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RECOMBINATION BREAK POINTS, PREMATURE STOP CODONS AND SINGLE NUCLEOTIDE INDELS OF HIV-1 GAG QUASISPECIES
JA Scott-Herridge1, RP Capina1, MG Mendoza1, X Mao1, B Sheardown1, I MacArthur1, J Kimani2, C Wachihi2, M Luo1, F Plummer1
1Winnipeg, MB; 2Nairobi, Kenya
Objectives: The gag gene encodes proteins for viral structural proteins such as the matrix (p17), capsid (p24), and nucleocapsid (p7) for HIV-1 virions. This study correlates recombination, premature stop codons and single nucleotide indels of the HIV-1 gag with disease progression in patients enrolled in the Pumwani Sex Worker Cohort.
Design: Proviral DNA was isolated from members of the Pumwani Sex Worker cohort from Nairobi, Kenya. The gag genes were PCR amplified, cloned, and sequenced to determine the quasispecies. Phylogenetic analysis was conducted using MEGA 3.1 while RIP 3.0 was used to detect recombination. SPSS 15.0 was used to correlate the frequency of premature stop codons, single nucleotide indels and recombination break points with disease progression.
Results: Phylogenetic analysis was conducted on 4698 quasispecies of HIV-1 gag from 170 patients, 16 of the patients have more than two samples each collected at different dates. Of these 170 patients 19 were classified as rapid progressors (RP), 34 as long term nonprogressors (LTNP) and the remainder as undefined controls. 58.5% of quasispecies were clade A1, 3.6% were clade B, 4.6% were clade C, 14.4% were clade D and 18.8% were recombinants.
Among the recombinant quasispecies 54.6% of the recombination occurs within the vicinity of protease cleavage sites (P=0.034). Within the p17/p24 protease cleavage site RPs displayed a higher frequency of recombination when compared to the control group (P=3.89E-21), whereas the situation of LTNPs is the opposite (P=2.94E-05). In the p24 region, the break point of recombinant quasispecies from LTNPs occurred mainly outside protease cleavage site (P=3.95E-05), the opposite was observed for RPs (P=3.76E-06).
LTNPs tend to have a higher prevalence of premature stop codons and single nucleotide indels that would produce a non-functional protein (P=0.077 and P=0.088 respectively), while no difference was observed in RPs.
Conclusions: The recombination break points, premature stop codons and single nucleotide indels of HIV-1 gag quasispecies correlated well with disease progression. Further correlation of recombination break points, mutations creating non-functional proteins and patient's HLA genotype will help to illustrate the host and viral interactions.