occasionally/rare, and Never: (1) Would there be harm for yourself or others (physically or otherwise) if you drive faster than the speed limit? (2) Would there be harm for yourself or others (physically or otherwise) if you drive the car without having a driving license? (3) Would there be harm for yourself or others (physically or otherwise) if you drive/ride the car without wearing a seat belt? (4) How often do you use your mobile phone while driving? (5) How much do you respect the traffic laws? (6) How often do you use a helmet when you rollerblade, skateboard, or ride a motorcycle? Demographic variables: Included in the analyses were the type of school (Government, Private, and International), grade (7th–12th), which was also used as proxy for age, and nationality (Saudi, Arab, and non-Saudi)

Attitudes toward unintentional injury behaviors: Six questions were asked to measure students' attitudes. These questions assessed whether the teen thinks there is the potential for harm to himself or others (physically or otherwise) across different driving scenarios. The available responses are Always, Occasionally/rare, and Never: (1) Would there be harm for yourself or others (physically or otherwise) if you drive faster than the speed limit? (2) Would there be harm for yourself or others (physically or otherwise) if you drive the car without having a driving license? (3) Would there be harm for yourself or others (physically or otherwise) if you drive/ride the car without wearing a seat belt? (4) How often do you use your mobile phone while driving? (5) How much do you respect the traffic laws? (6) How often do you use a helmet when you rollerblade, skateboard, or ride a motorcycle? Demographic variables: Included in the analyses were the type of school (Government, Private, and International), grade (7th–12th), which was also used as proxy for age, and nationality (Saudi, Arab, and non-Saudi) 2.3 Data analysis Univariate descriptive statistics were examined, and variables were regrouped as suggested by the distribution. Descriptive statistics were calculated and expressed either as the means and standard deviations or as percentages. To test for bivariate associations between the risk behavior (Tafheet) and the predictors, cross tabulation with chi-squared (χ2) analysis was conducted for categorical variables, and T-tests for continuous variables were used to test the group mean differences. Alpha was set at P < .05 for results to be considered statistically significant. For multivariable analysis, a logistic regression model was used, including variables that showed significant bivariate association with the risky behavior Tafheet. Adjusted odds ratios (OR) and 95% confidence intervals (CI) are presented. The missing data due to nonresponses or choice of a “don't know” option were set as missing, which resulted in missing data rates of 10–15% for some variables In the multivariate analysis. All analyses were conducted using SPSS Statistics (Version 22.0) software.

os (OR) and 95% confidence intervals (CI) are presented. The missing data due to nonresponses or choice of a “don't know” option were set as missing, which resulted in missing data rates of 10–15% for some variables In the multivariate analysis. All analyses were conducted using SPSS Statistics (Version 22.0) software. 3 Results The current analyses included 799 male students (7th through 12th grade) from governmental (34%), private (45%), and international schools (21%) in Riyadh. Almost all of them are of Islamic faith (99%). Nearly two-thirds of the participants were Saudi (71%), with 23% from other Arab nations and approximately 6% from other cultures. 3.1 Prevalence of attitudes favoring risky driving We tested for grade differences in attitudes favoring risky driving behavior. As noted in Table 1, significant grade differences are present across all four attitudes that may promote risky driving. In general, we observed a trend of increasing risk and rising grade in terms of pro-attitudes. This includes those who responded “never”, “rarely” or “occasionally” to questions inquiring whether the teen thinks there is potential harm to himself or others (physically or otherwise) across different driving scenarios.Table 1 Grade (age) differences in attitudes towards risky driving behaviors (N = 799).

ttitudes. This includes those who responded “never”, “rarely” or “occasionally” to questions inquiring whether the teen thinks there is potential harm to himself or others (physically or otherwise) across different driving scenarios.Table 1 Grade (age) differences in attitudes towards risky driving behaviors (N = 799). Grade (%) <=9th (n = 154) 10th (n = 223) 11th (n = 209) 12th (n = 211) Χ2 (df) sig Harmful to drive faster than the speed limit 30.0 (6)*** Always 33 33 20 24 Occasionally/rarely 41 56 69 61 Never 26 11 10 14 Harmful to drive a car without a driving license 20.5 (6)** Always 21 12 7 11 Occasionally/rarely 44 56 66 59 Never 35 32 27 30 Harmful to drive/ride the car without wearing a seat belt 18.3 (6)** Always 23 17 16 15 Occasionally/rarely 43 54 66 62 Never 34 29 18 23 How often do you use mobile while driving? 33.9 (6)*** Always 22 26 39 29 Occasionally/rarely 47 54 51 54 Never 31 20 10 17 How much do you think you respect traffic laws? 23.7 (6)*** Always 43 29 29 28 Occasionally/rarely 42 59 64 65 Never 15 12 7 7 Percentages were rounded to the closest integer. P-value sig * <= .05, ** <= .01, *** <= .001. For example, the number of teens who think it is not harmful to drive faster than speed limit (those who responded never, rarely or occasionally) is highest among 11th graders (χ2 = 30.0, df = 6, P < .01). A similar trend is also observed in other pro-attitudes. The rate of mobile phone use while driving (always) is also higher among 11th and 12th graders (χ2 = 33.9, df = 6, P < .001).

e faster than speed limit (those who responded never, rarely or occasionally) is highest among 11th graders (χ2 = 30.0, df = 6, P < .01). A similar trend is also observed in other pro-attitudes. The rate of mobile phone use while driving (always) is also higher among 11th and 12th graders (χ2 = 33.9, df = 6, P < .001). 3.2 Predictors of teen engagement in Tafheet More than three-quarters (77%) of the boys surveyed reported that they will engage in dangerous activities. Forty-four percent reported that they engaged in Tafheet, whereas nearly 40% of the students surveyed think it is a delinquent act and 12% believe it is a violent act. A positive attitude toward Tafheet was also voiced by 48% of the students, of whom 28% think it is a talent and 19% respond that it is a sport and a cool activity. On the other hand, nearly one half (46%) indicated that Tafheet should be prohibited. Twenty-six percent said it should be supported by special clubs, 20% said it requires special training, and 9% said it should be supported by conducting yearly tournaments.

alent and 19% respond that it is a sport and a cool activity. On the other hand, nearly one half (46%) indicated that Tafheet should be prohibited. Twenty-six percent said it should be supported by special clubs, 20% said it requires special training, and 9% said it should be supported by conducting yearly tournaments. Significant associations at the bivariate level were found between participation in Tafheet and the categories of “Do things even if they are dangerous”, “Harmful to drive a car without a driving license”, “Harmful to drive/ride the car without wearing a seat belt“, “How often do you use your mobile while driving?”, and “Respect traffic laws” (see Table 2). In general, a higher proportion of students engaged in Tafheet if they believed that there is no harm to themselves or others physically if they engaged in these activities. A higher proportion among those who believe Tafheet is a talent or a cool activity also practiced Tafheet.Table 2 Bivariate associations between Tafheet, attitudes, and driving risk behaviors among boys (N = 799).

hey believed that there is no harm to themselves or others physically if they engaged in these activities. A higher proportion among those who believe Tafheet is a talent or a cool activity also practiced Tafheet.Table 2 Bivariate associations between Tafheet, attitudes, and driving risk behaviors among boys (N = 799). Engaged in Tafheet (%) Yes No Χ2 (df) sig Type of school 5.3 (2) Government (n = 197) 40 60 Private (n = 266) 50 50 International (n = 138) 41 59 Grade 8.4 (4) 7/8 (n = 40) 50 50 9 (n = 73) 40 60 10 (n = 166) 52 48 11 (n = 160) 44 56 12 (n = 167) 37 63 Ethnic group .86 (2) Saudi (n = 424) 45 55 Arab (n = 143) 43 57 Neither (n = 37) 38 62 Do things even if they are dangerous 22.4 (1)*** Yes (n = 435) 51 49 No (n = 117) 27 73 Harmful to drive faster than the speed limit 1.9 (2) Always (n = 147) 46 54 Occasionally/rarely (n = 323) 43 57 Never (n = 86) 51 49 Harmful to drive a car without a driving license 9.6 (2)** Always (n = 64) 41 59 Occasionally/rarely (n = 316) 40 60 Never (n = 179) 54 46 Harmful to drive/ride the car without wearing a seat belt 12.7 (2)** Always (n = 91) 42 58 Occasionally/rarely (n = 316) 39 61 Never (n = 147) 57 43 How often do you use your mobile while driving? 13.9 (2)*** Always (n = 169) 56 44 Occasionally/rarely (n = 294) 44 56 Never (n = 83) 33 67 Use a helmet when you rollerblade, skateboard, or ride a motorcycle 3.5 (2) Always (n = 59) 44 56 Occasionally/rarely (n = 165) 41 59 Never (n = 238) 50 50 Respect traffic laws 42.05 (2)*** Always (n = 179) 31 69 Occasionally/rarely (324) 46 54 Never (n = 56) 80 20 Do you think “Tafheet” is a sort of 58.8 (4)*** Talent (n = 94) 65 35 Sport (n = 30) 37 63 Violence (n = 41) 20 80 Delinquency (n = 133) 23 77 Cool Activity (n = 33) 70 30 Do you think “Tafheet” should be 40.8 (3)*** Prohibited (n = 200) 24 76 Supported by special clubs (n = 118) 55 45 Supported by making it an early tournament (n = 37) 57 43 Supported by training and by providing safety equipment (n = 76) 51 49 Percentages were rounded to the closest integer. P-value sig * <= .05, ** <= .01, *** <= .001.

eet” should be 40.8 (3)*** Prohibited (n = 200) 24 76 Supported by special clubs (n = 118) 55 45 Supported by making it an early tournament (n = 37) 57 43 Supported by training and by providing safety equipment (n = 76) 51 49 Percentages were rounded to the closest integer. P-value sig * <= .05, ** <= .01, *** <= .001. A similar trend was observed in multivariate analysis (see Table 3) after controlling for grade which indicates that those who are willing to engage in activities even when they are dangerous, those who think there is no harm in not wearing seat belt, and those who use a mobile phone while driving are at nearly three times higher risk for practicing Tafheet. Those who never or sometimes respect traffic laws are also at higher risk for practicing Tafheet.Table 3 Adjusted odds ratios (OR) and 95% confidence intervals (CI) from regression model predicting Tafheeta.

lt, and those who use a mobile phone while driving are at nearly three times higher risk for practicing Tafheet. Those who never or sometimes respect traffic laws are also at higher risk for practicing Tafheet.Table 3 Adjusted odds ratios (OR) and 95% confidence intervals (CI) from regression model predicting Tafheeta. Adjusted OR 95% CI (lower, upper) Do things even if they are dangerous (ref: no) 2.70*** 1.55, 4.72 Think harmful to drive faster than the speed limit (ref: Always) Occasionally/rarely .52* .293, .93 Never .41* .18, .93 Think harmful to drive a car without a driving license (ref: Always) Occasionally/rarely .69 .32, 1.52 Never .84 .34, 2.08 Think harmful to drive/ride the car without wearing a seat belt (ref: Always) Occasionally/rarely 1.21 .63, 2.33 Never 2.77* 1.24, 6.21 Using mobile while driving (ref: Never) Always 2.96** 1.41, 6.23 Occasionally/rarely 2.10* 1.04, 4.25 Respect traffic laws (ref: Always) Occasionally/rarely 1.89** 1.12, 3.19 Never 8.78*** 3.59, 21.46 a Regression model controlled for grade. P-value sig * <= .05, ** <= .01, *** <= .001.

.63, 2.33 Never 2.77* 1.24, 6.21 Using mobile while driving (ref: Never) Always 2.96** 1.41, 6.23 Occasionally/rarely 2.10* 1.04, 4.25 Respect traffic laws (ref: Always) Occasionally/rarely 1.89** 1.12, 3.19 Never 8.78*** 3.59, 21.46 a Regression model controlled for grade. P-value sig * <= .05, ** <= .01, *** <= .001. 4 Discussion The current study examined the risky driving behaviors and attitudes among school youth that promote participation in the dangerous sport of “Tafheet”. Our study undeniably points to pro-attitudes and behaviors that include daring personality, not wearing a seat belt and using a mobile phone while driving. The results indicate most notably that youth do not respect traffic laws. Research suggests that when considered alone, human factors have been found to contribute to 57% of the accidents in developed countries. When also considering vehicular and environmental factors, human factors account for approximately 92% of these accidents [13]. Approximately 80% of accidents reported in the KSA during 1994 were attributed to driver-related factors [14].

ors have been found to contribute to 57% of the accidents in developed countries. When also considering vehicular and environmental factors, human factors account for approximately 92% of these accidents [13]. Approximately 80% of accidents reported in the KSA during 1994 were attributed to driver-related factors [14]. One of the common contributing factors to RTAs is not wearing a seat belt. Fewer than 2% of the drivers involved in accidents in the KSA during 2010 were wearing seat belts, compared to 85% in the USA [15]. Most industrialized and many developing countries passed laws requiring the use of motor vehicle seat belts much earlier than Saudi Arabia did. Despite the fact that wearing a seat belt saves lives, the KSA had no safety belt use laws until the year 2000, when seat belt use became compulsory for the driver and front-seat passengers [16]. Despite the passage of this law, no public awareness campaigns were conducted to educate drivers on the importance of wearing seat belts. The seat belt use rate in two Riyadh suburbs was found to be between 4 and 33% for drivers and 41–87% for front-seat passengers. However, those low rates are encouraging when compared with usage rates before enactment of the law [16]. Seat belt use is rarely practiced even today due to lack of strict enforcement. The Kingdom is therefore in dire need of strategies to strictly enforce the law and to conduct a campaign to increase public awareness. Such campaigns, especially when combined with law enforcement, have been proven to be effective in increasing seat belt usage rates in western countries. Such educational programs also must be implemented in KSA high schools to ensure safe driving.

enforce the law and to conduct a campaign to increase public awareness. Such campaigns, especially when combined with law enforcement, have been proven to be effective in increasing seat belt usage rates in western countries. Such educational programs also must be implemented in KSA high schools to ensure safe driving. The second most common factor in RTAs is speeding and not obeying traffic regulations, both of which are major concerns in Saudi Arabia. A study by Ansari et al [17] reported that over 65% of accidents occurred because of vehicles traveling at excess speed and/or drivers disobeying traffic signals. The drivers were responsible for nearly 80% of accidents. The proportion of teenage drivers involved was found to be three times that of the USA. Approximately one-fifth of drivers involved in accidents did not have a driver's license, and nearly 70% of accidents were attributable to speeding, and failure to obey traffic signals. Teenage drivers were involved in 2.05% of all accidents that occurred in the USA during 1993, whereas the rate in KSA was approximately 7%, about three times greater. Young age, lack of proper training, and poor driving education all contribute to motor vehicle accidents. Further, it is not unusual for a young male to drive even if he is under age 18, which is the legal age to drive in KSA. The central problem lies with youth having few or no outlets for organized and supervised extra-curricular activities. It is very common for young males to view driving, including Tafheet, as a sport. However, these youth lack proper education on safe driving, respect for the road, and obeying traffic rules before they begin to drive [18].

ies with youth having few or no outlets for organized and supervised extra-curricular activities. It is very common for young males to view driving, including Tafheet, as a sport. However, these youth lack proper education on safe driving, respect for the road, and obeying traffic rules before they begin to drive [18]. Studies show that the layout of Riyadh city also contributes to traffic confusion and accidents. As in many big cities, roundabouts are used extensively in Riyadh. Driving through roundabouts can be a dangerous experience, as many drivers there do not follow regulations. Approximately 90% of all drivers violate at least one traffic regulation when driving through these roundabouts. The most frequent violations include failure to signal when leaving or entering a roundabout and neglecting the right of way [19]. Drivers lack proper knowledge on roundabout driving regulations, as driver training and the licensing process do not include enough information on roundabouts.

ng through these roundabouts. The most frequent violations include failure to signal when leaving or entering a roundabout and neglecting the right of way [19]. Drivers lack proper knowledge on roundabout driving regulations, as driver training and the licensing process do not include enough information on roundabouts. Lack of public transportation and relatively low prices of automobiles and gas are main reasons for increased driving rates in Saudi Arabia. The economy of Saudi Arabia is petroleum-based, and KSA is a central player in the global oil market. Over the period 1990–2011, Saudi Arabia produced over 78 billion barrels of oil (approximately 13% of global supply), which accounted for approximately 75% of budget revenues and 90% of export earnings [20]. There were approximately 336 cars for every 1000 inhabitants in 2010 [21]. The country is large with a well-maintained highway structure, a well-designed local road system in large cities like Riyadh, and approximately 7 million registered cars [2]. Although the Kingdom has specified legal speed limits, driving at high speeds on the open wide roads is common and is a factor in many accidents.

The country is large with a well-maintained highway structure, a well-designed local road system in large cities like Riyadh, and approximately 7 million registered cars [2]. Although the Kingdom has specified legal speed limits, driving at high speeds on the open wide roads is common and is a factor in many accidents. As with the dangerous sport of Tafheet, the Saudi government adopted a tougher law in the past year to deter Arab drifting and joyriding. Previously, fatalities associated with drifting would have been classified as accidental. Now, they are classified as acts of criminal negligence, which carry a much heavier punishment. This is expected to help deter Arab drifters from performing reckless stunts. The punishments for Tafheet may include prison time or even a death sentence by beheading if deaths of other people result from the act [22], [23]. However, although the police receive complaints about drifting, the drifters are rarely caught, as the events are organized under the vigilance of illegal spotters who help disband vehicle activity before police arrive on the scene. Although the police are responsive to public complaints, investigations are futile as the spectators and drivers disperse quickly into normal moving traffic when the police arrive. Occasionally, traffic police are even chased away by both the drivers and spectators. Videos of Tafheet events are often posted on the Internet for public viewing. Accessibility to such videos further inspires thrill-seeking behavior among young males.

disperse quickly into normal moving traffic when the police arrive. Occasionally, traffic police are even chased away by both the drivers and spectators. Videos of Tafheet events are often posted on the Internet for public viewing. Accessibility to such videos further inspires thrill-seeking behavior among young males. The Saudi government has launched a road safety campaign targeting youth and has implemented the United Nations Global Plan for the Decade of Action for Road Safety 2011–2020. The Saudi Ministry of Interior also launched the Advanced Traffic Vehicle and Monitoring (ATVAM) project in 2011, known locally as Saher. The project is still in its infancy, but early signs are encouraging, and further impact on driver behavior is expected. Following introduction of this program, there has been a 19% decline in the number of traffic deaths, a rate 7.7% lower than in the same month in 2010. Arab News has also reported a 19% reduction in road deaths and a 30.4% reduction in injuries. In some provinces, e.g., Tabuk, the rate has dropped by an overwhelming 63%. Similar improvements in road safety have been recorded in other cities and provinces where the program is active [24].

han in the same month in 2010. Arab News has also reported a 19% reduction in road deaths and a 30.4% reduction in injuries. In some provinces, e.g., Tabuk, the rate has dropped by an overwhelming 63%. Similar improvements in road safety have been recorded in other cities and provinces where the program is active [24]. In addition to this major advancement to monitor RTAs, the Saudi government must consider promoting educational programs that should be mandated before youth graduate from high school. Our study underscores the desperate need for a standardized and effective educational curriculum to train and properly guide Saudi youth before they begin to drive. It is critical that children comprehend and develop positive attitudes towards respecting and obeying traffic laws. The Ministry of Education can facilitate intervention programs in each school to increase awareness and develop healthy attitudes about driving behavior. This approach has greater potential to change the current culture and to reduce the burden of traffic fatalities in the Kingdom.

respecting and obeying traffic laws. The Ministry of Education can facilitate intervention programs in each school to increase awareness and develop healthy attitudes about driving behavior. This approach has greater potential to change the current culture and to reduce the burden of traffic fatalities in the Kingdom. High school is an ideal setting for timely intervention. Our data show that 11th grade is the most crucial time to change attitudes that might lead to risky driving behaviors. Schools must examine strategies for changing teen driving behavior and identifying effective measures for improving the use of seat belts and for reducing other risky behaviors. A multifaceted and multilevel model based on ecological theory can be used for understanding how teens make choices about driving behaviors and for understanding the array of factors that can influence these choices [25]. The model can aid in generating recommendations for comprehensive intervention strategies that can be used in Arab communities to reduce disparities in risk behaviors, injury, disability, and death. Furthermore, healthcare providers and schools should consider counseling parents to discourage giving novice teen drivers unsupervised access to vehicles. In communities such as Riyadh, where teens require primary access to private vehicles due to limited public transportation options, greater efforts should be made to promote safe behaviors.

providers and schools should consider counseling parents to discourage giving novice teen drivers unsupervised access to vehicles. In communities such as Riyadh, where teens require primary access to private vehicles due to limited public transportation options, greater efforts should be made to promote safe behaviors. 4.1 Study strengths and drawbacks Our study suffers from classic drawbacks that affect any survey-based research. The cross-sectional nature of the study limits our ability to draw any causal conclusions. Given the self-reported nature of the questions, we must rely on respondents' honesty and memory recall. We also acknowledge that many of the risk behaviors measured might have been under-reported, leading to lower estimates of prevalence. Furthermore, students with inadequate literacy levels may not have reliably understood or responded to the questions. Other challenges include missing data due to nonresponse or choice of “don't know” responses. This issue may have resulted in missing at least 15% of potential responses for any given variable. It is important to note that the specific question assessing “Tafheet” does not distinguish between drivers and passengers, as it simply asks whether students ever participated in Tafheet. Accordingly, this question limits our ability to separate drivers from other participants. Additionally, the dichotomy of risky behavior (yes/no response to Tafheet) does not discriminate between those who regularly engage in Tafheet and those who may have experimented once or twice in their lifetimes. A more specific question with regard to frequency would have been appropriate to elicit risk level and to identify characteristics of drivers. Because the study was conducted only in the city of Riyadh, it may not be appropriate to extrapolate the results to adolescents within the whole Kingdom, particularly those in the rural provinces. Nonetheless, the study strengths include a large sample set randomly selected to represent students across age groups and across a variety of school types and locations in Riyadh, the largest city in the Kingdom, with a population at nearly 7 million [26].

n the whole Kingdom, particularly those in the rural provinces. Nonetheless, the study strengths include a large sample set randomly selected to represent students across age groups and across a variety of school types and locations in Riyadh, the largest city in the Kingdom, with a population at nearly 7 million [26]. 4.2 Policy implications According to the Arriyadh Development Authority, young people make up the majority of Riyadh city population [27]. Youth below 15 years of age constitute approximately 34% of the total Saudi native population (excluding expatriates). Therefore, the base of the population pyramid is wide and getting wider. The Saudi population as a whole is typically young compared to other countries, with a median age of 26.4 years.

opulation [27]. Youth below 15 years of age constitute approximately 34% of the total Saudi native population (excluding expatriates). Therefore, the base of the population pyramid is wide and getting wider. The Saudi population as a whole is typically young compared to other countries, with a median age of 26.4 years. Our study findings seem to suggest that male high school students do engage to some extent in risky driving behaviors and hold attitudes that support unsafe driving practices, regardless of whether they engage in Tafheet. Our study results clearly call for a strategic intervention plan to educate youth in school settings with special emphasis on the problems of speeding, disobeying traffic laws, driving without a driver's license, and the sport of Tafheet. Strict law enforcement and educational campaigns must be seriously considered to change the driving culture of Saudi Arabia. Strong enforcement of speeding laws should be enhanced, particularly on streets and at times when speeding and Tafheet are likely to take place. This includes weekends (i.e., Thursday thru Saturday) and national holidays. The use of public media can also help disseminate the messages of safe driving to reach a larger audience. Future research must explore the potential benefit of a multilevel systems approach that incorporates the school system, parents, and law enforcement to address unsafe youth attitudes and driving.

tional holidays. The use of public media can also help disseminate the messages of safe driving to reach a larger audience. Future research must explore the potential benefit of a multilevel systems approach that incorporates the school system, parents, and law enforcement to address unsafe youth attitudes and driving. Human subjects approval statement Ethical approval for the study was obtained from King Fahad Medical City Institutional Review Board along with permission from the Ministry of Education in Riyadh to administer the survey in schools. Conflict of interest None declared. Acknowledgments The authors would like to acknowledge all school principals and teachers who facilitated the survey and extended the needed support. High appreciation is also given to the teams who contributed their efforts to collecting the data, especially Basma and Othman. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

Perforations following ERCP accounts for less than 1%, however the mortality rate can reach up to 16% [1], [2]. Multiple risk factors have been implicated including Sphincter of Oddi dysfunction, papillary stenosis, anatomical alterations due to previous surgeries and prolonged ERCP duration [2]. The symptoms of perforation can range from asymptomatic to peritonitis and sepsis [1]. Consequently, the management differs. Based on the above, multiple attempts have been made by different authors to classify the types of injury and therefore recommend the most appropriate management [2]. One classification depends on the site of injury as follows: Type I: In this type the injury is in the lateral or medial wall of the duodenum. It usually causes large persistent pancreatic leak to the retroperitoneal or intraperitoneal space. It requires immediate diagnosis and surgical repair. Type II: In this type, the perforation occurs in the peri-ampullary area, usually during sphincterotomy. In this type, the retroperitoneal area is the main site for accumulation of the leakage. Type III: Refers to perforation occurring within the bile duct. It usually occurs during the insertion of a guide wire, treatment of bile duct stone or in biliary strictures. Type II and III can be managed either conservatively or surgically depending on the presenting symptoms and the progression of the disease. Type IV: retroperitoneal air accumulation. It considered as a non-true perforation that results from the use of compressed air to keep the lumen open. Therefore no surgical intervention is required.

Type II and III can be managed either conservatively or surgically depending on the presenting symptoms and the progression of the disease. Type IV: retroperitoneal air accumulation. It considered as a non-true perforation that results from the use of compressed air to keep the lumen open. Therefore no surgical intervention is required. Perforation injuries can be missed during an ERCP procedure despite careful observation as these patients are usually under anesthesia. Therefore, intraprocedural contrast medium leakage or free air detection plus a post procedural radiograph should be performed [1], [2]. If a perforation is suspected, patients should have a CT examination to confirm leakage, keeping in mind that fluid collection is more important than the presence of free air. Such a finding indicates continuous bile or pancreatic juice leakage through the perforation site. In addition to the above, endoscopic management has been introduced in recent years with successful outcomes. Clipping and spraying fibrin glue on the perforation sites have been used [2].

Perforation injuries can be missed during an ERCP procedure despite careful observation as these patients are usually under anesthesia. Therefore, intraprocedural contrast medium leakage or free air detection plus a post procedural radiograph should be performed [1], [2]. If a perforation is suspected, patients should have a CT examination to confirm leakage, keeping in mind that fluid collection is more important than the presence of free air. Such a finding indicates continuous bile or pancreatic juice leakage through the perforation site. In addition to the above, endoscopic management has been introduced in recent years with successful outcomes. Clipping and spraying fibrin glue on the perforation sites have been used [2]. In relation to our patient, Type II perforation is the most likely type. Our patient needed a large sphincterotome in order to be able to retrieve the large sized CBD stone. In addition the abnormal anatomy this patient had, secondary to his duodenal atresia repair, increased his risk for perforation. This is further supported by the subsequent course of this patient; he responded to the conservative management and his retroperitoneal air disappeared on follow up CT examination 10 days later. Subsequently, he was seen in the outpatient clinic a month later and he was asymptomatic. His Abdominal xray at the time of his visit was normal (Fig. 1).Figure 1 Retropritoneal air. ↑: sub-diaphragmatic air did not move between standing and lateral decubitus positions, A & B. →: retroperitoneal air outlining the right kidney. Figure 1

In relation to our patient, Type II perforation is the most likely type. Our patient needed a large sphincterotome in order to be able to retrieve the large sized CBD stone. In addition the abnormal anatomy this patient had, secondary to his duodenal atresia repair, increased his risk for perforation. This is further supported by the subsequent course of this patient; he responded to the conservative management and his retroperitoneal air disappeared on follow up CT examination 10 days later. Subsequently, he was seen in the outpatient clinic a month later and he was asymptomatic. His Abdominal xray at the time of his visit was normal (Fig. 1).Figure 1 Retropritoneal air. ↑: sub-diaphragmatic air did not move between standing and lateral decubitus positions, A & B. →: retroperitoneal air outlining the right kidney. Figure 1 Conflict of interest None to be declared by the authors. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

The publisher regrets that the original version of the above article contained an error. The error occurred in the heading of the paper. The paper carried an erroneous heading as “Case Report” instead of the correct heading which is “What's your diagnosis”. The publisher would like to apologize for any inconvenience caused. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Müllerian anomalies are congenital defects of the female reproductive tract resulting from failure in the development of the Müllerian ducts and their associated structures. The incidence of these anomalies has been estimated to be 1 in 70,000 females [1]. The cause of the anomalies is not known, but it is currently believed to be multifactorial. The patient might present in the neonatal period, during adolescence, or early adulthood, and the anomaly might affect the reproductive capacity. Transverse vaginal septum is a rare anomaly, and a low septum is even rarer. When clinically suspected, investigations leading to the diagnosis include imaging methods such as hysterosalpingography, ultrasonography (USG), magnetic resonance imaging (MRI), and cystoscopy. The management depends on the age at presentation, the site, and the thickness of the septum. Regular follow-up and strict adherence to vaginal dilatation are pre-requisites for good long-term results.

maging methods such as hysterosalpingography, ultrasonography (USG), magnetic resonance imaging (MRI), and cystoscopy. The management depends on the age at presentation, the site, and the thickness of the septum. Regular follow-up and strict adherence to vaginal dilatation are pre-requisites for good long-term results. 2 Case summary A 13-year-old pre-menarchal girl presented with history of episodic, colicky abdominal pain since 3 months of age. She had a history of anoplasty performed for ectopic anus in the neonatal period. There was a large, non-tender cystic lump arising from the pelvis. On rectal examination, a cystic bulge could be felt anteriorly up to the anal verge. The external genitalia were normal. There were three openings in the perineum with the anus placed slightly anteriorly (Fig. 1). The Tanner staging for both the external genitalia and the breasts was Stage II.Figure 1 Local examination showing three openings in the perineum with anus placed little anteriorly. No evidence of bulging membrane (as seen in imperforate hymen).

penings in the perineum with the anus placed slightly anteriorly (Fig. 1). The Tanner staging for both the external genitalia and the breasts was Stage II.Figure 1 Local examination showing three openings in the perineum with anus placed little anteriorly. No evidence of bulging membrane (as seen in imperforate hymen). Abdominal ultrasound suggested hematometrocolpos with fullness of the left pelvi-calyceal system. Micturating cystourethrography (MCU) showed grade 5 vesico-ureteric reflux (VUR) on the left side and a large, distended bladder. Perineal ultrasound revealed a low transverse vaginal septum of 8–10-mm thickness. An MRI of the pelvis showed hematometrocolpos and a low transverse vaginal septum. The distance between the lower end of the vagina and the introitus was 15 mm (Fig. 2). During examination under general anesthesia, the vaginal pit was shallow, but there was no bulging at the outlet. Vaginoscopy was attempted, but it was not possible as there was a complete low septum.Figure 2 MRI of the pelvis showing hematometrocolpos and a low transverse vaginal septum (white arrow). The uterus (blue arrow) distended upper vagina (black arrow) and the collapsed lower vagina (white arrow head) are seen. The distance between lower end of vagina and the introitus was 15 mm.

was a complete low septum.Figure 2 MRI of the pelvis showing hematometrocolpos and a low transverse vaginal septum (white arrow). The uterus (blue arrow) distended upper vagina (black arrow) and the collapsed lower vagina (white arrow head) are seen. The distance between lower end of vagina and the introitus was 15 mm. Intraoperatively, stay sutures were placed on the septum, and the septum was marked off (Fig. 3). It was aspirated and opened by a cruciate incision. The margins were then sutured (Fig. 4). Postoperatively, the vaginal form was kept.Figure 3 Intra-operative image showing the stay sutures and the cruciate incision. Figure 4 Intra-operative image showing completed suturing. A postoperative ultrasound showed no evidence of hematometrocolpos. The patient is awaiting management of ARM & VUR.

Intraoperatively, stay sutures were placed on the septum, and the septum was marked off (Fig. 3). It was aspirated and opened by a cruciate incision. The margins were then sutured (Fig. 4). Postoperatively, the vaginal form was kept.Figure 3 Intra-operative image showing the stay sutures and the cruciate incision. Figure 4 Intra-operative image showing completed suturing. A postoperative ultrasound showed no evidence of hematometrocolpos. The patient is awaiting management of ARM & VUR. 3 Discussion The female genital system develops from the paired Müllerian ducts and in close association with the urinary system and hindgut [1]. The paired Müllerian ducts develop from the coelomic epithelium lateral to the mesonephric ducts and cross medially to fuse in the midline [2]. The most cranial parts of the Müllerian ducts, which remain separate from the fallopian tubes, and the caudal segments proceed caudally to join the urogenital sinus, where they produce an elevation called the Müllerian tubercle [1], [2]. These fuse and form the uterus, cervix, and the proximal two-thirds of the vagina [2]. The sinovaginal bulbs are paired caudal ectodermal outgrowths that proliferate to form a cord of tissue that develops into the distal vaginal plate. This plate is later canalized in a caudal to cranial direction to form the distal vagina [2].

e and form the uterus, cervix, and the proximal two-thirds of the vagina [2]. The sinovaginal bulbs are paired caudal ectodermal outgrowths that proliferate to form a cord of tissue that develops into the distal vaginal plate. This plate is later canalized in a caudal to cranial direction to form the distal vagina [2]. The close proximity of the mesonephric and paramesonephric ductal systems helps to explain the common association of paramesonephric abnormalities and ipsilateral renal anomalies. Even mesodermally derived organs as cranial as the cervical vertebra and the tracheo-esohageal anlagen can be affected in association with congenital abnormalities of the mesonephric and paramesonephric ductal systems, as is observed in the vertebral defects, anal atresia, tracheo-esophageal fistula with esophageal atresia, radial and renal dysplasia (VATER) and Müllerian duct aplasia, renal aplasia, and cervicothoracic somite dysplasia (MURCS) associations [2]. The anomalies of the female reproductive system can be grouped into four main categories as per the American Fertility Society (AFS) classification [3]: 1) Those resulting from either hypoplasia or agenesis, 2) Those caused by vertical fusion, which can be obstructive or non-obstructive (canalization abnormalities resulting from abnormal contact of the mullerian structures with the urogenital sinus), 3) Those resulting from lateral fusion (duplication), and 4) Unusual configurations and combinations of defects.

The anomalies of the female reproductive system can be grouped into four main categories as per the American Fertility Society (AFS) classification [3]: 1) Those resulting from either hypoplasia or agenesis, 2) Those caused by vertical fusion, which can be obstructive or non-obstructive (canalization abnormalities resulting from abnormal contact of the mullerian structures with the urogenital sinus), 3) Those resulting from lateral fusion (duplication), and 4) Unusual configurations and combinations of defects. Delaunay first described the transverse vaginal septum in 1877 [4]. It is believed to arise from a failure in fusion or canalization (or both) of the urogenital sinus and Müllerian ducts. The cause is unknown although some cases might be the result of a female sex-limited autosomal recessive transmission [1] or exposure to certain agents in utero [5]. The septum is composed of fibrous connective tissue and vascular muscular elements. Although the lower surface is always covered with squamous epithelium, the upper surface can be covered by either vaginal squamous epithelium or columnar epithelium, with or without metaplasia [1].

posure to certain agents in utero [5]. The septum is composed of fibrous connective tissue and vascular muscular elements. Although the lower surface is always covered with squamous epithelium, the upper surface can be covered by either vaginal squamous epithelium or columnar epithelium, with or without metaplasia [1]. The estimated incidence is 1 in 70,000 females [2]. A complete transverse vaginal septum can be located at various levels in the vagina, but there is a higher frequency in the middle and upper third of the vagina. In one large series by Lodi in 1951, the distribution was 46% in the upper vagina, 40% in the middle vagina, and 14% in the lower vagina [6]. The septa are usually less than 1 cm thick and frequently have a small central or eccentric perforation [2], giving an impression of a vaginal vault without a cervix [3]. The septa are frequently accompanied by urinary tract abnormalities, such as unilateral renal agenesis, ectopia, or fusion [7], as well as musculoskeletal, gastrointestinal, and cardiac defects [1]. The most common genital anomalies associated are uterus bicollis with an obstructing partial vaginal septum and the unicornuate uterus with a rudimentary horn [7]. It has also been associated with bilateral tubal atresia [1]. Incomplete treatment of Transvaginal ultrasound (TVS) allows menstrual flow to escape periodically. The diagnosis can be made on transvaginal sonography, computed tomography, and magnetic resonance imaging [3].

unicornuate uterus with a rudimentary horn [7]. It has also been associated with bilateral tubal atresia [1]. Incomplete treatment of Transvaginal ultrasound (TVS) allows menstrual flow to escape periodically. The diagnosis can be made on transvaginal sonography, computed tomography, and magnetic resonance imaging [3]. The clinical presentation depends on whether it is complete or partial. With complete septum, the menstrual blood accumulates in the genital tract resulting in hematocolpos and hematometra. Such patients usually present at adolescence with cyclic lower abdominal pain, and occasionally, a lower abdominal mass (hematometra) can be palpable. Hematocolpos might not develop until puberty, but complete obstruction might cause serious compression of the surrounding structures in neonates and infants. A high transverse vaginal septum might cause cyclic hematuria if communicating with the urinary system. An incomplete septum allows partial egress of menstrual blood, and such patients complain of dysmenorrhea, hypomenorrhea, dyspareunia, foul-smelling vaginal discharge [2], [4], and dystochia. It might also be asymptomatic and appear as an incidental finding during routine gynecologic examination [4]. However, unlike an imperforate hymen, examination of the genitalia reveals no evidence of bulging at the introitus.

enorrhea, hypomenorrhea, dyspareunia, foul-smelling vaginal discharge [2], [4], and dystochia. It might also be asymptomatic and appear as an incidental finding during routine gynecologic examination [4]. However, unlike an imperforate hymen, examination of the genitalia reveals no evidence of bulging at the introitus. Transperineal, transvaginal, and vesical hydrosonography have been used in defining both the vaginal structure and pathological conditions; however, the fine details as to whether the septum is complete or incomplete, or the number and dimensions of vaginal compartments cannot be determined [1]. A transvaginal approach, coupled with catheter instillation of saline solution, provides a more accurate assessment [1]. MRI can help determine whether a cervix is present so that a high septum can be differentiated from congenital absence of the cervix [2].

dimensions of vaginal compartments cannot be determined [1]. A transvaginal approach, coupled with catheter instillation of saline solution, provides a more accurate assessment [1]. MRI can help determine whether a cervix is present so that a high septum can be differentiated from congenital absence of the cervix [2]. The treatment of a TVS is individualized, and the goal is to relieve cyclic abdominal pain and prevent development of endometriosis to preserve fertility [5], [8], [9]. Vaginal dilators are the preferred non-surgical choice for patients with small septa [5]. Dilatation techniques can be used in lieu of surgery, before surgery in order to improve outcomes, and after surgery to prevent strictures, scarring, or stenosis of surgical site [2]. A low transverse vaginal septum is treated by a transverse incision over the vault of the short vagina followed by anastomosis after identifying the cervix [5]. An indwelling stent or a soft foam rubber vaginal form is used in the immediate postoperative period followed by vaginal dilatations. Coital function is usually normal after surgery, and pregnancy rates are 40–50% for septum in the lower or middle third of the vagina. If the patient's pain from hematocolpos is manageable, surgery can be delayed with suppression of endometrial activity by a GnRH agonist or continuous oral contraceptives [2]. This treatment might allow time for dilation of the lower vaginal segment, potentially improving the ease of surgical repair [2], especially in cases of a high septum.

rom hematocolpos is manageable, surgery can be delayed with suppression of endometrial activity by a GnRH agonist or continuous oral contraceptives [2]. This treatment might allow time for dilation of the lower vaginal segment, potentially improving the ease of surgical repair [2], especially in cases of a high septum. Small vaginal septa (<1 cm in thickness) can be treated by excision with a simple end-to-end anastomosis of the vaginal epithelium or a Z-plasty [5]. Larger septa (>1 cm) might require pre-operative vaginal dilatation followed by a longitudinal Z-plasty technique to reduce stenosis and contraction of scarring at the site [5]. The Olbert balloon catheter technique has also been described to maximize the vaginal mucosa available for anastomosis and to avoid postoperative narrowing of the vagina [5]. Vaginal stenosis at the site of resection is the most common complication [2]. Postoperative vaginal dilation or a vaginal mold might help in decreasing the scarring and stenosis of the surgical site [5]. Postoperative vaginal dilation is critical to the success of the procedure, which is usually affected by the apprehension of the adolescents. Such apprehensive patients might be better served with a long-term mold or stent [5]. Daily guidance with patience is necessary to help these adolescent girls gain confidence for vaginal dilatation [5].

lation is critical to the success of the procedure, which is usually affected by the apprehension of the adolescents. Such apprehensive patients might be better served with a long-term mold or stent [5]. Daily guidance with patience is necessary to help these adolescent girls gain confidence for vaginal dilatation [5]. Adverse outcomes after surgery, such as dyspareunia, menstrual irregularities, fertility issues, and preterm labor, are common. Recognition and intervention at younger ages by draining the collected blood and possibly preventing endometriosis is necessary to preserve fertility [5], [8], [9]. Rock et al reported that patients were less likely to conceive after surgical correction, and even if they did so, there was a 50% chance of spontaneous abortion [9]. Therefore, these patients and their family members should be educated about these potential long-term complications and about regular follow-up while attempting to conceive. 4 Conclusions Müllerian anomalies are a diverse group of developmental disorders involving the internal female reproductive tract. Establishing an accurate diagnosis is hence essential for planning treatment and management strategies. Conflict of interest None to be declared by the authors. Source of funding None to be declared by the authors. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

ram-negative MDROs (in particular, beta-lactam resistant Gram-negative bacilli/extended-spectrum beta-lactamases [ESBLs] and carbapenem-resistant enterobacteriaceae [CRE]) within the pediatric population. This includes the areas of morbidity, mortality, length of stay (LOS), burden and cost to healthcare services [57]. As mentioned, the pediatric population heavily relies on direct hands-on-care. Therefore, MDRO pathogens can be spread easily via direct and indirect routes, are often difficult to treat and eradicate, are viable within the environment, and can remain on the skin whether as a result of colonization or as an infection for extended periods of time. Emphasizing preventive measures is important. These measures include hand hygiene, controlling environmental contamination, caregiver education, decolonization (if appropriate), appropriate PPE usage, and judicious antimicrobial controls [9], [27], [47], [57], [58], [59]. However, little consensus exists regarding appropriate isolation precaution standards for ESBLs and CRE in the context of known prolonged shedding among the immunocompromised pediatric population [60].

ation (if appropriate), appropriate PPE usage, and judicious antimicrobial controls [9], [27], [47], [57], [58], [59]. However, little consensus exists regarding appropriate isolation precaution standards for ESBLs and CRE in the context of known prolonged shedding among the immunocompromised pediatric population [60]. 2.1.9 Outside the healthcare setting 2.1.9.1 Preventative and protective measures IP&C practices outside the healthcare setting mainly revolve around ensuring child safety; preventing the acquisition of infections within the home, school or community; and preventing injuries that are both intentional and non-intentional. Guidance by caregivers is important in preparing the child during elective hospitalizations so that their physical, immunological and mental status is optimal in preparing them to cope with the healthcare setting [7]. In addition, circumstances resulting in an unexpected need for hospitalization may contribute to compromised states, which subsequently could lead to HAIs.

child during elective hospitalizations so that their physical, immunological and mental status is optimal in preparing them to cope with the healthcare setting [7]. In addition, circumstances resulting in an unexpected need for hospitalization may contribute to compromised states, which subsequently could lead to HAIs. Providing a safe environment outside the healthcare setting includes addressing the need for appropriate nutrition, clothing, shelter and education; minimizing the risk of injuries, such as falls, suffocation or choking; ingestion or absorption of poisons; scalding or burns; electrocution,; wearing of seat belts; safe play activities, including playgrounds, toys, and water activities that increase the risk of drowning (bathing or sports related); handling; care and exposure to pets and/or other animals with risks for bites and scratches; or exposure to abusive or neglectful situations, both physical and/or emotional [61], [62], [63], [64], [65].

Adherence to childhood immunization recommended schedules primarily against common vaccine-preventable infections can save lives through prevention and herd immunity [67]. Providing immunizations to children can heavily depend upon the child's age (could be too young to receive certain vaccines), their previous exposure to a certain disease(s) (with or without development of natural immunity), or parental attitude and/or beliefs that prohibit the administration of vaccines or other blood or blood products [7], [27], [28], [68]. 3 Conclusion It is important and the right of all children of any age group to have a safe and infection-free environment. Effective strategies, when applied appropriately, can control and prevent infections in the pediatric population. The literature review undertaken clearly supports the recommendation that IP&C programs need to be developed and tailored for specific pediatric age-groups. The factors that differ from adults need to be taken into consideration, including age, physical, and psycho-social factors that impact children's inability to follow IP&C standards; important role that the caregiver has in both prevention and transmission, and the need to address disease prevention strategies for those diseases that are known to occur most often among children, including those of an enteric and respiratory nature.

factors that impact children's inability to follow IP&C standards; important role that the caregiver has in both prevention and transmission, and the need to address disease prevention strategies for those diseases that are known to occur most often among children, including those of an enteric and respiratory nature. The authors identified areas that require further research and/or the need for more intensive study. These include studying the relationship between injuries or diseases that occur in the home or community and their impact on the healthcare settings; infection-related comparisons between different age groups; development of dedicated pediatric infection data-bases and guidelines, including the rates of CAUTI in NICUs; need to study the association and role of visitors in the transmission of infections within-the-healthcare setting as compared to the home environment; review of policies governing visitors within-the-healthcare setting; impact of viral shedding and need for resampling for clearance purposes, especially among the immunocompromised; and need for studies and guideline development on MDROs, especially ESBL and CRE, in relation to the pediatric population.

es, including playgrounds, toys, and water activities that increase the risk of drowning (bathing or sports related); handling; care and exposure to pets and/or other animals with risks for bites and scratches; or exposure to abusive or neglectful situations, both physical and/or emotional [61], [62], [63], [64], [65]. Safety measures take into account the child's age, developmental physical and cognitive abilities, individual characteristics, degrees of dependence, activities, and potential exposure to possible hazards and risk behaviors. As the child develops, their curiosity and wishes to experiment are not always matched by their capacity to understand or respond to danger, and therefore, adult supervision is essential. For the caregiver (or designee), their ability takes into account their level of judgment, setting, degree of verbal and physical intervention, and how much time the caregiver actively spends supervising [61], [62], [63], [64], [65]. Minimizing exposure to infectious organisms in the home, childcare/daycare/school settings or recreational centers will help reduce risk. Supervisory rules include the type/number of visitors with associated restrictions as applicable, preventing exposure to children with childhood illnesses being brought into the home or child care setting. Ensuring sufficient supervision is provided by the handling of pets or animal-assisted interventions/therapy. There is strong evidence suggesting that animals pose a risk for pathogen spread via the oral-fecal or skin/fur routes [7], [27], [32], [66].

1 Introduction Respiratory Syncytial Virus (RSV) is a single-stranded, non-segmented RNA negative-sense virus belonging to the Pneumovirinea subfamily of the Paramyxoviridae family. It has two subtypes, A and B, which are distinguished largely by differences in the viral attachment (G) protein and the nuclear (N) protein. During epidemics, either subtype may predominate, or both subtypes may circulate concurrently [1]. RSV is unstable in the environment and is readily inactivated by soap and water. The virus spreads through close contact with infected carriers or contaminated surfaces. Infection occurs when contaminated materials come in contact with the mucous membranes of the eyes, nose or mouth. It can remain infectious on surfaces or fomites for 4–7 h and can survive on unwashed hands [2], [3]. The main therapy for RSV in infants is supportive. Palivizumab (Synagis®), a human monoclonal antibody directed against the fusion protein F of RSV (conserved among isolates), is produced by recombinant DNA technology and was licensed for use in RSV prophylaxis in June 1998 by the United States Food and Drug Administration for the reduction of serious lower respiratory tract infection caused by RSV in children at increased risk of severe disease [4], [5], [6].

RSV (conserved among isolates), is produced by recombinant DNA technology and was licensed for use in RSV prophylaxis in June 1998 by the United States Food and Drug Administration for the reduction of serious lower respiratory tract infection caused by RSV in children at increased risk of severe disease [4], [5], [6]. This clinical practice policy statement was developed by the Ministry of Health and supported by the National Immunization Technical Advisory Group (NITAG) in Saudi Arabia based on all available national and recent international data for the use of Palivizumab for the prevention of severe LRTI caused by RSV in high-risk pediatric patients. The Saudi Pediatrics Infectious Diseases Society (SPIDS) and the Saudi Neonatology Society (SNS) have endorsed these RSV prophylaxis guidelines. These guidelines shall be reviewed and updated every 2 years as needed. The Ministry of Health laboratories will monitor changes in RSV seasons (see Table 1).Table 1 Summary of studies conducted in Saudi Arabia [14].

Society (SPIDS) and the Saudi Neonatology Society (SNS) have endorsed these RSV prophylaxis guidelines. These guidelines shall be reviewed and updated every 2 years as needed. The Ministry of Health laboratories will monitor changes in RSV seasons (see Table 1).Table 1 Summary of studies conducted in Saudi Arabia [14]. Year City/province Hospital No of samples Detection test HRSV positive No. (%) Type Ref. 1993 Riyadh KKUHa 127 IFAg 69 (54)k – Jamjoom et al 1998 Riyadh KFSHRCb 256 ND 73(28.5) – Al-Hajjar et al [9] 1998 Riyadh KKUH 1429 ND 412 (79) – Bakir et al [11] 2002 Riyadh KKUH 20 NDh 8 (40) – Kilani 2004 Mecca-Hajj KAUH-Jeddahc 500 IFA 4 (7.4) – Bakhly et al 2005 Abha ACHd 51 ELISAi/IFA 20 (40) – Al-shehri et al [10] 2005 Riyadh KKUH 4575 IFA 884 (19) – Sheir and Mona 2006 Al-Qassim BMPHe 282 IFA 128 (45) – Meqdam and Sobaih 2009 Riyadh KAMCf 10,617 IFA 733 (83) – Akhter et al [2] 2009 Riyadh KKUHa 200 RT-PCRj 70 (35) A(57%)B(42.9%) Al-majhdi et al [1] a King Khalid University Hospital. b King Faisal Specialist Hospital and Research Centre. c King Abdulaziz University Hospital. d Assir Central Hospital. e Buraidah Maternity and Pediatric Hospital. f King Abdulaziz Medical City. g Immunofluorescent assay. h Not defined. i Enzyme linked immunosorbent assay. j Reverse transcription polymerase chain reaction. k Percentage was calculated on the basis of respiratory samples of confirmed viral origin.

c King Abdulaziz University Hospital. d Assir Central Hospital. e Buraidah Maternity and Pediatric Hospital. f King Abdulaziz Medical City. g Immunofluorescent assay. h Not defined. i Enzyme linked immunosorbent assay. j Reverse transcription polymerase chain reaction. k Percentage was calculated on the basis of respiratory samples of confirmed viral origin. 2 Purpose of the guidelines 1. Implement national guidelines on RSV immunoprophylaxis to reduce variations across the country and limit Palivizumab to a specific high-risk population on the basis of available evidence, as well as expert opinion. 2. To be a resource for healthcare professionals (HCPs) involved in the management of an RSV Immunoprophylaxis Program. 3. Improve the utilization of resources and enhance cost-effective practices. 3 Epidemiology RSV is a highly contagious virus that causes serious global outbreaks. The virus results in significant morbidity and mortality in infants during the first year of life, and nearly all infants experience one or more RSV infections by the end of their second year [2]. The disease severity ranges from a mild upper respiratory tract infection (URTI) to a severe lower respiratory tract infections (LRTI). Globally, RSV is estimated to have caused 66,000 to 199,000 pneumonia deaths in children younger than 5 years in 2005. In the United States, the hospitalization rate is 2345 per 100,000 person-years for RSV compared to 151 for influenza, consistent with reports that RSV hospitalizes 1–2% of infants each winter [7], [8].

ildhood illnesses being brought into the home or child care setting. Ensuring sufficient supervision is provided by the handling of pets or animal-assisted interventions/therapy. There is strong evidence suggesting that animals pose a risk for pathogen spread via the oral-fecal or skin/fur routes [7], [27], [32], [66]. Adherence to childhood immunization recommended schedules primarily against common vaccine-preventable infections can save lives through prevention and herd immunity [67]. Providing immunizations to children can heavily depend upon the child's age (could be too young to receive certain vaccines), their previous exposure to a certain disease(s) (with or without development of natural immunity), or parental attitude and/or beliefs that prohibit the administration of vaccines or other blood or blood products [7], [27], [28], [68]. 3 Conclusion It is important and the right of all children of any age group to have a safe and infection-free environment. Effective strategies, when applied appropriately, can control and prevent infections in the pediatric population.

lobally, RSV is estimated to have caused 66,000 to 199,000 pneumonia deaths in children younger than 5 years in 2005. In the United States, the hospitalization rate is 2345 per 100,000 person-years for RSV compared to 151 for influenza, consistent with reports that RSV hospitalizes 1–2% of infants each winter [7], [8]. In Saudi Arabia, RSV was reported to be the main cause of LRTI in infants in more than one study [1], [9], [10], [11], [12], [13], accounting for up to 40% of all LRTIs in children aged <1 year and up to 83% in children aged <5 years. Most cases occur from November through March, but infections have been reported in other months in Saudi Arabia [6], [11]. Most RSV infections are mild and require minimal hospital stays; however, some children are severely affected, requiring pediatric ICU admission and a longer hospital stay. Risk factors for serious infection with RSV include prematurity; bronchopulmonary dysplasia (BPD); cyanotic congenital heart disease; and immunodeficiency diseases or immunosuppression caused by therapy [12]. 4 Definitions RSV season in this policy is defined as the period of time during which the prevalence of the infection increases. Although this varies among regions and nations according to seasonality, in Saudi Arabia the onset usually occurs in middle to late October and ends in early to mid-March [6]. Chronic lung disease (CLD) (also known as bronchopulmonary dysplasia) of prematurity was defined as oxygen dependency at 36 weeks corrected gestational age (GA) for babies delivered at less than 32 weeks GA [4].

4 Definitions RSV season in this policy is defined as the period of time during which the prevalence of the infection increases. Although this varies among regions and nations according to seasonality, in Saudi Arabia the onset usually occurs in middle to late October and ends in early to mid-March [6]. Chronic lung disease (CLD) (also known as bronchopulmonary dysplasia) of prematurity was defined as oxygen dependency at 36 weeks corrected gestational age (GA) for babies delivered at less than 32 weeks GA [4]. 5 Policy statement [4], [15], [16], [17], [18], [19], [20]] 1. RSV clinics shall dispense RSV prophylaxis from the middle of October until the middle of March. 2. Palivizumab prophylaxis shall be administered to all infants born before 29 weeks gestation who are younger than 12 months at the start of the RSV season. 3. Palivizumab prophylaxis must not be administered to healthy infants born at 29 weeks gestation or more who are free of CLD, as they experience a lower RSV hospitalization rate than preterm infants born at <29 weeks gestation. 4. Prophylaxis must be given in the first year of life for preterm infants with BPD. 5. Infants with certain hemodynamically significant heart diseases (a cyanotic heart disease under medical support and moderate to severe pulmonary hypertension) should also be given RSV prophylaxis.

3. Palivizumab prophylaxis must not be administered to healthy infants born at 29 weeks gestation or more who are free of CLD, as they experience a lower RSV hospitalization rate than preterm infants born at <29 weeks gestation. 4. Prophylaxis must be given in the first year of life for preterm infants with BPD. 5. Infants with certain hemodynamically significant heart diseases (a cyanotic heart disease under medical support and moderate to severe pulmonary hypertension) should also be given RSV prophylaxis. 6. Children with immunocompromised conditions, cystic fibrosis, pulmonary abnormalities or neuromuscular disease should not routinely be offered Palivizumab because of limited and inconclusive data. However, prophylaxis may be considered for children younger than 24 months of age with pulmonary abnormality or neuromuscular disease that impairs the ability to clear secretions from the upper airways, infants who are on home oxygen, who have had a prolonged hospitalization for severe pulmonary disease or who are severely immunocompromised. 7. Palivizumab should be administered up to a maximum of 5 monthly doses (15 mg/kg per dose administered intramuscularly once every 30 days) during the RSV season to infants who qualify for prophylaxis in the first year of life. A child with a history of a severe allergic reaction following a dose of Palivizumab should not receive additional doses. 8. Qualified infants born during the RSV season must receive fewer doses according to their month of birth. For example, infants born in January would receive their last dose in March.

7. Palivizumab should be administered up to a maximum of 5 monthly doses (15 mg/kg per dose administered intramuscularly once every 30 days) during the RSV season to infants who qualify for prophylaxis in the first year of life. A child with a history of a severe allergic reaction following a dose of Palivizumab should not receive additional doses. 8. Qualified infants born during the RSV season must receive fewer doses according to their month of birth. For example, infants born in January would receive their last dose in March. 9. Palivizumab prophylaxis should not be administered in the second year of life except for children who require supplemental oxygen at 36 weeks corrected gestational age who also continue to require medical intervention (supplemental oxygen, chronic corticosteroid treatment, or diuretic therapy) in the six months prior to the second season. 10. Monthly prophylaxis should be discontinued in any child who experiences a breakthrough RSV hospitalization. 11. Palivizumab prophylaxis is not recommended for the prevention of health care-associated RSV disease. 12. Injection Palivizumab should be stored in a refrigerator at 2 °C–8 °C. 13. To reduce the risk for RSV and other viral infections, all infants, especially preterm infants, should be offered breast milk. The parents should be instructed to avoid smoke exposure, attendance at large group childcare during the first winter season and contact with ill people. 14. It is recommended that household members should be immunized against influenza and practice good hand and cough hygiene.

13. To reduce the risk for RSV and other viral infections, all infants, especially preterm infants, should be offered breast milk. The parents should be instructed to avoid smoke exposure, attendance at large group childcare during the first winter season and contact with ill people. 14. It is recommended that household members should be immunized against influenza and practice good hand and cough hygiene. 15. Palivizumab is not approved or recommended for the treatment of RSV disease. 16. Palivizumab does not interfere with routine childhood immunizations. 6 Procedure 6.1 Preparation of palivizumab (synagis) [5]. 1. Obtain Palivizumab Injection from the refrigerator and dilute the powder using water for injection. 2. Use 1 ml of water for injection of a 100 mg vial of Palivizumab, and 0.6 ml for a 50 mg vial, for a final concentration of 100 mg/ml. 3. Slowly add the water along the inside wall of the vial to minimize foaming. Tilt the vial slightly and gently rotate the vial for 30 s. DO NOT SHAKE THE VIAL. 4. Leave the Palivizumab solution at room temperature for a minimum of 20 min until the solution clarifies. 5. Because the Palivizumab does not contain any preservatives, it must be administered within 3 h of preparation. 6.2 Administration of palivizumab [5]. 1. Verify the patient identity and data. 2. Calculate the dose to be administered according to the weight. The dose should be 15 mg/kg. 3. For example:

4. Leave the Palivizumab solution at room temperature for a minimum of 20 min until the solution clarifies. 5. Because the Palivizumab does not contain any preservatives, it must be administered within 3 h of preparation. 6.2 Administration of palivizumab [5]. 1. Verify the patient identity and data. 2. Calculate the dose to be administered according to the weight. The dose should be 15 mg/kg. 3. For example: Weight×15mg100mg=−mltobeadministered 4. Obtain the required amount of injection and administer intramuscularly in the anterolateral aspect of the thigh using standard aseptic technique. The gluteal muscle is not preferred as an injection site because of the risk of sciatic nerve damage. 5. Administer the injection volumes over 1 ml as a divided dose. 6. Palivizumab is contraindicated in patients with hypersensitivity to the active substance or other humanized monoclonal antibodies. 7. Keep all equipment needed for the treatment of severe hypersensitivity reactions ready before the administration of prophylaxis. 8. Do not reconstitute Palivizumab with any other diluents or medicinal components. 9. Educate the mother regarding adverse effects such as fever, nervousness and diarrhea, which are common post-administration of prophylaxis. All side effects should be reported to the Saudi Food and Drug Authority. 10. Document the patient details and the date of administration in the RSV log book and card.

8. Do not reconstitute Palivizumab with any other diluents or medicinal components. 9. Educate the mother regarding adverse effects such as fever, nervousness and diarrhea, which are common post-administration of prophylaxis. All side effects should be reported to the Saudi Food and Drug Authority. 10. Document the patient details and the date of administration in the RSV log book and card. It should be noted that Palivizumab does not interfere with the immune response to other live or inactivated vaccines and the childhood immunization schedule should be followed for all children, regardless of Palivizumab use. Furthermore, more effort should be made to monitor data for the seasonality of RSV circulation and disease burden in various regions of Saudi Arabia and to fully investigate the burden of RSV among Saudi children. Conflicts of interest No conflicts of interest are reported. Ethical approval None. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Infections within any healthcare institution can be avoidable when dealt with appropriately. If infections are ignored or proactive strategies are not applied, healthcare-associated infections (HAIs) will result in patient morbidity, mortality, and additional resource use. Healthcare workers (HCWs), families, visitors and contractors are susceptible to health and safety issues when infection prevention & control (IP&C) measures do not incorporate an all-inclusive approach [1], [2]. HAIs, including device related, surgical site infections (SSIs), transmissible/infectious diseases or pathogens of concern, including viral, bacterial, fungal, and multidrug resistant organisms (MDROs), do not discriminate between age, gender, religion, or ethnicity. If stringent measures are not applied, the pediatric population is at equal, if not at higher, risk of infection or injury compared to adolescents and/or adults. The pediatric/child population, as defined in this paper, include those children between (0–14) years of age [3]. This includes pre-term and term newborns (less than 6 months of age), as well as infants, toddlers, children and pre-teens (6 months of age to 14 years). This paper does not directly discuss issues related to adolescents (15–18 years) or adults (18 years and older).

clude those children between (0–14) years of age [3]. This includes pre-term and term newborns (less than 6 months of age), as well as infants, toddlers, children and pre-teens (6 months of age to 14 years). This paper does not directly discuss issues related to adolescents (15–18 years) or adults (18 years and older). An extensive literature review was undertaken to identify key IP&C areas that target pediatric-specific populations. The aim was to identify those IP&C related measures that take into account a child's developmental age; physiological, psycho-social and immunological development; risk of HAIs by compromised natural defense mechanisms, including procedural, device type and length of utilization causes; availability of specific technologies and disciplines; and the role the caregiver in providing a safe and infection-free environment [4]. Caregivers in this paper are defined as either familial (parent, relative, or guardian) or HCWs who provide direct or indirect care.

al, device type and length of utilization causes; availability of specific technologies and disciplines; and the role the caregiver in providing a safe and infection-free environment [4]. Caregivers in this paper are defined as either familial (parent, relative, or guardian) or HCWs who provide direct or indirect care. The device-associated (DA) infection rates reported in the National Healthcare Safety Network (NHSN) – 2013 report suggested that neonates (well-baby, step-down and neonatal intensive care units) have a 1.13/1000 device-days (DDs) infection rate, while the combined rate for children admitted to wards in critical care, step-down, and receiving hematology/oncology and hematopoietic stem cell transplants is 1.46/1000 DD [5]. Combining the provided pediatric-specific rates in the NHSN report, the findings suggest that the highest DA infections occur among hematology/oncology (2.12/1000 DDs) and hematopoietic stem cell transplant (2.3/1000 DDs) patients, with a combined rate of 2.14/1000 DDs. This compares to 1.31/1000 DDs for all other pediatric areas combined.

d pediatric-specific rates in the NHSN report, the findings suggest that the highest DA infections occur among hematology/oncology (2.12/1000 DDs) and hematopoietic stem cell transplant (2.3/1000 DDs) patients, with a combined rate of 2.14/1000 DDs. This compares to 1.31/1000 DDs for all other pediatric areas combined. These rates support the observation that severely immunocompromised hospitalized patients are at the highest risk of DA HAIs. In addition, differences in infection rates, sites, and pathogen distributions have been noted to differ depending on susceptibility, immune status, age-group, and setting. A mortality rate of 3% has been attributed to blood stream infections (BSIs) within the pediatric population, with a mortality rate as high as 11% in neonates alone, particularly in those with very low-birth weights [5], [6], [7].

been noted to differ depending on susceptibility, immune status, age-group, and setting. A mortality rate of 3% has been attributed to blood stream infections (BSIs) within the pediatric population, with a mortality rate as high as 11% in neonates alone, particularly in those with very low-birth weights [5], [6], [7]. To address the high rate of infections and need for corrective measures, IP&C teams have evolved over the years and have become an essential component in all healthcare institutions. They provide a service to prevent, control, or reduce avoidable complications and infections, as well as promote safe practices for all patients and staff [8], [9], [10]. When issues are identified, the IP&C team intervenes to ensure that safe quality care is provided. Yet, IP&C strategies, for the most part, specifically target adult healthcare institutions [11]. Therefore, specialized programs need to be developed and tailored to meet the needs of the pediatric population, taking into account age-related factors that prevent the child from adhering to IP&C standards, as well as taking into account the impact that caregivers have on prevention and transmission. Without these specialized IP&C programs, there is a higher risk for developing HAIs due to developmental stages, social dependencies, differing diagnostic strategies, and immature immune systems [10], [12], [13].

C standards, as well as taking into account the impact that caregivers have on prevention and transmission. Without these specialized IP&C programs, there is a higher risk for developing HAIs due to developmental stages, social dependencies, differing diagnostic strategies, and immature immune systems [10], [12], [13]. This review article specifically evaluates IP&C and management practices for the pediatric population. We divided the health needs of the child into two categories: within-the-healthcare and outside-of-the-healthcare setting. Each setting has individual issues that have an impact on pathogen, disease or injury acquisition; modes of transmission; and implementation of IP&C strategies. Refer to Table 1 for a summary of risk and mitigation strategies.Table 1 Infection prevention & control risk mitigation strategies. Table 1 Category Risk Mitigation strategy References 1 Institutional regulations a. IP&C Program/policies/management support/evidence-based guidance b. Monitoring IP&C compliance c. KPIs for IP&C a. Robust program that meets institutional and governmental strategic goals based on the available best practice guidelines, recommendations and accredited bodies b. Monitoring of tailored IP&C policies that meet institutional and governmental goals c. Compliance monitoring of IP&C targeted processes (standard precautions, hand hygiene, transmission-based precautions) to identify practice gaps and implement corrective actions a. [1], [9] b. [1], [5], [16], [17], [18], [19] c. [5], [20], [21], [28] 2 Surveillance a. IP&C team b. Planning surveillance for the pediatric population

b. Monitoring of tailored IP&C policies that meet institutional and governmental goals c. Compliance monitoring of IP&C targeted processes (standard precautions, hand hygiene, transmission-based precautions) to identify practice gaps and implement corrective actions a. [1], [9] b. [1], [5], [16], [17], [18], [19] c. [5], [20], [21], [28] 2 Surveillance a. IP&C team b. Planning surveillance for the pediatric population c. Monitoring surveillance outcomes a. Having a qualified IP&C team to undertake surveillance activities to detect trends/outbreaks/risks associated with healthcare interventions, and a tailored surveillance plan developed by undertaking an institutional risk assessment b. Enhanced IP&C measures and durations to reduce environmental contamination c. Use of appropriate resources, audits and measures to promote adherence to IP&C policies/procedures, decision-making and feedback outcomes to stakeholders including administrators a. [1], [7], [9], [14], [22] b. [1], [2], [9], [14], [22], [23], [24], [25], [26] c. [7], [11], [28] 3 Immune deficiencies a. Environmental impact on pediatric population b. Hygiene and the immune deficient pediatric patient c. Developmental stages and exposure to childhood diseases a. Maintain quality of care, including appropriate hygiene measures (intrinsic & extrinsic), minimizing or preventing contact will ill persons, use of non-live vaccinations b. Utilizing a protective environment to minimize potential environmental exposures through appropriate air handling, water quality, environmental surfaces and cleaning practices

a. Maintain quality of care, including appropriate hygiene measures (intrinsic & extrinsic), minimizing or preventing contact will ill persons, use of non-live vaccinations b. Utilizing a protective environment to minimize potential environmental exposures through appropriate air handling, water quality, environmental surfaces and cleaning practices c. Early detection and initiation of transmission-based precautions (contact/droplet/airborne) a. [7], [14], [23], [26], [27], [28], [29], [30], [32], [33] b. [28], [32], [33], [34] c. [3], [7], [11], [28] 4 Caregivers a. As a source of infection b. Poor compliance with hand hygiene c. Chain of infection d. Family-centered care and knowledge a. Appropriate immunization for caregivers (HCW and family members), implementation of specific HCW policies, maximization of immune status to childhood preventable diseases, appropriate staffing numbers and education b. Education of all caregivers (technique/reason why/when to) and adherence and compliance to policy c. Monitoring/compliance of hand, personal and environmental hygiene d. Strict policies/procedure on the family caregiver's role in the prevention of infections while visiting/staying, monitoring and education of family caregivers when lapses are identified a. [7], [11], [14], [31], [35], [36], [37], [38] b. [3], [7], [16], [28], [38], [39], [40], [41] c. [16], [28] d. [7], [11], [14], [26], [28], [30], [35], [37], [42], [43], [44] 5 Intensive care units for the neonate and child a. HAIs and/or other complications b. Compliance a. Use bundles of care for VAP, CLABSI, CAUTI, and SSI (pediatric-specific)

a. [7], [11], [14], [31], [35], [36], [37], [38] b. [3], [7], [16], [28], [38], [39], [40], [41] c. [16], [28] d. [7], [11], [14], [26], [28], [30], [35], [37], [42], [43], [44] 5 Intensive care units for the neonate and child a. HAIs and/or other complications b. Compliance a. Use bundles of care for VAP, CLABSI, CAUTI, and SSI (pediatric-specific) b. Monitoring for bundle compliance and reinforce education a. [5], [27], [46], [47], [48] b. [14], [16], [49] 6 Pediatric inpatient and outpatient settings a. Lack of awareness of basic hygiene needs b. Environmental consideration a. Reliant on caregivers supervisory skills and knowledge for day-to-day care, including all hygiene needs and bodily functions, education and reinforcement required to ensure pediatric safety b. Limit toys/type of toys available to those that are cleanable (non-plush), give dedicated toys whenever possible, instigate cleaning of toys between patients which is documented and monitored a. [9], [14], [27], [50] b. [7], [27], [51], [52], [53], [54] 7 Respiratory infections a. Identification b. Vaccination a. Early and accurate identification of infectious patients incorporates early isolation, treatment, reduced spread using segregation of symptomatic patients/appropriate use of PPE b. Appropriate vaccination where applicable a. [11], [29], [31] b. 27 8 Gastrointestinal infections a. Hygiene b. Environmental contamination a. Reduce fecal-oral spread by implementing appropriate IP&C measures (hand hygiene especially before/after handling diapers/cleaning procedures/use of PPE/use of transmission-based precautions)

b. Appropriate vaccination where applicable a. [11], [29], [31] b. 27 8 Gastrointestinal infections a. Hygiene b. Environmental contamination a. Reduce fecal-oral spread by implementing appropriate IP&C measures (hand hygiene especially before/after handling diapers/cleaning procedures/use of PPE/use of transmission-based precautions) b. Strict use of IP&C practices (contact transmission-based precautions/hand hygiene) environmental cleaning (may include enhanced cleaning practices or the use of disinfectants) a. [7], [14], [16], [24] b. [7], [14], [24], [27] 9 Multidrug resistant organisms a. Controlling spread a. Use of preventative IP&C practices: hand hygiene, use of PPE, reducing and controlling environmental contamination, education of caregivers, decolonization and antimicrobial controls a. [9], [27], [47], [57], [58], [59] 10 Preventative and protective measures outside the healthcare setting a. Unsafe environment a. Prevention of injury and providing a safe environment includes: all aspects of safety (home/water/road/slips/trips/falls/preventing abusive situations) as well as reducing the risk of infection by maintain cleanliness of environment (including toys), reduce exposure to childhood diseases, limit contact with animals and providing appropriate age-related immunizations a. [7], [27], [28], [32], [54], [62], [66], [67], [68]

a. Prevention of injury and providing a safe environment includes: all aspects of safety (home/water/road/slips/trips/falls/preventing abusive situations) as well as reducing the risk of infection by maintain cleanliness of environment (including toys), reduce exposure to childhood diseases, limit contact with animals and providing appropriate age-related immunizations a. [7], [27], [28], [32], [54], [62], [66], [67], [68] 2 Discussion Children may not always display symptoms or have the ability to implement and understand strategies that are instigated for their own care, and therefore, diligent attention and supervision is required by caregivers. Ensuring family-centered care that incorporates appropriate hygiene, nutrition, and administration of vaccines (depending on strict age-defined schedules) provides a basis for ensuring growth, development, and an immune response that is age appropriate [7], [14]. The forms of care provided within and outside the healthcare setting can either be independent of each other or work together to maximize the health and safety of the child. Although there are some similarities in IP&C strategies, pediatric populations cannot be considered miniature adults [3], [15].

opriate [7], [14]. The forms of care provided within and outside the healthcare setting can either be independent of each other or work together to maximize the health and safety of the child. Although there are some similarities in IP&C strategies, pediatric populations cannot be considered miniature adults [3], [15]. 2.1 Within-the-healthcare setting 2.1.1 Institutional regulations An IP&C program geared toward the needs of the pediatric population has to be robust, encompass the needs of all age groups, be supported by higher administration, and meet institutional strategic goals. If successful, it will meet growing international demands for medical care and increase capacities, waiting lists, patient flow, and the need for specialized and experienced staff in the field of pediatrics as well as improve efficiency, decision-making, and collaboration. A robust program will be able to protect and improve staff development, recruitment, retention, and promote internal and external relations.

ties, waiting lists, patient flow, and the need for specialized and experienced staff in the field of pediatrics as well as improve efficiency, decision-making, and collaboration. A robust program will be able to protect and improve staff development, recruitment, retention, and promote internal and external relations. Without higher management oversight and support, an IP&C program cannot function, meet its objectives or ensure maximum compliance to policies and recommendations [9]. Policies and recommendations that are evidence-based and utilize well recognized national and international authorities and directives (such as governmental ministries, recognized associations e.g., US-Centers of Disease Control and Prevention [US-CDC], European Center for Disease Prevention and Control [ECDC], Department of Health: Disease Control and Prevention in Australia, World Health Organization [WHO]), as well as approved accreditation bodies (e.g., governmental and private), provide reputable and evidence-based information [1]. Although no specific regulations were found in the reviewed literature for inclusion in this paper, specific guidelines geared toward the pediatric population will, if implemented correctly, have institutional and community-wide benefits.

governmental and private), provide reputable and evidence-based information [1]. Although no specific regulations were found in the reviewed literature for inclusion in this paper, specific guidelines geared toward the pediatric population will, if implemented correctly, have institutional and community-wide benefits. Monitoring compliance and performance of HCWs is a well-reported strategy for preventing the transmission of HAIs [1], [16], [17], [18]. This enables actions to be undertaken against the institution's documented policies and/or procedures and identifies issues that can be easily recognized and rectified [1], [19]. Although monitoring compliance is a well-known concept in relation to HAIs (outcome indicators) [5], no specific key performance indicator(s) (KPI) for IP&C practices (process indicators) have been established and approved internationally [20], [21]. The authors believe that to ensure compliance with basic IP&C principles (target processes such as hand hygiene, standard precautions and transmission-based precautions), robust KPIs need to be monitored, focusing on targeted processes. The results and recommendations need to be reported to key stakeholders, including higher management, which will subsequently help to reduce HAIs by ensuring that all HCWs are held responsible and accountable for their practice.

ssion-based precautions), robust KPIs need to be monitored, focusing on targeted processes. The results and recommendations need to be reported to key stakeholders, including higher management, which will subsequently help to reduce HAIs by ensuring that all HCWs are held responsible and accountable for their practice. 2.1.2 Surveillance Undertaking surveillance in pediatrics patients, including DA, SSI, dialysis-events (DEs), rates of MDROs, gastrointestinal, and respiratory illnesses, is important to provide a basis for action and decision-making. Furthermore, surveillance facilitates support and vital resources to be channeled into appropriate areas. This includes staffing numbers and levels of experience, early detection and intervention in outbreak situations [7], environmental controls, adequate supplies, and programs for education. The lack of a functional surveillance program may ultimately result in missed trends and underdetermine the magnitude of HAIs and risks associated with healthcare procedures [22]. It is vital that each healthcare facility determines the type and frequency of surveillance required for their patient population(s). This can be achieved by undertaking an IP&C risk assessment [1], [9], [14].

n missed trends and underdetermine the magnitude of HAIs and risks associated with healthcare procedures [22]. It is vital that each healthcare facility determines the type and frequency of surveillance required for their patient population(s). This can be achieved by undertaking an IP&C risk assessment [1], [9], [14]. Surveillance, whether it is targeted, total-house, or a combination of the two, is of paramount importance for identifying institutional problems, including outbreaks, as well as for directing initiatives [22]. Outbreaks of infectious pathogens can result in patient morbidity/mortality, environmental contamination, and/or HCW absenteeism. Implementation of appropriate measures to minimize or prevent transmission is of paramount importance. It is well documented that pediatric and the immunocompromised shed viruses for longer periods of time [23], [24], [25], [26], [27]. This alters and increases the potential duration of the preventative measures required and risks involved. Cohorting can be an effective preventative strategy in selected circumstances. It may be required during outbreaks when there is limited resources and increased occupancy. Consultation with experts in the IP&C and Infectious Disease fields is vital to ensure appropriate utilization of resources in decision-making and promoting adherence with standard and transmission-based precautions (contact, droplet or airborne) [7], [11], [28].

when there is limited resources and increased occupancy. Consultation with experts in the IP&C and Infectious Disease fields is vital to ensure appropriate utilization of resources in decision-making and promoting adherence with standard and transmission-based precautions (contact, droplet or airborne) [7], [11], [28]. 2.1.3 Immune deficiencies Within-the-healthcare setting, patients are placed at increased risk based on their compromised or naive immune systems/diseases or due to neutropenia [23], [26]. Certain diseases increase the risk of infections within-the-hospital setting by their nature alone. Those diseases that cause immunodeficiency or under-developed immune systems include certain syndromes, cancers, transplants, certain lung diseases, immaturity, and/or very low-birth weights of the newborn [7], [23], [27], [29]. Maintaining quality of care for the immunocompromised population includes routine practices, such as hand, environmental and personal hygiene measures; nutrition; dental care; minimizing/preventing contact with ill persons; using non-live vaccines, among other basic measures [30], [31], [32]. A physical protective environment used with this patient population encompasses air, water, and structural factors that reduce pathogenic materials in the environment [7], [14], [30].

tion; dental care; minimizing/preventing contact with ill persons; using non-live vaccines, among other basic measures [30], [31], [32]. A physical protective environment used with this patient population encompasses air, water, and structural factors that reduce pathogenic materials in the environment [7], [14], [30]. Environmental protection consists of positive air pressure; air hepa-filtration with a sufficient number of air changes per hour (ACH); absence of plants, carpets or difficult to clean surfaces, and windows; and gaps that are well-sealed to external elements. There needs to be routine and scheduled maintenance, as well as cleaning with a regulatory environmental protection agency (EPA) approved disinfectant, to ensure that the environment and equipment is free, at least to a minimal degree, of dust and pathogens [28], [32], [33], [34]. This includes the cleaning of medical and non-medical equipment prior to use. Water sources and transport systems, such as taps, showers, and ice machines, need to have frequent and routine maintenance and be tested for waterborne pathogens [14], [28], [32], [33].

l degree, of dust and pathogens [28], [32], [33], [34]. This includes the cleaning of medical and non-medical equipment prior to use. Water sources and transport systems, such as taps, showers, and ice machines, need to have frequent and routine maintenance and be tested for waterborne pathogens [14], [28], [32], [33]. Whether it is a within-the-healthcare or outside-of-the-healthcare setting, the stages of development and exposure to childhood illnesses have an impact on a child's risk of acquiring an infection [3], [7], [11], [28]. Breaking the chain of infection by utilizing standard and transmission-based precautions that are geared toward specific pathogen transmissibility ensures that control measures are individually tailored. The key is early detection and initiation of isolation or segregation with strict adherence [11].

, [7], [11], [28]. Breaking the chain of infection by utilizing standard and transmission-based precautions that are geared toward specific pathogen transmissibility ensures that control measures are individually tailored. The key is early detection and initiation of isolation or segregation with strict adherence [11]. 2.1.4 Caregivers Caregivers themselves can be a source of infection for a child [35], [36]. Close physical contact by HCWs or family caregivers provide a medium for transmission [7], [11], [31], [35], [37]. Poor IP&C practices, as discussed above, can be compounded by understaffing, poor knowledge and/or compliance, and overcrowding [11], [14]. To reduce risk, caregivers should be appropriately immunized with available vaccines, and non-immune personnel to childhood diseases should not provide direct care to an infectious patient as they themselves are at risk of disease acquisition and can contribute to subsequent spread [29], [30], [31], [38]. This is in compliance with standard and transmission-based precautions, which are a well-researched practices that break the chain of infection [28]. Hand hygiene by all direct and indirect caregivers has clear documented support in the reduction of HAIs [16], [39], [40]. Ensuring that all caregivers have the knowledge, education, skills, and physical resources to perform appropriate hand hygiene techniques and to know in which instances they should be applied (such as the WHO 5 Moments of Hand Hygiene) is vital and well-researched [3], [7], [16], [38], [40], [41].

AIs [16], [39], [40]. Ensuring that all caregivers have the knowledge, education, skills, and physical resources to perform appropriate hand hygiene techniques and to know in which instances they should be applied (such as the WHO 5 Moments of Hand Hygiene) is vital and well-researched [3], [7], [16], [38], [40], [41]. The family is a core component and an active part of the care-team in family-centered healthcare. Knowledge and compliance by family caregivers may be a challenge, thus it is important to recognize them as a potential source for infection spread. There are documented cases of family members and/or visitors being the source of infection [30], [37]. Strict polices and/or guidelines are required in addition to education activities and materials for parents/guardians/relatives staying or visiting a child while in the healthcare setting [9], [11], [26], [42], [43]. As a solution, education and monitoring of family and visitors is required and needs to be strongly reinforced [26], [35], [44]. Instructions to abide by set rules and regulations to prevent or reduce the acquisition and spread of pathogens is required. This includes adherence to hand hygiene, respiratory and cough etiquette, reducing socialization with other parents/patients, limiting visiting numbers, and limiting contact with ill or symptomatic persons [7], [11], [14], [28], [42], [43], [44].

to prevent or reduce the acquisition and spread of pathogens is required. This includes adherence to hand hygiene, respiratory and cough etiquette, reducing socialization with other parents/patients, limiting visiting numbers, and limiting contact with ill or symptomatic persons [7], [11], [14], [28], [42], [43], [44]. 2.1.4.1 Intensive care units for the neonate and child Premature infants may have immature defense mechanisms (including skin, gastrointestinal systems, lungs, etc.) or be born with severe medical conditions. These predispose neonates to HAIs and/or complications [14], [45]. Invasive devices, treatments, supports, or new technologies used in neonatal intensive care units (NICU) have, in modern times, prolonged the life of these pre-term infants, but each treatment modality has consequences and risk factors [45]. They independently increase the risk of morbidity and mortality as well as potential acquisition of pathogens. For this reason, extensive research concentrating on birthweight as a risk factor has centered on infection rates within NICUs [5], [46]. Strategies to prevent HAIs within the NICU should include bundles of care that are specifically created for this patient population [47]. Device related infections in the neonatal population include central and peripheral lines, as well as ventilator associated pneumonia (VAP). Due to limited use of foley catheters in the NICU, there is no benchmark or data available for catheter-associated urinary tract infections (CAUTI), but DA urinary tract infections can be a concern in this high-risk patient population [5].

central and peripheral lines, as well as ventilator associated pneumonia (VAP). Due to limited use of foley catheters in the NICU, there is no benchmark or data available for catheter-associated urinary tract infections (CAUTI), but DA urinary tract infections can be a concern in this high-risk patient population [5]. What separates pediatric intensive care units (PICUs) from NICUs is that the HCWs in a PICU need to be able to manage a variety of childhood and infectious diseases. Children of all ages are admitted into a PICU for multiple reasons, which include both immune-competent and immunocompromised statuses [27], [48]. Most, if not all, patients in PICUs have invasive devices inserted or have undergone invasive procedures (e.g., surgical) that require preventative measures. There needs to be individualized and unique considerations taken into account for the pediatric population, such as developmental age, levels of mobility, understanding, and appropriate products that are age-related, i.e., chlorhexidine gluconate (CHG) [27]. Adherence to bundles of care for central lines, VAP, CAUTI and SSI is paramount [5], [27], [46]. Each bundle component is vitally important and encompasses education and compliance [14], [49]. It is often difficult to determine which element has the greatest influence, and it would be unethical to undertake a study to evaluate this observation.

re for central lines, VAP, CAUTI and SSI is paramount [5], [27], [46]. Each bundle component is vitally important and encompasses education and compliance [14], [49]. It is often difficult to determine which element has the greatest influence, and it would be unethical to undertake a study to evaluate this observation. 2.1.5 Pediatric inpatient and outpatient clinic settings The younger the child, the less educated or aware and compliant to basic hygiene practices they are. This relates to both physical and emotional stages of growth. Basic hygiene practices refer to hand, respiratory, body, clothing and environmental cleanliness. Young infants and children have a tendency to drool, have incontinence, and undertake frequent mouthing of hands and objects regardless of their level of cleanliness [14]. These children are reliant on the adult caregiver's supervisory skills to ensure that all required safety, quality, and IP&C strategies are applied in their day-to-day care [27]. This includes feeding (i.e., food quality, quantity and preparation), toilet use and diapering, bathing and hand washing, and sharing of personal items, such as combs, brushes, coats, hats, toys, and shoes [9], [50]. Older children are more able to care for themselves to a certain degree and make conscious decisions regarding their care, though they are still reliant on adults to guide them and ensure their ultimate safety.

hand washing, and sharing of personal items, such as combs, brushes, coats, hats, toys, and shoes [9], [50]. Older children are more able to care for themselves to a certain degree and make conscious decisions regarding their care, though they are still reliant on adults to guide them and ensure their ultimate safety. Environmental considerations in inpatient and outpatient settings often include the use of toys, but toys can be a vehicle for pathogen transmission [27]. The use of toys within healthcare include therapeutic, recreational and educational purposes [7], [27]; use in waiting areas and play rooms or classrooms [51], [52], [53]. Of particular concern are toys that cannot be cleaned easily, are non-immersible, or due to/or lack of internal processes, are not on a cleaning schedule. Research supports that within healthcare settings, plush (soft) toys that are shared are of a higher risk for vector transmission [53], [54]. Strategies must include the use of non-plush toys (unless new and dedicated), dedicated toys where possible if they cannot be cleaned easily, cleaning between patients, or applying a strict cleaning regime that is documented and monitored [27], [53], [54].

shared are of a higher risk for vector transmission [53], [54]. Strategies must include the use of non-plush toys (unless new and dedicated), dedicated toys where possible if they cannot be cleaned easily, cleaning between patients, or applying a strict cleaning regime that is documented and monitored [27], [53], [54]. 2.1.6 Respiratory infections Respiratory infections, both upper and lower, account for the most common-occurring pediatric illness of infectious nature [29], [55]. Viral infections include influenza, respiratory syncytial virus (RSV), rhinovirus, parainfluenza, human meta-pneumovirus, adenovirus, bocavirus, and coronaviruses. Identification of contagious pathogens that cause respiratory infections has improved with diagnostic and rapid testing methods and, subsequently, has had an impact on early identification, isolation, and treatment [11]. Early and accurate identification of symptomatic children results in targeting early isolation, preventative measures, and improved compliance practices to reduce the risk of spread. This includes segregation, use of personal protective equipment (PPE), and promotion of available vaccines. Early identification of pathogens, implementation of appropriate droplet and contact precautions, and strict environmental and equipment cleaning prevent further transmission and potential outbreaks from occurring [29], [31].

segregation, use of personal protective equipment (PPE), and promotion of available vaccines. Early identification of pathogens, implementation of appropriate droplet and contact precautions, and strict environmental and equipment cleaning prevent further transmission and potential outbreaks from occurring [29], [31]. Vaccination plays a strong role in preventing respiratory and other serious infections in the pediatric population. Since the introduction of Haemophilus influenza type B, Streptococcus pneumonia, pertussis, and Neisseria meningitides, the rates of infections due to these diseases have declined significantly [27].

segregation, use of personal protective equipment (PPE), and promotion of available vaccines. Early identification of pathogens, implementation of appropriate droplet and contact precautions, and strict environmental and equipment cleaning prevent further transmission and potential outbreaks from occurring [29], [31]. Vaccination plays a strong role in preventing respiratory and other serious infections in the pediatric population. Since the introduction of Haemophilus influenza type B, Streptococcus pneumonia, pertussis, and Neisseria meningitides, the rates of infections due to these diseases have declined significantly [27]. 2.1.7 Gastrointestinal infections Gastrointestinal or diarrheal illnesses of a viral nature are often referred to as enteric infections and are common among the pediatric population. Enteric infections that commonly occur in children include Norovirus, enteric Adenovirus serotypes, Rotavirus, Enterovirus, and Clostridium difficile [7]. Non-viral enteric organisms of concern in children include MDROs, such as vancomycin resistant enterococcus (VRE); bacteria, such as salmonella and shigellosis; and parasites, such as cryptosporidium and pin-worm. These are often the source of fecal-oral spread due to poor hygienic measures among children and/or caregivers [14]. Prevention for this group of infectious pathogens heavily relies on the education of caregivers and, if possible, the child on key measures, such as hand hygiene, handling of soiled items (diaper, clothing, and equipment, etc.), and environmental cleaning with disinfectants to remove the presence of organisms [7], [24].

revention for this group of infectious pathogens heavily relies on the education of caregivers and, if possible, the child on key measures, such as hand hygiene, handling of soiled items (diaper, clothing, and equipment, etc.), and environmental cleaning with disinfectants to remove the presence of organisms [7], [24]. These and other organisms have the ability to remain viable in the environment for varying lengths of time, resulting in the occurrence of spread and, in some cases, extensive outbreaks [14], [27]. When suspected or confirmed utilization of contact precautions is advisable, the patient should be in a single room if possible or, alternatively, cohort cases should room together upon advisement by an IP&C team. This is in addition to strict adherence to hand and environmental hygiene. In instances of outbreak, consultation and implementation of mitigating strategies to contain and resolve the situation is required.

a single room if possible or, alternatively, cohort cases should room together upon advisement by an IP&C team. This is in addition to strict adherence to hand and environmental hygiene. In instances of outbreak, consultation and implementation of mitigating strategies to contain and resolve the situation is required. 2.1.8 Multidrug resistant organisms The importance and impact of MDROs cannot be under-estimated. These organisms pose a threat due to limited treatment options to first-line therapies [3]. There are strong concerns regarding the spread of MDROs within-the-healthcare setting. The pathogens of concern include methicillin resistant Staphylococcus aureus (MRSA) and VRE [27]. The concerns within-the-healthcare setting are in addition to the known occurrence of spread within the community, schools, and sports facilities [56]. At present, there is limited research about the attributable outcomes and effects of Gram-negative MDROs (in particular, beta-lactam resistant Gram-negative bacilli/extended-spectrum beta-lactamases [ESBLs] and carbapenem-resistant enterobacteriaceae [CRE]) within the pediatric population. This includes the areas of morbidity, mortality, length of stay (LOS), burden and cost to healthcare services [57].

me environment; review of policies governing visitors within-the-healthcare setting; impact of viral shedding and need for resampling for clearance purposes, especially among the immunocompromised; and need for studies and guideline development on MDROs, especially ESBL and CRE, in relation to the pediatric population. Ultimately, it is the caregiver's responsibility to oversee and ensure that safe care practices are being followed. This can be achieved by balancing and adhering to IP&C strategies within-the-healthcare and outside-of-the-healthcare setting. Each setting has the potential to influence and/or impact the other. As discussed in this paper, there is a comprehensive need for a proactive approach by all caregivers, oversight by an IP&C team, and strong support from management and other key stakeholders. Conflicts of interest The authors have no conflict of interest to report. Acknowledgments Mr. Keith D. Barron for his time and effort to assist in the article review. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Pott's puffy tumor (PPT) is a condition that is characterized by frontal bone osteomyelitis, which may be associated with sub periosteal abscess [1]. An English surgeon, Sir Percival Pott, first described Pott's puffy tumor in 1760 and explained it in his own words as “a puffy, circumscribed, indolent tumor of the scalp, and a spontaneous separation of the pericranium from the skull under such a tumor” [2]. Sir Pott initially thought that trauma was the only cause for this condition, but he later discovered that frontal sinus infection is the leading cause of PPT. Due to the wide use of broad spectrum antibiotics, its incidence has dramatically decreased [3], but the exact incidence is unknown. However, Clayman et al found that PPT may be seen in up to 0.5% of cases of acute and chronic sinusitis [4], [5]. The infection in PPT typically spreads from the frontal sinus into the frontal bone via direct extension or through septic emboli [3].

1 Introduction Pott's puffy tumor (PPT) is a condition that is characterized by frontal bone osteomyelitis, which may be associated with sub periosteal abscess [1]. An English surgeon, Sir Percival Pott, first described Pott's puffy tumor in 1760 and explained it in his own words as “a puffy, circumscribed, indolent tumor of the scalp, and a spontaneous separation of the pericranium from the skull under such a tumor” [2]. Sir Pott initially thought that trauma was the only cause for this condition, but he later discovered that frontal sinus infection is the leading cause of PPT. Due to the wide use of broad spectrum antibiotics, its incidence has dramatically decreased [3], but the exact incidence is unknown. However, Clayman et al found that PPT may be seen in up to 0.5% of cases of acute and chronic sinusitis [4], [5]. The infection in PPT typically spreads from the frontal sinus into the frontal bone via direct extension or through septic emboli [3]. PPT can affect all age groups and has a male: female ratio of 9:1. It is noted that adolescents are affected more frequently. This is thought to be due to high vascularity of the diploic venous system, and the accelerated growth of the frontal bone at this particular age [6], [7], [8]. The most common factors leading to PPT are frontal sinusitis and trauma [9]. Other rare etiologies include hematogenous spread of sinusitis, retrograde thrombophlebitis through the diploic veins, or even an insect bite [10], [11]. Signs and symptoms of PPT include forehead swelling and frontal headaches with or without fever. These clinical manifestations usually progress slowly, however, rapid progression may suggest the development of complications [12]. The most common cultured organisms are the same organisms causing chronic sinusitis: streptococci, staphylococci, and anaerobes [13].

swelling and frontal headaches with or without fever. These clinical manifestations usually progress slowly, however, rapid progression may suggest the development of complications [12]. The most common cultured organisms are the same organisms causing chronic sinusitis: streptococci, staphylococci, and anaerobes [13]. PPT is a serious condition because it is associated with high rate of intracranial complications such as meningitis, subdural and epidural empyema, cerebral abscess, and cavernous sinus thrombosis or superior sagittal sinus thrombosis [14], [15]. The diagnostic modality is the contrast enhanced CT scan of the brain and sinuses. The most important aspect in the management of PPT is early diagnosis, early use of proper antimicrobial therapy, and urgent surgical intervention to prevent serious complications [16], [17], [18]. 2 Case report A previously healthy 9-year-old girl presented with a one-month history of intermittent moderate frontal headaches after head trauma. She stated: “A swing fell over her forehead”. She subsequently developed forehead swelling, which was later diagnosed as frontal sinusitis. She made a complete recovery after a course of oral antibiotic. Two weeks later, the swelling reappeared, and she was treated with a course of both IV and oral antibiotics but without significant improvement. She denied any history of fever, nasal discharge, or cough.

ng, which was later diagnosed as frontal sinusitis. She made a complete recovery after a course of oral antibiotic. Two weeks later, the swelling reappeared, and she was treated with a course of both IV and oral antibiotics but without significant improvement. She denied any history of fever, nasal discharge, or cough. Upon examination, she had an undulating erythematous firm swelling over the right frontal area with mild right periorbital swelling (Figure 1, Figure 2). Normal pupillary reflexes and extra-ocular eye movements were intact. There were no meningeal signs that were elicited, and no neurological deficits were present.Figure 1 Frontal view of fluctuant firm swelling over the right frontal area with mild right periorbital swelling. Figure 1Figure 2 Lateral view of Fluctuant firm swelling over the right frontal area with mild right periorbital swelling. Figure 2 Laboratory tests showed a normal CBC and normal inflammatory markers. Brain imaging including a CT scan and MRI showed signs of frontal bone osteomyelitis with sequestrum formation and abnormal communication between the two frontal sinuses, both of which were filled with fluid (Fig. 3).Figure 3 Computed tomography of the brain (CT scan) showing frontal bone osteomyelitis with sequestrum formation and abnormal communication between the two frontal sinuses, both filled with fluid. Figure 3 A multidisciplinary approach was conducted that involved pediatric infectious diseases, ENT and neurosurgery. The patient was empirically treated with broad spectrum intravenous (IV) antibiotics Clindamycin and ceftriaxone.

Brain imaging including a CT scan and MRI showed signs of frontal bone osteomyelitis with sequestrum formation and abnormal communication between the two frontal sinuses, both of which were filled with fluid (Fig. 3).Figure 3 Computed tomography of the brain (CT scan) showing frontal bone osteomyelitis with sequestrum formation and abnormal communication between the two frontal sinuses, both filled with fluid. Figure 3 A multidisciplinary approach was conducted that involved pediatric infectious diseases, ENT and neurosurgery. The patient was empirically treated with broad spectrum intravenous (IV) antibiotics Clindamycin and ceftriaxone. Urgent surgical intervention was done by functional endoscopic sinus surgery (FESS) and a craniotomy with a flap to repair the defect (Fig. 4).Figure 4 Post functional endoscopic sinus surgery (FESS) and craniotomy with flap to repair the defect. Figure 4 Sterile culture from the frontal bone, which was taken intra-operatively, grew Aspergillus fumigatus. Bone histopathology showed fungal hyphae. Voriconazole was initiated with a high dose (9 mg/kg/dose) every 12 h. She was kept on the voriconazole for 6 months because of the chronic nature of the osteomyelitis. Laboratory tests were used to closely monitor for signs of voriconazole toxicity. Fortunately, there was no toxicity detected. She showed significant improvement with complete clinical and radiological resolution at the end of her six-month treatment.

ole for 6 months because of the chronic nature of the osteomyelitis. Laboratory tests were used to closely monitor for signs of voriconazole toxicity. Fortunately, there was no toxicity detected. She showed significant improvement with complete clinical and radiological resolution at the end of her six-month treatment. 3 Discussion Pott's puffy tumor is a very rare condition and nothing from previously reported cases detailed fungal species as a causative agent. However, the most commonly cultured organisms in PPT are Streptococci, mainly Streptococcus milleri which was found in approximately 50% of the reported cases, Staphylococci, anaerobes, namely Fusobacterium and Bacteroides, and less commonly Proteus and Pseudomonas [11], [19], [20], [21], [22], [23]. In our case, the surgical culture from the frontal bone grew A. fumigatus, which is abnormal to have grown as a fungal organism in an immunocompetent individual. In regards to diagnosis, computed tomography (CT) of the brain is the best diagnostic modality in such cases. It helps in the critical assessment of the sinuses as well as gauging the extent of the disease to rule out any intracranial complications [24], [25]. In our case, a brain CT scan showed signs of frontal bone osteomyelitis with sequestrum formation, which indicated a chronic infection, and an abnormal communication between the two frontal sinuses, which were both filled with fluid, which could be a posttraumatic occurrence.

any intracranial complications [24], [25]. In our case, a brain CT scan showed signs of frontal bone osteomyelitis with sequestrum formation, which indicated a chronic infection, and an abnormal communication between the two frontal sinuses, which were both filled with fluid, which could be a posttraumatic occurrence. Pott's puffy tumor is considered to be an emergency medical condition, and broad spectrum antibiotics should be initiated as soon as possible along with urgent surgical intervention which may include: sinus wash-outs, percutaneous aspiration, trepanation, endoscopic sinus surgery, or craniotomy [23]. Functional endoscopic sinus surgery is a safe and less invasive surgical procedure [26], [27]. The decision of whether or not a craniotomy is indicated depends on the size of the intracranial fluid collection [28]. In our case, a combined surgical approach was coordinated by ENT and neurosurgery. FEES along with craniotomy, aspiration of the sub periosteal collection, sequestrectomy, and frontal bone debridement were all conducted at the same time. Furthermore, antimicrobial therapy should be tailored according to the cultured organism [29]. In our case, we started voriconazole which is an antifungal agent that is known to be fungicidal against most of the Aspergillus species. Furthermore, a high dose of voriconazole is the best regimen for such condition [30].

In our case, a combined surgical approach was coordinated by ENT and neurosurgery. FEES along with craniotomy, aspiration of the sub periosteal collection, sequestrectomy, and frontal bone debridement were all conducted at the same time. Furthermore, antimicrobial therapy should be tailored according to the cultured organism [29]. In our case, we started voriconazole which is an antifungal agent that is known to be fungicidal against most of the Aspergillus species. Furthermore, a high dose of voriconazole is the best regimen for such condition [30]. We conclude that although PPT is rare, it still must be part of the differential diagnosis of forehead swelling and headache, especially if it was preceded by sinusitis or trauma. Broad spectrum antimicrobial therapy should be initiated immediately after surgical intervention. Adequate sampling should be attained for cultures, and serious consideration should be given to the possibility of fungal infections. Conflict of interest The authors declare that they have no competing interests. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Mesenteric cysts are rare intra-abdominal benign masses. Their incidence is estimated to be approximately 1/20,000 in the pediatric population [1]. More than half of mesenteric cysts involve the mesentery of the terminal ileum [2]. Mesenteric cysts usually present as an asymptomatic lump. Acute presentations are uncommon. The absence of obvious pathognomonic symptoms and signs makes diagnosis difficult. Abdominal ultrasound and computed tomography may aid in diagnosis. Complete surgical excision is the treatment of choice; the alternative treatment is excision or marsupialization of the cyst [1]. We describe three children with mesenteric cysts who presented with acute intestinal obstruction. 2 Patients and methods Of a total of 6 patients diagnosed with mesenteric cysts, 3 patients presented with symptoms and signs of acute intestinal obstruction (50%).

1 Introduction Mesenteric cysts are rare intra-abdominal benign masses. Their incidence is estimated to be approximately 1/20,000 in the pediatric population [1]. More than half of mesenteric cysts involve the mesentery of the terminal ileum [2]. Mesenteric cysts usually present as an asymptomatic lump. Acute presentations are uncommon. The absence of obvious pathognomonic symptoms and signs makes diagnosis difficult. Abdominal ultrasound and computed tomography may aid in diagnosis. Complete surgical excision is the treatment of choice; the alternative treatment is excision or marsupialization of the cyst [1]. We describe three children with mesenteric cysts who presented with acute intestinal obstruction. 2 Patients and methods Of a total of 6 patients diagnosed with mesenteric cysts, 3 patients presented with symptoms and signs of acute intestinal obstruction (50%). Case 1: A 4-year-old male presented with a 5-day history of abdominal pain and distention, fever, and intermittent episodes of non-bilious vomiting. On admission, the abdomen appeared distended and the patient reported generalized tenderness. No lump was palpable. Erect abdominal X-ray and abdominal ultrasound suggested the presence of a small bowel obstruction (Fig. 1). During emergency laparotomy by supra-umbilical right transverse incision, a mesenteric cyst of 10 × 6 cm in size containing serous fluid was uncovered between the leaves of the mesentery of the terminal ileum (Fig. 2). The small bowel proximal to the cyst was grossly distended. The cyst was deroofed, and the patency of the bowel was confirmed. Histopathology confirmed the diagnosis of a mesenteric cyst.Figure 1 X-ray of the patient revealing the intestinal obstruction.

tween the leaves of the mesentery of the terminal ileum (Fig. 2). The small bowel proximal to the cyst was grossly distended. The cyst was deroofed, and the patency of the bowel was confirmed. Histopathology confirmed the diagnosis of a mesenteric cyst.Figure 1 X-ray of the patient revealing the intestinal obstruction. Figure 2 Intra-operative image demonstrating a mesenteric cyst with serous fluid. Case 2: A 1.5-year-old male child was referred with complaints of abdominal pain, distention, constipation, and intermittent episodes of bilious vomiting for 3 days. Abdominal distention was apparent, and the patient presented with mild tenderness all over the abdomen. No lump could be felt. The rectum was empty on digital rectal examination. Blood tests revealed the presence of anemia. Erect X-ray of the abdomen and abdominal ultrasound suggested the presence of a small bowel obstruction. During emergency exploratory laparotomy by supra-umbilical right transverse incision, two mesenteric cysts 8 × 7 cm in size containing chylous fluid were detected in the mesentery of the small bowel approximately 20 cm and 30 cm distal to the duodeno-jejunal flexure. The proximal small bowel was dilated. The cysts were marsupialized, and excess cyst wall was excised. Histopathology confirmed the diagnosis of mesenteric cyst.

× 7 cm in size containing chylous fluid were detected in the mesentery of the small bowel approximately 20 cm and 30 cm distal to the duodeno-jejunal flexure. The proximal small bowel was dilated. The cysts were marsupialized, and excess cyst wall was excised. Histopathology confirmed the diagnosis of mesenteric cyst. Case 3: A 5-day-old female neonate presented with abdominal distention and multiple episodes of non-bilious vomiting for 2 days. Her abdomen was massively distended. The rectum was empty on digital rectal examination. Abdominal X-ray revealed multiple air fluid levels, and ultrasound suggested dilated small bowel loops with to and fro movements indicative of obstruction. During emergency surgical exploration by supra-umbilical right transverse incision, a mesenteric cyst 8 × 8 cm in size was detected at the terminal ileum. The cyst was completely excised. Post-operative recovery was uneventful. The diagnosis of mesenteric cyst was confirmed by histopathology. 3 Discussion Benivieni, an anatomist, first reported on the mesenteric cyst in 1507 [1]. Mesenteric cysts are rare intra-abdominal masses with an incidence of approximately 1:100,000 in adults and 1:20,000 in the pediatric population [1]. They may occur anywhere in the mesentery of the gastrointestinal tract from the duodenum to the rectum, but they are most commonly located in the mesentery of the ileum followed by localization in the sigmoid mesocolon [1].

an incidence of approximately 1:100,000 in adults and 1:20,000 in the pediatric population [1]. They may occur anywhere in the mesentery of the gastrointestinal tract from the duodenum to the rectum, but they are most commonly located in the mesentery of the ileum followed by localization in the sigmoid mesocolon [1]. Approximately one-third of mesenteric cysts occur in children younger than 15 years of age and are slightly more common in males [1]. The size of mesenteric cysts may vary from 2 cm to 35 cm [2]. In the current study, the three cysts that were associated with obstructive symptoms and signs were 8 cm or more in size. The other three cysts were smaller. The exact etiology of the development of mesenteric cysts is unknown. The most commonly accepted theory as proposed by Gross states that cysts result from benign proliferation of ectopic lymphatic tissue in the mesentery that lacks communication with the remainder of the lymphatic system [1].

Approximately one-third of mesenteric cysts occur in children younger than 15 years of age and are slightly more common in males [1]. The size of mesenteric cysts may vary from 2 cm to 35 cm [2]. In the current study, the three cysts that were associated with obstructive symptoms and signs were 8 cm or more in size. The other three cysts were smaller. The exact etiology of the development of mesenteric cysts is unknown. The most commonly accepted theory as proposed by Gross states that cysts result from benign proliferation of ectopic lymphatic tissue in the mesentery that lacks communication with the remainder of the lymphatic system [1]. Patients are usually asymptomatic unless complications arise. Symptoms are variable and are related to the size and the position of the cyst, with no pathognomonic signs observed in uncomplicated patients. Patients may also present with acute symptoms secondary to complications such as obstruction, rupture, hemorrhage into a cyst, infection, or abscess formation. Intestinal obstruction is a surgical emergency and requires urgent attention. It is usually produced by compression of the intestine adjacent to the cyst, volvulus or entrapment in the pelvis [3]. In a report of 17 cases of mesenteric cysts by Prakash A et al, nine patients presented with acute small intestinal obstruction [4].

l obstruction is a surgical emergency and requires urgent attention. It is usually produced by compression of the intestine adjacent to the cyst, volvulus or entrapment in the pelvis [3]. In a report of 17 cases of mesenteric cysts by Prakash A et al, nine patients presented with acute small intestinal obstruction [4]. Abdominal ultrasound and computed tomography are the methods of choice to diagnose mesenteric cysts pre-operatively [3]. However, the confirmation of this diagnosis may not be possible in all cases. The differential diagnosis of mesenteric cysts includes cystic lymphangioma, cystic teratoma, ovarian cyst, enteric duplication cyst, and hydatid cyst [5]. Mesenteric cysts are most commonly single and multilocular, and the cystic fluid is generally serous when the cyst involves the distal small bowel or colonic mesentery; cystic fluid is usually chylous when the cyst is located in the proximal small bowel mesentery [1]. Histopathological examination can confirm these observations [5]. Complete surgical excision is the preferred treatment for mesenteric cysts and leads to excellent outcomes. Surgery can be performed laparoscopically [6]. Bowel resection and anastomosis may be required along with excision of the mesenteric cysts [7]. In children, resection and anastomosis may be required in up to 50–60% of cases [1]. Rattan KN et al reported 8 cases of mesenteric cysts, two of which presented with intestinal obstruction, both were treated by exploratory laparotomy, and complete excision of the cyst required bowel resection [8].

esenteric cysts [7]. In children, resection and anastomosis may be required in up to 50–60% of cases [1]. Rattan KN et al reported 8 cases of mesenteric cysts, two of which presented with intestinal obstruction, both were treated by exploratory laparotomy, and complete excision of the cyst required bowel resection [8]. If enucleation or resection is not possible, then partial excision with marsupialization of the remaining cyst into the abdominal cavity is recommended [1]. The cyst lining could be sclerosed with electrocautery, tincture iodine, or OK432 after marsupialization to prevent recurrence [1]. 4 Conclusion Mesenteric cysts, though invariably asymptomatic, can occasionally be associated with an acute presentation. Mesenteric cysts can rarely cause acute intestinal obstruction in children, especially cysts that are larger in size. Surgical excision of the mesenteric cyst is the mainstay of treatment. Resection and anastomosis of the bowel may be required depending on individual patient anatomy. Ethical clearance Not applicable in this because the patients in this study were managed according to the existing protocols. No new investigations and intervention was done on the patient. Source of funding Nil. Conflict of interest The authors have no conflicts of interest relevant to this article to disclose. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Human immunodeficiency virus (HIV) in the adolescent community is an increasing concern. Globally, there are more than 2 million adolescents living with HIV [1]. Adolescents often engage in high-risk sexual behavior, and individuals aged 15–19 have the highest reported rates of sexually transmitted infections (STIs). Among high school students in 2013, 46.8% have had sexual intercourse, 34% were currently sexually active, and 48.9% of the currently sexually active students did not use a condom the last time they had sexual intercourse. Nationwide, while 85.3% of students had been taught about acquired immune deficiency syndrome (AIDS) or HIV infection in school, only 12.9% of students had ever been tested for HIV [2]. Centers for Disease Control and Prevention (CDC) estimates the 9961 youth aged 13–24 years who were diagnosed with HIV infections in 2013 represents 21% of the total diagnoses that year. Around the same time, there were an estimated 62,400 youth living with HIV infection, with over half undiagnosed [3]. In sub-Saharan Africa, only 10% of young men and 15% of young women aged 15–24 were aware of their HIV status. The World Health Organization (WHO) estimates that adolescent deaths from HIV are rising, which is assumed to be from the lack of support and care that they receive after childhood [1].

r half undiagnosed [3]. In sub-Saharan Africa, only 10% of young men and 15% of young women aged 15–24 were aware of their HIV status. The World Health Organization (WHO) estimates that adolescent deaths from HIV are rising, which is assumed to be from the lack of support and care that they receive after childhood [1]. During this time, adolescents are facing challenges with their own physical changes, brain development, and social milestones. This period of transition significantly impacts an adolescent's ability to take ownership of their health and successfully enter adult-centered care. In the 2013 CDC HIV Surveillance Report, the lowest percentage of HIV patients to establish linkage of care by age group was persons aged 13–24 years (73.4%) and the lowest percentage to be retained in HIV medical care was persons aged 25–34 years (46.7%) [4]. While many papers and medical organizations recognize the importance of capturing this patient population and preparing them for adulthood, very little has been published regarding outcome measurements of chronic diseases within this phase of adolescent transition. Several programs are being developed to address this very fragile period for young adult health, but understanding the current outcomes in chronic disease during adolescence is necessary for understanding the effectiveness of any transitional care program.

of chronic diseases within this phase of adolescent transition. Several programs are being developed to address this very fragile period for young adult health, but understanding the current outcomes in chronic disease during adolescence is necessary for understanding the effectiveness of any transitional care program. By reviewing the HIV Viral Load (VL) data from University Hospital in Newark, New Jersey, we assessed the trends in adolescent HIV VLs and virologic control in an urban community with a high HIV prevalence. Knowledge of these trends will allow physicians around the world to better recognize the challenges of transitioning young adults with HIV to adult-centered care and identify areas to target in the development of appropriate transitional care programs.

virologic control in an urban community with a high HIV prevalence. Knowledge of these trends will allow physicians around the world to better recognize the challenges of transitioning young adults with HIV to adult-centered care and identify areas to target in the development of appropriate transitional care programs. 2 Patients and methods All HIV RNA VLs from the Molecular Virology Laboratory at the University Hospital in Newark, New Jersey, from 2007 to 2010 were obtained. The study received expedited review and approval by the Rutgers New Jersey Medical School Institutional Review Board. Patients were divided into pediatric (age <13 years), adolescent (age 13–25 years), and adult (age >25 years) groups, all of whom were receiving treatment at University Hospital's HIV clinic. The clinic is located in the Francois-Xavier Bagnoud Center and is funded in part by the Ryan White Grant Program. Through social services, the clinic provides access to medication assistance programs, housing programs, and mental healthcare to all patients at point-of-care. Clinic staff consists of two board-certified pediatric infectious diseases physicians, two nurses, one advanced practice nurse, one psychologist, and two social workers. Patients are typically scheduled for 3-month follow-up visits; however, those patients requiring more intense therapy are given more frequent appointments. Most patients were retained in the clinic and were not transitioned out. Over 95% of pediatric patients were infected prenatally and many have grown up attending the clinic. During the time period of the study, all were offered highly active antiretroviral therapy (HAART) with the majority receiving protease inhibitors. Some patients were receiving complex multi-tablet regimes due to multiple past virologic failures with 3-class resistance.

ally and many have grown up attending the clinic. During the time period of the study, all were offered highly active antiretroviral therapy (HAART) with the majority receiving protease inhibitors. Some patients were receiving complex multi-tablet regimes due to multiple past virologic failures with 3-class resistance. The mean viral load was calculated as the average of each individual's viral load tests over a calendar year. Undetectable viral loads were assigned a value of half the lower limit of detection, and high viral loads were assigned the maximum limit of detection. Undetectable viral loads were defined as <400 copies/mL, which was the most frequent lower limit of detection used in 2001–2008. Patients were considered undetectable if they averaged an undetectable viral load over the calendar year. Univariate and multivariate analyses were performed to assess characteristics of patients by age and gender. 3 Results A minimum of 40 pediatric, 178 adolescent, and 1335 adult patients were identified per year. There were no statistically significant differences in the mean ages of the three groups over the four-year time period. Males comprised 37–47% of the pediatric patients, 48–57% of the adolescent patients, and 54–57% of the adults. Over 95% of the clinic's HIV patients were prenatally infected, and over 90% were actively accepting HAART.

3 Results A minimum of 40 pediatric, 178 adolescent, and 1335 adult patients were identified per year. There were no statistically significant differences in the mean ages of the three groups over the four-year time period. Males comprised 37–47% of the pediatric patients, 48–57% of the adolescent patients, and 54–57% of the adults. Over 95% of the clinic's HIV patients were prenatally infected, and over 90% were actively accepting HAART. Based on October 2010 performance measures, 75% of patients had two clinic appointments at least 3 months apart, 66% of patients had no less than two CD4 count measurements, 95% of patients with CD4 counts <200 received PCP prophylaxis, 91% of patients with AIDS received HAART, 67% received syphilis screening, and 81% received tuberculosis screening. Lower performance rates were recorded for cervical cancer screening in only 23% of eligible patients. Performance data did not include age and therefore could not be categorized into age cohorts.

hylaxis, 91% of patients with AIDS received HAART, 67% received syphilis screening, and 81% received tuberculosis screening. Lower performance rates were recorded for cervical cancer screening in only 23% of eligible patients. Performance data did not include age and therefore could not be categorized into age cohorts. The average viral loads from 2007 to 2010 were lowest for pediatric patients, next lowest for adolescents, and highest for adults (Table 1). This demonstrated a statistically significant increase in the mean viral load in adolescents when compared to pediatric patients (42% increase, P < .02) and in adults when compared to adolescents (195% increase, P < .0002). The pediatric population had the highest percentage of patients reaching undetectable VLs (58–72.6%). The adolescent group (34.9–54.5%) and the adult group (42.5–52.7%) were not significantly different over the four years (Fig. 1).Table 1 Average viral load by year and age group. 2007 2008 2009 2010 Pediatric 5357 7532 1885 12,248 Adolescent 37,683 27,117 10,836 17,404 Adult 57,120 53,252 36,580 51,344 Figure 1 Percent of patients reaching undetectable and average viral load by age grouping. In 3 of the 4 years, there was a statistically significant increase in the percentage of male adolescents reaching undetectable VL compared with female adolescents (Fig. 2). Of note, 2009 was the only year in which the mean viral load in females was lower than that of the males.Figure 2 Percent of adolescent patients reaching undetectable and average viral load by gender.

cant increase in the percentage of male adolescents reaching undetectable VL compared with female adolescents (Fig. 2). Of note, 2009 was the only year in which the mean viral load in females was lower than that of the males.Figure 2 Percent of adolescent patients reaching undetectable and average viral load by gender. Reviewing average VL by age demonstrates an increasing VL from age 12 through age 24, while the percentage of patients reaching undetectable VL peaks at 80% at age 8 and begins trending down through age 24 (Fig. 3).Figure 3 Percent of pediatric and adolescent patients reaching undetectable viral load and mean viral load by age.

average VL by age demonstrates an increasing VL from age 12 through age 24, while the percentage of patients reaching undetectable VL peaks at 80% at age 8 and begins trending down through age 24 (Fig. 3).Figure 3 Percent of pediatric and adolescent patients reaching undetectable viral load and mean viral load by age. 4 Discussion Data from University Hospital showed evidence of worsening virologic control by age group. Over the four years analyzed, the average viral load of pediatric patients was lower than that of the adolescent population, which in turn was lower than that of the adult population. Within the declining trend, the pediatric population had a significantly higher rate of patients averaging undetectable VLs compared with the adolescent and adult populations. The latter populations were not significantly different with regards to percentage of patients with undetectable VLs in comparison to each other. In a study by Ellen et al, almost 30% of youth with HIV in 14 cities were found to have significantly high VLs, associated with high rates of transmission [5]. This troubling trend highlights the potential challenges during adolescence that make treating and preventing disease more difficult. Agwu et al found a similar trend in patients with perinatally acquired HIV in clinics across the United States – older age, black race, and Hispanic ethnicity were associated with increased risk of advanced immunosuppression and detectable viremia [6]. During adolescence, patients are undergoing puberty, learning how to be autonomous young adults, and feeling challenged by increasing responsibilities with more complex external pressures [7]. While easily influenced by their environment, peers, and mentors, these young adults are developing behavioral foundations that will last their life time. These foundations subsequently impact their ability to independently manage their own health, including safer-sex practices, disease recognition, medication compliance, and prioritizing care.

their environment, peers, and mentors, these young adults are developing behavioral foundations that will last their life time. These foundations subsequently impact their ability to independently manage their own health, including safer-sex practices, disease recognition, medication compliance, and prioritizing care. When breaking this down further and examining mean VL by age from birth to 25 years of age, there is notable evidence of worsening virologic control and decreasing percentage of patients averaging undetectable VLs. The decrease in adolescent patients reaching undetectable viral load levels coincides with the average age of first sexual activity and could be related to the high rates of new HIV infections in this age group. Without proper counseling and care, these young adults are at significant risk of acquiring and perpetuating the spread of HIV. Additionally, given the challenges of an inner city environment, these patients must tackle their health concerns with limited resources and assistance. Yet another barrier to adolescents with HIV is its unique burden of stigma, relationship to poverty, infection rates within families, and its prevalence within ethnic minorities [8]. As Andiman states, “Stigma intersects with nearly every other factor and, as experienced in social networks, leads many youth to mistrust individuals outside of their closest social circles, sometimes including their [healthcare providers].” Especially in the adolescent period when self-perception and development of self-confidence is of high priority, having to confront and carry a stigma during this period can lead to self-doubt and conflict: “when adolescents fail to know or share their HIV status, they cannot become autonomous.” [7].

hcare providers].” Especially in the adolescent period when self-perception and development of self-confidence is of high priority, having to confront and carry a stigma during this period can lead to self-doubt and conflict: “when adolescents fail to know or share their HIV status, they cannot become autonomous.” [7]. Given these concerns, many programs and associations urge the creation of multifaceted transitional care initiatives and specifically target the transition of an adolescent in their early 20s. However, based on the virologic results in this urban population, declining virologic control was noted as young as 8 years of age. This would suggest that transitional care programs should be initiated at a much younger age and not necessarily focus on just the transfer of clinical care, but it should focus more on the psychosocial challenges of adolescence and young adulthood. Within the clinic studied, all patients are assessed annually for social work and mental health needs, have access to psychosocial assistance, and no services provided are restricted by age. Despite extensive social and mental health services, the decline in undetectable VLs in the adolescent population suggests identifying more specific adolescent needs. In a review of the Adolescent Trials Network for HIV/AIDS Interventions, researchers were surveyed on various aspects of their transitional care programs. Several clinics stated that transitions to adult care should occur between 22 and 24 years of age; however, discussions about when transitioning should begin was variable from as early as age 16 to as late as age 24. There were also discrepancies between how a “successful transition” was measured – “informants … were unable to accurately report an outcome” [9]. Some barriers to monitoring outcome included the lack of tracking mechanisms, while more anecdotal measures of success included emotional maturity, functional independence, available support systems, and stable health, and housing benefits [9]. Given the young age at which declines in virologic control was noted, these data would suggest a transitional care program starting at the age of 8 years and likely aimed at barriers seen in young adolescence and not just late adolescence.

endence, available support systems, and stable health, and housing benefits [9]. Given the young age at which declines in virologic control was noted, these data would suggest a transitional care program starting at the age of 8 years and likely aimed at barriers seen in young adolescence and not just late adolescence. The significant decline in virologic control by age observed in this study would also suggest that further investigations are needed to correlate adolescent perceptions of healthcare independence and virologic control. While virologic control over time has largely been affected by new advancements in antiretroviral therapy (ART), medication adherence, and understanding barriers to this remains tantamount to the success of any program. In a study looking at self-reported HIV-medication adherence of 12–19-year olds, only 28.3% of patients had taken all of their medications in the previous month, citing changes in one's day-to-day routine and forgetting to take medications as the most common barriers [10]. Another study performed at the National Institutes of Health (NIH) and National Cancer Institute (NCI), looked at changes in adolescent readiness over a 6-month period within a closing clinical research program. Demographics as well as CD4 cell count and HIV-1 RNA levels were measured 6 months apart in addition to completion of an anxiety scale and readiness questionnaire. After barriers to transition were addressed by the medical provider and social worker, the results showed no significant difference in CD4 cell counts and HIV-1 RNA levels, but there was a significant reduction in anxiety and increased confidence in their home community physician, social worker, and knowledge of disease and medications [11]. These studies suggest that adolescent education regarding their medications, side effects, and general care is key in developing a successful program in medication adherence and understanding, however, it may not result in better HIV outcomes. An early review of the Transition Readiness Assessment created by the national “Got Transition” program and given to adolescents in the pediatric subspecialty clinics at University Hospital showed good adolescent understanding of disease, emergencies, and medications, but it showed poor understanding of general healthcare navigation.

of the Transition Readiness Assessment created by the national “Got Transition” program and given to adolescents in the pediatric subspecialty clinics at University Hospital showed good adolescent understanding of disease, emergencies, and medications, but it showed poor understanding of general healthcare navigation. Even in early adolescence, however, virologic data from University Hospital indicates that more longitudinal studies are needed to objectively measure these transitional outcomes and correlate successful education with virologic control.

of the Transition Readiness Assessment created by the national “Got Transition” program and given to adolescents in the pediatric subspecialty clinics at University Hospital showed good adolescent understanding of disease, emergencies, and medications, but it showed poor understanding of general healthcare navigation. Even in early adolescence, however, virologic data from University Hospital indicates that more longitudinal studies are needed to objectively measure these transitional outcomes and correlate successful education with virologic control. Also concerning is that in 3 of the 4 years examined, the viral load in adolescent females and the number of female patients averaging undetectable VLs demonstrated worsening virologic control than that of their male counterparts. Additionally, based on the 2010 performance rates in the clinic, only 23% of adolescent females received a cervical cancer screening. In the 2009–2013 CDC data, rates of infection for female adults and adolescents actually decreased [12]; however, based on the above findings, one can question if the healthcare system within this area is able to deliver optimal care to females versus males. The Enhancing Communication to Improve HIV Outcomes (ECHO) study revealed that HIV providers had more negative perceptions of female personality characteristics than male and felt more frustrated with female patients than male. Women were also shown to be less educated or unemployed and more likely to report medication non-adherence and depressive symptoms [13]. In New Jersey, women had an HIV infection rate of 14.2 per 100,000 persons, compared with 9.3 in New York and 5.6 in Connecticut [14]. Adolescent girls and young women (15–24 years) are twice as likely to be at risk of HIV infection compared to boys and young men in the same age group worldwide [15]. Given this observed inequality, it is possible that transitional care programs aimed at treating this population may need to factor in gender inequalities psychologically, socially, or otherwise.

ars) are twice as likely to be at risk of HIV infection compared to boys and young men in the same age group worldwide [15]. Given this observed inequality, it is possible that transitional care programs aimed at treating this population may need to factor in gender inequalities psychologically, socially, or otherwise. It is also important for women to recognize that heterosexual contact accounted for 25% of all diagnosed HIV infections in the United States and 86% of those infections in adolescent and young adult women with HIV, and perinatally acquired infections accounted for an estimated 107 new infections in 2013 [14], [15]. Globally, HIV/AIDS is the leading cause of death among women aged 15–44 years [15]. While maternally acquired HIV has been dramatically reduced through the use of aggressive screening and treatment, scattered cases remain. University Hospital had at least one maternally acquired case of HIV in each year of this data set. Subsequently, with poor virologic control, females of child-bearing age in this study could represent a future increase in maternally acquired HIV cases in Newark.

ressive screening and treatment, scattered cases remain. University Hospital had at least one maternally acquired case of HIV in each year of this data set. Subsequently, with poor virologic control, females of child-bearing age in this study could represent a future increase in maternally acquired HIV cases in Newark. These data looked at viral load analysis performed in one lab at one urban university hospital during a four-year span and, consequently, is confounded by small sample sizes, particularly in the first few years of life. The population is also heterogeneous in terms of treatment, mode of acquisition, and stage of disease. While the majority of pediatric and adolescent patients in our study have prenatally acquired HIV, we were unable to separate out the small number of behaviorally acquired patients who may have different risk factors and behavioral patterns. Thus, a more in-depth study looking at the means of transmission should follow. Other demographic information would also be helpful in obtaining a closer look at the gender differences, as well as rates of medication compliance, clinic follow-up patterns, psychosocial barriers to care, and adolescent-perceived attitudes and knowledge of HIV and disease management. The recent addition of the Transition Readiness Assessment to adolescent care will reveal self-perceived adolescent understanding of disease and healthcare management. Extensive review of these assessments and other programs in adolescent care is required to further isolate barriers to optimal treatment. Additionally, we do not account for the increase in efficacious HIV medications and treatment strategies during the time period of this study.

g of disease and healthcare management. Extensive review of these assessments and other programs in adolescent care is required to further isolate barriers to optimal treatment. Additionally, we do not account for the increase in efficacious HIV medications and treatment strategies during the time period of this study. 5 Conclusion Pediatric patients develop a noticeable decrease in virologic control as they enter their adolescent years, which coincides with the onset of high-risk sexual activity. This increased viral load in infected patients puts others at higher risk and may contribute to adolescents having the fastest growing rate of new HIV infections. The decreased rate of virologic suppression in females of child-bearing age is also concerning as it presents an increased risk of vertical transmission. The development of adolescent HIV clinics, with dedicated services and transitional care programs focused on the unique needs of this population early in their adolescence, is one potential method of improving virologic control in this high-risk group. Acknowledgments No funding was secured for this study. Authors have no financial relationships relevant to this article to disclose and no conflicts of interest to disclose. Corresponding author affirms that all authors have contributed significantly to the reported research and production of the manuscript and that there are no prior publications or submissions with any overlapping information. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

Zika virus (ZIKV), an emerging single-stranded RNA mosquito born flavivirus that has been recognized in sporadic outbreaks in Africa since its first isolation from rhesus monkeys in the Zika forest (the virus carries the name) near Uganda in 1947. The first infection in humans was documented in Tanzania and Uganda in 1952. Further outbreaks have been reported in Africa, Americas, Asia, and Pacific islands. In 2015, ZIKV wide epidemic in susceptible Brazilian population have clearly demonstrated the effectiveness of its mosquito vector, Aedes genus, especially Aedes aegypti [1], [2]. The recent substation increase in the number of fetal cases, neonatal birth defects, and other neurological complications reported from Brazil have prompted the World Health Organization (WHO) to declare the Zika virus “A public health emergency of international concern” on February 1, 2016 [3]. The clinical manifestations of ZIKV during pregnancy are similar to those in nonpregnant women. Experimental laboratory-based and clinical studies have established a clear causal relation between maternal ZIKV infection and fetal defects [4]. The virus has a high efficiency to infect human neuronal progenitor cells and affect their development. ZIKV has the ability to cross the placenta, as demonstrated by the detection of the virus genome in the amniotic fluid infants who have confirmed microcephaly as well in fetal brain tissue in microcephalic fetus of an infected pregnant woman [4].

y to infect human neuronal progenitor cells and affect their development. ZIKV has the ability to cross the placenta, as demonstrated by the detection of the virus genome in the amniotic fluid infants who have confirmed microcephaly as well in fetal brain tissue in microcephalic fetus of an infected pregnant woman [4]. Although ZIKV has been linked to neonatal microcephaly, other abnormalities detected in prenatal sonography among ZIKV congenitally infected infant also included cerebral calcifications; fetal growth restriction and ocular abnormalities, which included bilateral macular and perimacular lesions as well as optic nerve abnormalities in most cases [5], [6], [7], [8], [9]. Prevalence and clinical spectrum of congenital ZIKV disease in newborns infected prenatally are unknown. Management of infants who have congenital ZIKV disease is primarily supportive, and there is neither available vaccine nor prophylactic medications [10]. However, the race is on to produce ZIKV vaccines. There are currently around 25 vaccine companies and organization groups are expediting vaccine research against ZIKV [11]. Healthcare providers should advice pregnant women to postpone their travel to endemic areas. If they have to travel, they should strictly follow steps to avoid mosquito bites [12]. Pediatricians should work closely with obstetricians to identify infant with maternal risk or diagnosed with ZIKV infection, and they have to standardize their approaches to diagnostic evaluation for congenital Zika virus infection [10], [12], [13].

ravel, they should strictly follow steps to avoid mosquito bites [12]. Pediatricians should work closely with obstetricians to identify infant with maternal risk or diagnosed with ZIKV infection, and they have to standardize their approaches to diagnostic evaluation for congenital Zika virus infection [10], [12], [13]. The list pathogens related to intrauterine infections continues to grow with the identification of ZIKV. The pattern of neurological abnormalities seen with congenital ZIKV infection is similar to what is seen with cytomegalovirus. The acronym for the common congenital infections that changed from TORCH to STORCH and CHEAPTORCHES need to be further expanding to ZiCHEAPTORCHES (Table 1) in order to include another important congenital infection, the Zika virus [14], [15], [16].Table 1 Congenital infection acronym: ZiCHEAPTORCHES Table 1Acronym Pathogens Zi Zika virus C Chickenpox and shingles H Hepatitis C, D, E E Enteroviruses A AIDS (HIV infection) P Parvovirus B19 T Toxoplasmosis O Other (Group B streptococcus, Listeria, Streptococcus, Candida, Lyme disease) R Rubella C Cytomegalovirus H Herpes simplex E Everything else sexually transmitted (Gonorrhea, Chlamydia infection, Ureaplasma urealyticum, human papillomavirus S Syphilis Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Ovarian masses are most commonly observed in adults; they rarely occur in children. The majority of the ovarian masses encountered in children or patients of premenarchal age are non-neoplastic lesions. The clinical signs and symptoms of ovarian masses are usually non-specific. Early management may be necessary to preserve fertility. Gynecological malignant conditions constitute approximately 3% of all types of cancer in children. Ovarian tumors in children account for only 1% of childhood malignancies. However, the true incidence of malignant ovarian tumors in the pediatric population is unknown [1]. 2 Patients and methods The records of 20 girls under the age of 12 years with adnexal masses who were treated at a tertiary referral center between May 2011 and November 2015 were reviewed and retrospectively analyzed based on their age at the time of admission, presenting complaints, clinical and radiological findings, tumor markers, management and follow-up.

irls under the age of 12 years with adnexal masses who were treated at a tertiary referral center between May 2011 and November 2015 were reviewed and retrospectively analyzed based on their age at the time of admission, presenting complaints, clinical and radiological findings, tumor markers, management and follow-up. All of the patients underwent pelvic ultrasound. CT scan was performed when necessary. Tumor markers—serum alpha fetoprotein (AFP), beta-human chorionic gonadotropin (β-HCG), and cancer antigen 125 (CA-125), were tested in ten patients. A thyroid function test was conducted in one patient because of associated precocious puberty. Complex lesions were surgically excised. A standard Pfannenstiel incision was made to remove the lesions. One patient was operated on laparoscopically. The patients with immature teratoma underwent complete surgical excision of tumor followed by chemotherapy; Bleomycin, Etoposide and Cisplatin (BEP regimen) were administered, and the patients' serum AFP levels were monitored. The diagnoses of all patients were histopathologically confirmed. Simple cysts measuring less than 6 cm were managed conservatively using ultrasound scans performed monthly for 3 months.

chemotherapy; Bleomycin, Etoposide and Cisplatin (BEP regimen) were administered, and the patients' serum AFP levels were monitored. The diagnoses of all patients were histopathologically confirmed. Simple cysts measuring less than 6 cm were managed conservatively using ultrasound scans performed monthly for 3 months. 3 Results The patients' age at the time of admission ranged between 3 days and 12 years. Four neonates were antenatally diagnosed, as confirmed by postnatal ultrasound scans. Abdominal lump (Fig. 1) (n = 12) and pain (n = 11) were the most common presenting complaints. Lesions were unilateral in 14 patients and bilateral in 1 patient. Tumor markers were sent in 10 patients and were normal in all of them.Fig. 1 Clinical photograph of an adnexal mass presenting as abdominal lump. Fig. 1 Four patients (20%) had antenatally diagnosed unilateral cystic ovarian lesions. On postnatal scan, 2 patients had a simple cyst measuring less than 6 cm, which was conservatively managed. These lesions resolved on follow-up ultrasound at 3 months. In one patient, a cyst measuring more than 6 cm was marsupialized. One antenatally diagnosed patient had a dermoid cyst and required oophorectomy.

ions. On postnatal scan, 2 patients had a simple cyst measuring less than 6 cm, which was conservatively managed. These lesions resolved on follow-up ultrasound at 3 months. In one patient, a cyst measuring more than 6 cm was marsupialized. One antenatally diagnosed patient had a dermoid cyst and required oophorectomy. Ten patients (50%) underwent complete surgical excision of the adnexal mass (Fig. 2, Fig. 3, Fig. 4, Fig. 5). Oophorectomy was performed in eight patients; based on histopathologic results, these patients had mature teratoma, and they required no additional treatment. Two patients were managed by salpingo-oophorectomy; histopathology revealed immature teratoma. The patients received adjuvant chemotherapy; Bleomycin, Etoposide and Cisplatin (BEP regimen) were administered and the patients' serum AFP levels were monitored. One patient had a large paraovarian cyst, which was laparoscopically excised (Fig. 6, Fig. 7).Fig. 2 CT image showing large left adnexal mass. Fig. 2Fig. 3 Intraoperative image of a left immature teratoma. Fig. 3Fig. 4 Excised specimen of the left immature teratoma. Fig. 4Fig. 5 Excised specimen of the left mature teratoma. Fig. 5Fig. 6 CT image of patient with right paraovarian cyst showing cystic lesion in right adnexa. Fig. 6Fig. 7 Intraoperative image showing right paraovarian cyst in a patient. Fig. 7

Fig. 2Fig. 3 Intraoperative image of a left immature teratoma. Fig. 3Fig. 4 Excised specimen of the left immature teratoma. Fig. 4Fig. 5 Excised specimen of the left mature teratoma. Fig. 5Fig. 6 CT image of patient with right paraovarian cyst showing cystic lesion in right adnexa. Fig. 6Fig. 7 Intraoperative image showing right paraovarian cyst in a patient. Fig. 7 One patient (5%) presented with bilateral multicystic ovarian masses and precocious puberty and had severe hypothyroidism on hormonal evaluation. She was diagnosed with Van Wyk–Grumbach syndrome and was administered thyroxin supplementation. The masses resolved significantly after 3 months. Five patients (25%) presented with acute abdomen and were diagnosed as torsion on USG and CT requiring emergency surgery. All of the patients had adnexal masses measuring greater than 8 cm, in addition to gangrene observed in the ipsilateral ovary. Three patients were managed by salpingo-oophorectomy and two required oophorectomy. Histopathologic results revealed teratoma in three patients, immature teratoma in one patient and simple cysts in one patient. The patient with immature teratoma received adjuvant chemotherapy (BEP regimen) and the patient's serum AFP levels were monitored. All of the patients had no complaints on follow-up (Table 1).Table 1 Presentation and management of 20 patients with adnexal masses.

Five patients (25%) presented with acute abdomen and were diagnosed as torsion on USG and CT requiring emergency surgery. All of the patients had adnexal masses measuring greater than 8 cm, in addition to gangrene observed in the ipsilateral ovary. Three patients were managed by salpingo-oophorectomy and two required oophorectomy. Histopathologic results revealed teratoma in three patients, immature teratoma in one patient and simple cysts in one patient. The patient with immature teratoma received adjuvant chemotherapy (BEP regimen) and the patient's serum AFP levels were monitored. All of the patients had no complaints on follow-up (Table 1).Table 1 Presentation and management of 20 patients with adnexal masses. Table 1Diagnosis and presentation Simple cyst Teratoma Other Total Lump Torsion Abdominal lump and pain Torsion Bilateral ovarian cysts with hypothyroidism Sub-type/size <6cm >6cm >6cm Mature Immature Mature Immature Conservative 2 1 3 Marsupialization 1 1 Excision 1 1 Oophorectomy 8 2 10 Salpingo-oophorectomy 1 2 1 1 5 Total 2 2 1 8 2 3 1 1 20 4 Discussion Adnexal masses may originate from the ovaries, fallopian tubes and other pelvic organs. The types of adnexal masses include tumors, inflammatory or functional cysts. Approximately one third of adnexal masses are ovarian tumors.

Table 1Diagnosis and presentation Simple cyst Teratoma Other Total Lump Torsion Abdominal lump and pain Torsion Bilateral ovarian cysts with hypothyroidism Sub-type/size <6cm >6cm >6cm Mature Immature Mature Immature Conservative 2 1 3 Marsupialization 1 1 Excision 1 1 Oophorectomy 8 2 10 Salpingo-oophorectomy 1 2 1 1 5 Total 2 2 1 8 2 3 1 1 20 4 Discussion Adnexal masses may originate from the ovaries, fallopian tubes and other pelvic organs. The types of adnexal masses include tumors, inflammatory or functional cysts. Approximately one third of adnexal masses are ovarian tumors. Malignant ovarian tumors are rare, particularly in patients under 5 years of age. Solid ovarian tumors are uncommon in the pediatric population; however, ovarian tumors when present, constitute a major source of anxiety for the patients and their family. Ovarian tumors must be considered in the differential diagnosis of young girls with abdominal pain, mass or other non-specific symptoms. In a large study, Templeman et al stated that malignant ovarian tumors in children and adolescents are rare, accounting for 0.9% of all malignancies in this age group [2]. Hassan et al confirmed that during the first two decades of life, ovarian tumors represent the most frequent tumors found in the female genital tract [3].

arge study, Templeman et al stated that malignant ovarian tumors in children and adolescents are rare, accounting for 0.9% of all malignancies in this age group [2]. Hassan et al confirmed that during the first two decades of life, ovarian tumors represent the most frequent tumors found in the female genital tract [3]. Historically, all ovarian masses discovered in infants, children, and adolescents were removed surgically. However, the identification of tumor markers and advances in radiologic imaging allow a more conservative approach to the management of these neoplasms, with ovarian preservation as the standard, except in cases of cancer. Abdominal pain was the most common symptom encountered. Ovarian masses often present with abdominal complaints that can mimic other diseases. A mobile, palpable abdominal mass was the most frequent physical finding. Primary ovarian cysts and tumors are uncommon in children. Two-thirds of malignant tumors in children are germ cell tumors. The majority of the malignant germ cell tumors are dysgerminoma, in contrast to adult ovarian tumors, where 90% are of epithelial cell origin and approximately 10% are non-epithelial. In our study, the most common malignant tumor was immature teratoma, and none of the patients had dysgerminoma. Although the true incidence of ovarian cysts in the fetus is unknown, they have been reported in 3%–7% of routine obstetric ultrasound analyses [4], [5]. Most of these cysts were resolved, which may explain why no antenatally detected ovarian cyst in our series required surgery.

Primary ovarian cysts and tumors are uncommon in children. Two-thirds of malignant tumors in children are germ cell tumors. The majority of the malignant germ cell tumors are dysgerminoma, in contrast to adult ovarian tumors, where 90% are of epithelial cell origin and approximately 10% are non-epithelial. In our study, the most common malignant tumor was immature teratoma, and none of the patients had dysgerminoma. Although the true incidence of ovarian cysts in the fetus is unknown, they have been reported in 3%–7% of routine obstetric ultrasound analyses [4], [5]. Most of these cysts were resolved, which may explain why no antenatally detected ovarian cyst in our series required surgery. Follicular cysts are commonly detected incidentally on antenatal ultrasound examination [6]. The etiology is unclear, but they most likely arise from ovarian stimulation by maternal and fetal gonadotropin [7]. The majority of fetal ovarian cysts are unilateral, although both ovaries may be involved. Follicular ovarian cysts in fetuses and neonates are common and increase in frequency with advancing gestational age and some maternal complications, such as diabetes mellitus, preeclampsia, and rhesus isoimmunization [7], [8]. In one autopsy series of 332 ovaries from stillbirths and neonatal deaths, one or more follicular cysts lined by granulosa epithelium and measuring greater than 1 mm in diameter were detected in 113 infants [8]. Among live births, the best estimate of the incidence of clinically significant ovarian cysts is 1 in 2500 [9].

one autopsy series of 332 ovaries from stillbirths and neonatal deaths, one or more follicular cysts lined by granulosa epithelium and measuring greater than 1 mm in diameter were detected in 113 infants [8]. Among live births, the best estimate of the incidence of clinically significant ovarian cysts is 1 in 2500 [9]. Occasionally, these cysts are further complicated by intracystic hemorrhages, ovarian torsion, or rarely, by a mass effect and respiratory distress or hydronephrosis. Spontaneous regression usually occurs by 4–6 months of age [10]. The differential diagnosis of a fetal cystic intraabdominal mass includes genitourinary tract disorders (e.g., reproductive tract anomalies, urinary tract obstruction, urachal cyst), gastrointestinal tract disorders (e.g., mesenteric or omental cyst, volvulus, colonic atresia, intestinal duplication), or miscellaneous disorders (e.g., choledochal, splenic, or pancreatic cyst; lymphangioma). Spontaneous regression of both simple and complex cysts often occurs either antenatally or postpartum by 6 months of age; therefore, the management is usually expectant. In one review of 66 published cases of simple cysts, 50% resolved by 1 month of age, 75% by 2 months of age, and 90% by 3 months of age [9]. The rate of malignancy is so low that it need not be considered in making therapeutic decisions. Ultrasound examination should be performed every 3–4 weeks antenatally.

expectant. In one review of 66 published cases of simple cysts, 50% resolved by 1 month of age, 75% by 2 months of age, and 90% by 3 months of age [9]. The rate of malignancy is so low that it need not be considered in making therapeutic decisions. Ultrasound examination should be performed every 3–4 weeks antenatally. The standard management of neonatal cysts consists of serial ultrasound examinations at birth and every 4–6 weeks thereafter until the cyst resolves, enlarges, has persisted for 4–6 months, or becomes symptomatic. The aspiration of simple cysts measuring larger than 4–5 cm is advised [7]. Surgical intervention is reserved for complex and symptomatic cysts and for cysts that increase in size and persist for more than 4–6 months [11], [12], [13]. The transumbilical approach has been reported to be both a feasible and safe approach for a broad spectrum of surgical procedures, including ovarian tumors, in neonates and infants. The cosmetic results have been reported to be excellent [14]. The transumbilical approach seems to be an attractive alternative for managing ovarian cysts in children in the absence of appropriate settings for laparoscopy [15]. Teratomas are the most common germ cell tumors observed in the majority of published series [16], [17]. This subgroup of tumors may be further divided into mature teratomas, which are benign or immature teratomas, which may be either malignant or benign.

The standard management of neonatal cysts consists of serial ultrasound examinations at birth and every 4–6 weeks thereafter until the cyst resolves, enlarges, has persisted for 4–6 months, or becomes symptomatic. The aspiration of simple cysts measuring larger than 4–5 cm is advised [7]. Surgical intervention is reserved for complex and symptomatic cysts and for cysts that increase in size and persist for more than 4–6 months [11], [12], [13]. The transumbilical approach has been reported to be both a feasible and safe approach for a broad spectrum of surgical procedures, including ovarian tumors, in neonates and infants. The cosmetic results have been reported to be excellent [14]. The transumbilical approach seems to be an attractive alternative for managing ovarian cysts in children in the absence of appropriate settings for laparoscopy [15]. Teratomas are the most common germ cell tumors observed in the majority of published series [16], [17]. This subgroup of tumors may be further divided into mature teratomas, which are benign or immature teratomas, which may be either malignant or benign. Teratomas are composed of recognizable tissues of ectodermal, mesodermal and endodermal origin, in any combination. Immature teratomas are common germ cell tumors comprising two or more germ cell layers (ecto-, meso- or endoderm) derived from a pluripotent malignant precursor cell. Mature teratomas account for approximately 15% of all ovarian tumors [18] whereas immature tumors are rare, representing less than 1% of ovarian tumors [19]. Mature teratomas are classified as cystic, solid or monodermal. Immature teratomas show only solid mass. In mature teratomas, the most commonly mature ectodermal elements such as skin, hair, sweat and sebaceous glands are noted, whereas in immature teratomas, tissues with partial somatic differentiation identical to that of fetal tissues are found. In this study, 7 patients had teratoma; of these, 5 patients (87.5%) had mature teratoma and two patients (13.33%) had immature teratoma.

ts such as skin, hair, sweat and sebaceous glands are noted, whereas in immature teratomas, tissues with partial somatic differentiation identical to that of fetal tissues are found. In this study, 7 patients had teratoma; of these, 5 patients (87.5%) had mature teratoma and two patients (13.33%) had immature teratoma. However, both malignant and benign teratomas can appear to be identical by ultrasound or CT findings. Germ cell tumors are serologically evaluated using tumor markers. In immature teratoma cases, AFP is widely used. It has been suggested that the AFP level in immature teratoma is not correlated to either stage or grade of the tumor. Thus, AFP plays a limited role in the evaluation and management of germ cell tumors [20], [21], [22], [23]. In this study, all of the patients underwent surgical resection. Two patients were histopathologically diagnosed with immature teratoma, and they received chemotherapy. Van Wyk–Grumbach syndrome is a rare condition characterized by breast development, uterine bleeding and multicystic ovaries in the presence of long-standing primary hypothyroidism.

However, both malignant and benign teratomas can appear to be identical by ultrasound or CT findings. Germ cell tumors are serologically evaluated using tumor markers. In immature teratoma cases, AFP is widely used. It has been suggested that the AFP level in immature teratoma is not correlated to either stage or grade of the tumor. Thus, AFP plays a limited role in the evaluation and management of germ cell tumors [20], [21], [22], [23]. In this study, all of the patients underwent surgical resection. Two patients were histopathologically diagnosed with immature teratoma, and they received chemotherapy. Van Wyk–Grumbach syndrome is a rare condition characterized by breast development, uterine bleeding and multicystic ovaries in the presence of long-standing primary hypothyroidism. The syndrome is characterized by juvenile hypothyroidism, delayed bone age, and isosexual precocious puberty with reversal to a prepubertal state following thyroid hormone replacement therapy [24]. The gonadotropin releasing hormone (GnRH)-dependent activation of the hypothalamic-pituitary-gonadal axis leads to central precocious puberty (CPP). The extrapituitary secretion of gonadotropins or secretion of gonadal steroids independent of pulsatile GnRH stimulation may lead to pseudoprecocious puberty or GnRH-independent sexual precocity [25].

rmone (GnRH)-dependent activation of the hypothalamic-pituitary-gonadal axis leads to central precocious puberty (CPP). The extrapituitary secretion of gonadotropins or secretion of gonadal steroids independent of pulsatile GnRH stimulation may lead to pseudoprecocious puberty or GnRH-independent sexual precocity [25]. Hypothyroidism should be considered, especially when young girls present with bilateral multicystic ovarian mass and vaginal bleeding accompanied by additional clinical presentations, such as cold intolerance, constipation, delayed bone age. Thyroxin replacement therapy should lead to complete resolution of symptoms and promote normal physical and mental development. Paraovarian cyst is a benign condition that is uncommon in children, is incidentally diagnosed, and occurs in the mesosalpinx between its two leaves. The cyst is believed to originate from mesothelium or the remnant of Mullerian duct and Wolffian duct [26]. Complications such as torsion, hemorrhage, rupture, and neoplastic transformation may occur. The preoperative diagnosis is difficult because of its close proximity to the ovary. Laparoscopy is a diagnostic as well as therapeutic approach.

iginate from mesothelium or the remnant of Mullerian duct and Wolffian duct [26]. Complications such as torsion, hemorrhage, rupture, and neoplastic transformation may occur. The preoperative diagnosis is difficult because of its close proximity to the ovary. Laparoscopy is a diagnostic as well as therapeutic approach. 5 Conclusion Ovarian lesions in children include a broad array of pathologic diagnoses that have variable clinical presentations. Most ovarian tumors are benign. Epithelial cysts and teratomas are the most common benign lesions and germ cell tumors are most commonly malignant. A laparoscopic approach has been reported to be attractive in the majority of the studies. With accurate staging, complete resection and chemotherapy for malignant tumors, patients have demonstrated excellent survival rates. In patients with bilateral ovarian lesion, endocrine causes should be ruled out. Ethical clearance Not applicable as this is a retrospective study. All patients in this study were managed according to the existing protocols. No new investigations and intervention was done on the patient. Source of funding No funding was obtained to conduct the study. Conflict of interest The authors have no conflict of interest to report. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction The incidence of methicillin-resistant Staphylococcus aureus (MRSA) infection has been steadily increasing. This has coincided with an increase in the number of patients presenting with serious invasive disease due to MRSA [1], [2]. The causative organisms are defined as Staphylococcus aureus strains with an oxacillin minimum inhibitory concentration (MIC) of at least 4 mcg/mL. MRSA is resistant to all beta-lactam agents, including cephalosporins (with the exception of ceftobiprole) [3]. Its resistance is derived from the mecA gene encoding for the low-affinity binding protein PBP-2a, which allows the organism to grow and divide in the presence of methicillin and other beta-lactam antibiotics [4], [5]. Due to its high resistance to previously mentioned antibiotics and the appearance of new strains, MRSA infection has become an increasing medical challenge [6], [7], [8], [9]. Infections caused by MRSA have been classified as either nosocomial (hospital acquired) or community acquired [2], [8], [10]. Infection is considered to be HA-MRSA if positive cultures result from samples drawn after 72 h of admission. Cases considered CA-MRSA include those in which positive cultures have been drawn outside the hospital or drawn within 72 h of admission or in cases in which MRSA was diagnosed in an outpatient setting [8], [11], [12].

n is considered to be HA-MRSA if positive cultures result from samples drawn after 72 h of admission. Cases considered CA-MRSA include those in which positive cultures have been drawn outside the hospital or drawn within 72 h of admission or in cases in which MRSA was diagnosed in an outpatient setting [8], [11], [12]. Although MRSA infection rates are not significant in countries such as the Netherlands, Denmark, and Sweden [13], [14], the threat has increased significantly in many other countries, such as the USA and Western European countries such as Great Britain, in which MRSA infection has become an epidemic [11], [12], [14], [15]. Furthermore, some Middle Eastern countries such as Iran have recorded high numbers of MRSA infections [16]. Children are at risk of acquiring MRSA infections [17], [18], [19]. The risk increases more when they have co-morbidities such as malignancies, recent surgeries, autoimmune diseases, previous antibiotic usage, and long-term hospitalization, with resultant exposure to potentially more dangerous strains of HA-MRSA [17], [19], [20], [21]. Moreover, the threat of MRSA infection has increased in children and has manifested more frequently in neonates and in countries such as the U.S [22], [23]. Risk factors for children are the same as for adults, with the addition of genetic diseases such as cystic fibrosis and congenital immunodeficiencies [17], [20], [24].

over, the threat of MRSA infection has increased in children and has manifested more frequently in neonates and in countries such as the U.S [22], [23]. Risk factors for children are the same as for adults, with the addition of genetic diseases such as cystic fibrosis and congenital immunodeficiencies [17], [20], [24]. The first report of MRSA in Saudi Arabia was published in 1994 by Zaman in the western region in Jeddah. Over a period of three years covering Zaman's report, he found that 7.5% of all Staphylococcus aureus infections were MRSA positive [25]. Thereafter, a few reports followed from Madani and others in 2002 in the setting of two tertiary care centers in Saudi Arabia. The studies included patients from adult and pediatric wards (both medical and surgical). They found that 33% of all Staphylococcus aureus cultured patients were MRSA positive [11], [12]. Despite these data, studies describing MRSA infections in the pediatric population in Saudi Arabia remain limited in number. Only one study [18] exclusively described pediatric MRSA infections in Saudi Arabia. Bukhari et al investigated 80 previously healthy pediatric patients with community acquired MRSA infection and found that 6% (five patients) had invasive CA-MRSA with serious complications that included osteomyelitis, deep vein thrombosis, and subdural empyema [18]. The growing risk of MRSA was reported by Bukharie and Abdelhadi from King Fahd University Hospital in Dammam, Saudi Arabia in a study showing that the prevalence of CA-MRSA infections increased from 9.9 per 10,000 admissions in 2001 to 67 per 10,000 admissions in 2008 [8], [13].

n thrombosis, and subdural empyema [18]. The growing risk of MRSA was reported by Bukharie and Abdelhadi from King Fahd University Hospital in Dammam, Saudi Arabia in a study showing that the prevalence of CA-MRSA infections increased from 9.9 per 10,000 admissions in 2001 to 67 per 10,000 admissions in 2008 [8], [13]. Rates of MRSA infection are increasing among the Saudi population, including children. The data on pediatric HA-MRSA are limited, but signs indicate that it is also becoming more frequent [2], [6], [7], [8], [11], [12], [13], [18], [26], [27]. These data demonstrates that MRSA infection in the pediatric population needs to be studied further. The aim of this study is to describe characteristics of pediatric HA-MRSA infection in a tertiary care hospital in King Abdulaziz Medical City, Riyadh, Saudi Arabia, and determine possible risk factors, in hope that it will add to the body of knowledge on this important infection in this region.

to be studied further. The aim of this study is to describe characteristics of pediatric HA-MRSA infection in a tertiary care hospital in King Abdulaziz Medical City, Riyadh, Saudi Arabia, and determine possible risk factors, in hope that it will add to the body of knowledge on this important infection in this region. 2 Patients and methods This is a case series retrospective chart review study that was conducted at King Abdulaziz Medical City in Riyadh, a tertiary care center with approximately 1000 beds. It included all patients who were 14 years of age or younger with a documented culture of MRSA from any site of the body between January 2009 and December 2011. Data on patients with MRSA were retrieved from the database of the Infection Prevention and Control Department at our institution, which performs approximately 900 polymerase chain reaction (PCR) screenings per month. The MRSA policy at KAMC-Riyadh is to screen all patients who are:• Admitted to the Pediatric intensive care unit (PICU). • Transferred from another hospital or treated in another hospital within the last six months. • Undergoing cardiac, orthopedic (including spine) surgery, preoperatively. • Hemodialysis patients on admission for the first dialysis treatment and for placement of any type of vascular access (i.e.: AV-fistula, permanent catheter, graft, or port access device). • Patients on continuous ambulatory peritoneal dialysis treatment for the first time when scheduled for catheter placement. • Known to be previously MRSA positive. • Roommates of MRSA-positive patients not on isolation precautions.

• Hemodialysis patients on admission for the first dialysis treatment and for placement of any type of vascular access (i.e.: AV-fistula, permanent catheter, graft, or port access device). • Patients on continuous ambulatory peritoneal dialysis treatment for the first time when scheduled for catheter placement. • Known to be previously MRSA positive. • Roommates of MRSA-positive patients not on isolation precautions. The sites to screen include:• Anterior nares. • Non-intact skin areas (e.g., tracheostomy, pressure sores, or surgical wounds). • The groin and axillae of neonates and pediatric patients awaiting liver or cardiac surgery.

• Patients on continuous ambulatory peritoneal dialysis treatment for the first time when scheduled for catheter placement. • Known to be previously MRSA positive. • Roommates of MRSA-positive patients not on isolation precautions. The sites to screen include:• Anterior nares. • Non-intact skin areas (e.g., tracheostomy, pressure sores, or surgical wounds). • The groin and axillae of neonates and pediatric patients awaiting liver or cardiac surgery. Presentations of MRSA infection that were included in the study were abscess, signs of shock (hypotension, tachycardia, etc.), discharge from any site of the body, fever, respiratory distress, localized swelling, etc. We considered only the first isolate of bacterial culture results. The time of culture was used to differentiate between CA-MRSA (less than 72 h of admission) and HA-MRSA (more than 72 h after admission). Patient data including age, gender, nationality, risk factors, clinical manifestation, culture site, unit of stay, diagnosis, and outcome were retrieved from patient charts into the data collection sheet. Data were then entered and analyzed using a software statistical package (SPSS version 20). Outcomes were divided into favorable (complete recovery) and unfavorable (recovery with complications or death). Patient confidentiality during data collection and entry was ensured using a coding system that prevented disclosure of names or medical record numbers. Statistical methods used to obtain our objectives included the mean, median, standard deviation, minimum, maximum age, and Chi-Square for categorical data. The results were then transferred to Microsoft Office Excel 2007 to create several charts and diagrams (Table 1, Fig. 1, Fig. 2, Fig. 3, Fig. 4, Fig. 5, Fig. 6, Fig. 7).

stical methods used to obtain our objectives included the mean, median, standard deviation, minimum, maximum age, and Chi-Square for categorical data. The results were then transferred to Microsoft Office Excel 2007 to create several charts and diagrams (Table 1, Fig. 1, Fig. 2, Fig. 3, Fig. 4, Fig. 5, Fig. 6, Fig. 7). 3 Results Two hundred patients were documented to have MRSA infection. After analyzing the data, we found that nearly one-quarter (22%, 39 patients) of MRSA-infected patients had HA-MRSA. The majority of patients in our study were males (59%, 23 patients), with 40% females (16 patients) (Table 1). The majority of our sample were Saudis (95%, 37 patients) (Table 1). We categorized the pediatric population studied into three age groups (below one year, between one and five years, and more than five years). Nearly half of the patients were below one year old (46%, 18 patients), with 26% (11 patients) between one and five years, and 28% (10 patients) above five years of age (Table 1). The mean age was 3.1 years of age, with a maximum of 12 years and a minimum of nine days. Most of our population had previous surgery (31%, 12 patients), previous hospitalization (26%, 10 patients), or a cardiac anomaly (23%, nine patients) (Fig. 1). Other co-morbidities included conditions such as asthma, burns, renal failure, or metabolic diseases (Fig. 1). 59% (23 patients) presented with fever. 38% (15 patients) presented with skin and soft tissue infection (SSTI) manifestations. And 23% (9 patients) presented with abscess (Fig. 2). Shock manifestations (hypotension, tachycardia, etc.) were found in 21% (8 patients) (Fig. 2). In 19 infected patients (52%), the site of HA-MRSA isolation was blood in 31% (12 patients), skin and soft tissue in 13% (4 patients), and bone or joint in 8% (3 patients) (Fig. 3). Other less frequently reported sites among our study population included CSF and lumbar drain catheter in two patients (5%) and tip of central venous catheter in one patient (3%) (Fig. 3). We found that 12 patients (31%) were admitted to the PICU, 10 patients (26%) were admitted to the pediatric ward, and five patients each (13% each) were admitted to the burn unit, NICU and cardiac CCU (Fig. 4). Twelve patients (31%) were diagnosed with septicemia, seven patients (18%) had pneumonia, and six patients (15%) had cellulitis (Fig. 5). We attempted to correlate the time of year with the rate of infection, but we were unable to establish a relationship.

were admitted to the burn unit, NICU and cardiac CCU (Fig. 4). Twelve patients (31%) were diagnosed with septicemia, seven patients (18%) had pneumonia, and six patients (15%) had cellulitis (Fig. 5). We attempted to correlate the time of year with the rate of infection, but we were unable to establish a relationship. Twenty-nine (74%) of the studied patients fully recovered (Fig. 6). Among the remaining patients (10%, four patients) recovered with complications such as vision impairment, and six patients (15%) died (Fig. 6). We tried to find an association between unfavorable outcomes and co-morbidities associated with HA-MRSA infection and found that unfavorable outcomes were associated with chronic pulmonary diseases such as asthma (P-Value = .03) (Fig. 7). Eighty percent of patients with unfavorable outcomes (eight patients) required PICU care and presented with shock manifestations and/or SSTI manifestations.Table 1 Patient demographics (n = 39) Number Percent Age category Up to 1 year 18 46% 1 to 5 years 11 28% 5 to 12 years 10 26% Gender Male 23 59% Female 16 41% Nationality Saudi 37 95% Non-Saudi 2 5% Fig. 1 Comorbidities and associated risk factors. Fig. 2 Clinical presentation (n = 39). Fig. 3 Site of isolation (n = 39). Fig. 4 Unit of stay (n = 39). Fig. 5 Presenting diagnosis (n = 39). Fig. 6 Outcome (n = 39). Fig. 7 Corelation between risk factors and outcomes (n = 39).

Number Percent Age category Up to 1 year 18 46% 1 to 5 years 11 28% 5 to 12 years 10 26% Gender Male 23 59% Female 16 41% Nationality Saudi 37 95% Non-Saudi 2 5% Fig. 1 Comorbidities and associated risk factors. Fig. 2 Clinical presentation (n = 39). Fig. 3 Site of isolation (n = 39). Fig. 4 Unit of stay (n = 39). Fig. 5 Presenting diagnosis (n = 39). Fig. 6 Outcome (n = 39). Fig. 7 Corelation between risk factors and outcomes (n = 39). 4 Discussion The threat of MRSA infection has been increasing on a national and global basis [2], [23]. MRSA infections are considered to be HA-MRSA if positive culture samples are drawn after 72 h of admission. Cases of CA-MRSA infection include patients from whom cultures have been withdrawn outside the hospital or for whom diagnosis occurred in an outpatient setting [8], [11], [12]. The epidemiological characteristics of HA-MRSA infections vary among geographic regions and from one year to another. A recently published study conducted in Pennsylvania by Casy et al indicated that between 2001 and 2010, the annual incidence of HA-MRSA has increased by 7% [28]. We tried to compare some of our research results to other studies conducted at a similar time in other regions. We found that the risk of acquiring HA-MRSA increased with younger age, with a median age of 3.1 years. The finding was correlated with a study conducted in Minnesota, USA, where the mean age of infection was 2.4 years [29].

are some of our research results to other studies conducted at a similar time in other regions. We found that the risk of acquiring HA-MRSA increased with younger age, with a median age of 3.1 years. The finding was correlated with a study conducted in Minnesota, USA, where the mean age of infection was 2.4 years [29]. We found that patients under one year of age had the highest rate of admission due to HA-MRSA (46%, 18 patients) (Table 1). This concurred with the findings of a study conducted by Gutierrez and others in California, USA, indicating that from 1985 to 2009, those under one year of age had a higher hospitalization rate than did children 1–2 years of age (OR 5.6) [23]. The PICU had the highest rate of admission in our study 31% (12 patients) (Fig. 4). This correlated with results of an MRSA review study conducted at Johns Hopkins Hospital in the United States by David MZ et al. This study found that from 2007 to 2008, 6% (72/1674) of patients in their PICU had MRSA colonization [30].

The PICU had the highest rate of admission in our study 31% (12 patients) (Fig. 4). This correlated with results of an MRSA review study conducted at Johns Hopkins Hospital in the United States by David MZ et al. This study found that from 2007 to 2008, 6% (72/1674) of patients in their PICU had MRSA colonization [30]. A 2013 study conducted in Pakistan to investigate nasal colonization among septicemia patients found that nasal screening for MRSA colonization can be helpful in determining the cause of septicemia. The same study detected MRSA infections in 100% of those on dialysis or with surgical site infections [31]. We reported that septicemia (31%, 12 patients) and pneumonia (18%, seven patients) were among the most common consequences of HA-MRSA infection in our study (Fig. 5). Similarly, a 2013 study from China by Wu X et al reported that children who acquired HA-MRSA had an aggressive infection course associated most frequently with pneumonia and septicemia. However, the same study indicated that CNS infections were common, whereas in our study, it was rare (6%, two patients) (Fig. 5) [32]. We did not find a correlation between time of year and infection rates in our study. In contrast, a study conducted in the USA to investigate the epidemiological change of MRSA infections found that the infection frequency was highest between the months of June to October. The same study reported that rates of blood stream and pneumonia-related admissions had not changed compared to data they had collected previously. Additionally, most patients in the study had strains of HA-MRSA [33]. Another study, conducted by Leonard M. Mermel et al, reported that children exhibited a seasonal pattern of HA-MRSA infection, whereas adult infections exhibited no such seasonality [34].

ad not changed compared to data they had collected previously. Additionally, most patients in the study had strains of HA-MRSA [33]. Another study, conducted by Leonard M. Mermel et al, reported that children exhibited a seasonal pattern of HA-MRSA infection, whereas adult infections exhibited no such seasonality [34]. We think that the spread of nosocomial can be reduced or even eliminated by implementing and enhancing general infection control and prevention measures, including hand hygiene. Pereira et al published results of a 2014 study that was conducted in Botucatu, Brazil which concluded that improving hygiene practices in high risk areas such as the PICU and NICU would help to prevent infection [35]. 5 Conclusion HA-MRSA is a serious infection with variable epidemiology that presents the need for further investigation within this geographic region. Young children and children with risk factors, especially those involving pulmonary disease, tend to have worse prognoses than those who do not. HA-MRSA is responsible for a high number of PICU and NICU admissions. It has been shown in our study to cause serious complications and adverse outcomes, especially for those who require PICU admission. This study emphasizes the need for surveillance of patients who are admitted with HA-MRSA infection. Strict Infection control measures must always be observed when interacting with patients who may be infected with HA-MRSA. Conflict of interest None. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Noonan syndrome ([NS1, OMIM 163950]) is a common autosomal dominant disorder characterized by short stature, congenital heart disease, facial dysmorphia and other features such as cryptorchidism, bleeding diathesis, skeletal malformations and mild cognitive delays with variable expressivity. The prevalence of this disorder is estimated to be 1/1000–2500 live births [1], [2], [3]. The varied clinical manifestations observed in Noonan syndrome patients (facial, skeletal cardiac, and hematological, among others) are a result of the involvement of the RAS MAP kinase molecular signaling pathway, as will be shown below.

alence of this disorder is estimated to be 1/1000–2500 live births [1], [2], [3]. The varied clinical manifestations observed in Noonan syndrome patients (facial, skeletal cardiac, and hematological, among others) are a result of the involvement of the RAS MAP kinase molecular signaling pathway, as will be shown below. The previous updates and reviews in the literature have thoroughly discussed Noonan syndrome, often focusing on diagnostic evaluations, clinical guidelines, and management with treatment options, which have amply helped clinicians provide the best care for Noonan syndrome children. The molecular aspects of this disorder have been approached in these reviews progressively with genetic advances. However, considering the fast advances and consecutive progress being made in the physiopathology and genetic studies of Noonan syndrome, it seems important to gather these findings in one paper to help the audience of clinicians and geneticists have a full view of recent advances in the molecular etiology of Noonan syndrome, as well as an authentic prevalence of the mutational rates of its causing-genes. Therefore, this review provides, in the first part, an update on the molecular aspect of the disease, in which we summarize the data concerning clinical features frequently observed, then focus on the molecular etiology, the inheritance pattern and the genetic counseling that should be given to patients. In the second part of this review, we establish and discuss the mutational rate reported up to now in most genes involved in Noonan syndrome.

data concerning clinical features frequently observed, then focus on the molecular etiology, the inheritance pattern and the genetic counseling that should be given to patients. In the second part of this review, we establish and discuss the mutational rate reported up to now in most genes involved in Noonan syndrome. 2 Review 2.1 Clinical features and diagnosis The diagnosis of Noonan syndrome is based primarily on the clinical features that have been established from the very beginning through several clinical studies that have meticulously defined the criteria and signs of diagnosis [1], [2], [3]. 2.1.1 Dysmorphic face The extreme variability of facial traits in Noonan syndrome from one individual to another makes the assessment of frequency difficult and not very meaningful. Furthermore, the dysmorphic signs could change within the same patient depending on his/her age, to be less perceptible in adulthood than earlier in childhood. The dysmorphology is characterized during the postnatal period by a tall forehead, low-set-posteriorly-rotated ears, a thickened helix, nerve deafness, hypertelorism, ptosis, down slanting palpebral fissures, epicanthal folds, deeply grooved philtrum, a high arched palate and triangular face, with a low posterior hairline and webbed neck. In adulthood, the facial features become more subtle, the eyes are less prominent, with a slightly elongated neck, wrinkled skin and high anterior hairline [3], [4], [5].

lpebral fissures, epicanthal folds, deeply grooved philtrum, a high arched palate and triangular face, with a low posterior hairline and webbed neck. In adulthood, the facial features become more subtle, the eyes are less prominent, with a slightly elongated neck, wrinkled skin and high anterior hairline [3], [4], [5]. 2.1.2 Congenital heart defect The cardiac features are well delineated and are estimated to be present in 50% up to 90% of Noonan syndrome patients [6], [7], [8]. The most common congenital heart defects (CHD) are pulmonic stenosis (50–60%), hypertrophic cardiomyopathy (HCM) (20%) and atrial septal defect (6–10%) [9], [10]. The other CHDs such as ventricular septal defect, atrioventricular canal defect, and aortic coarctation are observed less frequently [6], [8], [11]. Electrocardiographic abnormalities were reported in 87% of patients [3]. Electrocardiograms display wide QRS complexes with a predominant negative pattern in the left precordial leads, a left axis deviation and giant Q waves [9], [10], [11]. 2.1.3 Growth/short stature Weight and height are normal at birth. However, during childhood and at puberty, short stature becomes a prominent common sign of Noonan syndrome. In one series reported by Nora et al, the prevalence of Noonan children with height below the 3rd percentile is approximately 83% [12]. In another study, pubertal growth was found to be delayed by almost two years and the mean height was in the 3rd percentile, with female and male average heights of 151 cm and 161 cm respectively, and the average bone age delayed by two years [8].

children with height below the 3rd percentile is approximately 83% [12]. In another study, pubertal growth was found to be delayed by almost two years and the mean height was in the 3rd percentile, with female and male average heights of 151 cm and 161 cm respectively, and the average bone age delayed by two years [8]. 2.1.4 Skeletal defects A chest deformity characterized by superior pectus carinatum and inferior pectus excavatum is observed in up to 95% of Noonan syndrome patients. Half (50%) have cubitus valgus and 30% have a clinobrachydactyly. The other orthopedic features, such as thoracic scoliosis, talipes equinovarus or radiolunar synostosis, are observed less often [3], [7]. 2.1.5 Bleeding defect The coagulation defect is the most common hematologic disorder in Noonan syndrome patients. Approximately 55% of patients have mild to moderate abnormal bleeding, whereas only 3% have major abnormal bleeding [3]. 2.1.6 Genito-urinary Altered spermatogenesis and cryptorchidism are observed in 60–80% of patients [13]. Ten percent of patients have renal abnormalities that often include renal pelvis dilatation [3]. 2.1.7 Ophthalmological features Approximately 55% of Noonan syndrome patients have an abnormal ophthalmological test [3]. The frequent abnormalities are refractive errors (61–70%), strabismus (48–63%), amblyopia (33%) and anterior segment changes (63%) [14], [15]. 2.1.8 Other features Other features have also been reported, including lymphatic abnormalities, hematological malignancy, giant cell lesions and cognitive disability, among others [16], [17], [18], [19].

2.1.7 Ophthalmological features Approximately 55% of Noonan syndrome patients have an abnormal ophthalmological test [3]. The frequent abnormalities are refractive errors (61–70%), strabismus (48–63%), amblyopia (33%) and anterior segment changes (63%) [14], [15]. 2.1.8 Other features Other features have also been reported, including lymphatic abnormalities, hematological malignancy, giant cell lesions and cognitive disability, among others [16], [17], [18], [19]. 2.2 Genetic diagnosis and molecular etiology As shown above, Noonan syndrome is a heterogeneous disorder with various clinical features ranging from facial dysmorphology to short stature and bleeding defects up to congenital heart disease. Those features have been demonstrated by several studies to be the yield of diverse molecular and physiological mechanisms, some of which are well known, while others need to be further elucidated. The molecular etiology of Noonan syndrome has been established first by the study of two large Noonan syndrome families using linkage analysis [20], [21], [22]. These studies suggested that the concerned locus is located in the 12q24 region. Based on these data, and the data suggesting the involvement of SHP-2 in the signaling pathways that control semilunar valvulogenesis [23], and by means of a positional candidacy approach, Tartaglia et al [24] suggested PTPN11 to be a Noonan syndrome candidate gene, and confirmed this hypothesis by bidirectionally sequencing the fifteen exons of PTPN11, which have shown mutations in 50% of the studied cohort.

ays that control semilunar valvulogenesis [23], and by means of a positional candidacy approach, Tartaglia et al [24] suggested PTPN11 to be a Noonan syndrome candidate gene, and confirmed this hypothesis by bidirectionally sequencing the fifteen exons of PTPN11, which have shown mutations in 50% of the studied cohort. 2.2.1 PTPN11 (protein-tyrosine phosphatase, nonreceptor-type 11) The PTPN11 gene ([OMIM 176876]) (Fig. 1A) is organized into three domains: the N-amino terminal src-homology 2 domain (N-SH2) and the phosphotyrosine phosphatase (PTP) domain, which are the most commonly mutated domains; and the C-amino terminal src-homology 2 domain (C-SH2) and carboxy-terminal tail [25], [26].Figure 1 Organization of Noonan syndrome causing genes and domains, A. PTPN11 exons and SHP-2 domains, B. SOS1 exons and domains, C. KRAS coding exons and domains. In most cases, the exon 4a is spliced out. D. RIT1 coding exons and domains. E. RAF1 coding exons and domains with the localization of Ser259 and Ser621 residues that is critical for RAF1 auto-inhibition. F. BRAF coding exons and domains. G. MAP2K1 coding exons and domains. C-SH2: C-amino-terminal src-homology 2, CR: Conserved Region, CRD: Cysteine-Rich Domain, DH: Dbl Homology, HF: Histone-like Fold, HL: Helical Linker, N-SH2: N- amino-terminal src-homology 2, PH: Pleckstrin Homology, Pr: Proline-riche motif, PTP: protein-tyrosine phosphatase, RBD: RAS binding domain, REM: RAS Exchange Motif, Sw: Switch. Figure 1

2.2.1 PTPN11 (protein-tyrosine phosphatase, nonreceptor-type 11) The PTPN11 gene ([OMIM 176876]) (Fig. 1A) is organized into three domains: the N-amino terminal src-homology 2 domain (N-SH2) and the phosphotyrosine phosphatase (PTP) domain, which are the most commonly mutated domains; and the C-amino terminal src-homology 2 domain (C-SH2) and carboxy-terminal tail [25], [26].Figure 1 Organization of Noonan syndrome causing genes and domains, A. PTPN11 exons and SHP-2 domains, B. SOS1 exons and domains, C. KRAS coding exons and domains. In most cases, the exon 4a is spliced out. D. RIT1 coding exons and domains. E. RAF1 coding exons and domains with the localization of Ser259 and Ser621 residues that is critical for RAF1 auto-inhibition. F. BRAF coding exons and domains. G. MAP2K1 coding exons and domains. C-SH2: C-amino-terminal src-homology 2, CR: Conserved Region, CRD: Cysteine-Rich Domain, DH: Dbl Homology, HF: Histone-like Fold, HL: Helical Linker, N-SH2: N- amino-terminal src-homology 2, PH: Pleckstrin Homology, Pr: Proline-riche motif, PTP: protein-tyrosine phosphatase, RBD: RAS binding domain, REM: RAS Exchange Motif, Sw: Switch. Figure 1 Protein SHP-2 coded by PTPN11 was demonstrated to be involved in several developmental processes such as limb development, semilunar valvulogenesis, hemopoietic cell differentiation and mesodermal patterning [23], [27], [28], [29], and is widely expressed in several tissues such as the heart, muscles, and brain, in which SHP-2 modulates the cellular proliferation, migration or differentiation processes during the developmental stage [30]. SHP-2 is a key element of the signaling molecular RAS-MAP Kinase cascade (Fig. 2). The latter is triggered when cytokines, hormones or growth factors bind to membrane receptors [25], [26], [31]. Therefore, its disturbance may cause alterations of these tissues that lead to the aberrant phenotype shown in Noonan syndrome patients. This fact has been confirmed by De Rocca et al, has confirmed that the SHP-2 mutations in Noonan syndrome were associated with the inhibition of GH-induced IGF-1 via the hyperactivation of RAS/ERK1/2 in a mouse model, which lead to the short stature observed in Noonan syndrome [32].Figure 2 Localization of Noonan syndrome causing genes into the RAS-MAP kinase signal transduction pathway.

SHP-2 mutations in Noonan syndrome were associated with the inhibition of GH-induced IGF-1 via the hyperactivation of RAS/ERK1/2 in a mouse model, which lead to the short stature observed in Noonan syndrome [32].Figure 2 Localization of Noonan syndrome causing genes into the RAS-MAP kinase signal transduction pathway. Figure 2 On the molecular level, SHP-2 is a cytosolic phosphatase protein activated by the binding of a N-SH2 domain to phosphotyrosyl residue, which leads to the conformational change making the catalytic site (PTP) available to the phosphotyrosyl residue. In the absence of substrate, SHP-2 regains its inactive form in which N-SH2 interacts with PTP to hide the catalytic site [33]. Most of the PTPN11 mutations causing Noonan syndrome manifestations cluster in these two domains. The energy-based structured analysis suggests that PTPN11 mutations disrupt the inactive conformation, resulting in the emergence of the catalytic site which triggers the phosphotyrosyl phosphatase activity in the absence of the ligand [24]. Additionally, several screening studies were carried out to define PTPN11 mutations in other populations and to highlight putative novel mutations. PTPN11 mutations were significantly correlated to the manifestation of pulmonary stenosis observed in 45–70% of cases [34], [35]. Seven other genes from the RAS-MAP kinase pathway (Fig. 2) were demonstrated to be mutated and to cause Noonan syndrome.

Additionally, several screening studies were carried out to define PTPN11 mutations in other populations and to highlight putative novel mutations. PTPN11 mutations were significantly correlated to the manifestation of pulmonary stenosis observed in 45–70% of cases [34], [35]. Seven other genes from the RAS-MAP kinase pathway (Fig. 2) were demonstrated to be mutated and to cause Noonan syndrome. 2.2.2 SOS1 (Son of Sevenless homolog 1) At present, SOS1 ([OMIM 182530]) is considered the second molecular cause of Noonan syndrome. Roberts et al reported that SOS1 is responsible for approximately 20% of Noonan syndrome patients in the absence of PTPN11 mutations [36]. SOS1 is a guanine exchange factor (GEF) with a major role in the RAS-MAP kinase pathway. It is mapped on the 2p22-p21 region and consists of 23 exons (Fig. 1B) [37], [38] coding for multiple domains containing: histone-like folds domain (HF), Dbl homology domains (DH) and Pleckstrin homology domains (PH) with the following regulatory function; RAS exchanging motif (REM) and Cdc25 domains with catalytic function; helical linker (HL) relating PH and REM, and the PolyProline region [36], [39]. It was reported that Noonan syndrome-causing SOS1 was associated with a high prevalence of ectodermal abnormalities [36], [39], [40], [41]. In the RAS-MAP kinase molecular cascade, RAS protein is inactivated by hydrolyzing GTP to GDP via GTPase Activating Protein (GAP). To be reactivated, RAS protein needs to exchange bound GDP for GTP via SOS1.

It was reported that Noonan syndrome-causing SOS1 was associated with a high prevalence of ectodermal abnormalities [36], [39], [40], [41]. In the RAS-MAP kinase molecular cascade, RAS protein is inactivated by hydrolyzing GTP to GDP via GTPase Activating Protein (GAP). To be reactivated, RAS protein needs to exchange bound GDP for GTP via SOS1. It was suggested that the Dbl homology domain inhibits RAS-GEF activity by competing with RAS for binding to the REM-Cdc25 complex designated as an allosteric site, which stabilizes the inactive conformation [42]. Most mutations cluster in these domains and lead to gain-of-function in the RAS-MAP kinase pathway by disrupting the auto-inhibition of SOS1 RAS-GEF activity [36], [39], [43]. Pulmonic stenosis was more frequently observed in patients with SOS1 mutations (83.3%) than in those with PTPN11 mutations [34]. The RAS family (KRAS, HRAS, NRAS and RIT1) is a small GTPase protein family responsible of signal propagation through the RAS-MAP kinase cascade. It acts as a molecular switch cycling between an inactive GDP bound conformation and an active GTP bound conformation (Fig. 2) [44], [45].

Pulmonic stenosis was more frequently observed in patients with SOS1 mutations (83.3%) than in those with PTPN11 mutations [34]. The RAS family (KRAS, HRAS, NRAS and RIT1) is a small GTPase protein family responsible of signal propagation through the RAS-MAP kinase cascade. It acts as a molecular switch cycling between an inactive GDP bound conformation and an active GTP bound conformation (Fig. 2) [44], [45]. 2.2.3 KRAS (kirsten rat sarcoma viral oncogene homolog) The KRAS ([OMIM 190070]) gene is mapped to the 12p12 region, and consists of 6 exons coding for the P loop, and switch I and switch II domains (Fig. 1C) [46]. In response to a transduction signal generated from membrane receptor binding, KRAS regains its active bound GTP conformation via the guanine nucleotide exchange factor, and entails an activation cascade through the downstream effectors of RAS-MAP kinase (such as RAF1). KRAS has intrinsic GTPase activity, which, supported by RAS-GAPs (RAS-GTPase activating protein), hydrolyzes bound GTP to GDP to inactive itself [44], [45]. Lee et al and Schubbert et al have reported that KRAS mutations cause approximately 5% of Noonan syndrome cases in the absence of PTPN11 mutations [34], [47]. These mutations lead to a gain in the function effect through the RAS-MAP kinase pathway by altering the intrinsic GTPase activity of KRAS and creating insensitivity to RAS-GAPs [46], [47]. Noonan syndrome patients with KRAS mutations were reported to have severe phenotypes with severe mental retardation [46], [47], [48].

Lee et al and Schubbert et al have reported that KRAS mutations cause approximately 5% of Noonan syndrome cases in the absence of PTPN11 mutations [34], [47]. These mutations lead to a gain in the function effect through the RAS-MAP kinase pathway by altering the intrinsic GTPase activity of KRAS and creating insensitivity to RAS-GAPs [46], [47]. Noonan syndrome patients with KRAS mutations were reported to have severe phenotypes with severe mental retardation [46], [47], [48]. 2.2.4 NRAS (neuroblastoma RAS viral oncogene homolog) The NRAS ([OMIM 164790]) gene mapped on 1p13.2 comprises 6 coding exons [49]. Cirstea et al have reported that NRAS mutations are involved in less than 1% of Noonan syndrome cases [50]. 2.2.5 RIT1(Ric-like protein without Caax motif 1) Interestingly, in 2013, Aoki et al reported that mutations in RIT1 ([OMIM 609591]) cause Noonan syndrome [51]. RIT1 is a member of the RAS subfamily of small GTPases (Fig. 2) [52], and consists of 6 exons and is located in the 1q22 region (Fig. 1D).

2.2.4 NRAS (neuroblastoma RAS viral oncogene homolog) The NRAS ([OMIM 164790]) gene mapped on 1p13.2 comprises 6 coding exons [49]. Cirstea et al have reported that NRAS mutations are involved in less than 1% of Noonan syndrome cases [50]. 2.2.5 RIT1(Ric-like protein without Caax motif 1) Interestingly, in 2013, Aoki et al reported that mutations in RIT1 ([OMIM 609591]) cause Noonan syndrome [51]. RIT1 is a member of the RAS subfamily of small GTPases (Fig. 2) [52], and consists of 6 exons and is located in the 1q22 region (Fig. 1D). Aoki et al identified nine missense mutations in seventeen individuals from 180 patients (9%) with Noonan syndrome or related disorders and without mutations in known Noonan syndrome-causing genes, and observed that the frequency of RIT mutations in the Noonan syndrome cohort was seemingly similar to the frequency of RAF1 mutations. This finding was subsequently confirmed by Bertola et al, in 2014, who found the same prevalence (9%), and Gos et al, in 2014, who found a lower mutation rate (3.8%). Those mutation clusters in the G1, Switch I, and more frequently in Switch II domains, were proven to entail a significant activation of the RAS MAPK pathway by hyper-activating transcription factor ELK1. [51], [53], [54].

o found the same prevalence (9%), and Gos et al, in 2014, who found a lower mutation rate (3.8%). Those mutation clusters in the G1, Switch I, and more frequently in Switch II domains, were proven to entail a significant activation of the RAS MAPK pathway by hyper-activating transcription factor ELK1. [51], [53], [54]. Noonan syndrome patients with RIT1 mutations were characterized by a high incidence of congenital heart disease (94%), especially hypertrophic cardiomyopathy (71%) and pulmonic stenosis (65%). The frequency of hypertrophic cardiomyopathy among RIT1 mutation-positive patients is similar to that of RAF1 mutation-positive subjects with Noonan syndrome, which leads to the conclusion that RIT1 and RAF1 interact with each other and have the same effect on cardiac development [51], [55], [56]. The RAF family (ARAF, BRAF and RAF1) has an activation role upstream of the MEK-ERK cascade into the RAS-MAPK pathway (Fig. 2).

Noonan syndrome patients with RIT1 mutations were characterized by a high incidence of congenital heart disease (94%), especially hypertrophic cardiomyopathy (71%) and pulmonic stenosis (65%). The frequency of hypertrophic cardiomyopathy among RIT1 mutation-positive patients is similar to that of RAF1 mutation-positive subjects with Noonan syndrome, which leads to the conclusion that RIT1 and RAF1 interact with each other and have the same effect on cardiac development [51], [55], [56]. The RAF family (ARAF, BRAF and RAF1) has an activation role upstream of the MEK-ERK cascade into the RAS-MAPK pathway (Fig. 2). 2.2.6 RAF1 (v-raf-1 murine leukemia viral oncogene homolog 1) RAF1 ([OMIM 164760]) mutations cause 3–17% of Noonan syndrome cases. RAF1 consists of 17 exons coding for multidomain protein that act as a serine–threonine kinase (Fig. 1E) [55], [57], [58]. This protein comprises three conserved regions (CR): CR1 that contains the RAS binding domain (RBD) and cysteine-rich domain (CRD), which are both suggested to be involved in the negative regulation of RAF1 by direct physical interaction [59]; CR2, which is an important region where many RAF1-activating mutations and clusters lie. Eighty percent (80%) RAF1 mutations in Noonan syndrome have HCM. It contains Ser259 residue, which is critical to RAF1 auto-inhibition [55]; and the third conserved region (CR3) in the C-terminal region of RAF1 that is responsible for catalytic activity [55], [57]. The RAF1 inactive conformation is maintained via 14-3-3 protein dimers that bind to phosphorylated Ser259 and Ser621 in such a way that the catalytic site remains hidden [60]. Mutations in these conserved residues or in its flanking residues (such as Arg256 and Pro261, among others) prevent the 14-3-3 binding leading to an auto-inhibition fail and the activation of the RAS-MAP kinase cascade [57].

to phosphorylated Ser259 and Ser621 in such a way that the catalytic site remains hidden [60]. Mutations in these conserved residues or in its flanking residues (such as Arg256 and Pro261, among others) prevent the 14-3-3 binding leading to an auto-inhibition fail and the activation of the RAS-MAP kinase cascade [57]. Noonan syndrome-causing RAF1 mutations were strongly correlated to hypertrophic cardiomyopathy. The latter was observed in 95% of Noonan syndrome patients, and seems to have an allele specificity that is associated with Ser259 and Ser612 mutations (P < .0001) [57]. In 2015, De Iriarte Rodrıguez et al proved that reduced levels of RAF1 in mice may lead to hearing impairments [61]. 2.2.7 BRAF (V-Raf murine sarcoma viral oncogene homolog B1) BRAF ([OMIM 164757]), a member of the RAF family, was proven to be involved in Noonan syndrome pathogenesis by enhancing ERK activation [55], [62]. However, the frequency of these mutations reported in a large cohort screening of Noonan syndrome patients were lower than those observed in RAF1, approximately 1.7–1.9%. These mutations cluster predominantly in conserved regions of CR1 and CR3 (Fig. 1F), but modestly increase kinase activity compared to RAF1 [34], [63]. 2.2.8 MAP2K1 (mitogen activated protein kinase 1) MAP2K1 ([OMIM 176872]) is mapped to the 15q22 region and comprises 11 exons encoding the MEK protein, which is a dual-specificity kinase with a major protein kinase domain that activates the extracellular-signal-regulated (Erk) mitogen-activated protein (MAP) kinases (Fig. 1G) [64].

(mitogen activated protein kinase 1) MAP2K1 ([OMIM 176872]) is mapped to the 15q22 region and comprises 11 exons encoding the MEK protein, which is a dual-specificity kinase with a major protein kinase domain that activates the extracellular-signal-regulated (Erk) mitogen-activated protein (MAP) kinases (Fig. 1G) [64]. Previous studies confirmed that MEK plays a crucial role in the embryonic developmental process, especially in cell migration and placental development [64]. In 2007, Nava et al identified MAP2K1 mutations in 4.2% of Noonan syndrome patients who are negative for PTPN11 and SO1 mutations [65]. This finding proved that mutations in the MEK protein kinase domain increase MEK basal kinase activity. This hyperactivity entails an increased phosphorylation of the downstream targets (such as ERK1 and ERK2) [66]. Recently, with the progress of molecular screening tools, whole genome sequencing (WGS) and whole exome sequencing (WES) have become increasingly used in the screening of developmental diseases. As a result, several novel Noonan syndrome-causing genes were highlighted in the last two years. 2.2.9 SOS2 (Son of Sevenless homolog 2) In 2015, Yamamoto et al reported that SOS2 ([OMIM 601247]), the homolog of SOS1, is responsible of 4% of Noonan syndrome cases with no mutation in the genes previously associated with Noonan syndrome. SOS1 and SOS2 are 70% homologous. The reported mutations cluster in the DH domain, which plays a pivotal role in the stabilization of inactive conformation. Similar to SOS1, these mutations were associated with ectodermal defects [67].

rome cases with no mutation in the genes previously associated with Noonan syndrome. SOS1 and SOS2 are 70% homologous. The reported mutations cluster in the DH domain, which plays a pivotal role in the stabilization of inactive conformation. Similar to SOS1, these mutations were associated with ectodermal defects [67]. 2.2.10 LZTR1 (leucine-zipper-like transcription regulator 1) In the same study, Yamamoto et al identified pathogenic mutations in a candidate gene not associated with the RAS/MAPK pathway, the LZTR1 gene ([OMIM 600574]). The prevalence of these mutations in the same studied cohort was 8% [67]. The LZTR1 gene is located in 22q11.21 and consists of 21 exons and encodes a protein member of the BTB-kelch superfamily. It was suggested that this gene may have a crucial role in the control of fundamental cellular processes (such as the cell cycle and the regulation of chromatin conformation) [68], [69]. However, it is worth mentioning that LZTR1 is already known to be associated with the Schwannomatosis, a form of neurofibromatosis [70].

The LZTR1 gene is located in 22q11.21 and consists of 21 exons and encodes a protein member of the BTB-kelch superfamily. It was suggested that this gene may have a crucial role in the control of fundamental cellular processes (such as the cell cycle and the regulation of chromatin conformation) [68], [69]. However, it is worth mentioning that LZTR1 is already known to be associated with the Schwannomatosis, a form of neurofibromatosis [70]. 2.2.11 A2ML1 (α-2-macroglobulin (A2M)-like-1) Vissers et al carried out exome sequencing in one Noonan syndrome case-parent trio and found a de novo mutation affecting a highly conserved residue of A2ML1 ([OMIM 610627]), encouraging them to screen an additional cohort of 155 Noonan syndrome patients. This study showed the involvement of A2ML1 mutations in approximately 1% of Noonan syndrome subjects negative for the other NS major genes [71]. A2ML1 is a member of the α-macroglobulin superfamily, localized in 12p13 region with 35 coding exons. It acts as a protease inhibitor upstream of the MAPK pathway [72]. Mutations in A2ML1 are said to cause developmental defects, but the ways in which mutation affects the MAP kinase pathway needs to be further elucidated [71]. Other genes carrying rare variants in a Noonan syndrome population were also highlighted recently, in particular, RASA2, MAP3K8 and SPRY [73].

2.2.11 A2ML1 (α-2-macroglobulin (A2M)-like-1) Vissers et al carried out exome sequencing in one Noonan syndrome case-parent trio and found a de novo mutation affecting a highly conserved residue of A2ML1 ([OMIM 610627]), encouraging them to screen an additional cohort of 155 Noonan syndrome patients. This study showed the involvement of A2ML1 mutations in approximately 1% of Noonan syndrome subjects negative for the other NS major genes [71]. A2ML1 is a member of the α-macroglobulin superfamily, localized in 12p13 region with 35 coding exons. It acts as a protease inhibitor upstream of the MAPK pathway [72]. Mutations in A2ML1 are said to cause developmental defects, but the ways in which mutation affects the MAP kinase pathway needs to be further elucidated [71]. Other genes carrying rare variants in a Noonan syndrome population were also highlighted recently, in particular, RASA2, MAP3K8 and SPRY [73]. 2.2.12 Other causes Some studies have suggested other causes of some Noonan syndrome manifestations that are not related to a particular gene. For instance, wide-spaced nipples, cryptorchidism and some traits of facial dysmorphia such as hypertelorism, down slanting palpebral fissures, ptosis and low-set posteriorly rotated ears are suggested to be the result of tissue migration disruption or organ displacement caused by lymphedema at the intrauterine stage. Webbing of the neck and prominence of the trapezius may result from cystic hygroma at early intrauterine stages [74].

anting palpebral fissures, ptosis and low-set posteriorly rotated ears are suggested to be the result of tissue migration disruption or organ displacement caused by lymphedema at the intrauterine stage. Webbing of the neck and prominence of the trapezius may result from cystic hygroma at early intrauterine stages [74]. Recently, it was suggested that the impairment of testicular function responsible for delayed puberty or infertility is due to Sertoli and Leydig cell dysfunction rather than cryptorchidism [75]. The coagulation defect in Noonan syndrome was explained by the fact that the genes causing Noonan syndrome interact with the regulation process of the genes involved in the coagulation pathway [76]. 2.3 Assessment of the mutation rate In the second part of this review, we gathered and discussed mutational screening data reported in previous studies, to produce valid frequencies and authentic conclusions about mutational rates in Noonan syndrome ([NS, OMIM 163950]). 2.3.1 Search strategy The data gathered were extracted from original articles published up to now (December 2015) in the “PubMed”, “ScienceDirect” and “Wiley Online Library” databases using the following key words:- “Noonan” in ([Abstract]). - “Noonan” in ([Title/Abstract]) AND “PTPN11” in ([Title]), - “Noonan” in ([Title/Abstract]) AND “SOS1” in ([Title]), and the same goes for the other genes (RAF1, KRAS, BRAF, NRAS and RIT1).

2.3.1 Search strategy The data gathered were extracted from original articles published up to now (December 2015) in the “PubMed”, “ScienceDirect” and “Wiley Online Library” databases using the following key words:- “Noonan” in ([Abstract]). - “Noonan” in ([Title/Abstract]) AND “PTPN11” in ([Title]), - “Noonan” in ([Title/Abstract]) AND “SOS1” in ([Title]), and the same goes for the other genes (RAF1, KRAS, BRAF, NRAS and RIT1). 2.3.2 Inclusion criteria From all of the articles gathered, we considered only studies comprising genetic screening of patient's series. We also considered clinical or endocrinological Noonan syndrome studies based initially on genetic screening. The genes considered in this part of the paper should have been screened by at least 3 studies meeting the inclusion criteria. On the other hand, to determine the exact frequency of Noonan syndrome mutations, we excluded all data obtained from the screening of cohorts initially presenting with a particular condition (i.e., the mutational screening of Noonan syndrome patients with cardiac hypertrophy). The data gathered from the 82 included studies are summarized in Table 1 [77], [78], [79], [80], [81], [82], [83], [84], [85], [86], [87], [88], [89], [90], [91], [92], [93], [94], [95], [96], [97], [98], [99], [100], [101], [102], [103], [104], [105], [106], [107], [108].Table 1 Summary of the prevalence of Noonan syndrome-causing genes through previous studies.

s are summarized in Table 1 [77], [78], [79], [80], [81], [82], [83], [84], [85], [86], [87], [88], [89], [90], [91], [92], [93], [94], [95], [96], [97], [98], [99], [100], [101], [102], [103], [104], [105], [106], [107], [108].Table 1 Summary of the prevalence of Noonan syndrome-causing genes through previous studies. Table 1Genes N° of mutated subjects/total of subjects (% of mutations) % of gene in genetic etiology First year of gene study N° of studies References PTPN11 814/1917 (42.5%) 52.6 2001 29 [19], [24], [34], [35], [58], [77], [83], [84], [85], [86], [87], [88], [89], [90], [91], [92], [93], [94], [95], [96], [97], [98], [99], [100], [101], [102], [103], [104], [105] SOS1 242/1472 (16.4%) 20.3 2007 16 [19], [34], [36], [39], [43], [58], [78], [79], [97], [98], [102], [103], [104], [105], [106], [107] RIT1 31/383 (8%) 10 2013 4 [51], [53], [54], [73] RAF1 76/961 (8%) 10 2007 11 [34], [55], [56], [57], [58], [78], [79], [98], [103], [105], [107] KRAS 31/1087 (2.8%) 3.4 2006 11 [46], [47], [48], [58], [65], [79], [89], [98], [102], [103], [105], [108] BRAF 19/815 (2.3%) 2.8 2009 7 [34], [63], [78], [98], [103], [105], [108] NRAS 10/1213 (0.8%) 1 2010 4 [50], [80], [81], [82]

961 (8%) 10 2007 11 [34], [55], [56], [57], [58], [78], [79], [98], [103], [105], [107] KRAS 31/1087 (2.8%) 3.4 2006 11 [46], [47], [48], [58], [65], [79], [89], [98], [102], [103], [105], [108] BRAF 19/815 (2.3%) 2.8 2009 7 [34], [63], [78], [98], [103], [105], [108] NRAS 10/1213 (0.8%) 1 2010 4 [50], [80], [81], [82] 2.3.3 Mutational rate of Noonan syndrome-causing genes Through the analyzed data (Table 1), we found that PTPN11 is the most studied gene in Noonan syndrome populations (29 studies vs. 16 studies or less for other genes). This may be because PTPN11 was the first gene of the RAS MAPK pathway to be highlighted in 2001, whereas the second gene involved (KRAS) was found 5 years later, followed by SHP-2. Indeed, the second gene, KRAS, was known beforehand to be involved in malignancy disorders through somatic mutations, while its germline mutations were associated for the first time with Noonan syndrome in 2006. Then, in 2007, three genes were highlighted, (SOS1, RAF1 and MAP2K1), after which the other RAS/MAP kinase genes, BRAF, NRAS and RIT1, were proven to be involved in Noonan syndrome in more recent, in 2009, 2010 and 2013, respectively. It should be noted that the chronology of the gene identification does not consistently occur in the “more involved” to “less involved” direction. Indeed, RIT1, which was discovered in 2013, was proven to be involved in 8% of cases. This frequency is much higher than the frequency of some genes (BRAF, KRAS and NRAS) that were found first, while their incidences do not exceed 3% (2.3%, 2.8% and 0.8%, respectively).

the “more involved” to “less involved” direction. Indeed, RIT1, which was discovered in 2013, was proven to be involved in 8% of cases. This frequency is much higher than the frequency of some genes (BRAF, KRAS and NRAS) that were found first, while their incidences do not exceed 3% (2.3%, 2.8% and 0.8%, respectively). The first most common genetic cause of Noonan syndrome is still PTPN11 mutations that are observed in 42.5% of Noonan syndrome patients. Those mutations constitute 52.6% of all mutations detected up to now (Table 1). The second most-involved gene is SOS1, which is altered in 16.4% of patients. On the other hand, we found that RIT1 and RAF1 have the same prevalence (8%) and are the third most-involved genes. Taken together, PTPN11, SOS1, RAF1 and RIT1 cover 93% of reported mutations (Table 1). Therefore, these genes should be systematically considered in the genetic diagnosis of Noonan syndrome. Among the RAS subfamily members of the RAS/MAP kinase pathway reported in Noonan syndrome, RIT1 seems to be the most involved gene compared with KRAS and NRAS, which have the lowest frequency among all reported cases (2.8% and 0.8%, respectively); however, among RAF family members, RAF1 is the most involved gene compared to BRAF (8% versus 2.3%, respectively). Among these two families, neither HRAS nor ARAF were reported to cause Noonan syndrome. These divergences should attract more attention from researchers.

g all reported cases (2.8% and 0.8%, respectively); however, among RAF family members, RAF1 is the most involved gene compared to BRAF (8% versus 2.3%, respectively). Among these two families, neither HRAS nor ARAF were reported to cause Noonan syndrome. These divergences should attract more attention from researchers. It is worth mentioning that there were no mutations in the SHOC2, RRAS and CBL genes in Noonan syndrome patients. These genes were reported to be involved in Noonan-like syndrome (NLS) only. 2.4 Inheritance and genetic counseling Noonan syndrome is a common disorder that could occur as a sporadic condition or can be inherited in an autosomal dominant pattern. In the latter case, the affected parent who transmits the disorder is predominantly the mother. This may be explained by the adverse impact of Noonan syndrome on male fertility [7], [13], [108]. However, rare autosomal recessive cases of Noonan syndrome with consanguineous parents have also been reported [109]. When there is molecular evidence in one Noonan syndrome child, the clinical and molecular evaluation of parents becomes primordial in the determination of recurrence risks. In Noonan syndrome-causing genes with heterozygous mutations, the recurrence risk in one genetically affected parent is 50% per each pregnancy, while in an unaffected parent with Noonan syndrome child, the recurrence risk in subsequent pregnancies is very low, approximately 1–5% owing to putative germline mosaicism.

s. In Noonan syndrome-causing genes with heterozygous mutations, the recurrence risk in one genetically affected parent is 50% per each pregnancy, while in an unaffected parent with Noonan syndrome child, the recurrence risk in subsequent pregnancies is very low, approximately 1–5% owing to putative germline mosaicism. The Noonan syndrome patient and their family should be made aware of the consequences of some Noonan syndrome features that may arise or complicate with adulthood, such as fertility problems in cryptorchidism cases and cardiac and hematological complications, which need rigorous follow-up [110]. The de-novo PTPN11 mutations are known to be more recurrent in Noonan syndrome. Tartaglia et al suggested that de-novo mutations have a predominantly paternal origin. They also demonstrated that the paternal average age in sporadic Noonan cases was significantly higher than that of the general population [111]. 3 Conclusion Noonan syndrome is a common multigenic disease. Its clinical side has been profoundly discussed by several researchers and diagnostic guides as support has been developed. This review completes the efforts of researchers focusing on the genetic etiology side of this syndrome and explains the physiopathological intervention of different genes in the manifestation of Noonan syndrome traits. In the second part, this paper provides pediatricians and geneticists with the mutational prevalence of genes involved in Noonan syndrome, which was deduced from the analysis of most studies carried out so far.

plains the physiopathological intervention of different genes in the manifestation of Noonan syndrome traits. In the second part, this paper provides pediatricians and geneticists with the mutational prevalence of genes involved in Noonan syndrome, which was deduced from the analysis of most studies carried out so far. The result of this prevalence analysis leads to interesting conclusions, in particular, the high mutation rates observed in PTPN11, SOS1, RAF1 and RIT1, which cover 93% of reported mutations, suggesting that those genes should be systematically considered in the genetic diagnosis of Noonan syndrome. Secondly, we draw attention to the significant differences observed between mutational rates in genes of the same family (genes of the same level of the cascade). The explanation of this divergence and the other points discussed in this review may lead to new insights in the genetic research field of Noonan syndrome. Conflict of interest The authors have no conflict of interest to report. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.

1 Introduction Methotrexate inhibits dihydrofolate reductase and was initially developed as an anti-cancer treatment in the 1940s [1]. The most commonly described side-effects of high dose methotrexate (HDMTX) therapy are myelosuppression, oral mucositis, and acute liver toxicity with transient elevation of transaminase levels [1]. These adverse effects increase the risk of infection which can delay the scheduled therapy. There is a difference between methotrexate (MTX) associated myelosuppression and toxicity. MTX-associated toxicity is related to several factors including drug dose, the duration of administration, patient risk factors, and genetic factors [2], [3]. In addition, the criteria used to assess toxicity are variable. For example, a study showed the most common toxicity was hematologic toxicity, and 64–87% of the patients developed grade 3 and higher toxicity [3], [4]. The incidence of transient high levels of transaminase was reported as 64% [5]. There are currently no data regarding the use of HDMTX pharmacokinetic and toxicity information to predict hepatic and hematologic toxicity in children with acute lymphoblastic leukemia. Csordas et al [6] determined there is no correlation between MTX level and hematologic toxicity. However, Rask et al [4] reported there is a relationship between elevated serum MTX levels and hematologic toxicity. In this study, we aimed to determine the hepatic and hematologic toxicity frequency and assessed whether there is a significant relationship between treatment toxicity and MTX42 levels in children taking different doses of HDMTX.

1 Introduction Methotrexate inhibits dihydrofolate reductase and was initially developed as an anti-cancer treatment in the 1940s [1]. The most commonly described side-effects of high dose methotrexate (HDMTX) therapy are myelosuppression, oral mucositis, and acute liver toxicity with transient elevation of transaminase levels [1]. These adverse effects increase the risk of infection which can delay the scheduled therapy. There is a difference between methotrexate (MTX) associated myelosuppression and toxicity. MTX-associated toxicity is related to several factors including drug dose, the duration of administration, patient risk factors, and genetic factors [2], [3]. In addition, the criteria used to assess toxicity are variable. For example, a study showed the most common toxicity was hematologic toxicity, and 64–87% of the patients developed grade 3 and higher toxicity [3], [4]. The incidence of transient high levels of transaminase was reported as 64% [5]. There are currently no data regarding the use of HDMTX pharmacokinetic and toxicity information to predict hepatic and hematologic toxicity in children with acute lymphoblastic leukemia. Csordas et al [6] determined there is no correlation between MTX level and hematologic toxicity. However, Rask et al [4] reported there is a relationship between elevated serum MTX levels and hematologic toxicity. In this study, we aimed to determine the hepatic and hematologic toxicity frequency and assessed whether there is a significant relationship between treatment toxicity and MTX42 levels in children taking different doses of HDMTX. 2 Patients and methods This study included 48 children diagnosed with ALL at Eskişehir Osmangazi University Faculty of Medicine, Pediatric Hematology/Oncology Department between January 2010 and April 2015. The age, gender, leukemia immune phenotype, and the administered HDMTX doses were retrospectively recorded from the file records. The patient hemoglobin (Hb), absolute neutrophil count (ANC), platelet (PLT) count, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels were recorded before MTX administration and on the 7th day following drug infusion.

dministered HDMTX doses were retrospectively recorded from the file records. The patient hemoglobin (Hb), absolute neutrophil count (ANC), platelet (PLT) count, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels were recorded before MTX administration and on the 7th day following drug infusion. The patients were stratified into three risk groups. The first risk group was defined using the following criteria: (1) Standard risk (SR) patients have an initial leukocyte count < 20 × 109/L and age ≥1 year or <6 years and absolute blast count in the peripheral blood on day 8 after 7 days of prednisolone treatment <1 × 109/L and M1 (<5% blasts) or M2 (≥5% to <25% blasts) marrow on day 15, and M1 marrow on day 33 (all criteria must be fulfilled). The second group (2) was the high risk (HR) group and is defined as at least one of the following: absolute blast count in the peripheral blood on day 8 after 7 days of prednisolone treatment >1 × 109/L, M3 (>25% blasts) marrow on day 15, M2 or M3 marrow on day 33, t (9; 22) (BCR-ABL), t (4; 11) (MLL-AF4), or hypodiploidy ≤ 45. The third group (3) was the Intermediate risk (IR) grouped and was defined as the patients who were classified as neither SR nor HR. The HR patients were not included in the study because their consolidation treatment is different.

or M3 marrow on day 33, t (9; 22) (BCR-ABL), t (4; 11) (MLL-AF4), or hypodiploidy ≤ 45. The third group (3) was the Intermediate risk (IR) grouped and was defined as the patients who were classified as neither SR nor HR. The HR patients were not included in the study because their consolidation treatment is different. All patients received 4 HDMTX infusions at 2-week intervals on days 8, 22, 36, and 50 of the consolidation phase. The patients also received oral 6-mercaptopurine (6-MP) at a dose of 25 mg/m2/day continuously during the consolidation phase of chemotherapy in addition to HDMTX. Any concurrent treatment with trimoxazole was paused in all patients 2 days before and 3 days after the HDTMX infusion. We excluded patients whose absolute neutrophil count was <1 × 109/L with a platelet count <75 × 109/L or had liver function tests above the upper limits before the HDMTX infusion.

n addition to HDMTX. Any concurrent treatment with trimoxazole was paused in all patients 2 days before and 3 days after the HDTMX infusion. We excluded patients whose absolute neutrophil count was <1 × 109/L with a platelet count <75 × 109/L or had liver function tests above the upper limits before the HDMTX infusion. The patients received 3 different doses of HDMTX. The preB ALL SR patients received 2 g/m2 which is in accordance with the ALL-IC BFM 2009 protocol. The preB ALL IR and T ALL SR/IR patients received 5 g/m2 of HDTMX. According to BFM TRALL 2000 protocol, preB ALL SR/IR patients received 1 g/m2 of HDTMX. As per BFM TRALL 2000 protocol, the 1 g/m2 was given within 36 h and as per the ALLIC BFM 2009 protocol the doses of 2 and 5 g/m2 were administered via 24-h of infusion. Alkalization was used to maintain the urine pH ≥ 7 from −4 h through +72 h after the start of the MTX infusion. The urine was tested by dipstick. Briefly, a bolus infusion of 2 mmol/kg NaHCO3 with 2 ml/kg distilled water was delivered over 1 h. Thereafter, 500 ml 0.45% NaCl/5% dextrose +40 mmol NaHCO3 + 10 ml KCl 7.45% hydration fluid was administered during 4 h. The HDMTX infusion was started only when a urine pH was greater than 7. The bolus infusion was repeated if the urine pH was less than 7. All patients received a loading dose of methotrexate as 1/10 of the total MTX dose over 30 min. The remaining 9/10 dose was given over the following 23 h 30 min or 35 h 30 min for children treated with 2 g/m2, 5 g/m2, and 1 g/m2, respectively. We provided a total 3000 ml/m2 parallel hydration with 0.45% NaCl/5% dextrose containing NaHCO3 (180 mmol/m2/24 h) + KCl 7.45% (90 ml/m2/24 h) during the 72 h or 48 h to children treated with 2 g/m2, 5 g/m2, and 1 g/m2, respectively.

min or 35 h 30 min for children treated with 2 g/m2, 5 g/m2, and 1 g/m2, respectively. We provided a total 3000 ml/m2 parallel hydration with 0.45% NaCl/5% dextrose containing NaHCO3 (180 mmol/m2/24 h) + KCl 7.45% (90 ml/m2/24 h) during the 72 h or 48 h to children treated with 2 g/m2, 5 g/m2, and 1 g/m2, respectively. The patients also received standard calcium leucovorin administered via the intravenous route (IV) at a dose of 15 mg/m2 at 42, 48, and 54 h in patients receiving 1 g/m2, 2 g/m2, and 5 g/m2, respectively. A MTX level at 42 h > 1 μmol/L was defined as extended MTX elimination. These patients were treated with 30 mg/m2 of calcium leucovorin at 42 h. The MTX serum levels were monitored until they decreased below 0.25 μmol/L. The blood toxicity and the liver enzyme levels were classified based on the Common Terminology Criteria for Adverse Events 2010 guideline (Table 1) [7]. Myelotoxicity was defined as hemoglobin (Hb) < 10 g/L and absolute neutrophil count (ANC) <1 × 109/L or platelet (PLT) count <75 × 109/L. Transaminase levels of ≥5 times the upper limit were assessed as ≥ grade 3 hepatotoxicity. The reference range of our hospital for transaminases is 40 IU/L.Table 1 Toxicity criteria according to the Common Terminology Criteria for Adverse Events (CTCAE) 2010 guideline.

C) <1 × 109/L or platelet (PLT) count <75 × 109/L. Transaminase levels of ≥5 times the upper limit were assessed as ≥ grade 3 hepatotoxicity. The reference range of our hospital for transaminases is 40 IU/L.Table 1 Toxicity criteria according to the Common Terminology Criteria for Adverse Events (CTCAE) 2010 guideline. Table 1 Grade 1 Grade 2 Grade 3 Grade 4 Hb (g/L) LLN-10 8–10 <8 Life-threatening anemia ANC(×109/L) LLN-1.5 × 1 1.5–1 1–0.5 <0.5 PLT (×109/L) LLN-75 75–50 50–25 <25 AST and ALT (IU/L) >ULN-3 × ULN 3–5 × ULN 5–20 × ULN >20 × ULN Hb: Hemoglobin, ANC: Absolute neutrophil count, PLT: Platelet count, AST: Aspartate aminotransferase, ALT: Alanine aminotransferase, LLN: Lower limit of normal, ULN: Upper limit of normal. The myelotoxicity and hepatotoxicity frequency was investigated after repeated cycles, then, the toxicity frequencies for the three groups were compared. The MTX42 levels are the drug concentration values when the first calcium leucovorin treatment is provided. The MTX42 levels in patients with and without hepatic toxicity were compared in the patients receiving 2 and 5 g/m2. The correlation between MTX42h levels and Hb, AST, ALT, ANC, and PLT count were evaluated for each cycle. There was no evaluation conducted for the patients receiving 1 g/m2 as their MTX levels were not obtained.

The myelotoxicity and hepatotoxicity frequency was investigated after repeated cycles, then, the toxicity frequencies for the three groups were compared. The MTX42 levels are the drug concentration values when the first calcium leucovorin treatment is provided. The MTX42 levels in patients with and without hepatic toxicity were compared in the patients receiving 2 and 5 g/m2. The correlation between MTX42h levels and Hb, AST, ALT, ANC, and PLT count were evaluated for each cycle. There was no evaluation conducted for the patients receiving 1 g/m2 as their MTX levels were not obtained. 3 Statistical analysis The analyses were conducted using SPSS 21 software. The patients were grouped based on drug dose. The normality of distributions was evaluated with the Kolmogorov–Smirnov test. The mean pre- and post-infusion values were analyzed with the paired sample t-test and the Wilcoxon test. We used the Analysis of variance (ANOVA) for independent groups. The comparison of MTX42h levels was performed using the Mann Whitney U-test. The Pearson correlation test was used to evaluate the correlation between variables. All P < .05 values were considered statistically significant.

e t-test and the Wilcoxon test. We used the Analysis of variance (ANOVA) for independent groups. The comparison of MTX42h levels was performed using the Mann Whitney U-test. The Pearson correlation test was used to evaluate the correlation between variables. All P < .05 values were considered statistically significant. 4 Results In this study, there were 192 HDMTX infusions delivered to 48 leukemic children (1 g/m2: 17 children, 68 infusions; 2 g/m2: 14 children, 56 infusions; 5 g/m2: 17 children, 68 infusions) between 2 and 17 years of age. There were 25 females (52%) and 23 males (48%). The mean patient age of those who received 1 g/m2, 2 g/m2, and 5 g/m2 were 6.17 (2.94) years, 3.78 (3.06) years, and 8.23 (5.25) years, respectively. There were 46 cases with pre- B ALL and 2 cases with ALL. The clinical characteristics of the children are presented in Table 2.Table 2 Clinical characteristics of children. Table 2Dose 1 g/m2 (n = 17) 2 g/m2 (n = 14) 5 g/m2 (n = 17) Infusions 68 56 68 Gender (F/M) 5/12 7/7 13/4 SR/IR 13/4 13/1 –/17 Age 6.17 (2.94) 3.78 (3.06) 8.23 (5.25) Immunophenotype Pre B-ALL (17) Pre B-ALL (13) T-ALL (1) Pre-B ALL (16) T-ALL (1) SR: Standard risk, IR: Intermediate risk.

4 Results In this study, there were 192 HDMTX infusions delivered to 48 leukemic children (1 g/m2: 17 children, 68 infusions; 2 g/m2: 14 children, 56 infusions; 5 g/m2: 17 children, 68 infusions) between 2 and 17 years of age. There were 25 females (52%) and 23 males (48%). The mean patient age of those who received 1 g/m2, 2 g/m2, and 5 g/m2 were 6.17 (2.94) years, 3.78 (3.06) years, and 8.23 (5.25) years, respectively. There were 46 cases with pre- B ALL and 2 cases with ALL. The clinical characteristics of the children are presented in Table 2.Table 2 Clinical characteristics of children. Table 2Dose 1 g/m2 (n = 17) 2 g/m2 (n = 14) 5 g/m2 (n = 17) Infusions 68 56 68 Gender (F/M) 5/12 7/7 13/4 SR/IR 13/4 13/1 –/17 Age 6.17 (2.94) 3.78 (3.06) 8.23 (5.25) Immunophenotype Pre B-ALL (17) Pre B-ALL (13) T-ALL (1) Pre-B ALL (16) T-ALL (1) SR: Standard risk, IR: Intermediate risk. We detected a statistically significant reduction in Hb level and ANC (P < .05, P < .001, respectively) by comparing the mean laboratory data obtained prior to drug infusion and at day 7. There was no statistically significant difference in the PLT count. However, there was a statistically significant increase in ALT and AST levels (P < .001, for both) (Table 3).Table 3 The mean values of Hb, AST, ALT with PLT and ANC before and after 7 days of HDMTX infusion.

tained prior to drug infusion and at day 7. There was no statistically significant difference in the PLT count. However, there was a statistically significant increase in ALT and AST levels (P < .001, for both) (Table 3).Table 3 The mean values of Hb, AST, ALT with PLT and ANC before and after 7 days of HDMTX infusion. Table 3 Before infusion After 7 days of HDMTX infusion P Hb (g/L) 10.70 (1.07) 10.47 (1.28) <.05a ANC(×109L) 1663.22 (587.61) 1280.26 (583.54) <.001a PLT (×109L) 195875.00 (148812.50–237062.50) 182750.00 (142750.00–226500.00) >.05b ALT (IU/L) 24.25 (19.75–35.68) 51.50 (32.62–123.37) <.001b AST (IU/L) 26.50 (24.06–30.18) 40.37 (29.62–62.00) <.001b Hb: Hemoglobin, ANC: Absolute neutrophil count, PLT: Platelet count, ALT: Alanine aminotransferase, AST: Aspartate aminotransferase. a Paired sample t test. b Wilcoxon test. Neutropenia ≥ grade 3 developed in 23.5%, 19.6%, and 20.5% of the HDMTX infusions in the patients receiving 1, 2 and 5 g/m2, respectively. Myelotoxicity was observed in 12 (70.5%), 10 (71.4%), and 12 (70.5%) children following 24 (35.2%), 23 (41%), and 23 (33.8%) infusions in patients receiving 1 g/m2, 2 g/m2, and 5 g/m2, respectively. Hepatotoxicity ≥ grade 3 was detected in 5 (29.4%), 5 (35.7%), and 4 (23.5%) patients after 9 (13.2%), 7 (12.5%) and 8 (11.7%) infusions in children receiving 1 g/m2, 2 g/m2, and 5 g/m2, respectively (Table 4). There was no statistically difference in the frequency of myelotoxicity and hepatotoxicity grade ≥3 (Table 5).Table 4 Toxicity frequencies with regard to doses.

5 (35.7%), and 4 (23.5%) patients after 9 (13.2%), 7 (12.5%) and 8 (11.7%) infusions in children receiving 1 g/m2, 2 g/m2, and 5 g/m2, respectively (Table 4). There was no statistically difference in the frequency of myelotoxicity and hepatotoxicity grade ≥3 (Table 5).Table 4 Toxicity frequencies with regard to doses. Table 4 1 g/m2 2 g/m2 5 g/m2 İnfusion (n/%) Patient (n/%) İnfusion (n/%) Patient (n/%) İnfusion (n/%) Patient (n/%) ANC<1 × 109/L 16 (%23.5) 13 (%76.4) 11 (%19.6) 9 (%64.2) 14 (%20.5) 10 (%58.8) PLT<75 × 109/L 3 (%4.4) 3 (%17.6) 6 (%10.7) 2 (%14.2) 1 (%1.4) 1 (%5.8) Hb < 10 g/L & ANC<1 × 109/L 21 (%30.8) 9 (%52.9) 17 (%30.3) 8 (%57.2) 22 (%32.4) 11 (%64.7) Myelotoxicity 24 (%35.2) 12 (%70.5) 23 (%41) 10 (%71.4) 23 (%33.8) 12 (%70.5) ALT 40–120 IU/L (Grade 1) 21 (%25) 13 (%76.4) 19 (%33.9) 11 (%78.5) 16 (%23.5) 9 (%52.9) ALT 120–200 (Grade 2) 5 (%7.3) 5 (%29.4) 5 (%8.9) 4 (%28.5) 3 (%4.4) 3 (%17.6) ALT 200–800 (Grade 3) 8 (%11.7) 4 (%23.5) 5 (%8.9) 4 (%28.5) 5 (%7.3) 4 (%23.5) ALT>800 (Grade 4) 1 1 1 1 – – Hepatotoxicity grade≥ 3 9 (%13.2) 5 (%29.4) 6 (%10.7) 5 (%35.7) 5 (%7.3) 4 (%23.5) Myelotoxicity: Hb level <10 g/L and ANC <1 × 109/L or PLT <75.000 × 109/L Hepatotoxicity grade ≥ 3: Transaminase levels ≥5 times higher than the upper limit of normal (≥200 IU/L). Table 5 Comparison of the frequency of myelotoxicity and hepatotoxicity grade ≥3. Table 5 1 g/m2 2 g/m2 5 g/m2 Pa Myelotoxicity 24 (%35.2) 21 (%37.5) 23 (%33.8) >.05 Hepatotoxicity grade≥ 3 9 (%13.2) 7 (%12.5) 8 (%11.7) >.05 a One way ANNOVA.

Table 4 1 g/m2 2 g/m2 5 g/m2 İnfusion (n/%) Patient (n/%) İnfusion (n/%) Patient (n/%) İnfusion (n/%) Patient (n/%) ANC<1 × 109/L 16 (%23.5) 13 (%76.4) 11 (%19.6) 9 (%64.2) 14 (%20.5) 10 (%58.8) PLT<75 × 109/L 3 (%4.4) 3 (%17.6) 6 (%10.7) 2 (%14.2) 1 (%1.4) 1 (%5.8) Hb < 10 g/L & ANC<1 × 109/L 21 (%30.8) 9 (%52.9) 17 (%30.3) 8 (%57.2) 22 (%32.4) 11 (%64.7) Myelotoxicity 24 (%35.2) 12 (%70.5) 23 (%41) 10 (%71.4) 23 (%33.8) 12 (%70.5) ALT 40–120 IU/L (Grade 1) 21 (%25) 13 (%76.4) 19 (%33.9) 11 (%78.5) 16 (%23.5) 9 (%52.9) ALT 120–200 (Grade 2) 5 (%7.3) 5 (%29.4) 5 (%8.9) 4 (%28.5) 3 (%4.4) 3 (%17.6) ALT 200–800 (Grade 3) 8 (%11.7) 4 (%23.5) 5 (%8.9) 4 (%28.5) 5 (%7.3) 4 (%23.5) ALT>800 (Grade 4) 1 1 1 1 – – Hepatotoxicity grade≥ 3 9 (%13.2) 5 (%29.4) 6 (%10.7) 5 (%35.7) 5 (%7.3) 4 (%23.5) Myelotoxicity: Hb level <10 g/L and ANC <1 × 109/L or PLT <75.000 × 109/L Hepatotoxicity grade ≥ 3: Transaminase levels ≥5 times higher than the upper limit of normal (≥200 IU/L). Table 5 Comparison of the frequency of myelotoxicity and hepatotoxicity grade ≥3. Table 5 1 g/m2 2 g/m2 5 g/m2 Pa Myelotoxicity 24 (%35.2) 21 (%37.5) 23 (%33.8) >.05 Hepatotoxicity grade≥ 3 9 (%13.2) 7 (%12.5) 8 (%11.7) >.05 a One way ANNOVA. The frequency of grade ≥3 hepatotoxicity was 2-fold higher than myelotoxicity after the 1st cycle in cases receiving 5 g/m2. The frequency of myelotoxicity increased in repeated cycles in patients receiving 5 g/m2. The frequency of myelotoxicity showed variability among repeated cycles in the patients receiving 1 g/m2 and 2 g/m2. There was a reduction in the frequency of hepatotoxicity observed for all three dose groups after repeated cycles (Fig. 1).Figure 1 Myelotoxicity and hepatotoxicity grade ≥3 in HDMTX cycles.

ving 5 g/m2. The frequency of myelotoxicity showed variability among repeated cycles in the patients receiving 1 g/m2 and 2 g/m2. There was a reduction in the frequency of hepatotoxicity observed for all three dose groups after repeated cycles (Fig. 1).Figure 1 Myelotoxicity and hepatotoxicity grade ≥3 in HDMTX cycles. Fig. 1 The data revealed 13 of 22 patients (59%) (5 patients; 2 g/m2, 8 patients; 5 g/m2) who developed myelotoxicity and 5 of 9 patients (55%) (4 patients; 2 g/m2 and 1 patient; 5 g/m2) who developed hepatic toxicity had extended MTX elimination. There was no difference in MTX42 levels between the patients with and without hematologic toxicity or the patients with and without hepatotoxicity (P > .05, all) (Table 6, Table 7). There was no correlation observed between MTX42 levels and the values of Hb, AST, ALT, ANC, and PLT count (P < .05, for all).Table 6 Comparison of MTX42h levels in patients with and without hematologic toxicity and distribution with reference to the cycles.

and without hepatotoxicity (P > .05, all) (Table 6, Table 7). There was no correlation observed between MTX42 levels and the values of Hb, AST, ALT, ANC, and PLT count (P < .05, for all).Table 6 Comparison of MTX42h levels in patients with and without hematologic toxicity and distribution with reference to the cycles. Table 6 2 g/m2 Pa 5 g/m2 Pa Hematologic toxicity+ Hematologic toxicity- Hematologic toxicity+ Hematologic toxicity- Cycles 1 1.04 (0.29–2.02) (n = 8) 0.53 (0.22–0.90) (n = 6) >.05 0.64 (0.52–1.53) (n = 3) 0.92 (0.52–1.17) (n = 14) >.05 Cycles 2 0.96 (0.39–1.08) (n = 5) 0.70 (0.28–0.83) (n = 9) >.05 0.97 (0.34–1.65) (n = 5) 0.54 (0.41–1.01) (n = 12) >.05 Cycles 3 0.31 (0.22–1.22) (n = 6) 0.50 (0.48–0.99) (n = 8) >.05 0.95 (0.38–1.62) (n = 6) 0.45 (0.28–1.06) (n = 11) >.05 Cycles 4 0.56 (0.16–0.95) (n = 4) 0.58 (0.33–2.23) (n = 10) >.05 0.68 (0.29–1.94) (n = 7) 0.47 (0.28–0.97) (n = 10) >.05 The first row shows MTX levels at 42 h and the second row shows number of patients. a Mann Whitney U- test. Table 7 Comparison of MTX42h levels in patients with and without hepatotoxicity and distribution with reference to the cycles.

Table 6 2 g/m2 Pa 5 g/m2 Pa Hematologic toxicity+ Hematologic toxicity- Hematologic toxicity+ Hematologic toxicity- Cycles 1 1.04 (0.29–2.02) (n = 8) 0.53 (0.22–0.90) (n = 6) >.05 0.64 (0.52–1.53) (n = 3) 0.92 (0.52–1.17) (n = 14) >.05 Cycles 2 0.96 (0.39–1.08) (n = 5) 0.70 (0.28–0.83) (n = 9) >.05 0.97 (0.34–1.65) (n = 5) 0.54 (0.41–1.01) (n = 12) >.05 Cycles 3 0.31 (0.22–1.22) (n = 6) 0.50 (0.48–0.99) (n = 8) >.05 0.95 (0.38–1.62) (n = 6) 0.45 (0.28–1.06) (n = 11) >.05 Cycles 4 0.56 (0.16–0.95) (n = 4) 0.58 (0.33–2.23) (n = 10) >.05 0.68 (0.29–1.94) (n = 7) 0.47 (0.28–0.97) (n = 10) >.05 The first row shows MTX levels at 42 h and the second row shows number of patients. a Mann Whitney U- test. Table 7 Comparison of MTX42h levels in patients with and without hepatotoxicity and distribution with reference to the cycles. Table 7 2 g/m2 Pa 5 g/m2 Pa Hepatotoxicity+ Hepatotoxicity- Hepatotoxicity+ Hepatotoxicity- Cycles 1 0.29 (0.22–2.02) (n = 8) 0.80 (0.38–1.06) (n = 6) >.05 0.77 (0.444–1.92) (n = 6) 0.88 (0.56–1.20) (n = 11) >.05 Cycles 2 0.86 (0.43–1.02) (n = 5) 0.70 (0.25–0.99) (n = 9) >.05 0.47 (0.28–0.99) (n = 2) 0.70 (0.44–1,12) (n = 15) >.05 Cycles 3 0.88 (0.31–1.2) (n = 3) 0.49 (0.26–0.86) (n = 11) >.05 0.37 (0.24–0.90) (n = 2) 0.68 (0.42–1,09) (n = 15) >.05 Cycles 4 0.94 (0.72–1.1) (n = 2) 0.40 (0.17–1.06) (n = 12) >.05 0.42 (0.26–1.48) (n = 2) 0.60 (0.28–0.99) (n = 15) >.05 The first row shows MTX levels at 42 nd h and the second row shows number of patients. a Mann Whitney U-test.

Table 7 2 g/m2 Pa 5 g/m2 Pa Hepatotoxicity+ Hepatotoxicity- Hepatotoxicity+ Hepatotoxicity- Cycles 1 0.29 (0.22–2.02) (n = 8) 0.80 (0.38–1.06) (n = 6) >.05 0.77 (0.444–1.92) (n = 6) 0.88 (0.56–1.20) (n = 11) >.05 Cycles 2 0.86 (0.43–1.02) (n = 5) 0.70 (0.25–0.99) (n = 9) >.05 0.47 (0.28–0.99) (n = 2) 0.70 (0.44–1,12) (n = 15) >.05 Cycles 3 0.88 (0.31–1.2) (n = 3) 0.49 (0.26–0.86) (n = 11) >.05 0.37 (0.24–0.90) (n = 2) 0.68 (0.42–1,09) (n = 15) >.05 Cycles 4 0.94 (0.72–1.1) (n = 2) 0.40 (0.17–1.06) (n = 12) >.05 0.42 (0.26–1.48) (n = 2) 0.60 (0.28–0.99) (n = 15) >.05 The first row shows MTX levels at 42 nd h and the second row shows number of patients. a Mann Whitney U-test. 5 Discussion The most commonly described side-effects of MTX therapy are myelosuppression, acute liver toxicity, nephrotoxicity, mucositis, and neurotoxicity [4], [8], [9], [10], [11], [12]. Shimasaki et al [3] reported that the only factor that affected HDMTX-associated toxicity development was high MTX levels. Additionally, genetic differences affecting the absorption, distribution, metabolism, and expression of the drug were also significantly involved. The chemotherapeutics used in combination with HDMTX may also affect toxicity. Previous studies reported that using HDMTX with increased doses of 6-MP may also increase hematologic and hepatic toxicity [13], [14].

ing the absorption, distribution, metabolism, and expression of the drug were also significantly involved. The chemotherapeutics used in combination with HDMTX may also affect toxicity. Previous studies reported that using HDMTX with increased doses of 6-MP may also increase hematologic and hepatic toxicity [13], [14]. Prior studies have reported that there are different ratios for the frequency of HDMTX toxicity. Kapoor et al [15] detected neutropenia ≥ grade 3 (ANC < 1 × 109/L) in 24.8% of the infusions in patients receiving HDMTX at 5 g/m2. In our study, neutropenia ≥ grade 3 developed in 23.5%, 19.6%, and 20.5% of the HDMTX infusions at doses of 1, 2 and 5 g/m2, respectively (Table 4). The toxicity rates in our study were similar to each other and were slightly lower than those reported in other studies. In the same study, transient elevation of transaminases occurred in 35% of cycles and most were grade 1 and 2 (32%), as defined by serum ALT levels up to 5 times above normal (grade 3 and 4 in 3 cycles) levels. In our study, hepatotoxicity grade ≥3 was detected in 13.2%, 12.5%, and 11.7% of the HDMTX infusions for patients 1, 2, and 5 g/m2, respectively (Table 4, Table 5). Our rates for hepatotoxicity ≥ grade 3 were higher. Tsurasawa et al [16] reported that the hepatic and gastrointestinal system toxicity was higher after first drug administration and was followed by a sharp reduction in toxicity incidence in the subsequent cycles. In contrast to non-hematologic toxicity, the incidence of hematologic toxicity varied slightly along with HDMTX cycles. Ridolfi et al [17] reported that there was no increase in hematologic and hepatic toxicity with repeated drug administrations. Rask et al [4] identified a significant correlation between leukopenia and cycle count. However, there is no significant correlation between leukopenia and clinical or pharmacokinetic parameters. Our study demonstrated the frequency of hematologic toxicity changed with repeated infusions. We found that the frequency of hematologic toxicity gradually increased in the group receiving 5 g/m2 (Fig. 1). It is thought that the increase in myelosuppression in subsequent cycles is related to the accumulation of cytotoxic metabolites of MTX and 6-MP [4].

y of hematologic toxicity changed with repeated infusions. We found that the frequency of hematologic toxicity gradually increased in the group receiving 5 g/m2 (Fig. 1). It is thought that the increase in myelosuppression in subsequent cycles is related to the accumulation of cytotoxic metabolites of MTX and 6-MP [4]. The frequency of hepatotoxicity was higher with the first infusion in all three doses, then, showed a subsequent reduction (Fig. 1). The reduction in hepatotoxicity after the first infusion could be explained by the significantly increased plasma folate concentration resulting from repeated leucovorin administrations during HDMTX cycles [18]. Holmboe et al [19] reported that osteosarcoma patients receiving HDMTX showed a relationship between the levels of peak ALT and 7-OH MTX, which is a metabolite of MTX. According to the results of their study, there is no relationship between the levels of peak ALT and peak MTX levels. Additionally, Csordas et al [6] did not identify any correlation between bone marrow toxicity and the levels of serum MTX. In the same study, the authors determined that there was no correlation between 7-OH MTX levels and hepatotoxicity or bone marrow toxicity. However, according to Rask et al, [4], an important risk factor for temporary increases in ALT levels is the extended presence of high serum MTX levels.

nd the levels of serum MTX. In the same study, the authors determined that there was no correlation between 7-OH MTX levels and hepatotoxicity or bone marrow toxicity. However, according to Rask et al, [4], an important risk factor for temporary increases in ALT levels is the extended presence of high serum MTX levels. In this study, the MTX42 levels in patients with myelotoxicity and hepatotoxicity were not different from patients without toxicity (Table 6, Table 7). However, MTX elimination was extended in 59% of the patients with hematologic toxicity and 55% of patients with hepatic toxicity. These results suggest the levels of MTX42 are not able to identify hematologic and hepatic toxicity risks.

epatotoxicity were not different from patients without toxicity (Table 6, Table 7). However, MTX elimination was extended in 59% of the patients with hematologic toxicity and 55% of patients with hepatic toxicity. These results suggest the levels of MTX42 are not able to identify hematologic and hepatic toxicity risks. Previous studies have shown using 6-MP in combination with HDMTX affects toxicity frequency. Frandsen et al [20] found patients receiving increased 6-MP (50–75 mg/m2) doses did not experience more toxicity than patients receiving 25 mg/m2 per day. Furthermore, Niekerk et al [13] reported that they compared patients receiving 5 g/m2 HDMTX with patients receiving different doses of 6-MP; the hepatic and hematologic toxicity were higher in the patients receiving 75 mg/m2 than those receiving 25 mg/m2. The authors speculated the cause of these results was the increasing cytotoxic effect because MTX increases both the bioavailability and enzymatic activation of 6-MP [14], [21]. The same researchers also found hepatotoxicity was higher in patients not receiving 6-MP than patients receiving 6-MP, and there was no logical explanation for this result. There is a strong correlation between the severity of myelosuppression, delayed treatment, and the dose of 6-MP [13], [22]. Levinsen et al [14] reported increased levels of 6-MP are related to both hematologic and hepatic toxicity.

6-MP than patients receiving 6-MP, and there was no logical explanation for this result. There is a strong correlation between the severity of myelosuppression, delayed treatment, and the dose of 6-MP [13], [22]. Levinsen et al [14] reported increased levels of 6-MP are related to both hematologic and hepatic toxicity. The patients received standard 6-MP doses (25 mg/m2). Therefore, the dose of 6-MP was not an important factor affecting the frequency of toxicity. However, it is important to consider the individual differences in the metabolism of 6-MP. 6 Conclusion Our results indicate that myelotoxicity occurred in approximately 35% of patients, and severe hepatic toxicity occurred in 10% of treated patients. There was no increase in toxicity due to dose administration. Although the frequency of myelotoxicity increased in patients receiving 5 g/m2, the hepatotoxicity frequency decreased. There is no difference between MTX42h levels in patients with and without toxicity. Furthermore, there is no correlation between blood count, AST-ALT levels, and MTX42h levels. These results suggest that MTX42h levels are not able to predict hematologic and hepatic toxicity. Thus, there is a need for further studies to confirm these results because our patient population is limited. Ethical clearance This study followed the Declaration of Helsinki on medical protocol and ethics and the regional Ethical Review Board of Eskişehir Osmangazi University Faculty of Medicine approved the study.

6 Conclusion Our results indicate that myelotoxicity occurred in approximately 35% of patients, and severe hepatic toxicity occurred in 10% of treated patients. There was no increase in toxicity due to dose administration. Although the frequency of myelotoxicity increased in patients receiving 5 g/m2, the hepatotoxicity frequency decreased. There is no difference between MTX42h levels in patients with and without toxicity. Furthermore, there is no correlation between blood count, AST-ALT levels, and MTX42h levels. These results suggest that MTX42h levels are not able to predict hematologic and hepatic toxicity. Thus, there is a need for further studies to confirm these results because our patient population is limited. Ethical clearance This study followed the Declaration of Helsinki on medical protocol and ethics and the regional Ethical Review Board of Eskişehir Osmangazi University Faculty of Medicine approved the study. Funding This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. Conflict of interest The authors have no conflict of interest to report. Peer review under responsibility of King Faisal Specialist Hospital & Research Centre (General Organization), Saudi Arabia.