Variabilty of HIV-1 near full length genomes among pediatric infections from São Paulo, Brazil

Abstract

Genetic variability is a major feature of the human immunodeficiency virus type 1 (HIV-1) and considered the key factor to frustrate efforts to halt the virus epidemic. Understanding such diversity is fundamental not only for design of successful drugs or vaccine but also for epidemiology and genetic diagnosis. In this study we aim to conduct an analysis of the complete genome of HIV-1 isolates from pediatric patients infected by vertical transmission in the State of Sao Paulo. The acheivement of this objective will allow us to optimize stratigies for HIV-1 diagnosis, to identify important signatures and polymorphisms throughout sequenced fragments and their association with the biological evolution of CD4 T cell counts or plasma viremia, to provide an estimate of cross clade conservation of the best-defined HIV-1 cytotoxic T-lymphocyte (CTL) epitopes by comparing the predicted protein sequences of our isolates to the reference optimal epitope sequences from a large number of HLA class 1 and II alleles covering more than 90% of HLA haplotypes in different populations, to define the structural pattern of recombinant strains, if exist, and determine their proportions, provide more HIV-1 Brazilian sequences characterized on more than one region of the genomes which are still rare in HIV database, to describe the classification stability of HIV-1 subtypes at the level of full genomes since most available studies to date have reported only the assessment of partial genomes and to screen the entire viral genomes for resistance to available drug therapies. HIV-1 proviral DNA from patients` peripheral blood mononuclear cells (PBMC) will be extracted and genomic DNA will be amplified by nested polymerase chain reactions (PCR) from five overlapped fragments according to our established methods. Amplified products will be purified and directly sequenced on both strands. All sequenced fragments will be edited, assembled and analyzed by different phylogentic and other appropriate tools. Information on evolution, cohort statistics, host genetic background, epitope mapping, clinical, immunological, and epidemiological data will be cross-referenced to viral sequences to allow for subsequent association studies subsequentes. (AU)