HIV-1 Envelope Subtype Research Articles

Genetic architecture of HIV-1 genes circulating in north India & their functional implications

This review presents data on genetic and functional analysis of some of the HIV-1 genes derived from HIV-1 infected individuals from north India (Delhi, Punjab and Chandigarh). We found evidence of novel B/C recombinants in HIV-1 LTR region showing relatedness to China/Mynmar with 3 copies of Nfκb sites; B/C/D mosaic genomes for HIV-1 Vpr and novel B/C Tat. We reported appearance of a complex recombinant form CRF_02AG of HIV-1 envelope sequences which is predominantly found in Central/Western Africa. Also one Indian HIV-1 envelope subtype C sequence suggested exclusive CXCR4 co-receptor usage. This extensive recombination, which is observed in about 10 per cent HIV-1 infected individuals in the Vpr genes, resulted in remarkably altered functions when compared with prototype subtype B Vpr. The Vpu C was found to be more potent in causing apoptosis when compared with Vpu B when analyzed for subG1 DNA content. The functional implications of these changes as well as in other genes of HIV-1 are discussed in detail with possible implications for subtype-specific pathogenesis highlighted.

Relation Between Chemokine Receptor Use, Disease Stage, and HIV-1 Subtypes A and D

To determine whether there are differences in coreceptor use in subjects infected with HIV-1 envelope subtypes A and D that could explain the differences in progression rates between these subtypes in a rural Ugandan cohort. HIV-1 was subtyped in env by V3 sequencing or heteroduplex mobility assay. Coreceptor use was determined by the ability of the isolates to replicate in U87 CD4 cells expressing different coreceptors. The Fisher exact test was used to examine the relation between coreceptor use and subtype, clinical stage, and V3 charge. The Kruskall-Wallis nonparametric test was used to examine the association between median CD4 cell counts, coreceptor use, and subtype. Logistic regression was used to examine predicted coreceptor use at different CD4 groupings. Isolates from 66 participants were analyzed. Thirty-one were infected with subtype A, and 35 were infected with subtype D. Although this work was based on a small sample size, we found statistically significant differences. The probability of having an X4 virus was higher in subtype D infections than in subtype A infections among those with a non-AIDS clinical status (Fisher exact test, P = 0.040). Logistic regression analysis, in which we predicted X4 use by subtype and stratified by CD4 group, confirmed these findings among those with a CD4 count >200 cells/microL (likelihood ratio test, P = 0.003). R5 viruses were associated with higher median CD4 cell counts than X4 or X4/R5 (Kruskall-Wallis test, P = 0.0045). A V3 charge of +5 and greater was highly associated with X4 virus (Fisher exact test, P = 0.006). These subtype differences in coreceptor use may partially explain the faster progression rates we have previously reported in individuals infected with subtype D compared with subtype A. Our observations may have implications for the future use of coreceptor inhibitors in this population.

Novel engineered HIV-1 East African Clade-A gp160 plasmid construct induces strong humoral and cell-mediated immune responses in vivo

HIV-1 sequences are highly diverse due to the inaccuracy of the viral reverse transcriptase. This diversity has been studied and used to categorize HIV isolates into subtypes or clades, which are geographically distinct. To develop effective vaccines against HIV-1, immunogens representing different subtypes may be important for induction of cross-protective immunity, but little data exist describing and comparing the immunogenicity induced by different subtype-based vaccines. This issue is further complicated by poor expression of HIV structural antigens due to rev dependence. One costly approach is to codon optimize each subtype construct to be examined. Interestingly, cis-acting transcriptional elements (CTE) can also by pass rev restriction by a rev independent export pathway. We reasoned that rev+CTE constructs might have advantages for such expression studies. A subtype A envelope sequence from a viral isolate from east Africa was cloned into a eukaryotic expression vector under the control of the CMV-IE promoter. The utility of inclusion of the Mason–Pfizer monkey virus (MPV)-CTE with/without rev for driving envelope expression and immunogenicity was examined. Expression of envelope (gp120) was confirmed by immunoblot analysis and by pseudotype virus infectivity assays. The presence of rev and the CTE together increased envelope expression and viral infection. Furthermore the CTE+ rev construct was significantly more immunogenic then CTE alone vector. Isotype analysis and cytokine profiles showed strong Th1 response in plasmid-immunized mice, which also demonstrated the superior nature of the rev+CTE construct. These responses were of similar or greater magnitude to a codon-optimized construct. The resulting cellular immune responses were highly cross-reactive with a HIV-1 envelope subtype B antigen. This study suggests a simple strategy for improving the expression and immunogenicity of HIV subtype-specific envelope antigens as plasmid or vector-borne immunogens.

Effect of human immunodeficiency virus (HIV) type 1 envelope subtypes A and D on disease progression in a large cohort of HIV-1-positive persons in Uganda.

The effect of human immunodeficiency virus (HIV) type 1 envelope subtypes A and D on disease progression was investigated in 1045 adults in Uganda. At enrollment and every 6 months, a clinical history, examination, and laboratory investigations that included CD4 cell counts were done. HIV-1 envelope subtype was assessed mainly by peptide serology supplemented by heteroduplex mobility assay and DNA sequencing. A multivariate analysis of survival was performed to assess the prognostic value of HIV-1 subtype on death. A marginal general linear model also determined the effect of subtype on CD4 cell count during follow-up. Subtype D was associated with faster progression to death (relative risk, 1.29; 95% confidence interval, 1.07-1.56; P=.009) and with a lower CD4 cell count during follow-up (P=.001), compared with subtype A, after adjusting for CD4 cell count at enrollment. In Africa, envelope subtype D is associated with faster disease progression, compared with subtype A.

Relationship between HIV-1 Env subtypes A and D and disease progression in a rural Ugandan cohort.

To investigate the role of HIV-1 envelope subtypes on disease progression in a rural cohort of Ugandan adults where two major HIV-1 subtypes (A and D) exist. Participants of a clinical cohort seen between December 1995 and December 1998 had blood collected for HIV-1 subtyping. These included prevalent cases (people already infected with HIV at the start of the study in 1990) and incident cases (those who seroconverted between 1990 and December 1998). HIV-1 subtyping was carried out by heteroduplex mobility assay and DNA sequencing in the V3 env region. Disease progression was measured by the rate of CD4 lymphocyte count decline, clinical progression for the incident cases as time from seroconversion to AIDS or death, to first CD4 lymphocyte count < 200 x 10(6)/l and to the World Health Organization clinical stage 3. All analyses were adjusted for age and sex. One hundred and sixty-four individuals, including 47 prevalent and 117 incident cases, had V3 env subtype data of which 65 (40%) were subtyped as A and 99 as D. In the incident cases, 44 (38%) were subtyped as A and 73 as D. There was a suggestion that for most end-points A had a slower progression than D. The cumulative probability of remaining free from AIDS or death at 6 years post-seroconversion was 0.72 [95% confidence interval (CI), 0.50 to 0.85] for A and 0.58 (95% CI, 0.42 to 0.71) for D, and the adjusted hazard ratio of subtype D compared to A was estimated to be 1.39 (95% CI, 0.66 to 2.94; P = 0.39). The estimated difference in rates of decline in square root CD4 lymphocyte counts was -0.41 per year (95% CI, -0.98 to 0.15; P = 0.15). This study suggests that although subtype A may have a slower progression than D, HIV-1 envelope subtype is not a major factor in determining the progression of HIV-1 disease in a rural population in Uganda.

Clinical disease associated with HIV-1 subtype B' and E infection among 2104 patients in Thailand.

Two HIV-1 envelope subtypes have accounted for virtually all infections in Thailand: subtype B' (Thai B), found mainly in injection drug users (IDU), and subtype E, found in over 90% of sexually infected persons and an increasing proportion of IDU in recent years. It remains unclear whether there are differences in pathogenesis associated with these HIV-1 subtypes. From November 1993 to June 1996, demographic, risk, clinical, and laboratory data were collected by enhanced surveillance from HIV-infected inpatients (> or =14 years) at an infectious disease hospital near Bangkok. HIV-1 subtype was determined by V3-loop peptide enzyme immunoassay (EIA). Because of confounding, multivariate analyses were stratified by risk category and controlled for sex and age. The infecting HIV-1 subtype was determined for 2104 (94.9%) of 2217 HIV-infected patients with complete data: 284 (13.5%) were subtype B', 1820 (86.5%) were E. Specimens from 113 (5.1%) patients were non-reactive or dually reactive on peptide EIA and were excluded. Among IDU, 199 (67.2%) had subtype B', and 97 (32.7%) had E. IDU accounted for 70.1% (199/284) of patients with subtype B' and 5.3% (97/1820) of those with E. Patients infected with HIV-1 subtypes B' or E had similar spectrums of opportunistic infections (OI), levels of immunosuppression, and in-hospital mortality rates. Of patients who did not inject drugs, more patients infected with subtype E had extrapulmonary cryptococcosis than those with subtype B' (adjusted odds ratio, 2.6; 95% confidence interval, 1.28-5.37). HIV-1 subtypes B' and E seem to be associated with similar degrees of immunosuppression and, with one exception, with similar OI patterns. These data do not suggest an association between HIV-1 subtype and differences in pathogenicity.

HIV-1 subtype E incidence and sexually transmitted diseases in a cohort of military conscripts in northern Thailand.

To determine the rate of and risk factors for HIV-1 seroconversion and describe sexually transmitted disease (STD) prevalence rates for young men in northern Thailand. Data were collected from self-administered questionnaires and serologic testing at enrollment in a prospective study in 1991 and at follow-up after 6, 17, and 23 months on a cohort of 1115 men selected by lottery for military conscription. A total of 14 men seroconverted to HIV-1 envelope subtype E. The overall HIV-1 incidence rate was 1.1 (95% confidence interval [CI], 0.6-1.8) per 100 person-years (PY) of follow-up. However, the rate was 2.0/100 PY for conscripts from the upper northern subregion of Thailand compared with 0.5/100 PY from other regions (adjusted rate ratio [RR] = 2.69; 95% CI, 0.8-12.2). On multivariate analyses, the behavioral factors associated with HIV-1 seroconversion were frequency of sex with female sex workers (FSWs; p = .04), receptive anal sex (adjusted RR = 6.73; 95% CI, 1.8-21.7), and large amount of alcohol consumption (adjusted RR = 3.12; 95% CI, 1.0-10.9). Genital ulceration was the STD most strongly associated with seroconversion. The prevalence of serologic reactivity to syphilis, Haemophilus ducreyi, and herpes simplex virus type 2 increased with greater frequency of sex with FSWs and was generally higher for men from the upper north. Young men in northern Thailand are at high risk for HIV-1, primarily through sex with FSWs; and other STDs are highly associated with HIV-1 incidence. As HIV-1 infection extends into the general population, intervention programs are needed to address the problem of sexual transmission apart from commercial sex venues.

The evolving molecular epidemiology of HIV-1 envelope subtypes in injecting drug users in Bangkok, Thailand: implications for HIV vaccine trials.

To genetically characterize HIV-1 strains in injecting drug users (IDU) in Bangkok, Thailand in 1994, and compare these with strains found earlier in Thai IDU; such information is essential for HIV-1 vaccine development and evaluation. Peripheral blood mononuclear cells were collected from 84 IDU attending 14 drug treatment clinics in Bangkok in 1994. DNA was amplified using a nested polymerase chain reaction (PCR) procedure and sequenced directly (without cloning) from the PCR products. The V3 and flanking regions (345 nucleotides) of the env gene were analyzed using a neighbor-joining tree. Only one (1%) strain was a typical subtype B virus, 69 (82%) were genetically distinct subtype B' viruses (Thai B), and 14 (17%) were subtype E strains (Thai A). Persons with recently acquired infection were more likely to have subtype E viruses (P < 0.001) than those in our 1991 survey, who were more likely to have subtype B' viruses. Pairwise intra-subtype differences within subtypes E and B' were 5.3 and 4.3%, respectively, compared with 3.4 and 3.5% among strains collected in 1991 in Thailand. The genetic diversity within subtypes B' and E in Thailand and the proportion of new infections due to subtype E viruses among Bangkok IDU are increasing significantly. These data highlight the importance of monitoring the molecular epidemiology of HIV-1 in populations being considered for HIV-1 vaccine trials.

Rapid Genetic Characterization of HIV Type 1 Strains from Four World Health Organization-Sponsored Vaccine Evaluation Sites Using a Heteroduplex Mobility Assay

To assist in the preparation for the testing of vaccines against human immunodeficiency virus (HIV) we, as part of the World Health Organization Network for HIV Isolation and Characterization (WHO-NHIC), evaluated the genotypic variation of HIV-1 in cohorts from Brazil, Rwanda, Thailand, and Uganda. Here we report the results from a pilot study of 65 HIV-1-infected individuals. In all cases in which viral envelope gene fragments could be amplified by polymerase chain reaction, subtypes could be assigned using a heteroduplex mobility assay (HMA)1 by comparison with HIV-1 strains representing six HIV-1 envelope subtypes. All subtype classifications matched those found by envelope gene sequencing. Phylogenetic relationships were further clarified by heteroduplex formation between samples within each subtype. A relatively homogeneous subtype E virus population predominated over subtype B viruses in the sample set from Thailand. Viruses from the other countries were also limited to one or two subtypes but were more divergent within each subtype. All samples from Rwanda (13/13) and some from Uganda (3/16) were of subtype A; all Brazilian samples were of subtype B, except for one belonging to subtype C; most samples from Uganda (13/16) clustered with the subtype D. Analysis by HMA is therefore applicable for screening of HIV-1 genotypes in countries under consideration for large-scale vaccine trials. It should be generally useful when samples containing at least one variable genetic locus need to be rapidly classified by genotype and/or analyzed for epidemiological clustering.