HETERODUPLEX MOBILITY ASSAY AND PHYLOGENETIC ANALYSIS OF V3 REGION SEQUENCES OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 ISOLATES FROM GULU, NORTHERN UGANDA

We analyzed peripheral blood mononuclear cells from 19 asymptomatic se ropositive pregnant women from the district of Gulu in northern Uganda . A 700-bp fragment of the human immunodeficiency virus type 1 (HIV-1) env gene, including the V3-V5 region, was successfully amplified by P CR from 10 samples (52.6%) and was subsequently subjected to both a he teroduplex mobility assay for genetic screening and subtyping and DNA sequence analysis (approximately 300 bp) for nucleotide comparison and phylogenetic studies. The results show the presence of HIV-1 A and D subtypes (or clades) in this rural area, with the prevalence of the A subtype (8 of 10) being greater than that of the D (2 of 10) subtype, which is unlike what was previously reported for Uganda. By pairwise c omparison analysis, the percentage of sequence divergence among sample s within each subtype is low (the average intrasubtype divergence is 1 5.8%), but it is significantly higher between the two subtypes (the av erage intersubtype divergence is 23%). At the amino acid level, the tw o HIV-1 subtypes show distinct tetramers at the apex of the V3 loop an d, in particular, GPGQ in clade A and GPGR in clade D. In addition, 10 of the 19 viral samples (52.6%) have been isolated in vitro. Nine of the samples have been classified as rapid/high, which reflects a high in vitro replication capacity for the HIV 1 field isolates from this c ountry, even for those obtained from seropositive asymptomatic individ uals. These observations, despite being made on the basis of a limited sample size, show a modest degree of genetic divergence among samples isolated in the last 4 years in this country by comparison with those based on the 1990 data on HIV-1 isolates from Kampala. The results re ported here are, therefore, extremely relevant for Uganda, which is on e of the selected World Health Organization field sites for future HIV -1 vaccine evaluation programs.