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Este projecto de investigação aborda três aspectos da infecção por HIV na República Democrática de São Tomé e Príncipe (STP), nomeadamente: (i) epidemiologia dos vírus circulantes; (ii) subtipos de HIV-1 e sua filogenia; (iii) resistência a fármacos antirretrovirais.
Após a aprovação pela Comissão de Ética local, a epidemiologia dos vírus circulantes foi estudada numa amostragem efectuada nos distritos de Água Grande e Mé Zóchi, em 2007. Envolveu 1018 participantes de ambos os sexos dos 14 aos 65 anos, e consistiu num questionário anónimo com colheita de amostras aleatorizadas de sangue venoso. Estas foram testadas com “Determine™ HIV ½” (Abbott Diagnostics Il., USA) e, quando positivo, retestadas com “VIDAS™ HIV DUO” (BioMérieurx SA). Só cinco casos foram positivos em ambos os testes, quatro de sexo masculino e um feminino.
Os subtipos de HIV-1 e a sua filogenia foram estudados no plasma de 93 amostras de conveniência de pessoas entre os 7 e os 63 anos, de ambos os sexos. Estas amostras foram colhidas em diferentes instituições de Saúde do país, de 2004 a 2007. Dois fragmentos do gene da polimerase (pol) – um sendo a totalidade da protease (PR, codões 1-99) e o outro, parte da transcriptase reversa (RT, codões 1-335) – foram amplificados e sequenciados com “ViroSeq HIV-1 System” e o sequenciador “ABI 3100”. As sequências cDNA resultantes da PR e da RT foram analisadas com os algoritmos de subtipagem “HIVSeq” e “REGA Subtyping Tool”. Foram desenhadas árvores filogenéticas por comparação com as sequências de referência do grupo M. As discrepâncias encontradas entre os dois algoritmos foram analisadas com o software Simplot v3.2. Algumas amostras foram classificadas como subtipos puros (A, C, D, F, G, H, J) mas nenhum subtipo B foi encontrado. As amostras, na sua maioria (56%), eram recombinantes (CRF02_AG, CRF02_AG/G, CRF02_AG/H, A/CRF02_AG, K/CRF02_AG, CRF01_AE/CRF02_AG), sugerindo transmissão local. Os subtipos A, G e os seus recombinantes representaram mais do que ¾ do total.
Em pessoas infectadas, nas mesmas sequências de cDNA quer de doentes nunca tratados quer de doentes sob tratamento, o padrão de resistências genotípicas aos ARVs foi avaliado com o “Stanford HIV Drug Resistance Algorithm” (beta test). As mutações de resistência encontradas foram: uma de tipo PIs (primária – F53L), oito de tipo NRTIs (M41L, T69N/S, M184I/V, T215Y/S e K219Q) e seis de tipo NNRTIs (A98G, K103N/S, V179E, Y181C, G190A). Foram também encontrados vários polimorfismos nos sítios da PR e da RT.Embora a prevalência do HIV-1 fosse ainda relativamente baixa (0,5%), a grande diversidade genética dos vírus de subtipos não-B em circulação em STP, pode comprometer quer a fiabilidade dos testes serológicos quer a eficácia de hipotéticas futuras vacinas para os países da África Central e da costa Ocidental.
This research project aims to study three aspects of HIV infection in the Democratic Republic of São Tomé e Príncipe (STP), namely: (i) the epidemiology of circulating viruses; (ii) HIV-1 subtypes and their phylogeny; (iii) antiretroviral drug resistance patterns. The circulating viruses’ epidemiology was studied, after the local Ethics Committee approval, by way of a survey in the districts of Agua Grande and Mé Zóchi, during 2007. It involved 1018 participants of both genders from 14 to 65 years old, and consisted in a randomized anonymous questionnaire and venous blood sampling. The latter were screened with “DetermineTM HIV ½” (Abbott Diagnostics, Il., USA) and, if positive, tested with “VIDAS® HIV DUO” (BioMérieurx SA). Only five cases (0.5 %) were positive on both tests four from males and one from a female. The HIV-1 subtypes and their phylogeny were studied in 93 convenience blood samples of people aged 7 to 63 of both genders. These samples were collected in the country’s different Health institutions from 2004 to 2007. Two fragments of polymerase gene (pol) - one consisting of the full protease (PR, codons 1-99) and the other, part of the reverse transcriptase (RT, codons 1-335) - were amplified and sequenced with “ViroSeq HIV-1 System” and “ABI 3100” sequencer. The resulting cDNA from PR and RT were analyzed with the subtyping algorithms “HIVSeq” and “REGA Subtyping Tool”. Phylogenetic trees were designed using as comparison the reference’s sequences of group M. The discrepancies found between the two algorithms were analyzed with software Simplot v3.2. Some samples were classified as pure subtypes (A, C, D, F, G, H, J) while no subtype B was found. The majority (56%) of the samples were recombinants (CRF02_AG, CRF02_AG/G, CRF02_AG/H, A/CRF02_AG, K/CRF02_AG, CRF01_AE/CRF02_AG), suggesting regional transmission. Subtypes A, G and their recombinants represent more than ¾ of the total. In the same cDNA sequences, from both treated and treatment-naïve infected people, the genotypic ARVs resistance profile was assessed using “Stanford HIV Drug Resistance Algorithm” (beta test). The resistance mutations found were: one PIs type (primary – F53L), eight NRTIs type (M41L, T69N/S, M184I/V, T215Y/S and K219Q) and six NNRTIs type (A98G, K103N/S, V179E, Y181C, G190A). Several polymorphisms on the PR and RT sites were also found. Although HIV-1 prevalence was still relatively low (0,5%), the great genetic diversity of the non-B subtypes viruses circulating in STP may compromise both the reliability of the serological tests and the effectiveness of hypothetical future vaccines for the Central Africa and West coast.
This research project aims to study three aspects of HIV infection in the Democratic Republic of São Tomé e Príncipe (STP), namely: (i) the epidemiology of circulating viruses; (ii) HIV-1 subtypes and their phylogeny; (iii) antiretroviral drug resistance patterns. The circulating viruses’ epidemiology was studied, after the local Ethics Committee approval, by way of a survey in the districts of Agua Grande and Mé Zóchi, during 2007. It involved 1018 participants of both genders from 14 to 65 years old, and consisted in a randomized anonymous questionnaire and venous blood sampling. The latter were screened with “DetermineTM HIV ½” (Abbott Diagnostics, Il., USA) and, if positive, tested with “VIDAS® HIV DUO” (BioMérieurx SA). Only five cases (0.5 %) were positive on both tests four from males and one from a female. The HIV-1 subtypes and their phylogeny were studied in 93 convenience blood samples of people aged 7 to 63 of both genders. These samples were collected in the country’s different Health institutions from 2004 to 2007. Two fragments of polymerase gene (pol) - one consisting of the full protease (PR, codons 1-99) and the other, part of the reverse transcriptase (RT, codons 1-335) - were amplified and sequenced with “ViroSeq HIV-1 System” and “ABI 3100” sequencer. The resulting cDNA from PR and RT were analyzed with the subtyping algorithms “HIVSeq” and “REGA Subtyping Tool”. Phylogenetic trees were designed using as comparison the reference’s sequences of group M. The discrepancies found between the two algorithms were analyzed with software Simplot v3.2. Some samples were classified as pure subtypes (A, C, D, F, G, H, J) while no subtype B was found. The majority (56%) of the samples were recombinants (CRF02_AG, CRF02_AG/G, CRF02_AG/H, A/CRF02_AG, K/CRF02_AG, CRF01_AE/CRF02_AG), suggesting regional transmission. Subtypes A, G and their recombinants represent more than ¾ of the total. In the same cDNA sequences, from both treated and treatment-naïve infected people, the genotypic ARVs resistance profile was assessed using “Stanford HIV Drug Resistance Algorithm” (beta test). The resistance mutations found were: one PIs type (primary – F53L), eight NRTIs type (M41L, T69N/S, M184I/V, T215Y/S and K219Q) and six NNRTIs type (A98G, K103N/S, V179E, Y181C, G190A). Several polymorphisms on the PR and RT sites were also found. Although HIV-1 prevalence was still relatively low (0,5%), the great genetic diversity of the non-B subtypes viruses circulating in STP may compromise both the reliability of the serological tests and the effectiveness of hypothetical future vaccines for the Central Africa and West coast.
Descrição
Palavras-chave
Doenças infecciosas HIV-1 Epidemiologia Controle São Tomé e Príncipe
Contexto Educativo
Citação
Editora
Instituto de Higiene e Medicina Tropical