Transferência tecnológica durante a pandemia de covid-19: relato do primeiro treinamento prático presencial no Brasil / Technology transfer during the COVID-19 pandemic: report on the first face-to-face practical training course in Brazil / Transferencia de tecnología durante la epidemia de COVID-19 en Brasil: informe de la primera formación práctica presencial

Abstract The objective of this report was to describe the first face-to-face course aimed at training public health professionals in performing real-time genomic surveillance during the pandemic period. Experience report on a theoretical-practical course focusing on genomic research and surveillance, including mobile sequencing technologies, bioinformatics, phylogenetics and epidemiological modeling. There were 162 participants in the event and it was the first major face-to-face training course conducted during the COVID-19 epidemic in Brazil. No cases of SARS-CoV-2 infection was detected among the participants at the end of the event, suggesting the safety and effectiveness of all safety measures adopted. The results of this experience suggest that it is possible to conduct professional training safely during pandemics, as long as all safety protocols are followed.

Resumen Este estudio tuvo como objetivo describir el primer curso presencial para capacitar a los profesionales de la salud pública para llevar a cabo la vigilancia genómica en tiempo real durante los períodos de pandemia. Este es un informe de experiencia en un curso teórico-práctico centrado en la investigación y vigilancia genómica, que incluye secuenciación móvil, bioinformática, filogenética y tecnologías de modelado epidemiológico. Este evento contó con la asistencia de 162 participantes y fue la primera gran capacitación presencial realizada durante la epidemia de COVID-19 en Brasil. No se detectó infección por SARS-CoV-2 al final del evento en ningún participante, lo que sugiere la seguridad y efectividad de todas las medidas de seguridad adoptadas. Por lo tanto, los resultados del evento sugieren que es posible realizar entrenamientos profesionales de manera segura durante pandemias, siempre y cuando se sigan todos los protocolos de seguridad.

Resumo O relato descrebeu o primeiro curso presencial visando capacitar profissionais de saúde pública na realização de vigilância genômica em tempo real, durante períodos pandêmicos. Relato de experiência sobre um curso teórico-prático com foco em pesquisa e vigilância genômica, incluindo tecnologias de sequenciamento móvel, bioinformática, filogenética e modelagem epidemiológica. O evento contou com 162 participantes e foi o primeiro grande treinamento presencial realizado durante a epidemia de covid-19 no Brasil. Não foi detectada infecção pelo SARS-CoV-2 ao final do evento em nenhum participante, sugerindo a segurança e efetividade de todas as medidas de segurança adotadas. Os resultados do evento sugerem que é possível executar capacitação profissional com segurança durante pandemias, desde que seguidos todos os protocolos de segurança.

Distribuição espacial de dengue, chikungunya e Zika e os determinantes socioeconômicos em um município da Bahia / Spatial distribution of dengue, chikungunya and Zika and the socioeconomic determinants in a municipality of Bahia

Introdução: desde 1996, o município de Feira de Santana, na Bahia, apresenta sucessivas epidemias de dengue, sendo as mais importantes, do ponto de vista de incidência da doença, as relatadas em 2002 e 2009, com elevados números de casos registrados, hospitalizações e óbitos. Em 2015, houve destaque no registro de casos das três arboviroses, sendo que além da dengue, observase a introdução da chikungunya em 2014 e da Zika em 2015. O índice alto de infestação do Aedes aegypti nas áreas urbana e rural do município pode ter contribuído para a dispersão do vírus. Objetivo: analisar a distribuição espacial da incidência das três arboviroses nas áreas urbana (bairros) e rural (distritos) do município do estudo e avaliar a relação entre os casos das arboviroses e os determinantes socioeconômicos. Metodologia: estudo descritivo realizado em Feira de Santana, no período de 2009 a 2017. Os dados foram distribuídos com auxílio do QGIS por bairros e distritos, apresentados em mapas temáticos. Utilizou-se o programa estatístico Stata versão 14 para análise de dados. Resultados: o estudo mostrou que os bairros localizados na periferia do município e os distritos apresentaram as maiores incidências e as piores condições socioeconômicas, além da co-circulação das três arboviroses na mesma área geográfica e no mesmo período. Conclusão: faz-se necessário ter uma rede de diagnóstico e de vigilância epidemiológica e entomológica consolidada a fim de melhorar o controle destas arboviroses e aprimorar a assistência à saúde.

Introduction: Since 1996, the municipality of Feira de Santana, in Bahia, has presented successive dengue epidemics, the most important, from the point of view of the incidence of the disease, those reported in 2002 and 2009, with high numbers of registered cases, hospitalizations and deaths. In 2015, there was a highlight in the case report of the three arboviruses, and in addition to dengue, there was the introduction of chikungunya in 2014 and Zika in 2015. The high rate of Aedes aegypti infestation in urban and rural areas of the municipality can have contributed to the spread of the virus. Objective: to analyze the spatial distribution of the incidence of the three arboviruses in the urban (neighborhoods) and rural (districts) areas of the municipality of the study and to evaluate the relationship between the cases of arboviruses and the socioeconomic determinants. Methods: a descriptive study carried out in Feira de Santana, from 2009 to 2017. The data were distributed with the aid of QGIS by neighborhoods and districts, presented in thematic maps. The statistical program Stata version 14 was used for data analysis. Results: The study showed that the neighborhoods located on the outskirts of the municipality and the districts had the highest incidences and the worst socioeconomic conditions, in addition to the co-circulation of the three arboviruses in the same geographical area and in the same period. Conclusion: it is necessary to have a consolidated epidemiological and entomological diagnostic and surveillance network in order to improve the control of these arboviruses and improve health care.

Genomic detection of a virus lineage replacement event of dengue virus serotype 2 in Brazil, 2019

BACKGROUND Despite efforts to mitigate the impact of dengue virus (DENV) epidemics, the virus remains a public health problem in tropical and subtropical regions around the world. Most DENV cases in the Americas between January and July 2019 were reported in Brazil. São Paulo State in the southeast of Brazil has reported nearly half of all DENV infections in the country. OBJECTIVES To understand the origin and dynamics of the 2019 DENV outbreak. METHODS Here using portable nanopore sequencing we generated20 new DENV genome sequences from viremic patients with suspected dengue infection residing in two of the most-affected municipalities of São Paulo State, Araraquara and São José do Rio Preto. We conducted a comprehensive phylogenetic analysis with 1,630 global DENV strains to better understand the evolutionary history of the DENV lineages that currently circulate in the region. FINDINGS The new outbreak strains were classified as DENV2 genotype III (American/Asian genotype). Our analysis shows that the 2019 outbreak is the result of a novel DENV lineage that was recently introduced to Brazil from the Caribbean region. Dating phylogeographic analysis suggests that DENV2-III BR-4 was introduced to Brazil in or around early 2014, possibly from the Caribbean region. MAIN CONCLUSIONS Our study describes the early detection of a newly introduced and rapidly-expanding DENV2 virus lineage in Brazil.

Genetic characterisation of Langerin gene in human immunodeficiency virus-1-infected women from Bahia, Brazil

Studies on human genetic variations are a useful source of knowledge about human immunodeficiency virus (HIV)-1 infection. The Langerin protein, found at the surface of Langerhans cells, has an important protective role in HIV-1 infection. Differences in Langerin function due to host genetic factors could influence susceptibility to HIV-1 infection. To verify the frequency of mutations in the Langerin gene, 118 samples from HIV-1-infected women and 99 samples from HIV-1-uninfected individuals were selected for sequencing of the promoter and carbohydrate recognition domain (CRD)-encoding regions of the Langerin gene. Langerin promoter analysis revealed two single nucleotide polymorphisms (SNPs) and one mutation in both studied groups, which created new binding sites for certain transcription factors, such as NFAT5, HOXB9.01 and STAT6.01, according to MatInspector software analysis. Three SNPs were observed in the CRD-encoding region in HIV-1-infected and uninfected individuals: p.K313I, c.941C>T and c.983C>T. This study shows that mutations in the Langerin gene are present in the analysed populations at different genotypic and allelic frequencies. Further studies should be conducted to verify the role of these mutations in HIV-1 susceptibility.

Diversidade genética do vírus da imunodeficiência humana tipo 1 (HIV-1) em mulheres infectadas de uma cidade do nordeste do Brasil / Genetic diversity of human immunodeficiency virus type-1 (HIV-1) in infected women from a northeast city of Brazil

OBJETIVO: descrever a diversidade genética dos isolados de HIV-1 de mulheres soropositivas acompanhadas em um centro de referência. MÉTODOS: estudo transversal, no qual foram incluídas 96 mulheres com dois testes sorológicos ELISA e um teste confirmatório Western Blot. Das amostras de sangue periférico, foram determinadas a carga viral pelo kit b-DNA e a contagem de linfócitos T CD4 e T CD8 pela citometria de fluxo excalibur. A extração e purificação do DNA pró-viral foi realizada pela reação em cadeia da polimerase (PCR), utilizando o kit QIAamp Blood (Qiagen Inc., Chatsworth, CA, USA). O sequenciamento da região pol foi realizado em 52 isolados com o (3100 Genetic Analyzer, Applied Biosystems Inc., Foster City, CA) e a genotipagem foi investigada pela ferramenta Rega (Rega Subtyping Tool). O padrão de resistência aos antirretrovirais (ARV) foi inferido pelo algoritmo do banco de dados Stanford HIV Resistance. Os estágios clínicos das participantes foram definidos como A, B ou C segundo os critérios do Center for Diseases Control (CDC). Para a análise estatística dos dados, foram utilizados os testes do χ2 para as variáveis categóricas e o teste t de Student para as variáveis numéricas. RESULTADOS: a média de idade da amostra, o tempo médio de doença e de tratamento foram: 33,7; 3,8 e 2,5 anos, respectivamente. A média da carga viral foi log10 2,3 cópias/mL; a dos linfócitos T CD4 e T CD8 foi 494,9 células/µL e 1126,4 células/µL. Sobre o estágio clínico, 30 mulheres estavam no estádio A, 47 no B e 19 no C. O sequenciamento dos 52 isolados encontrou 33 do subtipo B, quatro do F, um do C e 14 do recombinante BF. A análise da resistência aos ARV mostrou 39 (75,0 por cento) isolados susceptíveis, 13 (25,0 por cento) resistentes aos inibidores da transcriptase reversa (INTR) e três (5,7 por cento) aos inibidores da protease (IP). CONCLUSÕES: Houve grande diversidade do HIV-1 e elevado percentual de isolados resistentes aos ARV na amostra estudada.

PURPOSE: to describe the genetic diversity of HIV-1 isolates from serum positive women followed up at a reference center. METHODS: transversal study, including 96 women with two ELISA serological tests and a Western Blot confirmatory test. The viral charge was determined by the b-DNA kit, and the counting of T CD4 and T CD8 lymphocytes, by the Excalibur flow cytometry, from the samples of peripheral blood. The extraction and purification of pro-viral DNA was performed by the polymerase (PCR) chain reaction, using the QIAamp Blood kit (Qiagen Inc., Chatsworth, CA, U.S.A.). Sequencing of the pol region was done in 52 isolates with the 3100 Genetic Analyzer (Applied Biosystems Inc., Foster City, CA), and the genotyping was assessed by the Rega Subtyping Tool. The resistance pattern to anti-retrovirals (ARV) was inferred by the algorithm from the Stanford HIV Resistance data bank. Participants' clinical stages were defined as A, B or C, according to the criteria established by the Center for Diseases Control (CDC). For statistical analysis, the χ2 test was used for the categorical variables and the Student's t test, for the numerical variables. RESULTS: The average age of the sample, the disease and treatment average duration were respectively: 33.7 years old, 3.8 and 2.5 years. The viral charge average was log10 2.3 copies/mL; the T CD4 e T CD8 lymphocytes, 494.9 cells/µL and 1126.4 cells/µL. Concerning the clinical stage, 30 women were in stage A, 47 in B and 19 in C. Sequencing from the 52 isolates found 33 of B subtype, 4 of F, 1 of C and 14 of BF recombinant. The analysis of resistance to ARV has shown 39 (75.0 percent) susceptible isolates, 13 (25.0 percent) resistant to reversal transcriptase inhibitors (RTIN), and 3 (5.7 percent) resistant to protease inhibitor (PI). CONCLUSIONS: There has been a large variety of HIV-1 and a high percentage of isolates resistant to ARV in the studied sample.

Mapping the molecular characteristics of Brazilian human T-cell lymphotropic virus type 1 Env (gp46) and Pol amino acid sequences for vaccine design

This study was carried out to evaluate the molecular pattern of all available Brazilian human T-cell lymphotropic virus type 1 Env (n = 15) and Pol (n = 43) nucleotide sequences via epitope prediction, physico-chemical analysis, and protein potential sites identification, giving support to the Brazilian AIDS vaccine program. In 12 previously described peptides of the Env sequences we found 12 epitopes, while in 4 peptides of the Pol sequences we found 4 epitopes. The total variation on the amino acid composition was 9 and 17 percent for human leukocyte antigen (HLA) class I and class II Env epitopes, respectively. After analyzing the Pol sequences, results revealed a total amino acid variation of 0.75 percent for HLA-I and HLA-II epitopes. In 5 of the 12 Env epitopes the physico-chemical analysis demonstrated that the mutations magnified the antigenicity profile. The potential protein domain analysis of Env sequences showed the loss of a CK-2 phosphorylation site caused by D197N mutation in one epitope, and a N-glycosylation site caused by S246Y and V247I mutations in another epitope. Besides, the analysis of selection pressure have found 8 positive selected sites (w = 9.59) using the codon-based substitution models and maximum-likelihood methods. These studies underscore the importance of this Env region for the virus fitness, for the host immune response and, therefore, for the development of vaccine candidates.

Re-mapping the molecular features of the human immunodeficiency virus type 1 and human T-cell lymphotropic virus type 1 Brazilian sequences using a bioinformatics unit established in Salvador, Bahia, Brazil, to give support to the viral epidemiology studies

The analysis of genetic data for human immunodeficiency virus type 1 (HIV-1) and human T-cell lymphotropic virus type 1 (HTLV-1) is essential to improve treatment and public health strategies as well as to select strains for vaccine programs. However, the analysis of large quantities of genetic data requires collaborative efforts in bioinformatics, computer biology, molecular biology, evolution, and medical science. The objective of this study was to review and improve the molecular epidemiology of HIV-1 and HTLV-1 viruses isolated in Brazil using bioinformatic tools available in the Laboratório Avançado de Sáude Pública (Lasp) bioinformatics unit. The analysis of HIV-1 isolates confirmed a heterogeneous distribution of the viral genotypes circulating in the country. The Brazilian HIV-1 epidemic is characterized by the presence of multiple subtypes (B, F1, C) and B/F1 recombinant virus while, on the other hand, most of the HTLV-1 sequences were classified as Transcontinental subgroup of the Cosmopolitan subtype. Despite the high variation among HIV-1 subtypes, protein glycosylation and phosphorylation domains were conserved in the pol, gag, and env genes of the Brazilian HIV-1 strains suggesting constraints in the HIV-1 evolution process. As expected, the functional protein sites were highly conservative in the HTLV-1 env gene sequences. Furthermore, the presence of these functional sites in HIV-1 and HTLV-1 strains could help in the development of vaccines that pre-empt the viral escape process.

Identification and characterization of previously described epitopes in HIV-1 subtypes B, C, F and BF in Brazil

Genetic analysis of HIV-1 is essential to improve treatment strategies and select epitopes for vaccine programs. The objective of this study was to determine whether known CD4+ and CD8+ epitopes were present in Brazilian HIV-1 strains. We used previously described CD8+ and CD4+ epitopes from the Los Alamos laboratory to search for these epitopes in the Brazilian sequences using the HIVbase program and we compared the frequency results with the analyses using physical-chemical profile tools from Network Protein Sequence Analysis (NPSA), and the SYFPEITHI program. Furthermore, this analysis was carried out with the Prosite tool using the GeneDoc program and ds/dn analyses using the Synonymous Nonsynonymous Analysis Program (SNAP). The HIVbase epitope mapping demonstrated that 30 CD8+ and 6 CD4+ epitopes were present in the Brazilian sequences at a high frequency. Only two of these epitopes were heavily glycosylated. Interestingly, ds/dn analyses showed evidence of purifying selective pressure. These types of analyses could be useful for the assessment of possible vaccine efficiency in populations.

Prevalência da infecção pelo vírus linfotrópico humano de células T e fatores de risco associados à soropositividade em doadores de sangue da cidade de Rio Branco, AC, Brasil (1998-2001) / Seroprevalence of human T cell lymphotropic virus infection and associated factors of risk in blood donors of Rio Branco city, AC, Brazil (1998-2001)

Para cada doador de sangue soropositivo (ELISA, Abbott®) para HTLV-I/II, de dezembro de 1998 a março de 2001, também foram selecionados dois soronegativos. As amostras séricas foram re-testadas pelo ELISA (Murex®) e aquelas que permaneceram soropositivas foram testadas pelo Western Blot e pela PCR. Das 11.121 amostras séricas, 73 (0,66 por cento) foram positivas (Abbott®), mas somente 12 (0,11 por cento) permaneceram positivas (Murex®), enquanto que as 146 soronegativas foram confirmadas, apesar de ser sofrível o índice de concordância entre os dois ELISA. O Western Blot confirmou as 12 amostras como soropositivas: 8 (0,07 por cento) HTLV-I; duas (0,02 por cento) HTLV-II e duas (0,02 por cento) indeterminadas - sendo pela PCR uma pelo HTLV-I e a outra pelo HTLV-II. Em conclusão, nessa população da Amazônia Ocidental foi muito baixa a soroprevalência de HTLV-I/II, apesar de ser esperada maior prevalência do HTLV-II devido a grande miscigenação racial indígena.

Estudo do polimorfismo genético dos vírus linfotrópicos de células-T humanas dos tipos I e II (HTLV-I e HTLV-II) em Salvador, Bahia, Brasil e em tribos indígenas brasileiras / Study of genetic polymorphism of lymphotropic virus in human T-cells of types I and II (HTLV-I and HTLV-II) in Salvador, Bahia, Brazil and Brazilian indigenous tribes

Poucos estudos analisaram a diversidade genética de HTLV em Salvador que têm características sócio-demográficas similares a algumas cidades africanas e apresenta a mais elevada prevalência para o HTLV-I (1,35 por cento) em doadores de sangue do Brasil. Nós investigamos a real prevalência nesta população, 1,76 por cento (23/1385). As taxas de infecção foram 1,2 por cento e 2,0 por cento para homens e mulheres, respectivamente, que aumentou com a idade. Quando nós analisamos uma coorte constituída por gestantes, encontramos uma freqüência de 0,9 por cento (57 de 6754). Através de análise filogenética de parte do gene LTR viral, nós investigamos na população geral, gestantes, indivíduos com HAM/TSP e UDls os subtipos de HTLV-I circulantes em Salvador. Demonstramos que a maioria das amostras pertence ao grupo da América Latina, subgrupo A (subtipo a). RFLP de fragmentos gênicos da globina β em 34 destes indivíduos demonstrou que 29.4 por cento dos cromossomos são do haplótipo tipo CAR (Banto), sugerindo que Salvador recebeu também africanos do sul da África durante o tráfico de escravos. Assim, nossos resultados corroboram as hipóteses de múltiplas introduções pós-Colombianas do subgrupo A em Salvador. Nós também analisamos o polimorfismo do HTLV-II em UDls de Salvador e demonstramos que a maioria deles está relacionada à variante molecular Brasileira do subtipo IIa. Assim, demonstramos pela primeira vez no Brasil a presença de um subtipo IIa, de UDls, relacionado aos isolados IIa da América do Norte/Europa. Para investigar aspectos da epidemiologia molecular das infecções causadas pelos HTLV-I, HTLV-II, e HIV-1 em populações indígenas brasileiras, testamos 683 e 321 soros de índios das tribos Tiriyo e Waiampi, respectivamente. As infecções pelo HIV-1 e HTLV-II foram detectadas com prevalências muito baixas entre os Tiriyos, enquanto somente HTLV-I estava presente, com baixa prevalência, entre o Waiampis. Análise filogenética do gene HTLV-Il isolado dos Tiriyos mostrou que estas seqüências se agrupam mais próximas dos isolados de HTLV-II de UDls que vivem em áreas urbanas do sul do Brasil, do que aos isolados da tribo Kayapo. Isto confirma que a maioria das cepas brasileiras de HTLV-II compreende um grupo filogenético com um considerável grau de diversidade.