Comparative performances of seven quantitative Reverse - Transcription Polymerase Chain Reaction assays (RT-qPCR) for detecting SARS-CoV-2 infection in samples from individuals suspected of COVID-19 in São Paulo, Brazil

Introduction Brazil is the second largest country with COVID-19 positive cases worldwide. Due to the potent spread of the virus and the scarcity of kits and supplies, the Brazilian Ministry of Health has granted authorization for the use of kits available during this emergency, without an accurate evaluation of their performance. This study compared the performance and cost-effectiveness of seven molecular assays/kits available in São Paulo, Brazil, for SARS-CoV-2 diagnosis Materials and methods A total of 205 nasopharyngeal/oropharyngeal samples from suspected cases of COVID-19, were tested using the following assays: (i) GeneFinder COVID-19 plus RealAmp kit; (ii) 2019-nCoV RNA PCR-Fluorescence Probing, Da An Gene Co.; (iii) in-house RT-qPCR SARS-CoV-2 IAL; (iv) 2019-nCoV kit, IDT; (v) molecular SARS-CoV-2 (E) kit, Bio-Manguinhos; (vi) Allplex 2019-nCoV modified Assay, Seegene Inc, and (vii) Biomol one-step COVID-19 kit, IBMP. The criteria for determining a SARS-CoV-2 true positive result included the cycle threshold cut-off values, the characteristics of exponential/linear curves, the gene target diversity, and a positive result in at least two assays Results The overall sensitivity of the assays listed were GeneFinder 83.6%, Da An Gene 100.0%, IAL 90.4%, IDT 94.6%, Bio-Manguinhos 87.7%, Allplex 97.3%, and IBMP 87.7%. The minor sensitive gene target was RdRP. Although all assays had a Cohen's Kappa index ≥0.893, the best tests used multiplex assays identifying N-gene and/or E-gene targets Conclusion All assays tested accurate for diagnosis, but considering cost-effectiveness (cost, time consumption, number of samples tested, and performance), the in-house IAL assay was ideal for COVID-19 diagnosis in São Paulo, Brazil.

Loop-mediated isothermal amplification (LAMP) assay for rapid and accurate confirmatory diagnosis of HTLV-1/2 infection

Laboratory diagnosis of human T-lymphotropic viruses (HTLV) 1 and 2 infection is performed by serological screening and further confirmation with serological or molecular assays. Thus, we developed a loop-mediated isothermal nucleic acid amplification (LAMP) assay for the detection of HTLV-1/2 in blood samples. The sensitivity and accuracy of HTLV-1/2 LAMP were defined with DNA samples from individuals infected with HTLV-1 (n = 125), HTLV-2 (n = 19), and coinfected with HIV (n = 82), and compared with real-time polymerase chain reaction (qPCR) and PCR-restriction fragment length polymorphism (RFLP). The overall accuracy of HTLV-1/2 LAMP (95% CI 74.8­85.5%) was slightly superior to qPCR (95% CI 69.5­81.1%) and similar to PCR-RFLP (95% CI 79.5­89.3%). The sensitivity of LAMP was greater for HTLV-1 (95% CI 83.2­93.4%) than for HTLV-2 (95% CI 43.2­70.8%). This was also observed in qPCR and PCR-RFLP, which was associated with the commonly lower HTLV-2 proviral load. All molecular assays tested showed better results with samples from HTLV-1/2 mono-infected individuals compared with HIV-coinfected patients, who present lower CD4 T-cell counts. In conclusion, HTLV-1/2 LAMP had similar to superior performance than PCR-based assays, and therefore may represent an attractive alternative for HTLV-1/2 diagnosis due to reduced working time and costs, and the simple infrastructure needed.

A cross-sectional study assessing the pharyngeal carriage of Neisseria meningitidis in subjects aged 1-24 years in the city of Embu das Artes, São Paulo, Brazil

ABSTRACT Meningococcal carriage is a prerequisite for invasive infection. This cross-sectional study assessed the pharyngeal carriage prevalence in healthy subjects aged 1-24 years in Embu das Artes city, São Paulo, Brazil. Pharyngeal swabs were examined for the presence of Neisseria meningitidis. The isolates were tested for different serogroups using agglutination and polymerase chain reaction. A logistic regression model assessed any independent association between Neisseria meningitidis carriage and various risk factors. A total of 87/967 subjects (9%, 95% Confidence Interval (CI): 7.3-11.0) tested positive for N. meningitidis: 6.2% (95% CI: 3.8-9.4) in 1-4 years, 8.5% (95% CI: 5.1-13.0) in 5-9 years, 12.5% (95% CI: 7.8-18.6) in 10-14 years, 12.6% (95% CI: 7.4-19.7) in 15-19 years and 9% (95% CI: 4.9-14.9) in 20-24 years age groups. Highest carriage prevalence was observed in adolescents 10-19 years old. Serogroup C was predominant (18.4%) followed by serogroup B (12.6%). The 15-19 years age group showed a significant association between number of household members and carriers of N. meningitidis. This cross-sectional study is the first in Brazil to evaluate meningococcal carriage prevalence and associated factors in a wide age range.

Comparative performances of serologic and molecular assays for detecting human T lymphotropic virus type 1 and type 2 (HTLV-1 and HTLV-2) in patients infected with human immunodeficiency virus type 1 (HIV-1)

ABSTRACT The present study evaluated several techniques currently available (commercial kits and in-house assays) for diagnosing human T lymphotropic viruses types 1 and 2 in two groups of patients enrolled at HIV/AIDS specialized care services in São Paulo: Group 1 (G1), n = 1608, 1237 male/371 female, median age 44.3 years old, majority using highly active antiretroviral therapy (HAART); G2, n = 1383, 930 male/453 female, median age of 35.6 years old, majority HAART naïve. Enzyme immunoassays [(EIA) Murex and Gold ELISA] were employed for human T lymphotropic viruses types 1 and 2 screening; Western blotting (WB), INNO-LIA (LIA), real-time PCR pol (qPCR), and nested-PCR-RFLP (tax) were used to confirm infection. Samples were considered human T lymphotropic viruses types 1 and 2 positive when there was reactivity using at least one of the four confirmatory assays. By serological screening, 127/2991 samples were positive or borderline, and human T lymphotropic virus infection was confirmed in 108 samples (three EIA-borderline): 56 human T lymphotropic virus type 1 [G1 (27) + G2 (29)]; 45 human T lymphotropic virus type 2 [G1 (21) + G2 (24)]; one human T lymphotropic virus type 1 + human T lymphotropic virus type 2 (G2); six human T lymphotropic virus [G1 (2) + G2 (4)]. Although there were differences in group characteristics, human T lymphotropic viruses types 1 and 2 prevalence was similar [3.1% (G1) and 4.2% (G2), p = 0.113]. The overall sensitivities of LIA, WB, qPCR, and PCR-RFLP were 97.2%, 82.4%, 68.9%, and 68.4%, respectively, with some differences among groups, likely due to the stage of human T lymphotropic virus infection and/or HAART duration. Indeterminate immunoblotting results were detected in G2, possibly due to the seroconversion period. Negative results in molecular assays could be explained by the use of HAART, the occurrence of defective provirus and/or the low circulating proviral load. In conclusion, when determining the human T lymphotropic virus infection, the findings highlight that there is a need to consider the blood samples with borderline results in screening assays. Of all the tested assays, LIA was the assay of choice for detecting human T lymphotropic virus type 1 and human T lymphotropic virus type 2 in human immunodeficiency virus type 1-infected patients.

Comparative performances of serologic and molecular assays for detecting human T lymphotropic virus type 1 and type 2 (HTLV-1 and HTLV-2) in patients infected with human immunodeficiency virus type 1 (HIV-1)

The present study evaluated several techniques currently available (commercial kits and in-house assays) for diagnosing human T lymphotropic viruses types 1 and 2 in two groups of patients enrolled at HIV/AIDS specialized care services in São Paulo: Group 1 (G1), n = 1608, 1237 male/371 female, median age 44.3 years old, majority using highly active antiretroviral therapy (HAART); G2, n = 1383, 930 male/453 female, median age of 35.6 years old, majority HAART naïve. Enzyme immunoassays [(EIA) Murex and Gold ELISA] were employed for human T lymphotropic viruses types 1 and 2 screening; Western blotting (WB), INNO-LIA (LIA), real-time PCR pol (qPCR), and nested-PCR-RFLP (tax) were used to confirm infection. Samples were considered human T lymphotropic viruses types 1 and 2 positive when there was reactivity using at least one of the four confirmatory assays. By serological screening, 127/2991 samples were positive or borderline, and human T lymphotropic virus infection was confirmed in 108 samples (three EIA-borderline): 56 human T lymphotropic virus type 1 [G1 (27) + G2 (29)]; 45 human T lymphotropic virus type 2 [G1 (21) + G2 (24)]; one human T lymphotropic virus type 1 + human T lymphotropic virus type 2 (G2); six human T lymphotropic virus [G1 (2) + G2 (4)]. Although there were differences in group characteristics, human T lymphotropic viruses types 1 and 2 prevalence was similar [3.1% (G1) and 4.2% (G2), p = 0.113]. The overall sensitivities of LIA, WB, qPCR, and PCR-RFLP were 97.2%, 82.4%, 68.9%, and 68.4%, respectively, with some differences among groups, likely due to the stage of human T lymphotropic virus infection and/or HAART duration. Indeterminate immunoblotting results were detected in G2, possibly due to the seroconversion period. Negative results in molecular assays could be explained by the use of HAART, the occurrence of defective provirus and/or the low circulating proviral load. In conclusion, when determining the human T lymphotropic virus infection, the findings highlight that there is a need to consider the blood samples with borderline results in screening assays. Of all the tested assays, LIA was the assay of choice for detecting human T lymphotropic virus type 1 and human T lymphotropic virus type 2 in human immunodeficiency virus type 1-infected patients. (AU)

Short communication: failures in detecting HTLV-1 and HTLV-2 in patients infected with HIV-1

Changes in retrovirus acquisition/transmission behaviors have been reported in Brazil, with a concerning increase in HIV-1-infected individuals aged 15-39 years. In São Paulo, HIV-1/HTLV-1 and HIV-1/HTLV-2 coinfections have been associated with intravenous drug use and failure to detect HTLV-1/2 (human T cell lymphotropic virus types 1 and 2) with immunosuppression and the use of highly active antiretroviral therapy (HAART). Negative results for HTLV serologic [western blotting (WB)] and molecular [real-time PCR pol (qPCR)] confirmatory assays have been reported, whereas the best sensitivity has been found for INNO-LIA (LIA). In this study, we expand our previous data by analyzing a group of young patients (n = 1,383; median age 35.6 years) who recently acquired HIV by sexual contact, the majority of whom were HAART naïve, and comparing the performances of four HTLV confirmatory assays LIA, WB, qPCR, and PCR-RFLP (tax). We confirmed HTLV infection in 58 (4.2%) blood samples 29 HTLV-1, 24 HTLV-2, 1 HTLV-1+HTLV-2, and 4 HTLV. LIA, WB, qPCR, and PCR-RFLP sensitivities were 94.8%, 82.8%, 79.2%, and 74.5%, respectively. Associations of HTLV infection with female gender (OR = 2.28, 1.31-4.00) and age >40 years (p < .0001) were detected. The results confirm the low sensitivities of molecular assays and the best performance of LIA in detecting HTLV-1/2 in such patients. We hypothesize that the negative PCR results are due to the presence of defective provirus and/or low proviral load circulating in such patients, with inconclusive WB coinciding with the seroconversion period. Corroborating the associations obtained, repeated exposure is required for HTLV sexual transmission/acquisition, which is more efficient from male to female

Performance of an in-house real-time polymerase chain reaction for identification of Mycobacterium tuberculosis isolates in laboratory routine diagnosis from a high burden setting

Abstract Brazil is one of the high burden countries for tuberculosis, and a rapid diagnosis is essential for effective control of the disease. In the present study, an in-house real-time polymerase chain reaction (PCR) assay targeting the mpt64 gene for identification of Mycobacterium tuberculosis complex isolates was evaluated under routine diagnosis conditions in a reference laboratory. From May 2011 to July 2012, 1,520 isolates of mycobacteria were prospectively submitted for phenotypic and/or PRA-hsp65 identification and to real-time PCR. The mpt64 real-time PCR showed 99.7% sensitivity and 96% specificity and detected 79.4% of the cases missed by phenotypic and PRA-hsp65 identification. The in-house real-time PCR assay showed high sensitivity and specificity and was successfully implemented in the routine diagnosis of tuberculosis in a reference laboratory from a high burden setting.

Comparação de testes laboratoriais para o diagnóstico de infecção por vírus linfotrópicos de células T humanas do tipo 1 (HTLV-1)e tipo 2 (HTLV-2) em pacientes infectados por HIV-1 / Comparison of laboratorial tests for the diagnosis of human T-lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) in HIV-1 infected patients

O presente estudo pesquisou o melhor algoritmo de testes laboratoriais para efetuar o diagnóstico de infecção por vírus linfotrópicos de células T humanas dos tipos 1 (HTLV-1) e 2 (HTLV-2) em pacientes HIV-1 positivos. Amostras de sangue de 1.608 pacientes do CRT DST/Aids-SP foram analisadas quanto à presença de anticorpos específicos usando-se dois ensaios de triagem (EIA Murex HTLV-I+II e Gold ELISA HTLV-I/II), dois confirmatórios [HTLV Blot 2.4 (Western Blot – WB) e INNO-LIA HTLV I/II (Line ImmunoAssay - LIA)] e um molecular (PCR em tempo real pol). Na triagem foram detectados 51(Murex) e 49 (Gold ELISA) soros reagentes. Pelo WB, 23 soros confirmaram infecção por HTLV-1, 12 HTLV-2, seis HTLV e nove apresentaram perfis indeterminados. O LIA detectou 24 soros HTLV-1 positivos, 20 HTLV-2 e seis HTLV. A PCR evidenciou segmento pol de HTLV-1 em 18 e HTLV-2 em 12 amostras de sangue. Pelos testes confirmatórios, em 50 pacientes foi confirmada a infecção por HTLV: 25 HTLV-1 (1,55 %), 21 HTLV-2 (1,31 %) e quatro HTLV (0,25 %). As sensibilidades do LIA, WB e PCR foram de 96 %, 76 % e 60 %, respectivamente. Considerando-se apenas o custo, o melhor algoritmo diagnóstico para população infectada pelo HIV-1 foi o uso da PCR seguida do LIA...

Evolution of bacterial meningitis diagnosis in Sao Paulo State-Brazil and future challenges / Evolucao do diagnostico das meningites bacterianas no Estado de Sao Paulo-Brasil e os futuros desafios

Bacterial meningitis (BM) is a severe disease and still represents a serious public health problem with high rates of morbidity and mortality. The most common cases of BM around the world, mainly in Brazil, have been caused by Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae type b. Bacterial culture is the gold-standard technique for BM confirmation, but approximately 50% of suspected cases are not culture-confirmed, due to problems related to improper transportation and seeding or previous antibiotic treatment. Immunological methods present low sensitivity and have possibility of cross-reactions. Real time PCR (qPCR) is a molecular technique and has been successful used for BM diagnosis at Instituto Adolfo Lutz in São Paulo State, Brazil, since 2007. The incorporation of qPCR in the Public Health surveillance routine in our state resulted in diminishing 50% of undetermined BM cases. Our efforts are focused on qPCR implementation in the BM diagnostic routine throughout Brazil.

A meningite bacteriana (MB) é uma doença grave e ainda representa um sério problema de saúde pública, com altas taxas de morbidade e mortalidade. Os casos mais comuns de MB em todo o mundo, principalmente no Brasil, tem sido causados por Neisseria meningitidis, Streptococcus pneumoniae e Haemophilus influenzae tipo b. Cultura bacteriana é a técnica padrão-ouro para a confirmação de MB, mas cerca de 50% dos casos suspeitos não são confirmados por cultura, devido a problemas relacionados ao transporte inadequado e semeadura ou antibioticoterapia prévia. Métodos imunológicos apresentam baixa sensibilidade e têm possibilidade de reações cruzadas. PCR em tempo real (qPCR) é uma técnica molecular e tem sido utilizada com êxito para o diagnóstico de MB no Instituto Adolfo Lutz, em São Paulo, Brasil, desde 2007. A incorporação da qPCR na rotina de vigilância em Saúde Pública em nosso estado resultou na diminuição de 50% dos casos de MB indeterminadas. Nossos esforços estão focados na implementação da qPCR na rotina diagnóstica de MB em todo o Brasil.

Ocorrência de resultados falso-negativos na reação de PCR em Tempo Real (PCR-TR) no diagnóstico da doença meningocócica / False-negative results in Real-Time PCR (RT-PCR) formeningococcal disease diagnosis

Factors responsible for false-negative results (F-N) in RT-PCR assay for detecting N. meningitidis in serumand CSF samples were investigated. As the meningococcal disease should be rapidly treated because ofits high mortality and epidemic potential, the F-N in diagnostic testing may cause treatment failuresand/or on disease restraint in community. Thus, it is crucial to learn the factors which cause F-N in RTPCRassays. The F-N were induced by inhibition, low quantity of target DNA in extracted samples, andinadequate temperature employed at PCR annealing procedure. As bacterial DNA concentration in samplesmight be highly variable, the ideal sample volume to be extracted could not be defined. As previouslyrecommended for N. meningitidis gene-grouping by RT-PCR assay, the annealing temperature at 60 °Cwas not suitable for B and W135 genogroups. Altogether, these factors induced F-N in 31 samples; therefore,30 % of N. meningitidis detected by RT-PCR were classified as non-genogrouped. The inhibitors and/orthe low amount of target DNA induced F-N on RT-PCR, independently of the specimen volume used forextracting DNA. However, adjustments on the PCR annealing temperature and amount of extracted DNAadded into the reaction might avoid the occurrence of the majority of F-N.

Avaliação de resultados discrepantes obtidos na execução de PCR em tempo real em amostras de pacientes com suspeita clínica de meningite bacteriana / Inconsistent results in real-time PCR in analyzing samples from patients with suspected bacterial meningitis

O princípio básico para obter resultado confiável é a compatibilidade entre as réplicas e suareprodutibilidade. Na rotina diagnóstica por PCR em tempo real (PCR-TR), em que centenas de amostrassão processadas, a obtenção de resultados com Cts tardios ou réplicas que diferem entre si por mais de trêsunidades, são inevitáveis. Das 3.000 amostras processadas em 2010, em duplicata, na rotina diagnósticadas meningites bacterianas por PCR-TR na pesquisa de N. meningitidis, S. pneumoniae e H. influenzae,157 (5,2 %), apresentaram inconsistência entre as réplicas (diferença entre Cts maior do que 3) e/ou altosvalores de Cts; e os ensaios foram retestados. O presente trabalho investigou estes resultados obtidos, osbenefícios destas repetições e as possíveis razões da ocorrência dos resultados discrepantes. Verificouseque, apenas 18 (11 %) das amostras submetidas à repetição, apresentaram resultados positivos. Erroshumanos inerentes à pipetagem, como o uso de pipetas não calibradas, a baixa concentração de DNAalvo nas amostras, a degradação da sonda ou mesmo a possível contaminação aleatória são fatores quecontribuem para induzir resultados discrepantes. A realização do ensaio de PCR-TR com amostras emduplicata e a repetição de ensaios com resultados discordantes é um artifício eficiente para avaliar e definirestes resultados.

Fast test for assessing the susceptibility of Mycobacterium tuberculosis to isoniazid and rifampin by real-time PCR

Mycobacterium tuberculosis is the bacterium that causes tuberculosis (TB), a leading cause of death from infectious disease worldwide. Rapid diagnosis of resistant strains is important for the control of TB. Real-time polymerase chain reaction (RT-PCR) assays may detect all of the mutations that occur in the M. tuberculosis 81-bp core region of the rpoB gene, which is responsible for resistance to rifampin (RIF) and codon 315 of the katG gene and the inhA ribosomal binding site, which are responsible for isoniazid (INH). The goal of this study was to assess the performance of RT-PCR compared to traditional culture-based methods for determining the drug susceptibility of M. tuberculosis. BACTEC TM MGIT TM 960 was used as the gold standard method for phenotypic drug susceptibility testing. Susceptibilities to INH and RIF were also determined by genotyping of katG, inhA and rpoB genes. RT-PCR based on molecular beacons probes was used to detect specific point mutations associated with resistance. The sensitivities of RT-PCR in detecting INH resistance using katG and inhA targets individually were 55% and 25%, respectively and 73% when combined. The sensitivity of the RT-PCR assay in detecting RIF resistance was 99%. The median time to complete the RT-PCR assay was three-four hours. The specificities for tests were both 100%. Our results confirm that RT-PCR can detect INH and RIF resistance in less than four hours with high sensitivity.

Perfil mutacional dos genes responsáveis pela resistência de Mycobacterium tuberculosis a rifampicina (rpoB) e isoniazida (katG e inhA) em cepas isoladas no Estado de São Paulo e avaliação do uso da PCR em tempo real para a detecção rápida destas mutações / Mutational profile of genes responsible for Mycobacterium tuberculosis resistance to rifampicin (rpoB) and isoniazid (katG and inhA) in strains isolated from the São Paulo State and evaluation the use of real-time PCR for rapid detection

INTRODUÇÃO. A detecção rápida de isolados resistentes contribui para prevenir a transmissão e orientar na escolha inicial das drogas para o tratamento da tuberculose. A PCR em tempo real (RT-PCR) detecta todas as mutações do Mycobacterium tuberculosis (MTB) que ocorrem na região de 81pb do gene rpoB, códon 315 do katG e sítio de ligação ribossomal inhA, responsáveis pela resistência à rifampicina (RIF) e isoniazida (INH), respectivamente. OBJETIVO. O objetivo deste estudo foi avaliar a sensibilidade, especificidade e rapidez do método de RT-PCR em determinar a susceptibilidade de isolados de MTB à RIF e INH em comparação ao método tradicional de susceptibilidade às drogas. MATERIAL E MÉTODOS. Foram analisados 988 isolados de MTB do Estado de São Paulo, o método BACTEC™MGIT™ 960 foi utilizado como padrão de referência para os testes de susceptibilidade às drogas. Na RT-PCR a susceptibilidade à INH e/ou RIF foi determinada pelo padrão de reatividade de diferentes sondas para regiões específicas dos genes katG e inhA e rpoB, respectivamente. O sequenciamento dos fragmentos dos genes amplificados foi utilizado como contraprova. RESULTADOS. A sensibilidade da RT-PCR na detecção de resistência à INH pelos genes katG e inhA, individualmente, foram de 55% e 25%, respectivamente e 73% quando combinadas; e para a RIF foi de 99%. A especificidade para ambos os testes foi de 100%. Todas as mutações no gene katG foram no códon 315 e 87% das mutações no sítio de ligação ribossomal inhA foram na posição -15. A maioria das mutações no gene rpoB ocorreram nos códons 531 (68%) seguidos pelo códon 526 (23%). CONCLUSÃO. Nossos resultados confirmam que a RT-PCR pode detectar a resistência à RIF e INH em menos de 4 horas com alta sensibilidade. O perfil mutacional dos isolados analisados foi similar aos descritos na literatura.

Implantação e otimização da PCR em tempo real para o diagnóstico da influenza A (H1N1) pandêmica no Instituto Adolfo Lutz e perspectivas para 2010 / Implementation and optimization of real time PCR for diagnosis of pandemic influenza A (H1N1) at Instituto Adolfo Lutz and perspectives for 2010

Os autores apresentam sua experiência no diagnóstico laboratorial de InfluenzaA (H1N1) em 37.240 amostras clínicas obtidas de pacientes com suspeita de gripe, encaminhadas ao Instituto Adolfo Lutz para análise. Eles apresentam os algoritmos de testes moleculares empregados, comparam a eficiência dos mesmos quanto à sensibilidade, especificidade e custo e, finalmente sugerem um novo algoritmo para ser usado em caso de nova epidemia de Influenza A (H1N1) em 2010.

The authors present their experience with the molecular diagnosis of Influenza A (H1N1) with 37.240 clinicalsamples obtained from individuals suspected of flu, sent to Instituto Adolfo Lutz for analysis. They show the used algorithms, compare their efficiency in terms of sensitivity, specificity and cost, and suggest a new algorithm to be employed in case of an outbreak of Influenza A (H1N1) in 2010.

Occurrence of influenza B/Hong Kong-Like strains in Brazil, during 2002

Through the influenza virus surveillance from January to October 2002, influenza B/Hong Kong-like strains circulating in the Southeast and Centre East regions of Brazil have been demonstrated. This strain is a variant from B/Victoria/02/88 whose since 1991 and until recently have been isolated relatively infrequently and have been limited to South-Eastern Asia. A total of 510 respiratory secretions were collected from patients 0 to 60 years of age, with acute respiratory illness, living in the Southeast and Centre East regions of Brazil, of which 86 (17.13 percent) were positive for influenza virus. Among them 12 (13.95 percent) were characterized as B/Hong Kong/330/2001; 3 (3.49 percent) as B/Hong Kong/1351/2002 a variant from B/Hong Kong/330/2001; 1 (1.16 percent) as B/Sichuan/379/99; 1 (1.16 percent) as B/Shizuoka/5/2001, until now. The percentages of cases notified during the surveillance period were 34.88 percent, 15.12 percent, 15.12 percent, 4.65 percent, 15.12 percent, 13.95 percent, in the age groups of 0-4, 5-10, 11-15, 16-20, 21-30, 31-50, respectively. The highest proportion of isolates was observed among children younger than 4 years but serious morbidity and mortality has not been observed among people older than 65 years, although B influenza virus component for vaccination campaign 2002 was B/Sichuan/379/99 strain. This was probably due to the elderly protection acquired against B/Victoria/02/88. In addition, in influenza A/Panama/2007/99-like (H3N2) strains 22 (25.58 percent) were also detected, but influenza A(H1N1) has not been detected yet

Outbreak of influenza type A (H1N1) in Iporanga, Säo Paulo State, Brazil

From June to July 1999 an outbreak of acute respiratory illness occurred in the town of Iporanga. Out of a total of 4,837 inhabitants, 324 cases were notified to the Regional Surveillance Service. Influenza virus was isolated from 57.1 percent of the collected samples and 100 percent seroconversion to influenza A (H1N1) was obtained in 20 paired sera tested. The isolates were related to the A/Bayern/07/95 strain (H1N1). The percentages of cases notified during the outbreak were 28.4 percent, 29.0 percent, 20.7 percent, 6.2 percent and 15.7 percent in the age groups of 0-4, 5-9, 10-14, 15-19 and older than 20 years, respectively. The highest proportion of positives was observed among children younger than 14 years and no cases were notified in people older than 65 years, none of whom had been recently vaccinated against influenza. These findings suggest a significant vaccine protection against A/Bayern/7/95, the H1 component included in the 1997-98 influenza vaccine for elderly people. This viral strain is antigenically and genetically related to A/Beijing/262/95, the H1 component of the 1999 vaccine. Vaccines containing A/Beijing/262/95 (H1N1) stimulated post-immunization hemagglutination inhibition antibodies equivalent in frequency and titre to both A/Beijing/262/95-like and A/Bayern/7/95-like viruses. Thus, this investigation demonstrates the effectiveness of vaccination against influenza virus in the elderly