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Ultra-Deep Sequencing of HIV-1 near Full-Length and Partial Proviral Genomes Reveals High Genetic Diversity among Brazilian Blood Donors.

Pessôa R, Loureiro P, Esther Lopes M, Carneiro-Proietti AB, Sabino EC, Busch MP, Sanabani SS - PLoS ONE (2016)

Bottom Line: To this end, a high-throughput sequencing-by-synthesis protocol and instrument were used to characterize the near full length (NFLG) and partial HIV-1 proviral genome in 259 HIV-1 infected blood donors at four major blood centers in Brazil: Pro-Sangue foundation (São Paulo state (SP), n 51), Hemominas foundation (Minas Gerais state (MG), n 41), Hemope foundation (Recife state (PE), n 96) and Hemorio blood bank (Rio de Janeiro (RJ), n 70).Of the 259 samples studied, 208 (80%) NFLGs and 49 (18.8%) partial fragments were de novo assembled into contiguous sequences and successfully subtyped.Based on this work, subtype B appears to be the prevalent subtype followed by a high proportion of intersubtype recombinants that appeared to be arising continually in this country.

View Article: PubMed Central - PubMed

Affiliation: Clinical Laboratory, Department of Pathology, LIM 03, Hospital das Clínicas (HC), School of Medicine, University of São Paulo, São Paulo, Brazil.

ABSTRACT

Background: Here, we aimed to gain a comprehensive picture of the HIV-1 diversity in the northeast and southeast part of Brazil. To this end, a high-throughput sequencing-by-synthesis protocol and instrument were used to characterize the near full length (NFLG) and partial HIV-1 proviral genome in 259 HIV-1 infected blood donors at four major blood centers in Brazil: Pro-Sangue foundation (São Paulo state (SP), n 51), Hemominas foundation (Minas Gerais state (MG), n 41), Hemope foundation (Recife state (PE), n 96) and Hemorio blood bank (Rio de Janeiro (RJ), n 70).

Materials and methods: A total of 259 blood samples were obtained from 195 donors with long-standing infections and 64 donors with a lack of stage information. DNA was extracted from the peripheral blood mononuclear cells (PBMCs) to amplify the HIV-1 NFLGs from five overlapping fragments. The amplicons were molecularly bar-coded, pooled, and sequenced by Illumina paired-end protocol.

Results: Of the 259 samples studied, 208 (80%) NFLGs and 49 (18.8%) partial fragments were de novo assembled into contiguous sequences and successfully subtyped. Of these 257 samples, 183 (71.2%) were pure subtypes consisting of clade B (n = 167, 65%), C (n = 10, 3.9%), F1 (n = 4, 1.5%), and D (n = 2, 0.7%). Recombinant viruses were detected in 74 (28.8%) samples and consist of unique BF1 (n = 41, 15.9%), BC (n = 7, 2.7%), BCF1 (n = 4, 1.5%), CF1 and CDK (n = 1, 0.4%, each), CRF70_BF1 (n = 4, 1.5%), CRF71_BF1 (n = 12, 4.7%), and CRF72_BF1 (n = 4, 1.5%). Evidence of dual infection was detected in four patients coinfected with the same subtype (n = 3) and distinct subtype (n = 1).

Conclusion: Based on this work, subtype B appears to be the prevalent subtype followed by a high proportion of intersubtype recombinants that appeared to be arising continually in this country. Our study represents the largest analysis of the viral NFLG ever undertaken worldwide and provides insights into the understanding the genesis of the HIV-1 epidemic in this particular area of South America and informs vaccine design and clinical trials.

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Related in: MedlinePlus

Summary of the subtype distribution of viruses from the 257 long-standing HIV-1 infected blood donors analyzed.The number of viruses that belong to each subtype is indicated in the relevant bar chart section. The regions of origin are color-coded and indicated by arrows.
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pone.0152499.g001: Summary of the subtype distribution of viruses from the 257 long-standing HIV-1 infected blood donors analyzed.The number of viruses that belong to each subtype is indicated in the relevant bar chart section. The regions of origin are color-coded and indicated by arrows.

Mentions: A total of NINE different unique HIV subtypes and three different BF1 CRF variants were identified through NFLGs and partial genome analysis. Fig 1 shows the regional distributions of HIV-1 subtypes and illustrates the substantial HIV genetic diversity in Brazil. In particular, there are at least five subtypes circulating in MG, seven in PE, seven in SP, and eight in RJ. The genetic subtype distribution did not differ significantly among the four regions. Approximately 82.2% (74 of 90) of non-subtype B strains in this study were identified as recombinant variants, representing 28.8% (74 of 257) of the total successfully sequenced samples. Subtype B is predominant in the four regions (167 of 257 [65%]); the lowest prevalence was observed in SP (29 of 50 [58%]) and RJ (42 of 70 [60%]), and the highest prevalence was seen in MG (29 of 41 [71%]). BF1 recombinants including the newly described CRF 70 and 71 BF1 [26] were identified in 27% (26 of 96) of subjects from PE, 21.4% (15 of 70) from RJ, 24% (12 of 50) from SP, and 19.5% (8 of 41) subjects from MG. Subclade F1 was observed at low prevalence in SP, PE, and RJ, where it represents 4% (2 of 50), 1% (1 of 96), and 1.4% (1 of 70) of the circulating strains, respectively. Subtype C was also seen at lower prevalence in all regions, 8% in SP. 7.4% in MG, 2.8% in RJ and 1% in PE (Fig 1). Two sequences belong to subtype D were detected in RJ. A detailed description of the NFLG of these two sequences has already been published elsewhere [26].


Ultra-Deep Sequencing of HIV-1 near Full-Length and Partial Proviral Genomes Reveals High Genetic Diversity among Brazilian Blood Donors.

Pessôa R, Loureiro P, Esther Lopes M, Carneiro-Proietti AB, Sabino EC, Busch MP, Sanabani SS - PLoS ONE (2016)

Summary of the subtype distribution of viruses from the 257 long-standing HIV-1 infected blood donors analyzed.The number of viruses that belong to each subtype is indicated in the relevant bar chart section. The regions of origin are color-coded and indicated by arrows.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4816342&req=5

pone.0152499.g001: Summary of the subtype distribution of viruses from the 257 long-standing HIV-1 infected blood donors analyzed.The number of viruses that belong to each subtype is indicated in the relevant bar chart section. The regions of origin are color-coded and indicated by arrows.
Mentions: A total of NINE different unique HIV subtypes and three different BF1 CRF variants were identified through NFLGs and partial genome analysis. Fig 1 shows the regional distributions of HIV-1 subtypes and illustrates the substantial HIV genetic diversity in Brazil. In particular, there are at least five subtypes circulating in MG, seven in PE, seven in SP, and eight in RJ. The genetic subtype distribution did not differ significantly among the four regions. Approximately 82.2% (74 of 90) of non-subtype B strains in this study were identified as recombinant variants, representing 28.8% (74 of 257) of the total successfully sequenced samples. Subtype B is predominant in the four regions (167 of 257 [65%]); the lowest prevalence was observed in SP (29 of 50 [58%]) and RJ (42 of 70 [60%]), and the highest prevalence was seen in MG (29 of 41 [71%]). BF1 recombinants including the newly described CRF 70 and 71 BF1 [26] were identified in 27% (26 of 96) of subjects from PE, 21.4% (15 of 70) from RJ, 24% (12 of 50) from SP, and 19.5% (8 of 41) subjects from MG. Subclade F1 was observed at low prevalence in SP, PE, and RJ, where it represents 4% (2 of 50), 1% (1 of 96), and 1.4% (1 of 70) of the circulating strains, respectively. Subtype C was also seen at lower prevalence in all regions, 8% in SP. 7.4% in MG, 2.8% in RJ and 1% in PE (Fig 1). Two sequences belong to subtype D were detected in RJ. A detailed description of the NFLG of these two sequences has already been published elsewhere [26].

Bottom Line: To this end, a high-throughput sequencing-by-synthesis protocol and instrument were used to characterize the near full length (NFLG) and partial HIV-1 proviral genome in 259 HIV-1 infected blood donors at four major blood centers in Brazil: Pro-Sangue foundation (São Paulo state (SP), n 51), Hemominas foundation (Minas Gerais state (MG), n 41), Hemope foundation (Recife state (PE), n 96) and Hemorio blood bank (Rio de Janeiro (RJ), n 70).Of the 259 samples studied, 208 (80%) NFLGs and 49 (18.8%) partial fragments were de novo assembled into contiguous sequences and successfully subtyped.Based on this work, subtype B appears to be the prevalent subtype followed by a high proportion of intersubtype recombinants that appeared to be arising continually in this country.

View Article: PubMed Central - PubMed

Affiliation: Clinical Laboratory, Department of Pathology, LIM 03, Hospital das Clínicas (HC), School of Medicine, University of São Paulo, São Paulo, Brazil.

ABSTRACT

Background: Here, we aimed to gain a comprehensive picture of the HIV-1 diversity in the northeast and southeast part of Brazil. To this end, a high-throughput sequencing-by-synthesis protocol and instrument were used to characterize the near full length (NFLG) and partial HIV-1 proviral genome in 259 HIV-1 infected blood donors at four major blood centers in Brazil: Pro-Sangue foundation (São Paulo state (SP), n 51), Hemominas foundation (Minas Gerais state (MG), n 41), Hemope foundation (Recife state (PE), n 96) and Hemorio blood bank (Rio de Janeiro (RJ), n 70).

Materials and methods: A total of 259 blood samples were obtained from 195 donors with long-standing infections and 64 donors with a lack of stage information. DNA was extracted from the peripheral blood mononuclear cells (PBMCs) to amplify the HIV-1 NFLGs from five overlapping fragments. The amplicons were molecularly bar-coded, pooled, and sequenced by Illumina paired-end protocol.

Results: Of the 259 samples studied, 208 (80%) NFLGs and 49 (18.8%) partial fragments were de novo assembled into contiguous sequences and successfully subtyped. Of these 257 samples, 183 (71.2%) were pure subtypes consisting of clade B (n = 167, 65%), C (n = 10, 3.9%), F1 (n = 4, 1.5%), and D (n = 2, 0.7%). Recombinant viruses were detected in 74 (28.8%) samples and consist of unique BF1 (n = 41, 15.9%), BC (n = 7, 2.7%), BCF1 (n = 4, 1.5%), CF1 and CDK (n = 1, 0.4%, each), CRF70_BF1 (n = 4, 1.5%), CRF71_BF1 (n = 12, 4.7%), and CRF72_BF1 (n = 4, 1.5%). Evidence of dual infection was detected in four patients coinfected with the same subtype (n = 3) and distinct subtype (n = 1).

Conclusion: Based on this work, subtype B appears to be the prevalent subtype followed by a high proportion of intersubtype recombinants that appeared to be arising continually in this country. Our study represents the largest analysis of the viral NFLG ever undertaken worldwide and provides insights into the understanding the genesis of the HIV-1 epidemic in this particular area of South America and informs vaccine design and clinical trials.

Show MeSH
Related in: MedlinePlus