Genomas completos e parciais de formas recombinantes BF1 e BC do HIV-1 circulantes nos estados de Goiás, Mato Grosso, Mato Grosso do Sul, Tocantins, Maranhão e Piauí

• Autor(a) principal: Reis, Mônica Nogueira da Guarda
• Data de Publicação: 2017
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da UFG
• Texto Completo: http://repositorio.bc.ufg.br/tede/handle/tede/9772
• topic: HIV-1; Formas recombinantes; Genoma completo; Recombinants forms; Full length genome; CIENCIAS BIOLOGICAS::MICROBIOLOGIA

Background and objectives: One of the most striking features of HIV-1 is its extensive genetic polymorphism which allows its classification into four groups (M, N, O and P). The pandemic group M can be classified into nine subtypes (A-D, F-H, J, K), six sub-subtypes (A1-A4, F1-F2), dozens of circulating recombinant forms (CRFs) and countless unique recombinant forms. The molecular epidemiology of HIV-1 in Brazil is complex and dynamic and has been characterized by the cocirculation of subtypes B, F1 and C and BF, BC, BFC and CF recombinants. Previous studies on pol sequences of HIV-1 isolates from six Brazilian States (Goiás, Mato Grosso, Mato Grosso do Sul, Tocantins, Maranhão e Piauí) have shown great variability of BF, BC and FC recombinant viruses. This thesis describes the molecular characterization of these mosaic genomes including reclassification in pol and generation of full length/near full length/ partial genomes of representative isolates. The results of this thesis are presented as articles, one of them is already published and two other are presented as manuscripts to be submitted. Methods: Proviral DNA was extracted from whole blood and four overlapping fragments that compose HIV-1 whole genome were amplified by “nested”-PCR. The generated sequences were aligned (BioEdit 7.2.0) and phylogenetic analysis performed (MEGA 6, Neighbor-Joining, Kimura 2 parameters). The recombination profiles were identified by point analysis, phylogenetic analysis of fragments, and Bootscan analysis (SIMPLOT v3.5.1). The time of the most recent common ancestor (TMRCA) of HIV-1 BF clades was estimated (Bayesian Markov Chain Monte Carlo, BEAST v1.8). Results: Article 1/Plos One: Among 828 HIV-1 isolates from six Brazilian States, phylogenetic analysis of pol identified 87 BF recombinant isolates: 48% (42/87) grouped into five different clusters (Cluster #1-5), 21% (18/87) was CRF_BF-like and 31% (27/87) were classified as URFs_BF. Among 22 isolates that composed the largest BF cluster (#5), we have obtained six full length genomes, one near full length genome and four partial genomes. These 11 isolates, obtained from patients that did not have any epidemiological link shared identical recombination profile in their genomes, allowing the description of a new CRF. This recombinant named CRF90_BF1 is circulating in Goiás, Mato Grosso e Tocantins states. Among 20 BF isolates from the other four clusters we have obtained: three full length genomes, four near full length genomes and five partial genomes. Analysis of these sequences characterized three URFs (Clusters #1, #2 e #3), circulating in the Central West region. The estimated TMRCA for these BF clades was the beginning of the 90’s. BLAST search analysis identified other sequences sharing the same recombination pattern among isolates from the North and South regions, suggesting that they may represent other potential, unidentified CRFs. A moderate rate of CRF28/CRF29_BF-like isolates was seen (16.1%, 14/87) in Goiás, Mato Grosso and Mato Grosso do Sul states. Manuscripts 1 and 2: BC and BF recombinant isolates were also detected (2.9%; 24/828): BC (2,3%, 19/828), BFC (0,4%, 3/828) and CF (0,2%, 2/828). Among these 24 isolates, 19 grouped into six clusters while five (BRGO4156, BRMT2509, BRMT3086, BRMS43, BRPI34) did not cluster. Six clusters were identified (Clusters #1-6): two clusters contained 7 isolates with recombination profiles similar to already identified CRFs_BC (29.2%, 7/24). Cluster #4/CRF31_BC-like comprised 5 isolates (20.8%) and two isolates were Cluster #5/CRF60_BC-like (8.3%). Most patients infected with BC and FC isolates was antirretroviral naïve and (23/24) and 21.7% (5/23) had mutations associated with transmitted drug resistance. Conclusions: Our analyses of isolates from Central West, North and Northeast regions allowed the characterization of the new CRF90_BF1 and different URFs_BF and BC from already described ones. Also isolates sharing recombination profiles with already described ones were also observed (CRF28/29_BF-like, CRF31_BC-like, CRF60_BC-like). Altogether our results indicate that coinfection/ superinfection and intersubtype recombination may be more prevalente that reported. Also, our results indicat the continuing generation of recombinants in the Brazilian epidemic that is taking place in urban centers located away from the epicenter of the epidemic. Continued surveillance studies including full/near full genomes in larger sample sizes are necessary to identify emerging and disseminating recombinants and their implication in the transmission and control of HIV-1 in Brazil.