title: Ultra-Deep Sequencing of HIV-1 near Full-Length and Partial Proviral Genomes Reveals High Genetic Diversity among Brazilian Blood Donors.
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).
  
  
  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).
  
  
  CONCLUSIONS
  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.
authors.affiliation: Clinical Laboratory, Department of Pathology, LIM 03, Hospital das Clínicas (HC), School of Medicine, University of São Paulo, São Paulo, Brazil.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Pessôa
authors.affiliation: Pernambuco State Center of Hematology and Hemotherapy-HEMOPE, Recife, Pernambuco, Brazil.
authors.forename: Paula
authors.initials: P
authors.lastname: Loureiro
authors.affiliation: Hemorio, Rio de Janeiro, Brazil.
authors.forename: Maria
authors.initials: M
authors.lastname: Esther Lopes
authors.affiliation: Minas Gerais State Center of Hematology and Hemotherapy-HEMOMINAS, Belo Horizonte, Minas Gerais, Brazil.
authors.forename: Anna B F
authors.initials: AB
authors.lastname: Carneiro-Proietti
authors.affiliation: Department of Infectious Disease/Institute of Tropical Medicine, University of São Paulo, Sao Paulo, Brazil.
authors.forename: Ester C
authors.initials: EC
authors.lastname: Sabino
authors.affiliation: Blood Systems Research Institute, San Francisco, California, United States of America.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Busch
authors.affiliation: Clinical Laboratory, Department of Pathology, LIM 03, Hospital das Clínicas (HC), School of Medicine, University of São Paulo, São Paulo, Brazil.
authors.forename: Sabri S
authors.initials: SS
authors.lastname: Sanabani
mesh_terms: D001483:Base Sequence; D001782:Blood Donors; D001938:Brazil / Q000453:epidemiology; D004247:DNA / Q000737:chemistry / Q000235:genetics; D015723:Gene Library; D014644:Genetic Variation; D016679:Genome, Viral; D005838:Genotype; D015658:HIV Infections / Q000453:epidemiology; D015497:HIV-1 / Q000145:classification / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D010802:Phylogeny; D011533:Proviruses / Q000235:genetics; D012367:RNA, Viral / Q000097:blood / Q000737:chemistry; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D017423:Sequence Analysis, RNA
chemical_list: D012367:RNA, Viral; D004247:DNA