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ive care unit), all funded by the Brazilian National Health System. Patients followed at INI have free-of charge access to all available facilities. A longitudinal database maintains in-hospital and outpatient clinical information on patients receiving HIV care. Cohort procedures and results were published eslwhere.6,7 The present study included HIV-infected adults (≥18 years of age at cohort enrollment), enrolled in the INI cohort between 01 January 1986 and 01 December 2013, who were alive and in active care (at least one medical visit) after 01 January 2007. Follow-up started on 01 January 2007 or the date of cohort enrollment, whichever occurred last, and it ended on 31 December 2013, date of death, or last clinical visit (medical visit, CD4, HIV viral load or any blood exam) whichever occurred first. Lost to follow-up was defined as not having a clinical visit after 01 January 2013 for those known not to be deceased. Information regarding vital status was exhaustedly checked using the patients’ medical charts and by linkage with the Rio de Janeiro mortality database (up to 31 December 2013) using a previously validated algorithm.8

Introduction Combination antiretroviral therapy (ART) has led to a reduction in the rates of hospitalization among HIV-infected patients.1,2 Immunological improvement and gain in life expectancy achieved as a result of ART also modified causes of hospitalizations, and, in most recent years, non-AIDS events surpassed AIDS-related as the main cause of hospitalization in high income settings.2–4 Simultaneously, duration of hospitalizations (i.e. length of stay) and in-hospital mortality1,5 among HIV-infected patients decreased over time. Nevertheless, there is a need to assess hospitalizations, length of stay, and inhospital mortality in late ART era since they provide updated information on morbidity and health care utilization among HIV-infected patients, which are essential to evaluate health care provision, guide health policies and project its associated costs. In this study, we sought to assess trends in hospitalization rates, length of stay, and in-hospital mortality in a cohort of HIV-infected patients from Rio de Janeiro, from 2007 through 2013.

nfected patients, which are essential to evaluate health care provision, guide health policies and project its associated costs. In this study, we sought to assess trends in hospitalization rates, length of stay, and in-hospital mortality in a cohort of HIV-infected patients from Rio de Janeiro, from 2007 through 2013. Methods Instituto Nacional de Infectologia Evandro Chagas (INI, formerly known as Instituto de Pesquisa Clínica Evandro Chagas/IPEC) is a reference center for research and care of HIV-infected patients, in Rio de Janeiro, Brazil, since 1986. INI provides primary, specialty and tertiary care for HIV-infected patients and it includes an outpatient facility, an emergency department, a day-clinic, and an inpatient care unit (comprising an intensive care unit), all funded by the Brazilian National Health System. Patients followed at INI have free-of charge access to all available facilities. A longitudinal database maintains in-hospital and outpatient clinical information on patients receiving HIV care. Cohort procedures and results were published eslwhere.6,7

p was defined as not having a clinical visit after 01 January 2013 for those known not to be deceased. Information regarding vital status was exhaustedly checked using the patients’ medical charts and by linkage with the Rio de Janeiro mortality database (up to 31 December 2013) using a previously validated algorithm.8 The primary cause of a hospitalization was inferred from discharge reports. All diagnoses listed in the discharge report were classified using the 10th Edition of the International Classification of Disease (ICD-10), into 24 different categories.9 Since some ICD-10 codes could be allocated to several categories, we considered a hierarchical classification protocol with a decreasing order of priority as follows: AIDS-events, non-AIDS malignancies, infections, and then systemic events.9 To determine the primary cause of a hospitalization, one or more ICD-10 codes listed in the discharge reports were hierarchically classified as follows: AIDS-defining diseases, non-AIDS cancer, cardiovascular disease, bacterial infections, fungal infections, viral infections, parasitic infections, digestive diseases, renal diseases, respiratory diseases, neurologic diseases, endocrine diseases, hematological diseases, psychiatric diseases, viral hepatitis, non-viral hepatitis, dermatological diseases, rheumatologic diseases, trauma, gynecologic disease, toxicities, others, and sings and symptoms.

tic infections, digestive diseases, renal diseases, respiratory diseases, neurologic diseases, endocrine diseases, hematological diseases, psychiatric diseases, viral hepatitis, non-viral hepatitis, dermatological diseases, rheumatologic diseases, trauma, gynecologic disease, toxicities, others, and sings and symptoms. Socio-demographic and clinical features were compared among included patients by study period (2007–2009, 2010–2011, 2012–2013) using Kruskal–Wallis test for continuous variables and Chi-square for categorical variables. Annual hospitalization rates, defined as the number of hospitalizations divided by the person-years (PY) of follow-up, were calculated per 100 PY; Poisson regression models were used to estimate trends in hospitalization rates. Length of stay (LOS) was calculated by subtracting hospital admission date from date of discharge and adding 1; linear regression models were used to estimate trends in LOS. In-hospital mortality, defined as the number of hospitalizations that ended in death divided by the total number hospitalizations, were calculated; logistic regression models were used to estimate trends of in-hospital mortality.

e of discharge and adding 1; linear regression models were used to estimate trends in LOS. In-hospital mortality, defined as the number of hospitalizations that ended in death divided by the total number hospitalizations, were calculated; logistic regression models were used to estimate trends of in-hospital mortality. Results A total of 3991 patients, enrolled from June 1986 until November 2013, were followed from 01 January 2007 until 31 December 2013, accounting for 17,822 PY of follow-up. One hundred and eighty nine patients (4.7%) were deemed loss to follow up, yielding a loss to follow up rate of rate of 1.06/100 PY. The study population aged slightly through the years and the proportion of patients with 60 years or more increased from 5.1% in 2007–2009 to 7.1% in 2012–2013 (p < 0.001, Table 1). Likewise, the median CD4 counts (419 cells/mm3 in 2007–2009 to 542 cells/mm3 in 2012–2013, p < 0.001), the proportion of patients on ART (80.9% in 2007–2009 vs. 90.8% in 2012–2013, p < 0.001) and the proportion of patients with a HIV viral load under 400 copies/mL (54% in 2007–2009 vs. 69.5% in 2012–2013, p < 0.001) significantly increased through the years.

ls/mm3 in 2007–2009 to 542 cells/mm3 in 2012–2013, p < 0.001), the proportion of patients on ART (80.9% in 2007–2009 vs. 90.8% in 2012–2013, p < 0.001) and the proportion of patients with a HIV viral load under 400 copies/mL (54% in 2007–2009 vs. 69.5% in 2012–2013, p < 0.001) significantly increased through the years. During the study period, there were 1861 hospitalizations, yielding an overall hospitalization rate of 10.44/100 PY (95% confidence interval [CI] 9.98–10.93/100 PY). Hospitalization rates decreased annually (from 10.52/100 PY in 2007 to 7.28/100 PY in 2013, per year incidence rate ratio [IRR] 0.92, 95% CI 0.89–0.95) mainly due to a decrease of AIDS-related hospitalizations (from 5.17/100 PY in 2007 to 2.78/100 PY in 2013, per year IRR 0.88, 95% CI 0.84–0.92). Non-AIDS related hospitalization also decreased with a borderline significant trend (from 5.34/100 PY in 2007 to 4.49/100 PY in 2013, per year IRR 0.96, 95% CI 0.92–1.00; Table 2). Moreover, throughout the years the proportion of non-AIDS related hospitalizations gradually increased and accounted for the majority of the hospitalizations in the last three years of the study period. Bacterial infections (53.4%, n = 507), cardiovascular diseases (18.6%, n = 177), and viral infections (10.3%, n = 98) represented the three most common causes on non-AIDS hospitalizations during the study.

ally increased and accounted for the majority of the hospitalizations in the last three years of the study period. Bacterial infections (53.4%, n = 507), cardiovascular diseases (18.6%, n = 177), and viral infections (10.3%, n = 98) represented the three most common causes on non-AIDS hospitalizations during the study. Following the trends of hospitalization rates, the overall LOS decreased significantly over the study period (median of 15 days in 2007 vs. 11 days in 2013, p-value for trend < 0.001) as well as the LOS of non AIDS-related hospitalizations (median of 11 days in 2007 vs. 8 days in 2013, p-value for trend = 0.038) and of AIDS-related hospitalizations (median of 19 days in 2007 vs. 16 days in 2013, p-value for trend = 0.036). Overall, in-hospital mortality decreased during the study period (from 13.4% in 2007 to 8.1% in 2013, per calendar year increase odds ratio 0.92, 95% CI 0.85–1.00), as well as in-hospital mortality of non-AIDS related hospitalizations (from 14.7% in 2007 to 5.6% in 2013, per calendar year increase odds ratio 0.84, 95% CI 0.74–0.96). In-hospital mortality of AIDS related hospitalizations remained stable throughout the study period and, overall, it was 1.66 times higher than in-hospital mortality of non-AIDS related hospitalizations (11.6% vs. 7.0%, p < 0.001; Table 2).

to 5.6% in 2013, per calendar year increase odds ratio 0.84, 95% CI 0.74–0.96). In-hospital mortality of AIDS related hospitalizations remained stable throughout the study period and, overall, it was 1.66 times higher than in-hospital mortality of non-AIDS related hospitalizations (11.6% vs. 7.0%, p < 0.001; Table 2). Discussion In this study, we have shown that among HIV-infected patients living in a middle-income setting in the late ART era, hospitalization rates have decreased through the years, mostly due to a decrease in the rate of AIDS related hospitalizations. Consequently, non-AIDS hospitalizations became more common than AIDS related in the last three years of the study period. Decreases in hospitalization rates in late ART era have been described for both high-4 and middle-income settings.10 This shift in hospitalizations causes (from AIDS related to non-AIDS related) follows the reduction in AIDS-defining diseases incidence11 and mortality12 already demonstrated in our cohort, highlighting an increased relevance of non-communicable events among HIV-infected patients. Changes in the study population characteristics through the study period have likely contributed to this scenario. Our results show that, over time, the cohort population aged, while CD4 counts, the proportion of virologic suppressed patients, and ART use among patients have increased.

ents among HIV-infected patients. Changes in the study population characteristics through the study period have likely contributed to this scenario. Our results show that, over time, the cohort population aged, while CD4 counts, the proportion of virologic suppressed patients, and ART use among patients have increased. LOS decreased throughout the study though it remained high mainly due to AIDS related hospitalizations. Overall, our estimated median LOS (13 days) surpasses the one reported for a US multicentric HIV study (median of 5 days)4 but is closer to that reported by a national Portuguese study (median 11 days).13 Additionally, similarly to other studies,4,14 we found that hospitalizations due to AIDS were associated with longer LOS (median of 9 vs. 18 days for non-AIDS and AIDS-related hospitalizations, respectively). In-hospital mortality also decreased over the study period, although in-hospital mortality of AIDS-related hospitalizations remained quite stable through the years. In-hospital mortality was almost two times higher in AIDS-related hospitalizations than in non-AIDS-related hospitalizations. The overall 9.2% in-hospital mortality rate found in our study is higher than the 2.6% rate previously reported in a tertiary hospital in New York between 2004 and 2008,15 but is lower than the rate observed by Akinkuotu et al. in Malawi (24%).16

AIDS-related hospitalizations than in non-AIDS-related hospitalizations. The overall 9.2% in-hospital mortality rate found in our study is higher than the 2.6% rate previously reported in a tertiary hospital in New York between 2004 and 2008,15 but is lower than the rate observed by Akinkuotu et al. in Malawi (24%).16 Disparities in hospitalization rates, LOS, and in-hospital mortality among the studies (particularly, when comparing high- vs. low- and middle-income settings) can be explained by several factors that range from hospital structure, hospital setting, type of health care system, as well as by the burden of diseases, in particular of AIDS-defining illnesses. In this context, tuberculosis burden might play a key role. In settings with high burden, tuberculosis is a leading cause of hospitalization among HIV-infected patients and is related to high in-hospital mortality (24.9% in a meta-analysis including 66 studies).17 In addition, tuberculosis is also associated with longer LOS both among the general and the HIV-infected population.18,19 In our study cohort, tuberculosis accounted for 43% of all AIDS related hospitalizations and yielded an inhospital mortality of 10.9% (data not shown). Additionally, LOS of tuberculosis related hospitalization was significantly longer than non-tuberculosis hospitalizations (median of 18 days vs. 12 days, respectively, p < 0.001, data not shown).

ulosis accounted for 43% of all AIDS related hospitalizations and yielded an inhospital mortality of 10.9% (data not shown). Additionally, LOS of tuberculosis related hospitalization was significantly longer than non-tuberculosis hospitalizations (median of 18 days vs. 12 days, respectively, p < 0.001, data not shown). There are several limitations that need to be highlighted in the present study. First, our study casuistic is from a single cohort that has access to an outpatient as well as an infectious diseases hospital located in Rio de Janeiro, and our results may not reflect those for other HIV-infected populations in Brazil. Second, although patients have a free of charge access to Evandro Chagas hospital we cannot rule out the possibility of hospitalizations in other hospitals within the city, implying that our rates may have been somewhat underestimated. Finally, Evandro Chagas hospitalizations are restricted to non-surgical and non-obstetrics procedures, and therefore our rates do not represent the entire sort of events that can happen to an HIV-infected patient.

zations in other hospitals within the city, implying that our rates may have been somewhat underestimated. Finally, Evandro Chagas hospitalizations are restricted to non-surgical and non-obstetrics procedures, and therefore our rates do not represent the entire sort of events that can happen to an HIV-infected patient. In summary, we demonstrated that, in a middle-income setting, hospitalizations rates are decreasing over time and that non-AIDS hospitalizations are currently more frequent than AIDS related ones. We also showed that in our setting we still struggle with long LOS and high in-hospital mortality. Studies addressing predictors of LOS and in-hospital mortality, mainly in low- and middle-income settings are needed and will be of utmost importance to guide health policies and assistance protocols in order to reduce health costs and inhospital mortality. Funding BG and PML acknowledge funding from the National Council of Technological and Scientific Development (CNPq) and the Research Funding Agency of the State of Rio de Janeiro (FAPERJ). This work was supported in part by the NIH-funded Caribbean, Central and South America network for HIV epidemiology (CCASAnet), a member cohort of the International Epidemiologic Databases to Evaluate AIDS (leDEA) (U01AI069923). Conflicts of interest The authors declare no conflicts of interest. Table 1 Study population characteristics by study periods.

BG and PML acknowledge funding from the National Council of Technological and Scientific Development (CNPq) and the Research Funding Agency of the State of Rio de Janeiro (FAPERJ). This work was supported in part by the NIH-funded Caribbean, Central and South America network for HIV epidemiology (CCASAnet), a member cohort of the International Epidemiologic Databases to Evaluate AIDS (leDEA) (U01AI069923). Conflicts of interest The authors declare no conflicts of interest. Table 1 Study population characteristics by study periods. 2007–2009 (n = 2639) 2010–2011 (n = 3117) 2012–2013 (n = 3605) p-Value Sex 0.678 Male 1699 (64.4) 2020 (64.8) 2359 (65.4) Female 940 (35.6) 1097 (35.2) 1246 (34.6) Age in yearsa Median (IQR) 41.7 (34.1, 48.6) 42 (34.2, 49.4) 42.5 (34.5, 50.5) 0.004 ≤30 372 (14.1) 414 (13.3) 466 (12.9) <0.001 31–40 807 (30.6) 925 (29.7) 1051 (29.2) 41–50 885 (33.5) 1065 (34.2) 1124 (31.2) 51–60 440 (16.7) 525 (16.8) 709 (19.7) >60 135 (5.1) 188 (6) 255 (7.1) Race/ethnicity White 1454 (55.1) 1619 (51.9) 1807 (50.1) <0.001 Non White 1185 (44.9) 1498 (48.1) 1798 (49.9) Educational level 0.075 Up to 9 years 1340 (50.8) 1555 (49.9) 1730 (48) More than 9 years 1299 (49.2) 1562 (50.1) 1875 (52) HIV exposure category 0.028 Heterosexual 1420 (53.8) 1636 (52.5) 1830 (50.8) MSM 948 (35.9) 1146 (36.8) 1363 (37.8) IDU 52 (2) 50 (1.6) 47 (1.3) Other/unknown 219 (8.3) 285 (9.1) 365 (10.1) Chronic hepatitis Bb 169 (6.4) 188 (6) 198 (5.5) 0.308 Chronic hepatitis Cc 278 (10.5) 301 (9.7) 309 (8.6) 0.03 CD4 count (cells/mm3)d Median (IQR) 419 (254, 616) 532 (336, 772) 542 (358, 775) <0.001 >500 970 (36.8) 1664 (53.4) 1924 (53.4) <0.001 500–351 581 (22) 554 (17.8) 676 (18.8) ≤350 996 (37.7) 801 (25.7) 831 (23.1) Missing 92 (3.5) 98 (3.1) 174 (4.8) HIV viral load (copies/mL)d <0.001 ≤400 1424 (54) 2050 (65.8) 2504 (69.5) >400 1073 (40.7) 952 (30.5) 956 (26.5) Missing 142 (5.4) 115 (3.7) 145 (4) ART usee 2136 (80.9) 2687 (86.2) 3273 (90.8) <0.001 MSM, men who have sex with men; IDU, injectable drug use; ART, combination antiretroviral therapy.

) 98 (3.1) 174 (4.8) HIV viral load (copies/mL)d <0.001 ≤400 1424 (54) 2050 (65.8) 2504 (69.5) >400 1073 (40.7) 952 (30.5) 956 (26.5) Missing 142 (5.4) 115 (3.7) 145 (4) ART usee 2136 (80.9) 2687 (86.2) 3273 (90.8) <0.001 MSM, men who have sex with men; IDU, injectable drug use; ART, combination antiretroviral therapy. a Age at the end of each period. b Defined as having a positive HBsAg antigen. c Defined as having a positive anti-HCV serology. d The closest result to the midpoint of each period. e Defined as ART start before the end of each period. Table 2 Number of hospitalizations, hospitalization rates and length of stay, and in-hospital mortality, stratified by AIDS- and non-AIDS-related causes by year, 2007–2013, INI cohort. Person-years 2007 1778 2008 2057 2009 2336 2010 2556 2011 2825 2012 3040 2013 3229 Total 17,822 Test for trend p-Value Hospitalizations, n (%) All causes 187 284 290 274 320 271 235 1861 AIDS-relatede 92 (49.2) 152 (53.5) 157 (54.1) 148 (54.0) 144 (45.0) 128 (47.2) 90 (38.3) 911 (49) <0.001a Non-AIDS related 95 (50.8) 132 (46.5) 133 (45.9) 126 (46.0) 176 (55.0) 143 (52.8) 145 (61.7) 950 (51) <0.001a

Total 17,822 Test for trend p-Value Hospitalizations, n (%) All causes 187 284 290 274 320 271 235 1861 AIDS-relatede 92 (49.2) 152 (53.5) 157 (54.1) 148 (54.0) 144 (45.0) 128 (47.2) 90 (38.3) 911 (49) <0.001a Non-AIDS related 95 (50.8) 132 (46.5) 133 (45.9) 126 (46.0) 176 (55.0) 143 (52.8) 145 (61.7) 950 (51) <0.001a Hospitalizations, rate/100 PY (95% CI) All causes 10.52 (9.11, 12.14) 13.80 (12. 29, 15.51) 12.42 (11.07, 13.93) 10.72 (9.52, 12.07) 11.33 (10.15, 12.64) 8.91 (7.91, 10.04) 7.28 (6.40, 8.27) 10.40 (9.98, 10.93) 0.92 (0.89–0.95)b AIDS-relatede 5.17 (4.22, 6.35) 7.39 (6.30, 8.66) 6.72 (5.75, 7.86) 5.79 (4.93, 6.80) 5.10 (4.33, 6.00) 4.21 (3.54, 5.01) 2.78 (2.27, 3.43) 5.11 (4.79, 5.45) 0.88 (0.84–0.92)b Non-AIDS related 5.34 (4.37, 6.53) 6.42 (5.41, 7.61) 5.69 (4.80, 6.75) 4.93 (4.14, 5.88) 6.23 (5.37, 7.22) 4.70 (3.99, 5.54) 4.49 (3.81, 5.28) 5.33 (5.00, 5.68) 0.96 (0.92–1.00)b Length of stay, median (IQR) All causes 15 (8, 25) 15 (8, 27) 14 (8, 26) 15 (7, 26) 12 (7, 21) 12 (6, 21) 11 (7, 20) 13 (7, 23) −1.00 (−1.54 to −0.47)c AIDS-relatede 19 (12, 31.2) 19 (11, 30.2) 19 (11, 33) 21 (10, 36) 18 (11, 31) 15 (9, 26) 16 (9, 25) 18 (11, 31) −0.92 (−1.78 to −0.06)c Non-AIDS related 11 (7, 22) 9 (6, 19) 10 (6, 16) 9 (5, 17) 8 (5, 14) 8 (5, 16) 8 (5, 16) 9 (6, 16) −0.65 (−1.26 to −0.04)c

21) 12 (6, 21) 11 (7, 20) 13 (7, 23) −1.00 (−1.54 to −0.47)c AIDS-relatede 19 (12, 31.2) 19 (11, 30.2) 19 (11, 33) 21 (10, 36) 18 (11, 31) 15 (9, 26) 16 (9, 25) 18 (11, 31) −0.92 (−1.78 to −0.06)c Non-AIDS related 11 (7, 22) 9 (6, 19) 10 (6, 16) 9 (5, 17) 8 (5, 14) 8 (5, 16) 8 (5, 16) 9 (6, 16) −0.65 (−1.26 to −0.04)c In-hospital mortality, n (%) All causes 25 (13.4) 24 (8.5) 30 (10.3) 25 (9.1) 19 (5.9) 30 (11.1) 19 (8.1) 172 (9.2) 0.92 (0.85–1.00)d AIDS-relatede 11 (12) 13 (8.6) 22 (14) 19 (12.8) 9 (6.3) 21 (16.4) 11 (12.2) 106 (11.6) 1.00 (0.90–1.12)d Non-AIDS related 14 (14.7) 11 (8.3) 8 (6.0) 6 (4.8) 10 (5.7) 9 (6.3) 8 (5.6) 66 (6.9) 0.84 (0.74–0.96)d PY, person-years; 95% CI, 95% confidence interval; IQR, inter-quartile range. a p-Value estimated using Chi-squared test for trend in proportions. b Per calendar year increase, incidence rate ratio and 95% confidence interval estimated using Poisson regression. c Per calendar year increase, linear coefficient and 95% confidence interval estimated using linear regression. d Per calendar year increase, odds ratio and 95% confidence interval, estimated using logistic regression. e Defined as presenting any AIDS event (CDC, 1994) during hospitalization.

Introduction One reason for the increase of Sexually transmitted diseases (STDs) in many developing countries is the lack of access to effective and reliable health care services.1 factors such as being young and Sexually active, urban migration with sociocultural changes, multiple Sexual partnerships without the use of condoms, and high prevalence of resistance against antimicrobial drugs also contribute to this increase.2,3 The analysis of studies performed in several countries4 showed that people with STDs, even the non-ulcerative kind, presented a risk three to 10 times higher of being infected by the human immunodeficiency virus (HIV), depending on the STD type and etiology. People who live with HIV/AIDS (PLHA) have a high rate of prior STDs5,6 and, if PLHA acquire any form of STD, the HIV viral load in the genital secretions increases,7,8 causing its infectivity also to increase considerably. Among ulcerative STDs, it has been recently shown that genital herpes can be considered the main co-factor for the higher proportion of new HIV infections.9 In heteroSexual partners, the higher the HIV plasma concentration the higher the risk of HIV transmission,10 and a study of HIV infected men with urethritis-associated STD showed a HIV average concentration in the seminal plasma eight times higher.11 However, some studies showed no correlation between the HIV viral load in the plasma and in the semen.12

r the HIV plasma concentration the higher the risk of HIV transmission,10 and a study of HIV infected men with urethritis-associated STD showed a HIV average concentration in the seminal plasma eight times higher.11 However, some studies showed no correlation between the HIV viral load in the plasma and in the semen.12 Some STDs, as human papilloma virus (HPV) infection, generally present a higher prevalence among HIV-infected women when compared to HIV-negative women,13–16 and this persistent infection constitutes a higher risk for the development of cervical intraepithelial neoplasia (CIN).17 According to the Brazilian Ministry of Health, 544,846 cases of AIDS were reported from 1980 to June 2009, and HIV infection is having an increasing impact on reproductive health in Brazil. During this time, the male to female ratio has been progressively reduced from 26.7:1 in 1985 to 1.5:1 currently.18 Although the number of accumulated cases among males is higher than in females, women represent the population where the fastest epidemic growth is observed in the country.18 Women experience different constraints for the exercise of Sexuality, making it difficult for them to incorporate protection practices, and health care services are not always able to deal with this situation, which in turn, increases their vulnerability.19 since women are asymptomatic for most STDs, the percentage in which these infections are involved in the HIV transmission is unknown.

aking it difficult for them to incorporate protection practices, and health care services are not always able to deal with this situation, which in turn, increases their vulnerability.19 since women are asymptomatic for most STDs, the percentage in which these infections are involved in the HIV transmission is unknown. The propose of this study was to describe the epidemiological profile, risk behavior, and frequency of prior STDs among women living with AIDS, aiming at collecting data that could be used in the implementation of prevention and assistance programs for women. Methodology This cross-sectional study was performed in the Centro de Referência e Treinamento em DST/AIDSs in the city of São Paulo, Brazil (CRT-DST/AIDSs). Out of approximately 4,000 HIV/AIDS patients receiving care at this center 1,100 were women. Data were abstracted from medical records of HIV-infected women. Women scheduled for a gynecological appointment in the outpatient gynecological clinics of the reference center between July 1, 2008 and May 31, 2009 were included in the study. Social, demographic, behavioral, and clinical characteristics such as age, educational level, marital status, age at first Sexual intercourse, number of Sexual partners, parity, use of drugs, time elapsed since the diagnosis of HIV infection, CD4+ count, and HIV viral load determination, among others, were selected for analysis.

mographic, behavioral, and clinical characteristics such as age, educational level, marital status, age at first Sexual intercourse, number of Sexual partners, parity, use of drugs, time elapsed since the diagnosis of HIV infection, CD4+ count, and HIV viral load determination, among others, were selected for analysis. The study included women who used highly active antiretroviral drugs, and excluded those who did not have the results of CD4+ count and/or HIV viral load, and those whose gynecological consultation had not been described. The reports of previous STDs were analyzed, as well as the results of laboratory exams and of cervical oncotic cytology. The variable race, self-reported, considered as black those women who declared themselves as black or brown. Educational level and the human development index (HDI) were used as indicators of social and economic level due to their association with several lifestyles characteristics. The data collected from the medical records were linked to the HDI database of the city of São Paulo for the year 2000, through the deterministic record linkage method, which made nominal identification between the two databases. History of prior STDs (syphilis, herpes, gonorrhea, Chlamydia, Trichomonas, hepatitis b, and HPV) was considered as dependent variable for the purpose of this analysis. The risks of prior STDs were computed for all independent variables of interest mentioned above.

The reports of previous STDs were analyzed, as well as the results of laboratory exams and of cervical oncotic cytology. The variable race, self-reported, considered as black those women who declared themselves as black or brown. Educational level and the human development index (HDI) were used as indicators of social and economic level due to their association with several lifestyles characteristics. The data collected from the medical records were linked to the HDI database of the city of São Paulo for the year 2000, through the deterministic record linkage method, which made nominal identification between the two databases. History of prior STDs (syphilis, herpes, gonorrhea, Chlamydia, Trichomonas, hepatitis b, and HPV) was considered as dependent variable for the purpose of this analysis. The risks of prior STDs were computed for all independent variables of interest mentioned above. The clinical and epidemiological information and the laboratorial findings were coded and stored in a database created for this purpose. The statistical program STATA 10.0 was used for data storage and analysis.

History of prior STDs (syphilis, herpes, gonorrhea, Chlamydia, Trichomonas, hepatitis b, and HPV) was considered as dependent variable for the purpose of this analysis. The risks of prior STDs were computed for all independent variables of interest mentioned above. The clinical and epidemiological information and the laboratorial findings were coded and stored in a database created for this purpose. The statistical program STATA 10.0 was used for data storage and analysis. The analysis was performed using data exploratory techniques to check the distribution patterns and trends of the main variables. Then, univariate analysis was performed to check the association between variables. The chi-square test (χ2) was used to assess the difference between proportions and odds ratio calculated with the respective 95% confidence intervals. student’s t-test and analysis of variance were used to assess differences between means. The variables were selected for stepwise logistic regression analysis based on a p-value equal to or lower than 0.25 in the likelihood ratio test. The importance of the variables for the final model was assessed with the likelihood ratio test, considering p < 0.05. The project was approved by the Ethics committee of Centro de Referência e Treinamento em DST/AIDSs of São Paulo.

The analysis was performed using data exploratory techniques to check the distribution patterns and trends of the main variables. Then, univariate analysis was performed to check the association between variables. The chi-square test (χ2) was used to assess the difference between proportions and odds ratio calculated with the respective 95% confidence intervals. student’s t-test and analysis of variance were used to assess differences between means. The variables were selected for stepwise logistic regression analysis based on a p-value equal to or lower than 0.25 in the likelihood ratio test. The importance of the variables for the final model was assessed with the likelihood ratio test, considering p < 0.05. The project was approved by the Ethics committee of Centro de Referência e Treinamento em DST/AIDSs of São Paulo. Results A total of 598 women out of the 710 scheduled for the gynecological consultation in the study period were included. One-hundred and twelve (15.8%) were excluded for the following reasons: 33 (4.6%) women with no information about prior STDs; 29 (4.1%) did not have previous or current gynecological consultations; 23 (3.2%) had never been on antiretroviral drugs; 18 (2.5%) had a negative HIV serology; and nine (1.3%) had acquired HIV through vertical transmission (VT) and never had Sexual activity. Prior STDs was observed in 364 women (60.9%; 95% CI: 56.9%−64.8%).

1%) did not have previous or current gynecological consultations; 23 (3.2%) had never been on antiretroviral drugs; 18 (2.5%) had a negative HIV serology; and nine (1.3%) had acquired HIV through vertical transmission (VT) and never had Sexual activity. Prior STDs was observed in 364 women (60.9%; 95% CI: 56.9%−64.8%). The proportions of prior STDs were 44.0% (263/598) for HPV; 17.9% (107/598) for herpes; 6.2% (37/598) for syphilis; 3.0% (18/598) for Trichomonas; 2.2% (13/598) for hepatitis b; 0,8% (5/598) for gonorrhea; and 0.7% (4/598) for Chlamydia. It should be pointed out that 21.7% of the patients mentioned two or more prior STDs, in addition to HIV infection. The socioeconomical, behavioral and clinical characteristics of the women are presented in Table 1. The majority of women was less than 40 years old (73.6%) at the time of HIV diagnosis, and most of them were white (67.2%). As to their educational level, 50% had completed more than 8 years of schooling, being 56.1% among the white women and 37.6% among the black women. More than 60% were not married and had no stable partner. Table 2 describes the behavioral and clinical characteristics of the studied population, comparing women with and without a history of prior STDs. More than half of them (53.2%) began their Sexual life after 15 years of age; 35.1% mentioned six or more Sexual partners throughout life; and 2.7% declared themselves as Sex workers. A total of 43.6% had been diagnosed with HIV infection for nine or more years, varying from 1 to 22 years.

hout a history of prior STDs. More than half of them (53.2%) began their Sexual life after 15 years of age; 35.1% mentioned six or more Sexual partners throughout life; and 2.7% declared themselves as Sex workers. A total of 43.6% had been diagnosed with HIV infection for nine or more years, varying from 1 to 22 years. Use of intravenous drugs was admitted by 2.2% of the women; 13.2% reported being non-injectable drug users and, among them, 41.8% used more than one drug. Factors associated to prior STDs included in the logistic regression model are presented in Table 3. These factors and their correspondent risks were: HDI of the place of residence < 0.50 (ORaj = 5.5; 95% CI: 2.8–11.0); race other than white (ORaj = 5.2; 95% CI: 2.5–11.0); age of the first Sexual intercourse up to 15 years (ORaj = 4.4; 95% CI: 2.3–8.3); follow-up time of the HIV infection of nine years or more [ORaj = 4.2 (95% CI: 2.3–7.8); number of Sexual partners in lifetime between three and five (ORaj = 2.2; 95% CI: 1.1–4.6)], and six or more (ORaj = 3.9; 95% CI: 1.9–8.0)]; being a Sex worker (ORaj = 1.9; 95% CI: 1.1–3.1). Discussion The purpose of this study was to describe the epidemiological profile, risk behaviors, and the frequency of prior STDs in women living with AIDS receiving care at the Centro de Referência e Treinamento em DST/AIDSs in São Paulo. A high prevalence of prior STDs was found in the population studied. In Brazil, there are few available studies that analyzed STD prevalence in PLHA.

cal profile, risk behaviors, and the frequency of prior STDs in women living with AIDS receiving care at the Centro de Referência e Treinamento em DST/AIDSs in São Paulo. A high prevalence of prior STDs was found in the population studied. In Brazil, there are few available studies that analyzed STD prevalence in PLHA. Two studies were carried out in the state of PeRNAmbuco. One assessed STD prevalence in 399 PLHA in a sample made up of heteroSexuals (33%), homoSexuals (23%), and biSexuals (11%), 75% males, in which syphilis was the most frequent disease, with a prevalence of 8.8%, followed by 5.8% of genital herpes and 4.3% of genital candidiasis.20 the second study21 assessed genital infection in women, showed frequencies of HPV, Chlamydia trachomatis, and Trichomonas vaginalis of 20.0%, 2.2%, and 2.2%, respectively. Grinsztejn et al.22 found a HPV prevalence of 48% in a sample of 634 HIV-infected women in Rio de Janeiro. Studies conducted in Jamaica and taiwan5,6 have shown rates of prior STDs in PLHA of 51.1% and 43.1%, respectively. In the present study, prior STD, in addition to presenting a higher proportion (60.9%), was significantly associated with the district of residence, race other than white, first Sexual intercourse under the age of 16, three or more Sexual partners in life, being a Sex worker, and having been diagnosed with the HIV infection for nine years or more. these results are supported by other studies in Brazil, showing that the AIDS epidemic in the country affects predominantly (80%) individuals at the lower socioeconomic level.16,23

of 16, three or more Sexual partners in life, being a Sex worker, and having been diagnosed with the HIV infection for nine years or more. these results are supported by other studies in Brazil, showing that the AIDS epidemic in the country affects predominantly (80%) individuals at the lower socioeconomic level.16,23 The available data provide further evidence that other STDs facilitate HIV transmission through direct biological mechanisms, and that the early treatment of STDs should be a part of high-quality strategies for the prevention of HIV infection.24 Perhaps a high percentage of the participants of the present study have been infected, or have transmitted the HIV infection, due to a concomitant STD, since the majority of STDs are asymptomatic in women. The high proportion of women with history of prior HPV in this study (44.0%) draws the attention to the need of preventive actions in relation to gynecological cancers, since many studies have shown a strong association between HIV and HPV co-infection and the development of CIN and genital cancer.25–27 there is evidence that HIV-infected women have significantly higher CIN rates and a higher probability to evolve to invasive carcinoma than non-infected women.28–31

ecological cancers, since many studies have shown a strong association between HIV and HPV co-infection and the development of CIN and genital cancer.25–27 there is evidence that HIV-infected women have significantly higher CIN rates and a higher probability to evolve to invasive carcinoma than non-infected women.28–31 The present study found a history of prior genital herpes in 17.9% of the women, a higher rate than that reported by Saxton et al. (13.7%), in Ukraine.32 Abu-Raddad et al., showed that herpes simplex virus 2 (HsV-2) infection is a biological co-factor in HIV acquisition and transmission, and can facilitate the spread of HIV infection even among the population with a lower risk of infection who maintain stable Sexual partnerships.33 History of prior of syphilis was observed in 6.2% of the sample, a similar rate to that found by Hutton-rose et al. (7.3%),5 although lower than that found by Saxton et al. (1.9%),32 which may be explained by the lower age of their study population.

h a lower risk of infection who maintain stable Sexual partnerships.33 History of prior of syphilis was observed in 6.2% of the sample, a similar rate to that found by Hutton-rose et al. (7.3%),5 although lower than that found by Saxton et al. (1.9%),32 which may be explained by the lower age of their study population. In the present study, history of prior Chlamydia trachomatis infection was low (0.7%). Perhaps due to the non-availability of routine diagnostic work-up for this infection in Brazil, and its asymptomatic character in up to 80% of the women. Its importance is not well known by the population. This fact must be considered in HIV acquisition, because of the high prevalence of this infection among the Brazilian female population according to a multicenter trial conducted by the Ministry of Health,34 and another study with a nationally representative sample,35 with prevalences of 9.4% and 9.8%, respectively. The use of retrospective data is one of the limitations of this study. The temporality of the infections could not be assessed. However, both the good quality of the data of the medical records and the low rate of missing information may counteract this limitation. In attempting to give socially accepted answers, another limitation that may have occurred would be the inaccuracy of the information on the use of condoms, age at first Sexual intercourse, and number of Sexual partners.

data of the medical records and the low rate of missing information may counteract this limitation. In attempting to give socially accepted answers, another limitation that may have occurred would be the inaccuracy of the information on the use of condoms, age at first Sexual intercourse, and number of Sexual partners. Health care services for HIV positive women exist, and they are able to control successfully the infection and prevent the disease progression. Although this is promising, there is still much work to be done to identify innovative interventions that target the social, cultural, and environmental influences of the STD presence in this group. It is also necessary to provide better ways to access prevention programs of HIV infection, so that effective interventions can be more broadly used. Sexually active women need confidential, affordable, and supportive services to teach them how to protect themselves against diseases, including STDs and HIV infection. Considering that STDs are among the most well established risk factors for HIV infection, public health programs should be enforced with the implementation of control actions and tracking, diagnosis, and early treatment strategies, thus avoiding complications, reducing morbidity, and enhancing the Sexual and reproductive health of the population. Acknowledgements Supported by the National Institute of Health grant number 3D43tW000010–21S1 and National Institutes of Health grant AI06699 (JEG). Conflict of interest All authors declare to have no conflict of interest.

Considering that STDs are among the most well established risk factors for HIV infection, public health programs should be enforced with the implementation of control actions and tracking, diagnosis, and early treatment strategies, thus avoiding complications, reducing morbidity, and enhancing the Sexual and reproductive health of the population. Acknowledgements Supported by the National Institute of Health grant number 3D43tW000010–21S1 and National Institutes of Health grant AI06699 (JEG). Conflict of interest All authors declare to have no conflict of interest. Table 1 - Distribution of the sociodemographic characteristics of women living with AIDS according to history of prior STD at the Centro de Referência e Treinamento em DST/AIDSS, São Paulo, Brazil, 2008 to 2009

Acknowledgements Supported by the National Institute of Health grant number 3D43tW000010–21S1 and National Institutes of Health grant AI06699 (JEG). Conflict of interest All authors declare to have no conflict of interest. Table 1 - Distribution of the sociodemographic characteristics of women living with AIDS according to history of prior STD at the Centro de Referência e Treinamento em DST/AIDSS, São Paulo, Brazil, 2008 to 2009 Characteristics With STD (n = 364) Without STD (n = 234) Total (598) P n % n % n % Age at AIDS diagnosis 0.591 up to 40 years 265 72.8 175 74.8 440 73.6 > 40 years 99 27.2 59 25.2 158 26.4 Educational level (years) 0.956 None 4 1.1 2 0.9 6 1.0 1 to 4 years 29 8.0 19 8.1 48 8.0 5 to 8 years 151 41.5 91 38.9 242 40.5 9 to 11 years 135 37.1 87 37.2 222 37.1 12 or more 42 11.5 33 14.1 75 12.5 Unknown 3 0.8 2 0.9 5 0.8 Race (self-referred) < 0.001 White 202 55.5 200 85.5 402 67.2 Not white 162 44.5 32 13.7 194 32.4 Unknown 0 0.0 2 0.9 2 0.3 Marital status* 0.909 Single 102 28.0 63 26.9 165 27.6 Married/lives with partner 128 35.2 76 32.5 204 34.1 Separated/divorced 62 17.0 46 19.7 108 18.1 Widowed 69 19.1 47 20.1 116 19.4 Unknown 3 0.8 2 0.9 5 0.8 HDI of the residence district < 0.001 > 0.550 172 47.3 213 91.0 385 64.4 0–0.550 192 52.7 21 9.0 213 35.6 STD, sexually transmitted diseases; HDI, human development index; * five women with unknown data were excluded. Table 2 - Distribution of behavioral characteristics of women living with AIDS according to history of prior STD at the Centro de Referência e Treinamento em DST/AIDS, São Paulo, Brazil, 2008 to 2009

Characteristics With STD (n = 364) Without STD (n = 234) Total (598) P n % n % n % Age at AIDS diagnosis 0.591 up to 40 years 265 72.8 175 74.8 440 73.6 > 40 years 99 27.2 59 25.2 158 26.4 Educational level (years) 0.956 None 4 1.1 2 0.9 6 1.0 1 to 4 years 29 8.0 19 8.1 48 8.0 5 to 8 years 151 41.5 91 38.9 242 40.5 9 to 11 years 135 37.1 87 37.2 222 37.1 12 or more 42 11.5 33 14.1 75 12.5 Unknown 3 0.8 2 0.9 5 0.8 Race (self-referred) < 0.001 White 202 55.5 200 85.5 402 67.2 Not white 162 44.5 32 13.7 194 32.4 Unknown 0 0.0 2 0.9 2 0.3 Marital status* 0.909 Single 102 28.0 63 26.9 165 27.6 Married/lives with partner 128 35.2 76 32.5 204 34.1 Separated/divorced 62 17.0 46 19.7 108 18.1 Widowed 69 19.1 47 20.1 116 19.4 Unknown 3 0.8 2 0.9 5 0.8 HDI of the residence district < 0.001 > 0.550 172 47.3 213 91.0 385 64.4 0–0.550 192 52.7 21 9.0 213 35.6 STD, sexually transmitted diseases; HDI, human development index; * five women with unknown data were excluded. Table 2 - Distribution of behavioral characteristics of women living with AIDS according to history of prior STD at the Centro de Referência e Treinamento em DST/AIDS, São Paulo, Brazil, 2008 to 2009 Characteristics With STD (n = 364) Without STD (n = 234) Total (598) P n % n % n % Age at first sexual intercourse < 0.001 Older than 15 years 138 37.9 180 76.9 318 53.2 Up to 15 years 190 52.2 33 14.1 223 37.3 Unknown 36 9.9 21 9.0 57 9.5 No. of partners in lifetime < 0.001 1 to 2 34 9.3 68 29.1 102 17.1 3 to 5 104 28.6 38 16.2 142 23.7 6 or more 180 49.5 30 12.8 210 35.1 Unknown 46 12.6 98 41.9 144 24.1 Use of non-intravenous drugs 0.001 No 299 82.1 218 93.2 517 86.5 Yes 63 17.3 16 6.8 79 13.2 Unknown 2 0.5 0 0.0 2 0.3 Use of intravenous drugs 0.007 No 349 95.9 234 100.0 583 97.5 Yes 13 3.6 0 0.0 13 2.2 Unknown 2 0.5 0 0.0 2 0.3 STD, Sexually transmitted diseases.

49.5 30 12.8 210 35.1 Unknown 46 12.6 98 41.9 144 24.1 Use of non-intravenous drugs 0.001 No 299 82.1 218 93.2 517 86.5 Yes 63 17.3 16 6.8 79 13.2 Unknown 2 0.5 0 0.0 2 0.3 Use of intravenous drugs 0.007 No 349 95.9 234 100.0 583 97.5 Yes 13 3.6 0 0.0 13 2.2 Unknown 2 0.5 0 0.0 2 0.3 STD, Sexually transmitted diseases. Table 3 - Univariate and multivariate analysis of the factors associated with history of prior STD in women living with AIDS at the Centro de Referência e Treinamento em DST/AIDS, São Paulo, Brazil, 2008 to 2009 Characteristics ORbr 95%CI (ORbr) p ORaj 95%CI (ORaj) p HDI of the residence district > 0.550 1 - - 1 - - 0–0.550 11.3 6.9–18.5 < 0.001 5.5 2.8–11.0 < 0.001 Race (self-referred) White 1 - - 1 - - Not white 5.0 3.3–7.7 < 0.001 5.2 2.5–11.0 < 0.001 Age at first sexual intercourse Older than 15 years 1 - - 1 - - Up to 15 years 7.5 4.9–11.5 < 0.001 4.4 2.3–8.3 < 0.001 Time of HIV diagnosis (in years) 1 to 8 years 1 - - 1 - - 9 years and more 4.4 3.0–6.3 < 0.001 4.2 2.3–7.8 < 0.001 No. of sexual partners in life 1 to 2 1 - - 1 - - 3 to 5 5.5 3.1–9.5 < 0.001 2.2 1.1–4.6 0.045 6 and more 12.0 6.8–21.1 < 0.001 3.9 1.9–8.0 < 0.001 STD, sexually transmitted diseases; HDI, human development index.

Introduction Influenza is a highly infectious acute viral illness resulting in significant morbidity as well as healthcare resource utilization. In healthy individuals influenza is generally self-limiting, but can often cause complications.1, 2 There are 3 types of seasonal influenza viruses – A, B, and C. Influenza A causes moderate to severe illness and affects individuals of all age groups. Influenza B can cause disease of similar severity as influenza A, and even though the morbidity is higher in children, all age groups can be affected.3, 4 The influenza B virus is more stable than influenza A, with less antigenic drift and consequent immunologic stability, and does not undergo the process of antigenic shift. Influenza C is rarely reported as a cause of human illness, probably because most cases are subclinical.5 Both influenza A and B cause annual epidemics worldwide and are estimated to result in 3–5 million cases of severe illness, and 250,000–500,000 deaths.6

ability, and does not undergo the process of antigenic shift. Influenza C is rarely reported as a cause of human illness, probably because most cases are subclinical.5 Both influenza A and B cause annual epidemics worldwide and are estimated to result in 3–5 million cases of severe illness, and 250,000–500,000 deaths.6 Influenza vaccination is the most important prophylactic intervention against infection. Until the 2012–2013 influenza season, use of trivalent inactivated influenza vaccines, containing two influenza A strains (A[H1N1] and A[H3N2]) and one influenza B lineage (B/Yamagata or B/Victoria) was recommended for use in immunization programs by the World Health Organization (WHO).6 As influenza viruses undergo frequent changes in their surface antigens, new influenza vaccines are designed annually to match the circulating virus subtype expected for the next influenza season.7 The selection of the influenza B lineage is considered critical in determining the effectiveness of vaccination programs.7 Unfortunately, the correct prediction of the predominating circulating B lineage is quite difficult, often leading to inaccuracies in prediction, causing a mismatch between the recommended vaccine lineage and the circulating influenza B lineage. Prior studies have raised concerns that mismatches can result in lower vaccine effectiveness, due to the absence of cross-protection between antigenically distinct influenza B lineages, leading to more influenza cases,8, 9 and an increase in influenza-related medical resource utilization and costs.2, 10 In addition to this, Matias et al.11 have found that influenza B-associated mortality could serve as a surrogate marker of disease severity.

tection between antigenically distinct influenza B lineages, leading to more influenza cases,8, 9 and an increase in influenza-related medical resource utilization and costs.2, 10 In addition to this, Matias et al.11 have found that influenza B-associated mortality could serve as a surrogate marker of disease severity. In the Latin America and Caribbean region, seasonal influenza causes high morbidity placing a substantial economic burden on healthcare systems and society.12 Data on the burden of influenza disease for Brazil are limited, most likely due to underreporting.12 Between 2000 and 2008, data from the influenza surveillance system in Brazil revealed that influenza-like illness (ILI) led to a total of 4.39–16.92% of hospital consultations, and in 2008, of all positive reported influenza cases, 43.29% (95% CI: 37.59–49.13) were influenza B.12

d, most likely due to underreporting.12 Between 2000 and 2008, data from the influenza surveillance system in Brazil revealed that influenza-like illness (ILI) led to a total of 4.39–16.92% of hospital consultations, and in 2008, of all positive reported influenza cases, 43.29% (95% CI: 37.59–49.13) were influenza B.12 The Ministry of Health (MoH, “Ministério da Saúde”) of Brazil promotes annual national influenza vaccination campaigns. Over the years, there has been a gradual expansion of the recommended groups for annual influenza immunization in Brazil.13 Since 1999, influenza vaccination was introduced for elderly people aged above 65 years and other groups vulnerable to complications (patients with co-morbidities). In the year 2000, individuals 60 years or older were included for vaccination.14, 15 During 2011–2012, in addition to elderly people, vaccination was extended to children aged six months to those aged below two years, pregnant women, healthcare professionals, and indigenous people. In 2013, women after child birth, individuals with chronic disease and transplant, and individuals in detention facilities were included for annual influenza vaccination. In 2014, children aged 2–4 years were also included in the recommended target at-risk groups for vaccination. The information system of National Immunization Program for Brazil (“Programa Nacional de Imunização”)13 reported that across all target vaccination groups, overall mean vaccination coverage of 86.8% was reached in 2014.13 During all influenza vaccination campaigns in Brazil, trivalent vaccines were used according to WHO recommended vaccine composition for South hemisphere.16

r Brazil (“Programa Nacional de Imunização”)13 reported that across all target vaccination groups, overall mean vaccination coverage of 86.8% was reached in 2014.13 During all influenza vaccination campaigns in Brazil, trivalent vaccines were used according to WHO recommended vaccine composition for South hemisphere.16 Although high vaccination coverage levels have been reached in these target vaccination groups, little is known about the effectiveness of vaccination programs in Brazil.14, 17 A number of factors, in particular vaccine coverage, is known to influence the effectiveness of influenza vaccination programs. However, the Brazilian MoH data shows vaccine coverage to be high in almost all years since the introduction of vaccination. Importantly, the extent to which the vaccine recommended influenza B virus lineage matches the influenza virus lineage circulating in the population during an influenza season is known to impact the effectiveness of seasonal influenza vaccination programs.14 Data on laboratory surveillance of the influenza B virus in Brazil are limited, specifically data on the burden of disease and circulation patterns of influenza B lineages. The present integrative review of publicly available data aims to consolidate findings on the pattern of influenza B occurrence in Brazil to have a better understanding of influenza B epidemiology and its relevance to seasonal vaccine composition.

the burden of disease and circulation patterns of influenza B lineages. The present integrative review of publicly available data aims to consolidate findings on the pattern of influenza B occurrence in Brazil to have a better understanding of influenza B epidemiology and its relevance to seasonal vaccine composition. Material and methods Information sources and search strategy Epidemiological surveillance systems Different sources were used to retrieve information on epidemiological surveillance. We referred to international data sources to check WHO recommendations on the vaccine composition in the Southern hemisphere,18 and information on circulating influenza lineages for Brazil, the South America region and globally from the WHO/FluNet database which provides data through its network – Global Influenza Surveillance and Response System (GISRS) laboratories.19 Consolidation of available national epidemiological information on Influenza in the Epidemiological Bulletins of the Brazilian MoH was also performed. Bulletins available from 2009 to Epidemiological Week 35 of 2014 were considered.15 In Brazil, two types of surveillance exist: Sentinel surveillance of ILI and Universal Severe Acute Respiratory Syndrome (SARS) surveillance. ILI is defined as fever followed by cough or sore throat and symptoms onset within the last seven days. SARS is defined as fever followed by cough or sore throat and experiencing dyspnea, requiring hospitalization. Oxygen saturation levels lower than 95%, respiratory distress, or respiratory rate increase, are also considered in SARS. The sentinel ILI surveillance, which has a network of units distributed throughout the country's geographic regions, is used in the identification and characterization of circulating respiratory viruses in viral isolations. The universal SARS surveillance, which was implemented post-identification of the influenza A(H1N1) 2009 pandemic strain (pdm09), monitors hospitalized cases and deaths. In both systems, data are collected by means of standardized forms and entered into the online information systems: Influenza epidemiological surveillance system called SIVEP-Gripe for ILI cases, and the National Information System for Notifiable Diseases known as SINAN Influenza Web for SARS cases. Results from tests performed at the 27 Public Health Central Laboratories (“Laboratórios Centrais de Saúde Pública, Lacen”) are routinely included in these systems.

veillance system called SIVEP-Gripe for ILI cases, and the National Information System for Notifiable Diseases known as SINAN Influenza Web for SARS cases. Results from tests performed at the 27 Public Health Central Laboratories (“Laboratórios Centrais de Saúde Pública, Lacen”) are routinely included in these systems. The diagnostic kits currently available identify the influenza A(H1N1)pdm09 viral strain (determined by WHO), influenza A(H3N2), influenza A not subtyped, and influenza B virus. The antigenic characterization of the circulating influenza virus lineage is performed by three laboratories in Brazil (“Instituto Evandro Chagas” – Pará, “Instituto Adolfo Lutz”, São Paulo and “Instituto Oswaldo Cruz”, Rio de Janeiro). These laboratories are part of a network of 140 National Influenza Centers acknowledged by WHO as members of the GISRS.

e circulating influenza virus lineage is performed by three laboratories in Brazil (“Instituto Evandro Chagas” – Pará, “Instituto Adolfo Lutz”, São Paulo and “Instituto Oswaldo Cruz”, Rio de Janeiro). These laboratories are part of a network of 140 National Influenza Centers acknowledged by WHO as members of the GISRS. Online literature search and review of abstracts An integrative literature review was performed using online database tools such as PubMed/MEDLINE, Lilacs and Scielo. Searches on online databases were conducted using the following search strategy built for the PubMed database (via Medical Subject Headings [MeSH] controlled vocabulary) and adjusted for other databases according to their specificities: “(((((influenza [Title/Abstract]) OR influenzae)) OR ((orthomyxoviridae [MeSH Major Topic]) OR influenza, human [MeSH Major Topic]))) AND Brazil”. Studies on influenza B, with Brazil as the place of study and those with virological influenza B information were considered. Articles published between 2007 and 2014 were considered and a language constraint was not applied. Articles and abstracts with general influenza reports or without subtyping (A or B) were not included. One reviewer screened titles and abstracts for relevance, as the defined inclusion criteria were restrictive. An additional research was conducted in abstracts of the Annals of Scientific Events related to the area of research. A total of 10 events were considered.20, 21, 22, 23, 24, 25, 26, 27, 28, 29 All authors agreed with the selected publications.

bstracts for relevance, as the defined inclusion criteria were restrictive. An additional research was conducted in abstracts of the Annals of Scientific Events related to the area of research. A total of 10 events were considered.20, 21, 22, 23, 24, 25, 26, 27, 28, 29 All authors agreed with the selected publications. Analyses Data on viral lineage isolation and characterization, vaccination campaign, viral lineage surveillance and strain match-mismatch from different sources were collected and consolidated. Statistical analysis was not performed, and thus all data are descriptive. Results Influenza viral strain data Epidemiologic surveillance systems Information on influenza A and B subtype patterns were obtained from the WHO/FluNet database (up to epidemiological week 35, 2014). Globally and for South America, 1,122,415 and 95,040 samples were recorded in the database (WHO/FluNet-GISRS), respectively; of these 45,856 and 937 samples were typed as influenza B. In South America, of 937 influenza B samples, only 279 were subtyped, of which 252 (90.3%) samples were of the B/Yamagata lineage and 27 (9.7%) were subtyped as the B/Victoria lineage. In comparison, the WHO/FluNet database recorded 7756 samples from Brazil of which 100 samples were positive for the influenza B virus. Among these no influenza B sample was subtyped.

nly 279 were subtyped, of which 252 (90.3%) samples were of the B/Yamagata lineage and 27 (9.7%) were subtyped as the B/Victoria lineage. In comparison, the WHO/FluNet database recorded 7756 samples from Brazil of which 100 samples were positive for the influenza B virus. Among these no influenza B sample was subtyped. Epidemiologic surveillance data showed that the influenza B disease burden was moderate in both South America and Brazil. Based on WHO/FluNet (September 07, 2014), in Brazil, between 2006 and 2014, influenza B circulated in all the years, with an average of 19% of all circulating influenza viruses (A and B), varying from 1.0% to 42.6% (2006: 21.8%; 2007: 27.5%; 2008: 42.6%; 2009: 1.0%; 2010: 14.0%; 2011: 15.5%; 2012: 8.9%; 2013: 30.6% and 2014: 10.2%). In South America, between 2006 and 2014, a similar trend was observed. Influenza B virus circulated in all the years, with an average 16% of influenza B detected in relation to all of the circulating influenza viruses (A and B), varying from 0.5% to 30.1% (2006: 14.0%; 2007: 12.6%; 2008: 30.1%; 2009: 0.5%; 2010: 16.9%; 2011: 4.2%; 2012: 28.3%; 2013: 19.6% and 2014: 14.0%).

d was observed. Influenza B virus circulated in all the years, with an average 16% of influenza B detected in relation to all of the circulating influenza viruses (A and B), varying from 0.5% to 30.1% (2006: 14.0%; 2007: 12.6%; 2008: 30.1%; 2009: 0.5%; 2010: 16.9%; 2011: 4.2%; 2012: 28.3%; 2013: 19.6% and 2014: 14.0%). Trends on the etiological distribution of influenza disease were studied from the Epidemiological Bulletins of the Brazilian MoH based on two types of surveillance systems. Data from the Sentinel Surveillance of ILI (2009–2014) shows a dynamic pattern of distribution of different respiratory viruses in Brazil through the years, by epidemiological week, with predominance of influenza and respiratory syncytial virus (RSV). Influenza virus circulation demonstrates a seasonal pattern with highest occurrence from April to June, while RSV circulation showed more intense activity in the first months of the years. Influenza B follows similar seasonal trends; however it seems to occur slightly later than influenza A. During the seasons of 2009, 2010, 2011 and 2012, influenza A and RSV predominated followed by influenza B. In the year 2013, the absolute numbers of influenza B typed samples in ILI cases was higher (n = 727; 36.2% [2013]) compared to the previous years (n = 128; 21.0% [2009], n = 213; 46.3% [2010], n = 191; 34.5% [2011], n = not available [2012]) and 2014 (n = 150; 12.7% [2014, up to epidemiological week 35]). There were regional differences in the seasonal patterns of influenza virus, however strain specific information is lacking.

3]) compared to the previous years (n = 128; 21.0% [2009], n = 213; 46.3% [2010], n = 191; 34.5% [2011], n = not available [2012]) and 2014 (n = 150; 12.7% [2014, up to epidemiological week 35]). There were regional differences in the seasonal patterns of influenza virus, however strain specific information is lacking. Data on virus circulation by age group (2013, Fig. 1 ) shows that influenza A(H1N1)pdm09 virus was predominant in individuals aged 30–59 years, especially among individuals aged 40–49 years of age (37.4%). On the other hand, influenza B virus was predominant in younger people, aged 5–29 years, with highest proportions observed in children aged 5–9 years and 10–19 years (34.9% and 34.6%, respectively).Fig. 1 Distribution of the respiratory viruses identified in sentinel units of ILI by age group, Brazil, 2013. Note: Data obtained from ILI Sentinel Surveillance, SIVEP/Gripe/SVS/MS (01 March, 2014); ILI, influenza-like illness The pattern of hospitalized SARS cases from the Universal SARS surveillance showed that in 2013, although influenza A(H1N1)pdm09 was predominant, 22.5% of SARS cases positive for influenza (1337/5935, with 85 deaths) were caused by the influenza B virus. Regional variation was observed between the regions – South region (29%) and North region (12%). In 2014 (up to epidemiological week 35) 6% of SARS cases positive for influenza (75/1307, with 9 deaths) were caused by the influenza B virus (Table 1 ).Table 1 Percentage of SARS cases positive for influenza according to identified virus type by region of residence and year, Brazil.

egion (29%) and North region (12%). In 2014 (up to epidemiological week 35) 6% of SARS cases positive for influenza (75/1307, with 9 deaths) were caused by the influenza B virus (Table 1 ).Table 1 Percentage of SARS cases positive for influenza according to identified virus type by region of residence and year, Brazil. North Northeast Southeast South Central-West Brazil Up to epidemiological week 52/2013 Influenza A 88.3 79.0 80.0 70.9 85.0 77.5 Influenza B 11.7 21.0 20.0 29.1 15.0 22.5 Up to epidemiological week 35/2014 Influenza A 86.5 89.8 94.9 94.5 94.6 94.3 Influenza B 13.5 10.2 5.1 5.5 5.4 5.7 Note: Data obtained from Sinan Influenza Web/SVS/MS; SARS, Severe Acute Respiratory Syndrome. Online literature search A total of 722 articles were identified, of which 602 articles were excluded after the review of titles as they were related to influenza other than influenza B. Subsequent to the screening of titles, abstracts of the 120 potentially relevant articles were reviewed and, as a result, 108 articles were excluded when the eligibility criteria were not met (Fig. 2 ). In all, 12 articles presenting evidence on influenza B virus circulation in Brazil for the years 1996–2013 were included.30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 Details for each of the studies are shown in Supplementary File 1.Fig. 2 Flowchart of article selection from 3 databases.

iteria were not met (Fig. 2 ). In all, 12 articles presenting evidence on influenza B virus circulation in Brazil for the years 1996–2013 were included.30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 Details for each of the studies are shown in Supplementary File 1.Fig. 2 Flowchart of article selection from 3 databases. Two studies detailed characteristics of an influenza outbreak in February 2012 on cruise ships.32, 35 Fernandes et al.35 reported findings from the first influenza outbreak detected by Brazilian public health authorities in a vessel cruising in South America. Of 11 hospitalized cases of acute respiratory illness, there were six cases with influenza B virus detected in the nasopharyngeal isolates.35 In another study by Borborema et al.32, it was shown that the influenza B virus was the cause of the outbreak (detected in seven individuals with respiratory illness). Other reports from the online literature review were primarily from retrospective descriptive studies. Freitas36 showed that influenza B was attributable to moderate levels of disease during 2000 and 2010 (3–17%), and its occurrence was notable in the South and Central-West regions. Paiva et al.37 confirmed the reemergence of the B/Victoria viruses in São Paulo, Brazil, during the years 1996–2012.

e descriptive studies. Freitas36 showed that influenza B was attributable to moderate levels of disease during 2000 and 2010 (3–17%), and its occurrence was notable in the South and Central-West regions. Paiva et al.37 confirmed the reemergence of the B/Victoria viruses in São Paulo, Brazil, during the years 1996–2012. Review of abstracts A total of three abstracts from Annals of Scientific Events, in their periodic issues from 2007 to 2014 were selected42, 43, 44 (Supplementary File 1). Perosa and Bellei44 studied influenza B circulation patterns during 12 seasons (2001–2013) for São Paulo city and identified that of 96 samples subtyped in patients (mean age: 23.9 years), the majority of samples had the B/Victoria lineage (58.3%) and the remaining 41.7% belonged to the B/Yamagata lineage.44 Oliveira et al.42 studied mismatch between vaccine and circulating influenza lineages in different regions of Brazil between 2001 and 2013 and observed varying levels of co-circulation (up to 100%) of influenza B lineages.42 A study by Paiva et al.43 showed that across different regions of Brazil, 16% of influenza in clinical specimens from sentinel units in 2013 were caused by the B/Victoria lineage while the remaining cases were caused by the influenza A virus.

rved varying levels of co-circulation (up to 100%) of influenza B lineages.42 A study by Paiva et al.43 showed that across different regions of Brazil, 16% of influenza in clinical specimens from sentinel units in 2013 were caused by the B/Victoria lineage while the remaining cases were caused by the influenza A virus. Influenza B lineage match-mismatch in Brazil Epidemiologic surveillance systems In Table 2 , based on WHO/FluNet reports, consolidated annual data on the lineages that were part of the vaccine, the proportion of predominant circulating lineages and the proportion of lineage mismatch is shown for South America and Brazil. Data for Brazil was available for nine years; of these, it was only possible to compare data from three years (2007, 2008 and 2013) which have information available on the circulating influenza B virus lineage. In Brazil, co-circulation of both influenza B lineages was detected in all these three influenza seasons, and a high level of mismatch (91.4%) was observed in one year (2013) (Table 2). In the same year for South America 52.0% (2013) of mismatch was observed.Table 2 Comparison of influenza virus vaccine lineage and circulating lineage in the population, South America and Brazil, 2006–2014 (WHO/FluNet).

e influenza seasons, and a high level of mismatch (91.4%) was observed in one year (2013) (Table 2). In the same year for South America 52.0% (2013) of mismatch was observed.Table 2 Comparison of influenza virus vaccine lineage and circulating lineage in the population, South America and Brazil, 2006–2014 (WHO/FluNet). Reference Predominant lineage (n; %)a Vaccine Recommendation % Mismatchb 2006 South America – Victoria – Brazil – Victoria – 2007 South America Victoria (5; 0.2) Victoria 28.6 Brazil Victoria (5; 6.3) Victoria 28.6 2008 South America Yamagata (46; 2.1) Yamagata 37.0 Brazil Yamagata (15; 4.6) Yamagata 42.3 2009 South America – Yamagata – Brazil – Yamagata – 2010 South America Victoria (103; 1.0) Victoria 1.9 Brazil – Victoria – 2011 South America Victoria (6; 0.1) Victoria 0 Brazil – Victoria – 2012 South America Yamagata (489; 4.5) Victoria 50.5 Brazil – Victoria – 2013 South America Victoria (364; 1.8) Yamagata 52.0 Brazil Victoria (32; 0.7) Yamagata 91.4 2014 South America Yamagata (252; 3.7) Yamagata 9.7 Brazil – Yamagata – Note: Data taken from WHO/FluNet (07 September, 2014).19 “–” indicates that data is not available for this year. a Percentages are representative of the predominating lineage which are estimated with the total number of samples positive for influenza B as the denominator. b % Mismatch = 100% − % Circulating B lineage match with vaccine.

Reference Predominant lineage (n; %)a Vaccine Recommendation % Mismatchb 2006 South America – Victoria – Brazil – Victoria – 2007 South America Victoria (5; 0.2) Victoria 28.6 Brazil Victoria (5; 6.3) Victoria 28.6 2008 South America Yamagata (46; 2.1) Yamagata 37.0 Brazil Yamagata (15; 4.6) Yamagata 42.3 2009 South America – Yamagata – Brazil – Yamagata – 2010 South America Victoria (103; 1.0) Victoria 1.9 Brazil – Victoria – 2011 South America Victoria (6; 0.1) Victoria 0 Brazil – Victoria – 2012 South America Yamagata (489; 4.5) Victoria 50.5 Brazil – Victoria – 2013 South America Victoria (364; 1.8) Yamagata 52.0 Brazil Victoria (32; 0.7) Yamagata 91.4 2014 South America Yamagata (252; 3.7) Yamagata 9.7 Brazil – Yamagata – Note: Data taken from WHO/FluNet (07 September, 2014).19 “–” indicates that data is not available for this year. a Percentages are representative of the predominating lineage which are estimated with the total number of samples positive for influenza B as the denominator. b % Mismatch = 100% − % Circulating B lineage match with vaccine. Online literature search and review of abstracts None of the articles from the online literature search in particular studied influenza B lineage match-mismatch in Brazil. However, from the reviewed abstracts, all three focused on B lineage match-mismatch in Brazil.42, 43, 44 Overall, mismatch levels in the range 0–100% were observed between the circulating influenza B virus lineage and the vaccine recommended lineage in Brazil and São Paulo city between 2000 and 2013 (Oliveira et al.42 and Perosa and Bellei44) (Fig. 3 ). Oliveira et al.42 showed that the B/Victoria and B/Yamagata lineages have co-circulated in Brazil since 2005. Key mismatches between the WHO recommended Southern hemisphere vaccine composition and the most prevalent circulating viruses in Brazil were observed in the years 2002 (100%), 2005 (67%), 2008 (65%), 2010 (72%) and 2013 (91%).42 A separate dataset on influenza B circulation from the São Paulo city also showed a high degree of mismatch in three seasons i.e. years 2002 (75%), 2010 (91%) and 2013 (100%)44 (Fig. 3). Moderate levels of regional variation across Brazil were evident (data not shown). Paiva et al.43 reported a complete mismatch (100%) between the circulating influenza B virus lineage and the vaccine recommended lineage in the 2013 influenza season in different regions of Brazil.Fig. 3 Circulation of Influenza B lineages according to season (year) and recommended vaccine lineage, Brazil and São Paulo city, 2001–2013. Note: Data taken (with permission) for Brazil from Oliveira et al.42 and for São Paulo city from Perosa and Bellei44; Samples for Brazil are estimated from regional data; Samples for the South (Rio Grande do Sul, Parana, Santa Catarina states), Southeast (Minas Gerais, Espirito Santo, Rio de Janeiro states) and Northeast (Bahia, Alagoas, Sergipe states) for which the samples were sequenced at WHO/National Influenza Center, Rio de Janeiro; Samples from other states were downloaded from the database of The Global Initiative on Sharing All Influenza Data; SP, São Paulo; Vic, B/Victoria; Yam, B/Yamagata.

io de Janeiro states) and Northeast (Bahia, Alagoas, Sergipe states) for which the samples were sequenced at WHO/National Influenza Center, Rio de Janeiro; Samples from other states were downloaded from the database of The Global Initiative on Sharing All Influenza Data; SP, São Paulo; Vic, B/Victoria; Yam, B/Yamagata. Discussion Data on influenza B disease burden for Brazil are limited, especially with regards to influenza B disease burden and subtypes, which are important when considering the use of influenza vaccines. In this review, we aimed to summarize the available evidence base on influenza B disease patterns in Brazil in the international and local public health databases, as well from the literature and plenary scientific events. The review of these data highlights important patterns of influenza B circulation in Brazil. Based on WHO/FluNet reports, moderate levels of influenza B burden (1.0–42.6%) were observed over nine years, with the exception of 2009 (1.0%), which could potentially have been due to the displacement of influenza B circulation resulting from the influenza A pandemic in 2009. This finding indicates the unpredictability of influenza B circulation.

oderate levels of influenza B burden (1.0–42.6%) were observed over nine years, with the exception of 2009 (1.0%), which could potentially have been due to the displacement of influenza B circulation resulting from the influenza A pandemic in 2009. This finding indicates the unpredictability of influenza B circulation. Based on WHO/FluNet reports, it was possible to compare data from only three years during 2006–2014 (2007, 2008 and 2013) which have information on the circulating influenza B virus lineages. Significant levels of co-circulation of both influenza B virus lineages during the three influenza seasons between 2006 and 2014 were observed. These data indicate that in 2013, a high degree of mismatch between the vaccine and the predominating circulating lineage (91.4%) occurred, and during the other two influenza seasons, a partial mismatch was reported.

both influenza B virus lineages during the three influenza seasons between 2006 and 2014 were observed. These data indicate that in 2013, a high degree of mismatch between the vaccine and the predominating circulating lineage (91.4%) occurred, and during the other two influenza seasons, a partial mismatch was reported. The three reviewed abstracts, which specifically report findings on influenza B mismatch, corroborate this unpredictable behavior of influenza B disease in Brazil for many other seasons for which data were not available in the International Epidemiological surveillance data. Significant levels of co-circulation of both influenza B lineages (B/Victoria and B/Yamagata) and lineage mismatch between the vaccine and circulating lineage was reported for Brazil in five influenza seasons (2002, 2005, 2008, 2010, and 2013).42, 43, 44 Previous studies suggest that even with a partial mismatch over the years, due to the unpredictability of influenza B lineage circulation, the disease burden (in terms of clinical cases, hospitalizations and health care resource utilization) and societal burden can be considerable.2, 9 For example, a mismatch in the 2007–2008 influenza season in the United States was estimated to have costed health care providers and society well over $100 million and $1 billion, respectively.45 In another study conducted in Taiwan, it was reported that an epidemic predominated by influenza B/Yamagata lineage occurred during the 2011–2012 season during which the trivalent influenza vaccine contained influenza B/Victoria lineage. Expectedly, the morbidity and mortality of this vaccine mismatched epidemic was substantial (influenza B: 60.7% and 66.9% of confirmed influenza cases and deaths, respectively; 87.2% of samples showed the presence of vaccine opposite lineage).46

ch the trivalent influenza vaccine contained influenza B/Victoria lineage. Expectedly, the morbidity and mortality of this vaccine mismatched epidemic was substantial (influenza B: 60.7% and 66.9% of confirmed influenza cases and deaths, respectively; 87.2% of samples showed the presence of vaccine opposite lineage).46 Surveillance data on the age distribution of influenza cases is available only for 2013. This data showed that influenza B virus was predominantly observed in younger individuals aged 5–29 years, with the highest proportions in children aged 5–9 years and 10–19 years. Even though this observation was restricted to only one year with 90% mismatch it is in line with previous reports wherein influenza B has been reported to affect more schoolchildren and the appearance of the new B lineage reinforced this pattern.4 An implication of this finding is that children and adolescents might benefit the most from the addition of a second influenza B lineage to the seasonal influenza vaccine. Moreover, it could be useful for the entire population as it is widely considered that vaccinating children can reduce influenza illness in family members and other susceptible populations in the community by reducing the risk of exposure and subsequent influenza infection and related complications.47 A similar finding by Heikkinen et al.10 reports a substantial population-level impact of outbreaks resulting from mismatched seasons in Finland with this impact predominating in children and adolescents.

ns in the community by reducing the risk of exposure and subsequent influenza infection and related complications.47 A similar finding by Heikkinen et al.10 reports a substantial population-level impact of outbreaks resulting from mismatched seasons in Finland with this impact predominating in children and adolescents. From the analysis of the Epidemiological Bulletins of the Brazilian MoH based on two types of surveillance systems in Brazil, it is shown that the proportions of ILI and SARS cases positive for influenza B were moderate. It was observed that despite high levels of vaccination coverage (91.6%) observed across all target groups in 2013,13 the proportion of registered SARS cases due to influenza B in 2013 was 22.5% (1337/5935; Universal SARS surveillance), a year in which 91.4% of the influenza B laboratory samples did not correspond to the lineage contained in the trivalent vaccine (WHO/FluNet). In the same year, 85 deaths due to influenza B were registered. This occurrence could have been due to an outbreak of SARS caused by influenza B. It was demonstrated by Matias et al.11 that influenza B-associated mortality could potentially be a valid indication of disease severity. Thus the evidence presented here for one year underscores the importance of a universal influenza vaccination strategy compared to targeted immunization as universal immunization at high vaccine coverage levels could potentially interrupt transmission reducing the occurrence of outbreaks and severe disease.

e severity. Thus the evidence presented here for one year underscores the importance of a universal influenza vaccination strategy compared to targeted immunization as universal immunization at high vaccine coverage levels could potentially interrupt transmission reducing the occurrence of outbreaks and severe disease. Our literature review confirms the re-emergence of B/Victoria lineage in Brazil during the years 2000–2002. These data are in line with that of Motta et al.48 which shows the re-emergence of the Victoria-lineage viruses in the Northern hemisphere and in the South and South East regions of Brazil where previously the B/Yamagata lineage was the predominating circulating lineage and the vaccine recommended lineage. Other studies confirm that the B/Yamagata was the major circulating lineage until the 1980s, when B/Victoria lineage viruses appeared; since then, drift variants of both influenza B lineages have been co-circulating worldwide.49, 50 As influenza B viruses can co-circulate during an epidemic allowing the re-emergence of old lineages due to re-assortment between the different strains,4, 48 there is a need to improve influenza laboratory-based surveillance in Brazil. Importantly these data also highlight the unpredictability of influenza B circulation, making it difficult to consistently predict which B lineage will predominate during a given influenza season.

ssortment between the different strains,4, 48 there is a need to improve influenza laboratory-based surveillance in Brazil. Importantly these data also highlight the unpredictability of influenza B circulation, making it difficult to consistently predict which B lineage will predominate during a given influenza season. Due to the little cross-protection conferred between the two antigenically distinct influenza B lineages, a vaccine lineage mismatch with the circulating lineage could result in additional burden in terms of health care resource utilization and can adversely impact the society. This burden could, in particular, be substantial during an outbreak. To avoid this additional burden, a plausible option would be to switch from the use of trivalent influenza vaccines to quadrivalent vaccines in national campaigns.51 Quadrivalent influenza vaccines are expected to offer broader protection against influenza B disease. The WHO recently updated recommendations beginning with the 2013–2014 influenza, now recommending the use of a quadrivalent vaccine composition with the addition of a second influenza B lineage as well as the two influenza A strains and one influenza B lineage contained in the trivalent vaccines.6, 52

luenza B disease. The WHO recently updated recommendations beginning with the 2013–2014 influenza, now recommending the use of a quadrivalent vaccine composition with the addition of a second influenza B lineage as well as the two influenza A strains and one influenza B lineage contained in the trivalent vaccines.6, 52 Some limitations of this review require consideration. Besides the inherent limitations of each of the studies included in the literature search and abstracts, there are other important limitations. Statistical and quality analysis of the articles was not performed. Also, the interpretation of data is constrained by the gaps in laboratory surveillance in which there were no influenza B data available for many years with respect to age, severity, and seasonality. As we present data over many years through which the surveillance system has gradually improved, the number of positive samples is still small, and the more recent years are likely to be over-represented. Additionally, sentinel surveillance data cannot be truly representative for the whole population as the system of sentinel units involves different levels of care. Moreover, there is no standard protocol for patient selection for sample collection which may have led to a selection bias of certain age groups. Despite these limitations, the surveillance system for influenza and other respiratory viruses which has improved over the years has proven useful to describe influenza B circulation patterns for Brazil and demonstrate that influenza B epidemiology is changing in line with observations reported from other countries.4

. Despite these limitations, the surveillance system for influenza and other respiratory viruses which has improved over the years has proven useful to describe influenza B circulation patterns for Brazil and demonstrate that influenza B epidemiology is changing in line with observations reported from other countries.4 Conclusions Influenza surveillance systems, and laboratory-based and epidemiological studies of influenza B in Brazil, while limited, have improved over the years. There is a need to strengthen and extend influenza surveillance for influenza B sample subtyping to determine the behavior pattern of influenza B lineages. The findings from this integrative data review provide evidence of the unpredictable nature of influenza B circulation in Brazil, the increasing frequency of co-circulation of both influenza B lineages, and mismatch between the circulating influenza B lineage and the composition of the seasonal influenza vaccine for the region. This data is suggestive of the additional benefit that quadrivalent influenza vaccines, containing both influenza B lineages (B/Yamagata and B/Victoria), could offer over the use of the currently available trivalent vaccines for the prevention of seasonal influenza in Brazil.

seasonal influenza vaccine for the region. This data is suggestive of the additional benefit that quadrivalent influenza vaccines, containing both influenza B lineages (B/Yamagata and B/Victoria), could offer over the use of the currently available trivalent vaccines for the prevention of seasonal influenza in Brazil. Authors contributions All authors participated to the conception/design of the review, performed or supervised the analysis, and interpreted the data. Otavio Cintra, Eliana Nogueira Castro de Barros, Laís Freitas and Erika Rossetto collected or assembled the data. Erika Rossetto wrote the preliminary report of the review findings. All authors read and approved the final manuscript. Conflicts of interest Eliana Nogueira Castro de Barros, Otavio Cintra and Romulo Colindres are employees of the GSK group of companies. Romulo Colindres and Otavio Cintra report ownership of stock options and/or restricted shares. Laís Freitas reports that she is working for GSK Vaccines, but is employed by Shift de Gestão em Serviços. Erika Rossetto has no conflict of interest. Appendix A Supplementary data The following are the supplementary data to this article: Acknowledgements The authors would like to thank Andreza Madeira Macario (epidemiologist to INOVATEC) for intellectual contribution to the study, Amrita Ostawal for medical writing services (consultant publications writer to GSK Vaccines) and Bruno Dumont (Business & Decision Life Sciences on behalf of GSK Vaccines) for editorial assistance and publication coordination.

reza Madeira Macario (epidemiologist to INOVATEC) for intellectual contribution to the study, Amrita Ostawal for medical writing services (consultant publications writer to GSK Vaccines) and Bruno Dumont (Business & Decision Life Sciences on behalf of GSK Vaccines) for editorial assistance and publication coordination. ☆ GlaxoSmithKline Biologicals SA funded this study and was involved in all stages of study conduct, including analysis of the data and funded all costs associated with the development and publication of this manuscript. Appendix A Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.bjid.2015.09.009.

Introduction Influenza (INF) A and B viruses are a frequent cause of respiratory tract infection in humans, affecting people of all ages with varied morbidity, sometimes leading to fatal outcomes. INF A viruses are divided into various subtypes based on the antigenic properties of surface glycoproteins, hemagglutinin and neuraminidase. INF B viruses are not subdivided into subtypes, but two genetically distinct lineages of influenza B viruses, namely B/Yamagata lineage and B/Victoria lineage have circulated globally since 1985 and co-circulated since 2001.1, 2, 3 People at the extremes of age are at increased risk of complications, hospitalization and influenza-associated death. The burden of influenza has long been recognized in the elderly due to the increased risk of death secondary to pneumonia or cardiac complications.4 In young and very young children, studies have demonstrated that the burden of influenza-associated acute lower respiratory infection (ALRI) is also substantial.5 A recent meta-analysis including 43 studies and data from approximately eight million children, estimated that 90 million new cases of influenza and 20 million cases of influenza-associated ALRI occurred worldwide in children younger than five years in 2008.6 However, few studies have considered the burden of influenza A and B in the outpatient setting.7

studies and data from approximately eight million children, estimated that 90 million new cases of influenza and 20 million cases of influenza-associated ALRI occurred worldwide in children younger than five years in 2008.6 However, few studies have considered the burden of influenza A and B in the outpatient setting.7 Seasonal influenza vaccination is the main strategy to prevent influenza and its complications. In Brazil, the trivalent influenza vaccine (TIV) is offered at no cost for individuals aged 60 years and over, children from six months to five years of age, pregnant and puerperal women (up to 45 days postpartum), health care workers, school teachers, indigenous populations, immunocompromised patients or with chronic conditions, adolescents and young adults aged 12–21 under socio-educational measures, and prisoners and prison staff. The quadrivalent vaccine (QIV), which includes also the B/Yamagata lineage is available only in private clinics. Influenza vaccination campaigns have been successful, with vaccine coverage above 70%. However, the impact of campaigns on reducing influenza burden is not well established since the information on the incidence and severity of influenza is limited. Data on the burden of influenza B in children and adolescents are even scarcer.

vaccination campaigns have been successful, with vaccine coverage above 70%. However, the impact of campaigns on reducing influenza burden is not well established since the information on the incidence and severity of influenza is limited. Data on the burden of influenza B in children and adolescents are even scarcer. During a prospective cohort study of dengue in children and adolescents living in the city of Araraquara, Sao Paulo, Brazil the investigators observed that patients with fever and respiratory symptoms such as cough, coryza and sore throat were more likely to test negative for dengue.8 We then recognized in this consolidated cohort, a window of opportunity to prospectively compare the frequency, epidemiology and morbidity of influenza A and B infections in outpatient children and adolescents with influenza-like illness (ILI).

ough, coryza and sore throat were more likely to test negative for dengue.8 We then recognized in this consolidated cohort, a window of opportunity to prospectively compare the frequency, epidemiology and morbidity of influenza A and B infections in outpatient children and adolescents with influenza-like illness (ILI). Material and methods Study population and sample size The consolidated dengue cohort was recruited between August 2014 and March 2015. A total of 3,514 children and adolescents aged two to 16 years were randomly selected from the population of Araraquara at the time of recruitment. From December 2016 to August 2018, subjects from the original cohort who presented ILI were included in the influenza cohort and prospectively followed during two consecutive influenza seasons. In 2017 and 2018, the winter started on June 21st in the southern hemisphere. Sample size was estimated based on the frequency of febrile episodes with respiratory symptoms observed in the first year of the original (dengue) cohort. During the 20-month period of the study, we hypothesized that 1,200 cases of febrile syndrome followed by respiratory symptoms would occur, of which 5% would be influenza with virological confirmation.9 Thus, at least 60 cases of proven influenza infection would be available for analysis.

the original (dengue) cohort. During the 20-month period of the study, we hypothesized that 1,200 cases of febrile syndrome followed by respiratory symptoms would occur, of which 5% would be influenza with virological confirmation.9 Thus, at least 60 cases of proven influenza infection would be available for analysis. Definitions ILI was defined by the presence of fever and two or more of the following signs and symptoms: cough, coryza, tonsillitis, pharyngitis, tachypnea, dyspnea, myalgia, headache, inappetence or prostration appearing within seven days. Proven influenza infection was defined by the occurrence of ILI with a laboratory confirmation of influenza A or B. Lymphopenia was defined as an absolute lymphocyte count (ALC) below 1.0 × 106 lymphocytes/mm3. Leukocytosis was defined as an elevated white blood cell (WBC) count greater than 11,000 per mm3.

seven days. Proven influenza infection was defined by the occurrence of ILI with a laboratory confirmation of influenza A or B. Lymphopenia was defined as an absolute lymphocyte count (ALC) below 1.0 × 106 lymphocytes/mm3. Leukocytosis was defined as an elevated white blood cell (WBC) count greater than 11,000 per mm3. Follow-up After enrollment, parents or guardians received a weekly phone call for fever surveillance. In the case of a temperature >37.5 °C and respiratory symptoms, a home visit was scheduled to confirm ILI and collection of oral and nasopharynx swabs. In addition to demographic data, the following information was assessed during home visits: type of respiratory symptoms, date of initial symptoms, presence of symptomatic household contacts, presence of comorbidities, need of hospitalization and information on influenza vaccination in that year. All information was transcribed to case report forms (CRF). Children with laboratory-confirmed influenza or with more pronounced symptoms were referred to a pediatrician by the field team. Oseltamivir was introduced at physician discretion.

need of hospitalization and information on influenza vaccination in that year. All information was transcribed to case report forms (CRF). Children with laboratory-confirmed influenza or with more pronounced symptoms were referred to a pediatrician by the field team. Oseltamivir was introduced at physician discretion. Diagnosis of influenza and other respiratory viruses Respiratory virus diagnosis was performed at the Virology Laboratory, Institute of Tropical Medicine (University of São Paulo School of Medicine). Influenza A or B was diagnosed by nucleic acid test (NAT), using the Cepheid (Xpert® Xpress Flu, Sunnyvale, CA, USA) platform, or the XGen Influenza multiplex kit (Mobius Life Science, Pinhais, PR, Brazil) according to manufacturer’s instructions. Due to its rapid turnaround time, the Cepheid platform was used first, to assure early introduction of oseltamivir, at physician discretion. Aliquots of respiratory samples were frozen and stored at −80° C for further diagnosis of other pathogens using a multiplex platform (XGen Respiratory Panel 21 pathogens multiplex kit (Mobius Life Science, Pinhais, PR, Brazil), according to manufacturer’s instructions.

roduction of oseltamivir, at physician discretion. Aliquots of respiratory samples were frozen and stored at −80° C for further diagnosis of other pathogens using a multiplex platform (XGen Respiratory Panel 21 pathogens multiplex kit (Mobius Life Science, Pinhais, PR, Brazil), according to manufacturer’s instructions. Sequencing of influenza B viruses Sequencing was proposed to evaluate the match between circulating influenza B viruses and the lineages included in the trivalent (TIV) and quadrivalent (QIV) influenza vaccines recommended during the study. Primers for the hemagglutinin (HA) region were designed for INF B (FluBHA_F1-TTGGAACCTCAGGRTCTTGC or FluBHA_F2 – TCCTATAATGCACGAYAGAACA and FluBHA_R-TGCAGGAGGTCTATATTTGGTTC). Fragments were sequenced using Sanger method and sequences were built and analyzed in the CLC GenomicsWorkbench (Qiagen-https://www.qiagenbioinformatics.com/products/clc-genomics-workbench/). The visualization and obtaining of the information described in the report were done with the aid of the CLC. Reference sequences of Influenza vaccine strains were obtained from the GISAID - Global Initiative on Sharing All Influenza Data (https://www.gisaid.org/). These were aligned together to worldwide references samples from 2015 to 2018 available at GenBank and also to sequences generated in this study.

he aid of the CLC. Reference sequences of Influenza vaccine strains were obtained from the GISAID - Global Initiative on Sharing All Influenza Data (https://www.gisaid.org/). These were aligned together to worldwide references samples from 2015 to 2018 available at GenBank and also to sequences generated in this study. We first looked for amino acid differences between our samples and the vaccine strains. Then we reconstructed a phylogenetic tree from partial HA gene (47 sequences with 616 nucleotides long) to investigate the epidemiology of the circulating strains. Phylogenetic reconstructions were performed using the Maximum Likelihood method in the PhyML program within the SeaView package10 using the best nucleotide model (HKY + I) chosen by ModelTEst.11 Influenza vaccines and policies In 2016, the components of influenza vaccine for the southern hemisphere were A/California/7/2009 (H1N1)pdm09, A/Hong Kong/4801/2014(H3N2) and B/Brisbane/60/2008 (B/Victoria lineage). The strain A/Michigan/45/2015(H1N1)pdm09 replaced the strain A/California/7/2009(H1N1)pdm09 in 2017. The quadrivalent influenza vaccine (QIV) also including the lineage B/Phuket/3073/2013-like virus (B/Yamagata lineage) and was only available in private clinics.

1/2014(H3N2) and B/Brisbane/60/2008 (B/Victoria lineage). The strain A/Michigan/45/2015(H1N1)pdm09 replaced the strain A/California/7/2009(H1N1)pdm09 in 2017. The quadrivalent influenza vaccine (QIV) also including the lineage B/Phuket/3073/2013-like virus (B/Yamagata lineage) and was only available in private clinics. Ethical issues The study was approved by the Ethics Committee of the Faculty of Medicine of the University of São Paulo (CAAE no. 25706913.6.1001.0065). The selected households were visited by the project’s field team. The parents or guardians were asked to sign additional informed consent forms, specific for the influenza study. The child and/or the adolescent were requested to sign the Term of Assent, in accordance with the current legislation. Statistical analysis Patient characteristics were summarized using descriptive statistics. Differences between categorical and continuous variables were calculated using Pearson’s chi square, Student’s t-test and Mann Whitney tests, as appropriate. In all statistical analyses, a two-sided p-value ≤ 0.05 was considered statistically significant (IBM SPSS statistics version 21 for Windows). Results Study population The study started on December 5, 2016, when the first case of ILI was included, and ended on August 30, 2018. Thus, the study cohort was followed for 20 consecutive months, including two winters. One hundred and seventy-nine patients were included, 99 males (55.3%) and 80 females (44.7%), median age 10 years, ranging from eight months to 19 years of age.

6, when the first case of ILI was included, and ended on August 30, 2018. Thus, the study cohort was followed for 20 consecutive months, including two winters. One hundred and seventy-nine patients were included, 99 males (55.3%) and 80 females (44.7%), median age 10 years, ranging from eight months to 19 years of age. Seasonality and clinical findings in ILI episodes The patients had 277 episodes of ILI during the study period, with a median of one episode per patient, ranging from one to four episodes. Thirty-three patients had two episodes of ILI, eight had three episodes and two had four episodes of ILI. As expected, an increase in the number of episodes of ILI was observed in winter months, although a second wave of ILI was observed in the spring of 2017 (arrow, Fig. 1 ). In addition to fever, the most frequent symptoms were headache, observed in 233 of the 277 ILI episodes (84.1%), somnolence (in 208 episodes, 75.1%), cough (in 204 episodes, 73.6%) and coryza (in 193 episodes, 69.7%). Hemogram was performed in 202 of the 277 episodes of ILI (73%). Median leukocyte and lymphocyte counts were 6,200 (1,800–31,300)/mm3 and 2,419 (708–4,602)/mm3, respectively. Leukocytosis was observed in 121 (60%) and lymphopenia in four (2%) of the ILI episodes. Children were referred to the pediatrician in 87 of the 277 episodes of ILI (31.4%). No patient with ILI needed hospital admission.Fig. 1 Seasonality of respiratory pathogens diagnosed during influenza-like illness episodes. Dotted line represents influenza-like illness episodes. Fig. 1

Seasonality and clinical findings in ILI episodes The patients had 277 episodes of ILI during the study period, with a median of one episode per patient, ranging from one to four episodes. Thirty-three patients had two episodes of ILI, eight had three episodes and two had four episodes of ILI. As expected, an increase in the number of episodes of ILI was observed in winter months, although a second wave of ILI was observed in the spring of 2017 (arrow, Fig. 1 ). In addition to fever, the most frequent symptoms were headache, observed in 233 of the 277 ILI episodes (84.1%), somnolence (in 208 episodes, 75.1%), cough (in 204 episodes, 73.6%) and coryza (in 193 episodes, 69.7%). Hemogram was performed in 202 of the 277 episodes of ILI (73%). Median leukocyte and lymphocyte counts were 6,200 (1,800–31,300)/mm3 and 2,419 (708–4,602)/mm3, respectively. Leukocytosis was observed in 121 (60%) and lymphopenia in four (2%) of the ILI episodes. Children were referred to the pediatrician in 87 of the 277 episodes of ILI (31.4%). No patient with ILI needed hospital admission.Fig. 1 Seasonality of respiratory pathogens diagnosed during influenza-like illness episodes. Dotted line represents influenza-like illness episodes. Fig. 1 Laboratory-confirmed influenza A and B Using the influenza platform (Cepheid Xpert® Xpress Flu) we identified 90 episodes of proven influenza infection, 73 influenza A (81.1%) and 17 by influenza B (18.9%). During laboratory work up with the multiplex platform to identify other respiratory pathogens, nine additional influenza infections were diagnosed (three influenza B and six influenza A). Thus, 90 of the 179 patients (50.3%) had 99 episodes of proven influenza infection, being 79 caused by influenza A (79.8%) and 20 by influenza B (20.2%). Nine patients had two episodes of influenza. Two patients had two episodes of INF A in the same year; four had two episodes of INF A in consecutive years; one patient had two episodes of INF B in the same year; and three patients had two separate episodes of influenza A and B in the same year (one patient), or in consecutive years (two patients).

es of influenza. Two patients had two episodes of INF A in the same year; four had two episodes of INF A in consecutive years; one patient had two episodes of INF B in the same year; and three patients had two separate episodes of influenza A and B in the same year (one patient), or in consecutive years (two patients). In general, the clinical presentations of influenza A and B infections were similar and regularly mild, as shown in Table 1 . Patients with influenza B were more likely to be male (85%, p = 0.01) and presented more prostration than patients infected with influenza A virus (p = 0.01). Among the 87 outpatient visits to the pediatrician, 61 were due to influenza (70.1%). Interestingly, the number of visits was significantly higher for INF A episodes in comparison with INF B (67.1% vs 40%; p = 0.0003). Oseltamivir for five days was introduced only in one case of influenza A.Table 1 Clinical and laboratory findings in influenza A and B infection (N = 99).

1 were due to influenza (70.1%). Interestingly, the number of visits was significantly higher for INF A episodes in comparison with INF B (67.1% vs 40%; p = 0.0003). Oseltamivir for five days was introduced only in one case of influenza A.Table 1 Clinical and laboratory findings in influenza A and B infection (N = 99). Table 1Variable Category Influenza A Influenza B p Value N = 79 (%) N = 20 (%) Median age (range) years 9 (4–17) 9 (5–14) NS Sex Female 37 (37.4) 3 (3.0) 0.01 Male 42 (42.4) 17 (17.2) Symptoms Coryza 67 (84.8) 20 (100) 0.06 Somnolence 66 (83.5) 19 (95) 0.19 Inappetence 62 (78.5) 15 (75) 0.74 Headache 72 (72.7) 18 (18.2) 0.87 Cough 71 (71.7) 16 (16.2) 0.23 Myalgia 49 (62) 13 (65) 0.80 Abdominal pain 46 (58.2) 9 (45) 0.23 Nausea 46 (58.2) 8 (40) 0.14 Prostration 38 (48.1) 16 (80) 0.01 Conjunctivitis 30 (38) 12 (60) 0.075 Arthralgia 30 (38) 5 (25) 0.28 Pharyngitis 25 (31.6) 5 (25) 0.56 Vomit 24 (30.4) 6 (30) 0.97 Tonsillitis 19 (24.1) 4 (20) 0.70 Diarrhea 16 (20.3) 4 (20) 0.98 Dyspnea 12 (15.2) 3 (15) 0.98 Rash 5 (6.3) 2 (10) 0.56 Lethargy 3 (3.8) 2 (10) 0.26 Tachypnea 1 (1.3) 1 (5) 0.29 Median lymphocyte count (range) 2,343 (708–4,602) 2,743.5 (1,888–4,484) 0.89 Leukocytosis 30 (38) 4 (20) 0.18 Lymphopenia 4 (5) 0 (0) 0.58 Outpatient visits 53 (67.1) 8 (40) 0.026

rrhea 16 (20.3) 4 (20) 0.98 Dyspnea 12 (15.2) 3 (15) 0.98 Rash 5 (6.3) 2 (10) 0.56 Lethargy 3 (3.8) 2 (10) 0.26 Tachypnea 1 (1.3) 1 (5) 0.29 Median lymphocyte count (range) 2,343 (708–4,602) 2,743.5 (1,888–4,484) 0.89 Leukocytosis 30 (38) 4 (20) 0.18 Lymphopenia 4 (5) 0 (0) 0.58 Outpatient visits 53 (67.1) 8 (40) 0.026 Other respiratory pathogens One hundred and thirty-eight of the 277 episodes of ILI (49.8%) tested negative by all techniques. In the remaining 139 episodes (50.2%), at least one respiratory pathogen was identified. Influenza A and B viruses were the most frequent agents identified (99 episodes) followed by human rhinovirus (22 episodes). Table 2 shows the respiratory pathogens diagnosed during the study. Fig. 1 shows the seasonality of ILI and of the respiratory pathogens. The second wave of ILI episodes observed in 2017 was mainly due to influenza B infections.Table 2 Respiratory pathogens identified in 139 episodes of influenza-like illness. Table 2Pathogen Number (%) Influenza A 76 (54.6) Influenza B 20 (14.4) Rhinovirus 16 (11.5) Influenza A/H1N1 2 (1.4) Influenza A + Rhinovirus 1 (0,07) Rhinovirus + enterovirus 4 (2.8) Rhinovirus + RSV 1 (0.07) Respiratory Syncytial Virus 4 (2.8) Adenovirus 4 (2.8) Parainfluenza 2 2 (1.4) Parainfluenza 2 + metapneumovirus A/B 1 (0,07) Metapneumovirus A/B 2 (1.4) Human coronavirus 2 (1.4) Mycoplasma pneumoniae 2 (1.4) Parecovirus 2 (1.4)

fluenza A + Rhinovirus 1 (0,07) Rhinovirus + enterovirus 4 (2.8) Rhinovirus + RSV 1 (0.07) Respiratory Syncytial Virus 4 (2.8) Adenovirus 4 (2.8) Parainfluenza 2 2 (1.4) Parainfluenza 2 + metapneumovirus A/B 1 (0,07) Metapneumovirus A/B 2 (1.4) Human coronavirus 2 (1.4) Mycoplasma pneumoniae 2 (1.4) Parecovirus 2 (1.4) Influenza vaccination Information on influenza vaccination was obtained during home visits, when respiratory samples were taken. In 49 of the 277 episodes of ILI (17.7%), the information about influenza vaccine was not available. In the remaining 228 episodes of ILI, 85 had received the trivalent vaccine (37.3%), and 143 had not been vaccinated (62.7%). Excluding the cases without vaccine information, no difference was seen in the occurrence of influenza in vaccinated and non-vaccinated patients (p = 0.47). According to the country recommendation, seven children were within the age range of influenza vaccine. In two of them, the vaccination status was unknown, and out of the remaining five, four (80%) had been vaccinated.

ference was seen in the occurrence of influenza in vaccinated and non-vaccinated patients (p = 0.47). According to the country recommendation, seven children were within the age range of influenza vaccine. In two of them, the vaccination status was unknown, and out of the remaining five, four (80%) had been vaccinated. Influenza B lineages From the 20 cases of influenza B, 12 sequences could be generated and were used for analysis. The amino acid analysis showed that while 11 strains were related to the Phuket (B/Yamagata lineage), one of the circulating virus (patient BR_776_June_2017) was similar to the Brisbane virus (Victoria lineage) (data not shown). This was also noticeable through the phylogenetic reconstruction (Fig. 2 ) where the two main clusters comprised strains related to either Brisbane or Phuket lineages sampled at the same location and years, indicating co-circulation of both strains. The phylogeny also revealed that there was little difference among the Brazilian, European and USA circulating influenza B viruses since they all clustered together with no particular geographic pattern. A slight temporal structure can be observed in the tree, with viruses sampled in September and October clustering together, and samples from November and December 2017 clustered together to viruses sampled in 2018 (from other countries). In comparison to the vaccine lineages (Brisbane_60 and Phuket_3073), we observed that six of the eleven patients (54.5%) in the Phuket branch of the phylogenetic tree had not received influenza vaccine, one (9%) had unknown vaccination status, and four (36.3%) had received the TIV, which contained only the Brisbane lineage of influenza B virus. The only patient with influenza B in the Brisbane branch had not been vaccinated.Fig. 2 Maximum likelihood phylogenetic tree of influenza B HA. Black dots demonstrate the clinical samples from this study. Yamagata lineage and Victoria lineage are indicated by Y and V letters aside the respective clusters. Bootstrap values above 80% are depicted in the key nodes.

sbane branch had not been vaccinated.Fig. 2 Maximum likelihood phylogenetic tree of influenza B HA. Black dots demonstrate the clinical samples from this study. Yamagata lineage and Victoria lineage are indicated by Y and V letters aside the respective clusters. Bootstrap values above 80% are depicted in the key nodes. Fig. 2 Discussion As the dengue epidemic subsided in the city of Araraquara, we detected fewer febrile episodes in the following years than anticipated for the influenza cohort. However, this fact did not affect the expected number of influenza cases to be analyzed in the study. We observed a high frequency of influenza A and B in children and adolescents with fever and respiratory symptoms. Among the 277 episodes of ILI, 99 (35.7%) were caused by influenza viruses, mostly influenza A (79.9%). The use of “influenza-like illness” as the case definition for respiratory sampling justifies this finding. Similar rates were found in a recently published study that detected 263 virologically confirmed episodes of influenza in 811 ILI episodes (36.7%), the majority also caused by influenza A viruses.12 In three consecutive influenza seasons, other authors detected 48% of influenza in ILI episodes in children and adolescents aged five to 14 years, a similar age range of the present cohort.13 Additionally, the use of robust diagnostic tests in multiplex platforms enhanced the likelihood of influenza diagnosis due to superior sensitivity of these new assays.14

ther authors detected 48% of influenza in ILI episodes in children and adolescents aged five to 14 years, a similar age range of the present cohort.13 Additionally, the use of robust diagnostic tests in multiplex platforms enhanced the likelihood of influenza diagnosis due to superior sensitivity of these new assays.14 It is well established that INF A and INF B co-circulate annually. In the USA, the proportion of circulating INF A and INF B viruses has varied from 65% to 88% and from 12% to 35%, respectively, in the last 10 years.15 Our findings (79.9% of INF A and 20.1% of INF B) reflect co-circulation of influenza A and B viruses in Brazil during the study period. In 2016, the proportion of INF A and B viruses was 69.7% and 30.3%, respectively; 62% and 38% in 2017; and 80.1% and 19.9% in 2018.16, 17, 18 It is difficult to predict which influenza virus will predominate in a given season. In the southern hemisphere spring of 2017, a second wave of ILI related to influenza B infections was observed. Influenza surveillance data from the Brazilian Ministry of Health showed that in 2016 and 2017, influenza B viruses circulated with greater activity in spring months. As early as 2018, this trend did not remain.16, 17, 18 In other countries, this second wave of INF B viruses were also observed in the season.13

bserved. Influenza surveillance data from the Brazilian Ministry of Health showed that in 2016 and 2017, influenza B viruses circulated with greater activity in spring months. As early as 2018, this trend did not remain.16, 17, 18 In other countries, this second wave of INF B viruses were also observed in the season.13 Concerning to the clinical findings, symptoms caused by influenza B virus were remarkably similar to those caused by influenza A virus, and included fever, headache, cough, somnolence, myalgia, abdominal pain, vomiting and conjunctivitis, as observed by other authors.19, 20 We found a higher proportion of males among INF B infections, which also caused more prostration than INF A infections. Few studies have shown that male sex may predispose to respiratory virus infection. A recent observational study has shown a significant association between male sex and adenovirus infection, but not with influenza or other respiratory viruses.21

F B infections, which also caused more prostration than INF A infections. Few studies have shown that male sex may predispose to respiratory virus infection. A recent observational study has shown a significant association between male sex and adenovirus infection, but not with influenza or other respiratory viruses.21 Early beliefs were that influenza B viruses caused less severe disease than influenza A viruses.22 Taking into account the hospitalization rates, morbidity or mortality of influenza viruses over the last decades, INF B infections are now considered less severe than those caused by A/H3N2 viruses, but more severe than A/H1N1 infections.15, 19, 23 Both influenza A and B viruses may cause rare neurological complications such as encephalitis and Reye’s syndrome, the latter more often associated with INF B infections.19 From 2010 to 2018, the frequency of INF B infections associated with pediatric deaths varied from 15% to 37% in the USA.15 In Brazil in 2017, 498 deaths of patients with severe acute respiratory syndrome (SARS) were due to influenza viruses, the vast majority caused by influenza A(H3N2) (277 deaths, 55.6%) and influenza B (154 deaths, 30.9%).17 Fortunately, in the present cohort no patient died or had severe influenza requiring hospitalization. Maybe individuals included in prospective cohort studies are more properly informed about the importance of early report of symptoms and advised to seek medical attention sooner than the general population.

Early beliefs were that influenza B viruses caused less severe disease than influenza A viruses.22 Taking into account the hospitalization rates, morbidity or mortality of influenza viruses over the last decades, INF B infections are now considered less severe than those caused by A/H3N2 viruses, but more severe than A/H1N1 infections.15, 19, 23 Both influenza A and B viruses may cause rare neurological complications such as encephalitis and Reye’s syndrome, the latter more often associated with INF B infections.19 From 2010 to 2018, the frequency of INF B infections associated with pediatric deaths varied from 15% to 37% in the USA.15 In Brazil in 2017, 498 deaths of patients with severe acute respiratory syndrome (SARS) were due to influenza viruses, the vast majority caused by influenza A(H3N2) (277 deaths, 55.6%) and influenza B (154 deaths, 30.9%).17 Fortunately, in the present cohort no patient died or had severe influenza requiring hospitalization. Maybe individuals included in prospective cohort studies are more properly informed about the importance of early report of symptoms and advised to seek medical attention sooner than the general population. Although we did not categorize the burden of ILI episodes, the frequency of visits to pediatricians was 70.1% in the case of influenza infections in comparison with 29.9% in other ILI episodes (p < 0.0001). As much as 57% of outpatient visits have been reported in laboratory-confirmed influenza episodes.12 In the present study, patients with INF A were more frequently referred to the pediatrician (67.1%) than those with INF B (40%). The above-mentioned belief that INF A is more severe than INF B may explain this finding.

As much as 57% of outpatient visits have been reported in laboratory-confirmed influenza episodes.12 In the present study, patients with INF A were more frequently referred to the pediatrician (67.1%) than those with INF B (40%). The above-mentioned belief that INF A is more severe than INF B may explain this finding. We also did not observe any difference in the frequency of influenza infections among vaccinated and non-vaccinated patients (34% vs 40%, p = 0.47). Seasonal influenza vaccination is the most effective way to prevent influenza and its complications.24 However, due to the changing nature of the influenza virus, influenza vaccine effectiveness (IVE) may vary over different influenza seasons and virus subtypes/lineages. A recent study conducted in the Netherlands from 2004 to 2014 showed that the adjusted IVE dropped from 40% when the vaccine (partially) matched the circulating viruses, to 20% in mismatched seasons. The authors also observed that IVE was particularly low when A(H3N2) was the predominant influenza virus subtype.25

ges. A recent study conducted in the Netherlands from 2004 to 2014 showed that the adjusted IVE dropped from 40% when the vaccine (partially) matched the circulating viruses, to 20% in mismatched seasons. The authors also observed that IVE was particularly low when A(H3N2) was the predominant influenza virus subtype.25 Concerning to influenza B lineages, a review of INF B infections conducted in 26 countries showed that a vaccine mismatch occurred in approximately 25% of the seasons.26 In a mismatched season, IVE may be suboptimal against INF B, potentially increasing the burden of the disease.2 In a recently published study, conducted in the state of Paraná, the authors observed that both B/Yam and B/Vic lineages co-circulated from to 2013 to 2016, with a frequency of 47% and 53%, respectively.27 In Brazil, previous studies have demonstrated the mismatch between vaccine and circulating lineages of influenza B.28 We observed a higher proportion of the B/Yamagata lineage circulating in the city of Araraquara during the study period. As this lineage was not included in the TIV provided by the Brazilian Ministry of Health, protection against INF B infections was certainly lower than expected. Among the 10 INF B/Yamagata lineage patients with known vaccination status, four (40%) had received the TIV and six (60%) had not been vaccinated.

tudy period. As this lineage was not included in the TIV provided by the Brazilian Ministry of Health, protection against INF B infections was certainly lower than expected. Among the 10 INF B/Yamagata lineage patients with known vaccination status, four (40%) had received the TIV and six (60%) had not been vaccinated. In the 12 cases of INF B that could be sequenced, one case of INF B/Victoria lineage was detected in a 5-year-old child. In the remaining 11 (median age 10, range six to 12 years) we identified the B/Yamagata lineage. Recent epidemiological studies have demonstrated that viruses of the B/Victoria lineage infect subjects at a relatively younger age than those of the B/Yamagata lineage.29, 30 As both INF B lineages have been circulating since 2001, and it is widely recognized that the QIV vaccine offers better protection than the TIV,19 it sounds paradoxical that to date there is no recommendation of preferential use of QIV over TIV. In conclusion, our study showed that influenza viruses are often detected in children and adolescents during ILI episodes. Morbidity of INF A and B infections was very similar, and therefore it is time to overcome old beliefs that INF A is more severe than INF B. The mismatch between the circulating INF viruses and the TIV offered in public health services in Brazil may also have contributed to the high frequency of influenza A and B in this population.

NF A and B infections was very similar, and therefore it is time to overcome old beliefs that INF A is more severe than INF B. The mismatch between the circulating INF viruses and the TIV offered in public health services in Brazil may also have contributed to the high frequency of influenza A and B in this population. Acknowledgments This study was supported by Sanofi-Aventis Farmacêutica Ltda (Sanofi FLU 55), contract number 104225 (FFM-USP). The authors thank Bárbara B. de Souza Pereira, Marta Inenami, Raphael Luiz de Holanda e Silva, Eliezer Robson, Mariana Cristina Câmara, Fernanda de Jesus Notário dos Santos and Maísa de Fátima Hortenci Corona for laboratory support and field assistance during the study.