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O Instituto Adolfo Lutz (IAL) foi criado em 1940 como resultado da unificação dos Institutos Bacteriológico e Bromatológico, um moderno laboratório voltado ao controle de doenças, inaugurando uma nova fase de laboratórios de saúde pública no estado de São Paulo. Os primeiros testes sorológicos oferecidos à população foram executados pelas "antigas" Seções de Sorologia e de Imunologia. Essas seções destacam-se no desenvolvimento científico do IAL pela realização de pesquisas, produção científica e inovação tecnológica, seguramente, fundamentais para a saúde pública no decorrer dos anos. O Centro de Imunologia do IAL (CIM-IAL) foi criado em 2010, com a unificação das Seções de Sorologia e Imunologia, quando ocorreu a reorganização institucional. O CIM-IAL contribuiu para importantes avanços científicos na área da saúde, reforçando sua capacidade de desenvolver pesquisas, executar e monitorar o diagnóstico e a vigilância de diferentes agravos. Este manuscrito tem como objetivo apresentar os principais acontecimentos que ressaltam o papel fundamental na busca de soluções para os problemas de saúde pública, desde a época das Seções de Sorologia e Imunologia até tornar-se o Centro de Imunologia. Na elaboração deste trabalho foram utilizadas bibliografias contendo dados históricos, científicos e relatos de profissionais da área.

A new phase of Public Health Laboratories in the state of São Paulo occurred in 1940, with the unification of Instituto Bacteriológico and Bromatológico, creating the Instituto Adolfo Lutz (IAL), a modern laboratory focused on solving problems in this area. The first diagnostic tests offered to the population were carried out by the "old" Serology and Immunology Sections. It's worth highlighting the importance of these sections in the scientific development of the IAL by carrying out research, scientific production and technological innovation, which have certainly been fundamental to public health over the years. The Immunology Center (CIM) of IAL was created in 2010, when organizational adaptation took place with the junction of the Serology and Immunology Sections. The CIM-IAL has undergone important advances in the health area, reinforcing its capacity to develop research, carry out and monitor the diagnosis and surveillance of different diseases. This manuscript aims to present the main events that highlight the fundamental role in the search for solutions to public health problems, from the time of the Serology and Immunology Sections until it became the CIM. In the preparation, bibliographies were used based on historical and scientific data and reports from professionals in the field.

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Introduction. Brazil was one of the most affected countries by the COVID-19 pandemic. Instituto Adolfo Lutz (IAL) is the reference laboratory for COVID-19 in São Paulo, the most populous state in Brazil. In April 2020, a secondary diagnostic pole named IAL-2 was created to enhance IAL's capacity for COVID-19 diagnosis.Hypothesis/Gap Statement. Public health laboratories must be prepared to rapidly respond to emerging epidemics or pandemics.Aim. To describe the design of IAL-2 and correlate the results of RT-qPCR tests for COVID-19 with secondary data on suspected cases of SARS-CoV-2 infection in the São Paulo state.Methodology. This is a retrospective study based on the analysis of secondary data from patients suspected of infection by SARS-CoV-2 whose clinical samples were submitted to real-time PCR after reverse transcription (RT-qPCR) at IAL-2, between 1 April 2020 and 8 March 2022. RT-qPCR Ct results of the different kits used were also analysed.Results. IAL-2 was implemented in April 2020, just over a month after the detection of the first COVID-19 case in Brazil. The laboratory performed 304,250 RT-qPCR tests during the study period, of which 98 319 (32.3 %) were positive, 205827 (67.7 %) negative, and 104 (0.03 %) inconclusive for SARS-CoV-2. RT-qPCR Ct values≤30 for E/N genes of SARS-CoV-2 were presented by 79.7 % of all the samples included in the study.Conclusion. IAL was able to rapidly implement a new laboratory structure to support the processing of an enormous number of samples for diagnosis of COVID-19, outlining strategies to carry out work with quality, using different RT-qPCR protocols.

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Brazil currently has the highest number of individuals infected with human T-lymphotropic virus 1- and 2- (HTLV-1 and HTLV-2) globally. At present, neither molecular protocols nor commercial assays are available for HTLV-1/-2 diagnosis or validated by the Brazilian Ministry of Health regulatory agency (ANVISA). We developed and validated two in-house multiplex quantitative real-time PCR for HTLV-1/-2 (mqPCR_HTLV) assays, targeting the pol and tax genes, for the simultaneous identification of HTLV-1, HTLV-2, and the albumin reference gene. The robustness of the assays was evaluated on two platforms using seven commercial master mix formulations. The reactions employed double plasmids (pHTLV1-Alb and pHTLV2-Alb) for the standard curve's construction and for expressing the detection limit of the assays. They were able to detect 10 and 10 copies of HTLV-1 and 10 and 70 copies of HTLV-2 for the tax and pol targets, respectively. High efficiency was obtained using both the platforms and all the reagents evaluated and were successfully reproduced by other analysts. DNA samples from HTLV-1/-2-infected and non-infected patients and from HIV/HTLV-coinfected patients were evaluated to determine the feasibility of their use in routine diagnosis. The mqPCR_HTLV (pol and tax) assays demonstrated an overall specificity of 100% and a sensitivity of 97.4% when testing samples from patients without HIV infection, and sensitivities of 77.1% (pol) and 74.6% (tax) in samples from HIV/HTLV-coinfected patients. In addition, they resolved the issue of HTLV western blotting (WB) indeterminate and WB-untyped results in 45.5 and 66.7% of cases, respectively. The developed mqPCR_HTLV (pol and tax) assays indicated their feasibility for efficient and reliable HTLV diagnosis in various core facility laboratories under different conditions and supplies.

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Brazil currently has the highest number of individuals infected with human T-lymphotropic virus 1- and 2- (HTLV-1 and HTLV-2) globally. At present, neither molecular protocols nor commercial assays are available for HTLV-1/-2 diagnosis or validated by the Brazilian Ministry of Health regulatory agency (ANVISA). We developed and validated two in-house multiplex quantitative real-time PCR for HTLV1/-2 (mqPCR_HTLV) assays, targeting the pol and tax genes, for the simultaneous identification of HTLV-1, HTLV-2, and the albumin reference gene. The robustness of the assays was evaluated on two platforms using seven commercial master mix formulations. The reactions employed double plasmids (pHTLV1-Alb and pHTLV2-Alb) for the standard curve's construction and for expressing the detection limit of the assays. They were able to detect 10 and 10 copies of HTLV-1 and 10 and 70 copies of HTLV-2 for the tax and pol targets, respectively. High efficiency was obtained using both the platforms and all the reagents evaluated and were successfully reproduced by other analysts. DNA samples from HTLV-1/-2-infected and non-infected patients and from HIV/HTLV-coinfected patients were evaluated to determine the feasibility of their use in routine diagnosis. The mqPCR_HTLV (pol and tax) assays demonstrated an overall specificity of 100% and a sensitivity of 97.4% when testing samples from patients without HIV infection, and sensitivities of 77.1% (pol) and 74.6% (tax) in samples from HIV/HTLV-coinfected patients. In addition, they resolved the issue of HTLV western blotting (WB) indeterminate and WB-untyped results in 45.5 and 66.7% of cases, respectively. The developed mqPCR_HTLV (pol and tax) assays indicated their feasibility for efficient and reliable HTLV diagnosis in various core facility laboratories under different conditions and supplies. (AU)

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Introduction. Invasive meningococcal disease is a major health problem, impacting morbidity and mortality worldwide. Exploratory genomics has revealed insights into adaptation, transmissibility and virulence to elucidate endemic, outbreaks or epidemics caused by Neisseria meningitidis serogroup W (MenW) strains.Gap Statement. Limited information on the genomics of Neisseria meningitis serogroup W ST11/cc11 is available from emerging countries, especially in contemporary isolates.Aim. To (i) describe the antigenic diversity and distribution of genetic lineages of N. meningitidis serogroup W circulating in Brazil; (ii) study the carriage prevalence of hypervirulent clones in adolescents students and (iii) analyse the potential risk factors for meningococcal carriage.Methodology. Using whole-genome sequencing, we analysed the genomic diversity of 92 invasive N. meningitidis serogroup W isolates circulating in Brazil from 2016 to 2019. A cross-sectional survey of meningococcal carriage was conducted in 2019, in the city of Florianópolis, Brazil, among a representative sample of 538 students.Results. A predominance (58.5 %, 41/82) of ST11/cc11 presenting PorB2-144, PorA VR1-5, VR2-2, FetA 1-1, and a novel fHbp peptide 1241 was found on invasive N. meningitidis W isolates, on the other hand, a high diversity of clonal complexes was found among carriage isolates. The overall carriage rate was 7.5 % (40/538). A total of 28 of 538 swab samples collected were culture positive for N. meningitidis, including four serogroup/genogroup B isolates (14.8 %;4/27), 1 serogroup/genogroup Y isolate (3.7 %;1/27), 22 (81.5 %; 22/27) non-groupable isolates. No MenW isolate was identified among carriages isolates.Conclusion. This report describes the emergence of the new MenW ST11/cc11 South America sublineage variant, named here, 2016 strain, carrying a novel fHbp peptide 1241, but its emergence, was not associated with an increased MenW carriage prevalence. Continuous surveillance is necessary to ascertain the role of this sublineage diversification and how its emergence can impact transmission.

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Introduction. Invasive meningococcal disease is a major health problem, impacting morbidity and mortality worldwide. Exploratory genomics has revealed insights into adaptation, transmissibility and virulence to elucidate endemic, outbreaks or epidemics caused by Neisseria meningitidis serogroup W (MenW) strains.Gap Statement. Limited information on the genomics of Neisseria meningitis serogroup W ST11/cc11 is available from emerging countries, especially in contemporary isolates.Aim. To (i) describe the antigenic diversity and distribution of genetic lineages of N. meningitidis serogroup W circulating in Brazil; (ii) study the carriage prevalence of hypervirulent clones in adolescents students and (iii) analyse the potential risk factors for meningococcal carriage.Methodology. Using whole-genome sequencing, we analysed the genomic diversity of 92 invasive N. meningitidis serogroup W isolates circulating in Brazil from 2016 to 2019. A cross-sectional survey of meningococcal carriage was conducted in 2019, in the city of Florianópolis, Brazil, among a representative sample of 538 students.Results. A predominance (58.5 %, 41/82) of ST11/cc11 presenting PorB2-144, PorA VR1-5, VR2-2, FetA 1-1, and a novel fHbp peptide 1241 was found on invasive N. meningitidis W isolates, on the other hand, a high diversity of clonal complexes was found among carriage isolates. The overall carriage rate was 7.5 % (40/538). A total of 28 of 538 swab samples collected were culture positive for N. meningitidis, including four serogroup/genogroup B isolates (14.8 %;4/27), 1 serogroup/genogroup Y isolate (3.7 %;1/27), 22 (81.5 %; 22/27) non-groupable isolates. No MenW isolate was identified among carriages isolates.Conclusion. This report describes the emergence of the new MenW ST11/cc11 South America sublineage variant, named here, 2016 strain, carrying a novel fHbp peptide 1241, but its emergence, was not associated with an increased MenW carriage prevalence. Continuous surveillance is necessary to ascertain the role of this sublineage diversification and how its emergence can impact transmission.

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Haemophilus influenzae (Hi) é um importante patógeno causador de meningites (MB) e pneumonias bacterianas (PB), principalmente em países onde a imunoprevenção é precária ou inexistente. O Hi é classificado em tipáveis (sorotipos a, b, c, d, e, f) e não tipáveis (HiNt), de acordo com a presença ou ausência da cápsula polissacarídica, respectivamente. A cápsula é o principal fator de virulência dos Hi e o gene bexA, responsável pela sua expressão, é comumente empregado na detecção molecular e vigilância das MB e PB causadas por Hi. Em 2010, o Instituto Adolfo Lutz (IAL) implantou a PCR em tempo real (qPCR) empregando esse alvo genético para a detecção de Hi. Entretanto, relatos de falha na detecção de alguns Hi encapsulados e HiNt motivaram a substituição do gene alvo para essa bactéria. Desta forma, em agosto de 2012, o IAL fez a substituição do bexA pelo alvo genético hpd no ensaio de qPCR, permitindo a detecção de Hi tipáveis e não tipáveis. Neste estudo, avaliamos o impacto da substituição do alvo genético na vigilância das MB e PB analisando o emprego do alvo genético bexA, no período de 2010 a julho de 2012, em comparação com o emprego do hpd, de agosto de 2012 a 2019. Esta substituição promoveu a melhoria na detecção de variantes não vacinais de Hi nas MB e PB em 37% e 23%, respectivamente, com predomínio de Hia e HiNt, contribuindo para o aprimoramento da vigilância laboratorial das doenças invasivas causadas por Hi. (AU)

Haemophilus influenzae (Hi) is an important pathogen pathogen causing bacterial Meningitis (BM) and bacterial pneumonia (BP), especially in countries where immunoprevention is poor or absent. Hi is differentiated into encapsulated (serotypes a, b, c, d, e, f), and unencapsulated (HiNt), according to the presence or lack of the polysaccharide capsule, respectively. The capsule is the main Hi virulence factor; the bexA gene, responsible for its expression, has been largely used for molecular detection and surveillance of BM and BP. In 2010, the Adolfo Lutz Institute (ALI) implemented real-time PCR (qPCR) using the bexA gene for detecting Hi; but reports on its failing to detect some encapsulated Hi and HiNt caused IAL to replace bexA with hpd as the target gene in the qPCR assay, extending Hi detection to both encapsulated and unencapsulated Hi. In this study, we assessed the impact of replacing the target gene on BM and BP surveillance, by analyzing the use of bexA target gene, within the period from 2010 to July 2012, compared with the use of hpd, from August 2012 to 2019. Adopting the hpd target gene in BM and BP surveillance improved the detection of non-vaccine Hi variants by 37% and 23%, respectively, predominantly Hia and HiNt; and it has contributed to improve laboratory surveillance of invasive Hi diseases. (AU)

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The gold standard for the laboratory diagnosis of COVID-19 is the reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) assay, which searches for SARS-CoV-2 target genes in nasopharyngeal/oropharyngeal (NP/OP) samples, and its performance depends on the quantity and quality of the RNA input. This study compared the performance and cost-effectiveness of three different kits/reagents for RNA extraction used in COVID-19 diagnosis in Sao Paulo, Brazil. A total of 300 NP/OP samples belonging to suspected cases of COVID-19 stored in a biorepository were randomly selected, and RNA was extracted using (i) automated extraction (Loccus, Extracta Kit FAST), (ii) manual extraction (BioGene Kit, Bioclin, Quibasa), and (iii) quick extraction methods (Lucigen, Quick DNA Extract Kit). Next, the samples were tested using RT-qPCR for SARS-CoV-2 with the Allplex 2019-nCoV modified assay and the Charité-Berlin protocol. All assays/kits were used according to the manufacturer's instructions. For the Allplex kit, the sensitivity in detecting SARS-CoV-2 with previously extracted RNA by different procedures was 100.0% for Loccus, 100.0% for BioGene and 91.9% for Quick. Using the Charité-Berlin protocol, the sensitivities were 81.4% for Loccus, 81.2% for BioGene and 60.7% for Quick. The least sensitive target gene and the gene most affected by RNA extraction procedures was the RNA-dependent RNA polymerase gene (Charité-Berlin protocol). No false-positive SARS-CoV-2 results were detected using RNA obtained from any of the different protocols. In conclusion, Loccus and BioGene RNA extractions were efficient for RT-qPCR assays, and although the BioGene procedure is less expensive, Loccus is the best choice because it allows the rapid handling of hundreds or thousands of samples, a desirable feature during pandemics. Although less sensitive, the Quick extraction is useful during outbreaks coupled with the Allplex amplification kit for SARS-CoV-2 diagnosis (κ = 0.925).

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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.

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The gold standard for the laboratory diagnosis of COVID-19 is the reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) assay, which searches for SARS­CoV-2 target genes in nasopharyngeal/oropharyngeal (NP/OP) samples, and its performance depends on the quantity and quality of the RNA input. This study compared the performance and cost-effectiveness of three different kits/reagents for RNA extraction used in COVID-19 diagnosis in Sao Paulo, Brazil. A total of 300 NP/OP samples belonging to suspected cases of COVID-19 stored in a biorepository were randomly selected, and RNA was extracted using (i) automated extraction (Loccus, Extracta Kit FAST), (ii) manual extraction (BioGene Kit, Bioclin, Quibasa), and (iii) quick extraction methods (Lucigen, Quick DNA Extract Kit). Next, the samples were tested using RT-qPCR for SARS-CoV-2 with the Allplex 2019-nCoV modified assay and the Charité-Berlin protocol. All assays/kits were used according to the manufacturer's instructions. For the Allplex kit, the sensitivity in detecting SARS-CoV-2 with previously extracted RNA by different procedures was 100.0% for Loccus, 100.0% for BioGene and 91.9% for Quick. Using the Charité-Berlin protocol, the sensitivities were 81.4% for Loccus, 81.2% for BioGene and 60.7% for Quick. The least sensitive target gene and the gene most affected by RNA extraction procedures was the RNA-dependent RNA polymerase gene (Charité-Berlin protocol). No false-positive SARS-CoV-2 results were detected using RNA obtained from any of the different protocols. In conclusion, Loccus and BioGene RNA extractions were efficient for RT-qPCR assays, and although the BioGene procedure is less expensive, Loccus is the best choice because it allows the rapid handling of hundreds or thousands of samples, a desirable feature during pandemics. Although less sensitive, the Quick extraction is useful during outbreaks coupled with the Allplex amplification kit for SARS-CoV-2 diagnosis (κ = 0.925).

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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.

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We aimed to investigate the nasopharyngeal colonization (NPC) by Streptococcus pneumoniae, Haemophilus influenzae, and Staphylococcus aureus in the elderly population and to assess the demographic factors associated with NPC. This was an observational cohort study in which outpatients aged ≥60 years were enrolled from April to August 2017, with a follow-up visit from September through December 2017. Nasopharyngeal (NP) swabs were collected, bacteria were detected and isolated, and isolates were subjected to phenotypic and molecular characterization using standard microbiological techniques. At enrolment, the rates of S. aureus, methicillin-resistant S. aureus (MRSA), H. influenzae, and S. pneumoniae among 776 elderly outpatients were 15.9%, 2.3%, 2.5%, and 2.2%, respectively. Toxin production was detected in 21.1% of methicillin-susceptible S. aureus, and three SCCmec types were identified: II/IIb, IVa, and VI. At the follow-up visit, all carriage rates were similar (p > 0.05) to the rates at enrolment. Most of S. pneumoniae serotypes were not included in pneumococcal conjugate vaccines (PCVs), except for 7F, 3, and 19A. All strains of H. influenzae were non-typeable. Previous use of antibiotics and 23-valent pneumococcal polysaccharide vaccination (p < 0.05) were risk factors for S. aureus and MRSA carriage; S. aureus colonization was also associated with chronic kidney disease (p = 0.021). S. pneumoniae carriage was associated with male gender (p = 0.032) and an absence of diabetes (p = 0.034), while not receiving an influenza vaccine (p = 0.049) and chronic obstructive pulmonary disease (p = 0.031) were risk factors for H. influenzae colonization. The frailty of study participants was not associated with colonization status. We found a higher S. aureus carriage rate compared with the S. pneumoniae- and H. influenzae-carriage rates in a well-attended population in a geriatric outpatient clinic. This is one of the few studies conducted in Brazil that can support future colonization studies among elderly individuals.

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BACKGROUND: The lack of information regarding the burden of acute bacterial meningitis in Latin America leads to a reduction in the estimated incidence rates of the disease, and impairs public health decisions on the use and follow-up of preventive interventions, particularly, the evaluation of existing vaccination policies. The use of the real-time PCR in diagnostic routine procedures has resulted in a substantial increase in confirmed bacterial meningitis cases. However, in resource-poor countries, these assays are only available in reference laboratories. Sample transportation to these laboratories is a critical constraint, as it requires specialized, high cost courier services. To overcome this barrier we evaluated the use of FTATM Elute filter paper cards for the conservation and processing of samples under normal environmental conditions, as they would be when transported from remote and under-equipped healthcare facilities to the reference centers. A total of 401 samples received in 2015 as part of Sao Paulo's national surveillance for routine diagnosis were selected for this study. METHODS: The sensitivity and specificity of real-time PCR were evaluated using fresh serum and cerebrospinal fluid (CSF) samples processed using our laboratory's standard DNA extraction, and processing the same samples after being dried and stored on FTATM card, and DNA extracted following the manufacturer's instructions. RESULTS: The sensitivities for detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from CSF dried and stored on FTATM cards were 98%, 92%, and 100%, respectively, and with serum samples were 73%, 88%, and 100%, respectively. When compared to our laboratory's standard methodology, results showed high concordance, with Kappa index ranges of 0.9877-1.00 for CSF, and 0.8004-1.00 for serum samples. CONCLUSION: The use of FTATM cards for CSF and serum conservation and transport represents a rapid, reliable, and cost-effective alternative that will allow obtaining valuable epidemiological information that would otherwise be lost.

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Background The lack of information regarding the burden of acute bacterial meningitis in Latin America leads to a reduction in the estimated incidence rates of the disease, and impairs public health decisions on the use and follow-up of preventive interventions, particularly, the evaluation of existing vaccination policies. The use of the real-time PCR in diagnostic routine procedures has resulted in a substantial increase in confirmed bacterial meningitis cases. However, in resource-poor countries, these assays are only available in reference laboratories. Sample transportation to these laboratories is a critical constraint, as it requires specialized, high cost courier services. To overcome this barrier we evaluated the use of FTATM Elute filter paper cards for the conservation and processing of samples under normal environmental conditions, as they would be when transported from remote and under-equipped healthcare facilities to the reference centers. A total of 401 samples received in 2015 as part of Sao Paulo's national surveillance for routine diagnosis were selected for this study. Methods The sensitivity and specificity of real-time PCR were evaluated using fresh serum and cerebrospinal fluid (CSF) samples processed using our laboratory's standard DNA extraction, and processing the same samples after being dried and stored on FTATM card, and DNA extracted following the manufacturer's instructions. Results The sensitivities for detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from CSF dried and stored on FTATM cards were 98%, 92%, and 100%, respectively, and with serum samples were 73%, 88%, and 100%, respectively.

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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.

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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.

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The present study aimed at investigating the best algorithm definition to be employed for diagnosing the human T lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) in HIV-1 infected/Aids patients. Blood samples of 1,608 HIV-1 infected patients from the AIDS Reference Center (CRT DST/AIDS-SP) were tested for the presence of HTLV-1/2 antibodies using two screening assays (EIA Murex HTLV-I+II, and Gold ELISA HTLV-I/II), and confirmed by two Blots [HTLV Blot 2.4 (Western Blot WB) and INNO-LIA HTLV I/II (Line ImmunoAssay - LIA)], and one molecular assay (pol real-time PCR). The screening tests detected 51(Murex) and 49 (Gold ELISA) reagents sera. WB , confirmed 23 HTLV-1, 12 HTLV-2, six HTLV, and nine showed HTLV indeterminate profiles. LIA confirmed 24 HTLV-1, 20 HTLV-2, and six HTLV. PCR detected 18 HTLV-1- and 12 HTLV-2-infected blood samples. By using any confirmatory assay, 50 patients confirmed HTLV infection: 25 HTLV-1 (1.55 %), 21 HTLV-2 (1.31 %) and four HTLV (0.25 %). The sensitivity of LIA, WB and PCR assays were 96 %, 76 % and 60 %, respectively. By considering the assays cost as the sole variable, the best testing algorithm for diagnosing HIV-1/HTLV-coinfection in HIV-1 infected patient was the use of PCR followed by LIA technique(AU)

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(AU)

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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...

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