Recent studies in the SIV-macaque model of HIV infection suggest that Nef-specific CD8+ T-cell responses may mediate highly effective immune control of viraemia. In HIV infection Nef recognition dominates in acute infection, but in large cohort studies of chronically infected subjects, breadth of T cell responses to Nef has not been correlated with significant viraemic control. Improved disease outcomes have instead been associated with targeting Gag and, in some cases, Pol. However analyses of the breadth of Nef-specific T cell responses have been confounded by the extreme immunogenicity and multiple epitope overlap within the central regions of Nef, making discrimination of distinct responses impossible via IFN-gamma ELISPOT assays. Thus an alternative approach to assess Nef as an immune target is needed. Here, we show in a cohort of >700 individuals with chronic C-clade infection that >50% of HLA-B-selected polymorphisms within Nef are associated with a predicted fitness cost to the virus, and that HLA-B alleles that successfully drive selection within Nef are those linked with lower viral loads. Furthermore, the specific CD8+ T cell epitopes that are restricted by protective HLA Class I alleles correspond substantially to effective SIV-specific epitopes in Nef. Distinguishing such individual HIV-specific responses within Nef requires specific peptide-MHC I tetramers. Overall, these data suggest that CD8+ T cell targeting of certain specific Nef epitopes contributes to HIV suppression. These data suggest that a re-evaluation of the potential use of Nef in HIV T-cell vaccine candidates would be justified.
Use of antiretroviral-based microbicides for HIV-1 prophylaxis could introduce a transmission barrier that inadvertently facilitates the selection of fitter viral variants among incident infections. To investigate this, we assessed the in vitro function of gag-protease and nef sequences from participants who acquired HIV-1 during the CAPRISA 004 1% tenofovir microbicide gel trial.
Methods and Results
We isolated the earliest available gag-protease and nef gene sequences from 83 individuals and examined their in vitro function using recombinant viral replication capacity assays and surface protein downregulation assays, respectively. No major phylogenetic clustering and no significant differences in gag-protease or nef function were observed in participants who received tenofovir gel versus placebo gel prophylaxis.
Results indicate that the partial protective effects of 1% tenofovir gel use in the CAPRISA 004 trial were not offset by selection of transmitted/early HIV-1 variants with enhanced Gag-Protease or Nef fitness.
The potential contribution of HLA-A alleles to viremic control in chronic HIV type 1 (HIV-1) infection has been relatively understudied compared with HLA-B. In these studies, we show that HLA-A*7401 is associated with favorable viremic control in extended southern African cohorts of >2100 C-clade–infected subjects. We present evidence that HLA-A*7401 operates an effect that is independent of HLA-B*5703, with which it is in linkage disequilibrium in some populations, to mediate lowered viremia. We describe a novel statistical approach to detecting additive effects between class I alleles in control of HIV-1 disease, highlighting improved viremic control in subjects with HLA-A*7401 combined with HLA-B*57. In common with HLA-B alleles that are associated with effective control of viremia, HLA-A*7401 presents highly targeted epitopes in several proteins, including Gag, Pol, Rev, and Nef, of which the Gag epitopes appear immunodominant. We identify eight novel putative HLA-A*7401–restricted epitopes, of which three have been defined to the optimal epitope. In common with HLA-B alleles linked with slow progression, viremic control through an HLA-A*7401–restricted response appears to be associated with the selection of escape mutants within Gag epitopes that reduce viral replicative capacity. These studies highlight the potentially important contribution of an HLA-A allele to immune control of HIV infection, which may have been concealed by a stronger effect mediated by an HLA-B allele with which it is in linkage disequilibrium. In addition, these studies identify a factor contributing to different HIV disease outcomes in individuals expressing HLA-B*5703.
Exclusive breastfeeding has been associated with a reduced risk of late vertical HIV transmission as compared to an infant diet composed of breast milk mixed with supplemental foods or liquids. Hypothesized mechanisms include increased infectivity of breast milk from mothers who practice mixed breastfeeding (MBF), or mechanisms such as increased gastrointestinal permeability in the infant caused by mixed feeding. It has been proposed that MBF may result in subclinical mastitis and higher breast milk HIV titers. However, little is known about the relationship between feeding strategy and breast milk viral load. We measured the HIV-1 concentration in breast milk in a sub-cohort of women enrolled in a mother-to-child HIV transmission prevention trial (the "Mashi" study). We report no observed relationship between MBF and measured breast milk viral RNA load. Our findings suggest that the increased transmission risk associated with higher breast milk HIV-1 RNA during MBF is unlikely.
exclusive; mixed; PMTCT; breast feeding; vertical HIV transmission
The CAPRISA004 trial demonstrated reduction of sexual HIV-1 acquisition in women using a vaginal microbicide containing tenofovir. A better understanding of the consequences of antiretroviral-containing microbicides for immune responses in individuals with intercurrent HIV-1 infection is needed for future trials combining the use of microbicides with HIV-1 vaccines. Investigation of immune responses in women who acquired HIV-1 whilst using tenofovir gel showed significantly higher (p=0.01) Gag-specific IFNγ+ CD4+ T-cell responses. The use of tenofovir containing gel around the time of infection can modulate HIV-1 immunity, and these immunological changes need to be considered in future trials combining vaccines and microbicides.
HIV-1; vaginal microbicide; tenofovir; HIV-1-specific CD4+ T cell help
The strongest genetic influence on immune control in HIV-1 infection is the HLA class I genotype. Rapid disease progression in B-clade infection has been linked to HLA-B*35 expression, in particular to the less common HLA-B*3502 and HLA-B*3503 subtypes but also to the most prevalent subtype, HLA-B*3501. In these studies we first demonstrated that whereas HLA-B*3501 is associated with a high viral set point in two further B-clade-infected cohorts, in Japan and Mexico, this association does not hold in two large C-clade-infected African cohorts. We tested the hypothesis that clade-specific differences in HLA associations with disease outcomes may be related to distinct targeting of critical CD8+ T-cell epitopes. We observed that only one epitope was significantly targeted differentially, namely, the Gag-specific epitope NPPIPVGDIY (NY10, Gag positions 253 to 262) (P = 2 × 10−5). In common with two other HLA-B*3501-restricted epitopes, in Gag and Nef, that were not targeted differentially, a response toward NY10 was associated with a significantly lower viral set point. Nonimmunogenicity of NY10 in B-clade-infected subjects derives from the Gag-D260E polymorphism present in ∼90% of B-clade sequences, which critically reduces recognition of the Gag NY10 epitope. These data suggest that in spite of any inherent HLA-linked T-cell receptor repertoire differences that may exist, maximizing the breadth of the Gag-specific CD8+ T-cell response, by the addition of even a single epitope, may be of overriding importance in achieving immune control of HIV infection. This distinction is of direct relevance to development of vaccines designed to optimize the anti-HIV CD8+ T-cell response in all individuals, irrespective of HLA type.
Certain immune-driven mutations in HIV-1, such as those arising in p24Gag, decrease viral replicative capacity. However, the intersubtype differences in the replicative consequences of such mutations have not been explored. In HIV-1 subtype B, the p24Gag M250I mutation is a rare variant (0.6%) that is enriched among elite controllers (7.2%) (P = 0.0005) and appears to be a rare escape variant selected by HLA-B58 supertype alleles (P < 0.01). In contrast, in subtype C, it is a relatively common minor polymorphic variant (10 to 15%) whose appearance is not associated with a particular HLA allele. Using site-directed mutant viruses, we demonstrate that M250I reduces in vitro viral replicative capacity in both subtype B and subtype C sequences. However, whereas in subtype C downstream compensatory mutations at p24Gag codons 252 and 260 reduce the adverse effects of M250I, fitness costs in subtype B appear difficult to restore. Indeed, patient-derived subtype B sequences harboring M250I exhibited in vitro replicative defects, while those from subtype C did not. The structural implications of M250I were predicted by protein modeling to be greater in subtype B versus C, providing a potential explanation for its lower frequency and enhanced replicative defects in subtype B. In addition to accounting for genetic differences between HIV-1 subtypes, the design of cytotoxic-T-lymphocyte-based vaccines may need to account for differential effects of host-driven viral evolution on viral fitness.
Genetic variation within the HLA-B locus has the strongest impact on HIV disease progression of any polymorphisms within the human genome. However, identifying the exact mechanism involved is complicated by several factors. HLA-Bw4 alleles provide ligands for NK cells and for CD8 T cells, and strong linkage disequilibrium between HLA class I alleles complicates the discrimination of individual HLA allelic effects from those of other HLA and non-HLA alleles on the same haplotype. Here, we exploit an experiment of nature involving two recently diverged HLA alleles, HLA-B*42:01 and HLA-B*42:02, which differ by only a single amino acid. Crucially, they occur primarily on identical HLA class I haplotypes and, as Bw6 alleles, do not act as NK cell ligands and are therefore largely unconfounded by other genetic factors. We show that in an outbred cohort (n = 2,093) of HIV C-clade-infected individuals, a single amino acid change at position 9 of the HLA-B molecule critically affects peptide binding and significantly alters the cytotoxic T lymphocyte (CTL) epitopes targeted, measured directly ex vivo by gamma interferon (IFN-γ) enzyme-linked immunospot (ELISPOT) assay (P = 2 × 10−10) and functionally through CTL escape mutation (P = 2 × 10−8). HLA-B*42:01, which presents multiple Gag epitopes, is associated with a 0.52 log10 lower viral-load set point than HLA-B*42:02 (P = 0.02), which presents no p24 Gag epitopes. The magnitude of this effect from a single amino acid difference in the HLA-A*30:01/B*42/Cw*17:01 haplotype is equivalent to 75% of that of HLA-B*57:03, the most protective HLA class I allele in this population. This naturally controlled experiment represents perhaps the clearest demonstration of the direct impact of a particular HIV-specific CTL on disease control.
HIV-1 drug resistance monitoring in resource-poor settings is crucial due to limited drug alternatives. Recent reports of the increased prevalence of CXCR4 usage in subtype C infections may have implications for CCR5 antagonists in therapy. We investigated the prevalence of drug resistance mutations and CXCR4 coreceptor utilization of viruses from HIV-1 subtype C-infected children. Fifty-one children with virological failure during highly active antiretroviral therapy (HAART) and 43 HAART-naive children were recruited. Drug resistance genotyping and coreceptor utilization assessment by phenotypic and genotypic methods were performed. At least one significant drug resistance mutation was present in 85.4% of HAART-failing children. Thymidine analogue mutations (TAMs) were detected in 58.5% of HAART-failing children and 39.0% had ≥3 TAMs. CXCR4 (X4) or dual (R5X4)/mixed (R5, X4) (D/M)-tropic viruses were found in 54.3% of HAART-failing and 9.4% of HAART-naive children (p<0.0001); however, the HAART-failing children were significantly older (p<0.0001). In multivariate logistic regression, significant predictors of CXCR4 usage included antiretroviral treatment, older age, and lower percent CD4+ T cell counts. The majority of genotypic prediction tools had low sensitivity (≤65.0%) and high specificity (≥87.5%) for predicting CXCR4 usage. Extensive drug resistance, including the high percentage of TAMs found, may compromise future drug choices for children, highlighting the need for improved treatment monitoring and adherence counseling. Additionally, the increased prevalence of X4/D/M viruses in HAART-failing children suggests limited use of CCR5 antagonists in salvage therapy. Enhanced genotypic prediction tools are needed as current tools are not sensitive enough for predicting CXCR4 usage.
Interleukin-10 (IL-10) is an immunoregulatory cytokine that influences the clinical outcome of chronic viral infections. Results show that IL-10-promoter genetic variants influence human immunodeficiency virus-1 pathogenesis possibly via regulating IL-10 plasma levels and the breadth of CD8+ T-cell immune responses.
Background. Interleukin-10 (IL-10) is a potent immunoregulatory cytokine. IL-10-promoter polymorphisms have been shown to affect human immunodeficiency virus type 1 (HIV-1) clinical outcomes but the underlying mechanisms are poorly understood.
Methods. We investigated the relationship between IL-10-promoter variants, plasma cytokine levels, immune responses and markers of disease outcome in antiretroviral-naïve HIV-1 chronically infected individuals from South Africa. Two IL-10-promoter single nucleotide polymorphisms (SNPs) were genotyped in 451 participants. Baseline plasma levels of select cytokines were measured for 112 individuals. Viral load, CD4+ T-cell counts and HIV-1-specific interferon-gamma CD8+ T-cell immune responses were measured at baseline. CD4+ T-cell counts were measured longitudinally and rates of CD4+ T-cell decline computed for 300 study subjects.
Results. The minor IL-10-1082G and -592A variants occurred at frequencies of 0.31 and 0.34, respectively. The -592AA genotype associated significantly with attenuated loss of CD4+ T cells (P = .0496). Individuals possessing -1082GG had significantly higher IL-10 levels compared to -1082AA/AG (P = .0006). The -592AA genotype was associated with greater breadth of virus-specific CD8+ T-cell responses compared to CC and CA (P = .002 and .004 respectively).
Conclusions. IL-10-promoter variants may influence the rate of HIV-1 disease progression by regulating IL-10 levels and the breadth of CD8+ T-cell immune responses.
This study addressed two questions: (1) What fraction of individuals maintain a sustained high HIV-1 RNA load after the acute HIV-1C infection peak? and (2) How long is a high HIV-1 RNA load maintained after acute HIV-1C infection in this subpopulation?
Plasma HIV-1 RNA dynamics were studied in 77 subjects with primary HIV-1C infection from African cohorts in Gaborone, Botswana, and Durban, South Africa. HIV-infected individuals who maintained mean viral load of ≥ 100,000 (5.0 log10) copies/ml after 100 days post-seroconversion (p/s) were termed Extended High Viremics. Individuals were followed longitudinally for a median (IQR) of 573 (226;986) days p/s.
The proportion of Extended High Viremics was 34% (95% CI: 23%–44%) during the period 100 to 300 days p/s and 19% (95% CI: 9%–29%) over the period of 200 to 400 days p/s. The median (IQR) duration of HIV-1 RNA load ≥ 100,000 copies/ml among Extended High Viremics was 271 (188;340) days p/s. For the subset with average viral load ≥ 100,000 copies/ml during 200–400 days p/s, the median (IQR) duration was 318 (282;459) days. The Extended High Viremics had a significantly shorter time to CD4 decline to 350 cells/μl (median: 88 vs. 691 days p/s for those not designated as Extended High Viremics; p<0.0001, Gehan-Wilcoxon test).
A high proportion of Extended High Viremics – individuals maintaining high plasma HIV-1 RNA load after acute infection – has been identified during primary HIV-1 subtype C infection. These Extended High Viremics likely contribute disproportionately to HIV-1 incidence.
HIV-1 subtype C; primary infection; viral HIV-1 RNA load; Southern Africa; HIV-1 transmission
Recent reports suggest that Natural Killer (NK) cells may modulate pathogenesis of primary HIV-1 infection. However, HIV dysregulates NK-cell responses. We dissected this bi-directional relationship to understand how HIV impacts NK-cell responses during primary HIV-1 infection.
Paired samples from 41 high-risk, initially HIV-uninfected CAPRISA004 participants were analysed prior to HIV acquisition, and during viraemic primary HIV-1 infection. At the time of sampling post-infection five women were seronegative, 11 women were serodiscordant, and 25 women were seropositive by HIV-1 rapid immunoassay. Flow cytometry was used to measure NK and T-cell activation, NK-cell receptor expression, cytotoxic and cytokine-secretory functions, and trafficking marker expression (CCR7, α4β7). Non-parametric statistical tests were used. Both NK cells and T-cells were significantly activated following HIV acquisition (p = 0.03 and p<0.0001, respectively), but correlation between NK-cell and T-cell activation was uncoupled following infection (pre-infection r = 0.68;p<0.0001; post-infection, during primary infection r = 0.074;p = 0.09). Nonetheless, during primary infection NK-cell and T-cell activation correlated with HIV viral load (r = 0.32'p = 0.04 and r = 0.35;p = 0.02, respectively). The frequency of Killer Immunoglobulin-like Receptor-expressing (KIRpos) NK cells increased following HIV acquisition (p = 0.006), and KIRpos NK cells were less activated than KIRneg NK cells amongst individuals sampled while seronegative or serodiscordant (p = 0.001;p<0.0001 respectively). During HIV-1 infection, cytotoxic NK cell responses evaluated after IL-2 stimulation alone, or after co-culture with 721 cells, were impaired (p = 0.006 and p = 0.002, respectively). However, NK-cell IFN-y secretory function was not significantly altered. The frequency of CCR7+ NK cells was elevated during primary infection, particularly at early time-points (p<0.0001).
Analyses of immune cells before and after HIV infection revealed an increase in both NK-cell activation and KIR expression, but reduced cytotoxicity during acute infection. The increase in frequency of NK cells able to traffic to lymph nodes following HIV infection suggests that these cells may play a role in events in secondary lymphoid tissue.
Natural killer (NK) cells play a critical role in the control of HIV-1 infection, and NK cells that respond to HIV-1 peptides have been recently described. However, the mechanisms by which NK cells recognize HIV-1 antigens are not fully understood. We investigated NK cell activation in response to HIV-1 peptides during early and chronic HIV-1 clade B infection using a whole-blood assay and multiparameter flow cytometry. Antibody-mediated NK cell activation in response to HIV-1 peptides was not detected in HIV-1-uninfected individuals. In contrast, 79% of individuals with chronic infection and 22% of individuals with early infection had detectable gamma interferon (IFN-γ) NK cell responses to HIV-1 antigens (P < 0.00001). IFN-γ- and tumor necrosis factor alpha (TNF-α)-producing NK cells most frequently targeted Env gp120 (median of 4% and range of 0 to 31% of all NK cells). NK cells rarely targeted other HIV-1 proteins such as Gag, Pol, and Nef. Antibody-mediated NK cell responses to peptides mapped predominantly to Env protein, required the presence of plasma or plasma IgG, and resulted in lower CD16 expression on NK cells, suggesting an antibody-mediated activation of NK cells. Further studies are needed to assess the consequences of these antibody-mediated NK cell responses for HIV-1 disease progression and vaccine-induced protection from infection.
It is widely documented that a complete switch from the predominant CCR5 (R5) to CXCR4 (X4) phenotype is less common for HIV-1 subtype C (HIV-1C) compared to other major subtypes. We investigated whether dualtropic HIV-1C isolates represented dualtropic, mixed R5 and X4 clones or both. Thirty of 35 functional HIV-1 env clones generated by bulk PCR amplification from peripheral blood mononuclear cells (PBMCs) infected with seven dualtropic HIV-1C isolates utilized CXCR4 exclusively. Five of 35 clones displayed dualtropism. Endpoint dilution of one isolate did not yield a substantial proportion of R5-monotropic env clones. Sequence-based predictive algorithms showed that env sequences from PBMCs, CXCR4 or CCR5-expressing cell lines were indistinguishable and all possessed X4/dualtropic characteristics. We describe HIV-1C CXCR4-tropic env sequence features. Our results suggest a dramatic loss of CCR5 monotropism as dualtropism emerges in HIV-1C which has important implications for the use of coreceptor antagonists in therapeutic strategies for this subtype.
HIV-1 subtype C; Dualtropism; Coreceptor usage; Envelope; V3 loop; CXCR4
Drug resistance poses a significant challenge for the successful application of highly active antiretroviral therapy (HAART) globally. Furthermore, emergence of HIV-1 isolates that preferentially utilize CXCR4 as a coreceptor for cell entry, either as a consequence of natural viral evolution or HAART use may compromise the efficacy of CCR5 antagonists as alternative antiviral therapy.
We sequenced the pol gene of viruses from 45 individuals failing at least six months of HAART in Durban, South Africa to determine the prevalence and patterns of drug resistance mutations. Coreceptor usage profiles of these viruses and those from 45 HAART-naive individuals were analyzed using phenotypic and genotypic approaches.
Ninety-five percent of HAART-failing patients had at least one drug resistance mutation. Thymidine analog mutations (TAMs) were present in 55% of patients with 9% of individuals possessing mutations indicative of the TAM1 pathway, 44% had TAM2 while 7% had mutations common to both pathways. Sixty percent of HAART-failing subjects had X4/dual//mixed-tropic viruses compared to 30% of HAART-naïve subjects (p<0.02). Genetic coreceptor usage prediction algorithms correlated with phenotypic results with 60% of samples from HAART-failing subjects predicted to possess CXCR4-using (X4/dual/mixed viruses) versus 15% of HAART-naïve patients.
The high proportion of TAMs and X4/dual/mixed HIV-1 viruses among patients failing therapy highlight the need for intensified monitoring of patients taking HAART and the problem of diminished drug options (including CCR5 antagonists) for patients failing therapy in resource-poor settings.
Coreceptor usage; viral tropism; antiretroviral drug resistance; HAART-failing patients; HAART-naïve patients
HLA class I genotype is a major determinant of the outcome of HIV infection, and the impact of certain alleles on HIV disease outcome is well studied. Recent studies have demonstrated that certain HLA class I alleles that are in linkage disequilibrium, such as HLA-A*74 and HLA-B*57, appear to function co-operatively to result in greater immune control of HIV than mediated by either single allele alone. We here investigate the extent to which HLA alleles - irrespective of linkage disequilibrium - function co-operatively.
We here refined a computational approach to the analysis of >2000 subjects infected with C-clade HIV first to discern the individual effect of each allele on disease control, and second to identify pairs of alleles that mediate ‘co-operative additive’ effects, either to improve disease suppression or to contribute to immunological failure. We identified six pairs of HLA class I alleles that have a co-operative additive effect in mediating HIV disease control and four hazardous pairs of alleles that, occurring together, are predictive of worse disease outcomes (q<0.05 in each case). We developed a novel ‘sharing score’ to quantify the breadth of CD8+ T cell responses made by pairs of HLA alleles across the HIV proteome, and used this to demonstrate that successful viraemic suppression correlates with breadth of unique CD8+ T cell responses (p = 0.03).
These results identify co-operative effects between HLA Class I alleles in the control of HIV-1 in an extended Southern African cohort, and underline complementarity and breadth of the CD8+ T cell targeting as one potential mechanism for this effect.
In the absence of HIV-1 Vif, cellular cytosine deaminases such as APOBEC3G, inhibit the virus by inducing hypermutations on viral DNA, among other mechanisms of action. We investigated the association of APOBEC3G mRNA levels and APOBEC3G genetic variants on HIV-1 susceptibility, and early disease pathogenesis using viral load and CD4+ T cell counts as outcomes.
Study subjects were 250 South African females at high risk for HIV-1C infection. We used quantitative real-time PCR to measure the expression of APOBEC3G in HIV−ve and HIV+ve primary infection samples. APOBEC3G variants were identified by DNA re-sequencing and TaqMan genotyping.
We found no correlation between APOBEC3G expression levels and plasma viral loads (r=0.053, p=0.596) or CD4+ T cell counts (r=0.030, p=0.762) in 32 seroconverters. However, APOBEC3G expression levels were significantly higher in HIV−ve individuals compared to HIV+ve individuals (p<0.0001), including matched pre- and post infection samples from the same individuals (n=13, p<0.0001). 25 single nucleotide polymorphisms (SNPs), nine of which were novel, were identified within APOBEC3G by re-sequencing followed by genotyping of 168 individuals. The H186R mutation, a codon changing variant in exon 4, was associated with high viral loads (p=0.0097) and decreased CD4+ T cell levels (p=0.0081).
These data suggest that APOBEC3G transcription is rapidly downregulated upon HIV-1 infection. During primary infection, APOBEC3G expression levels in PBMCs do not correlate with viral loads or CD4+ T cell counts. However, structural variation of APOBEC3G may significantly affect early HIV-1 pathogenesis, although the mechanism remains unclear and warrants further investigation.
APOBEC3G; HIV-1 C; Primary Infection; mRNA Expression; Polymorphisms; Host proteins
LEDGF/p75, encoded by the PSIP1 gene, interacts with HIV-1 integrase and targets HIV-1 integration into active genes. We investigated the influence of polymorphisms in PSIP1 on HIV-1 acquisition and disease progression in black South Africans.
Integrase binding domain (IBD) of LEDGF/p75 was sequenced in 126 participants. Four haplotype tagging SNPs, SNP1-SNP4 and one exonic SNP, SNP5 were genotyped in 195 HIV-1 seronegative, 52 primary and 403 chronically infected individuals using TaqMan assays. LEDGF/p75 expression was quantified by real-time RT-PCR. The impact of Q472L mutation on the interaction with HIV-1 IN was measured by AlphaScreen.
rs2277191A was more frequent among seropositives (p=0.06, Fisher's exact test), and among individuals followed longitudinally trended towards association with higher likelihood of HIV-1 acquisition (RH=2.21, p=0.08; Cox model) and it was also associated with rapid disease progression (RH=5.98, p=0.04; Cox model). rs12339417C was associated with slower decline of CD4+ T cell (p=0.02) and lower levels of LEDGF/p75 (p<0.01). Seroconverters had higher preinfection levels of LEDGF/p75 (p<0.01) but levels decreased after HIV infection (p=0.02).
Genetic variants of PSIP1 may affect HIV-1 outcomes. Further studies are needed to confirm the effect of genetic variation of PSIP1 on HIV-1 pathogenesis in different cohorts.
Background. Human immunodeficiency virus type 1 (HIV-1)–specific CD8+ responses contribute to the decline in acute peak viremia following infection. However, data on the relative immunogenicity of CD8+ T-cell epitopes during and after acute viremia are lacking.
Methods. We characterized CD8+ T-cell responses in 20 acutely infected, antiretroviral-naive individuals with HIV-1 subtype C infection using the interferon-γ enzyme-linked immunosorbent spot assay. Eleven of these had not fully seroconverted at the time of analysis. Viruses from plasma were sequenced within defined cytotoxic T-lymphocyte (CTL) cell epitopes for selected subjects.
Results. At approximately 28 days after estimated initial infection, CD8+ T-cell responses were directed against an average of 3 of the 410 peptides tested (range, 0–6); 2 individuals had no detectable responses at this time. At 18 weeks, the average number of peptides targeted had increased to 5 (range 0–11). Of the 56 optimal Gag CTL epitopes sequenced, 31 were wild-type in the infecting viruses, but only 11 of 31 elicited measurable CD8+ T-cell responses.
Conclusions. These data demonstrate that the majority of CD8+ responses are not elicited during acute HIV infection despite the presence of the cognate epitope in the infecting strain. There is a need to define factors that influence lack of induction of effective immune responses and the parameters that dictate immunodominance in acute infection.
HIV-1 attenuation resulting from immune escape mutations selected in Gag may contribute to slower disease progression in HIV-1-infected individuals expressing certain HLA class I alleles. We previously showed that the protective allele HLA-B*81 and the HLA-B*81-selected Gag T186S mutation are strongly associated with a lower viral replication capacity of recombinant viruses encoding Gag-protease derived from individuals chronically infected with HIV-1 subtype C. In the present study, we directly tested the effect of this mutation on viral replication capacity. In addition, we investigated potential compensatory effects of various polymorphisms, including other HLA-B*81-associated mutations that significantly covary with the T186S mutation. Mutations were introduced into a reference subtype B backbone and into patient-derived subtype C sequences in subtype B and C backbones by site-directed mutagenesis. The exponential-phase growth of mutant and wild-type viruses was assayed by flow cytometry of a green fluorescent protein reporter T cell line or by measurement of HIV-1 reverse transcriptase activity in culture supernatants. Engineering of the T186S mutation alone into all patient-derived subtype C sequences failed to yield replication-competent viruses, while in the subtype B sequence, the T186S mutation resulted in impaired replication capacity. Only the T186S mutation in combination with the T190I mutation yielded replication-competent viruses for all virus backbones tested; however, these constructs replicated slower than the wild type, suggesting that only partial compensation is mediated by the T190I mutation. Constructs encoding the T186S mutation in combination with other putative compensatory mutations were attenuated or defective. These results suggest that the T186S mutation is deleterious to HIV-1 subtype C replication and likely requires complex compensatory pathways, which may contribute to the clinical benefit associated with HLA-B*81.
Pediatric HIV-1 infection is characterized by rapid disease progression and without antiretroviral therapy (ART), more than 50% of infected children die by the age of 2 years. However, a small subset of infected children progresses slowly to disease in the absence of ART. This study aimed to identify functional characteristics of HIV-1-specific T cell responses that distinguish children with rapid and slow disease progression. Fifteen perinatally HIV-infected children (eight rapid and seven slow progressors) were longitudinally studied to monitor T cell polyfunctionality. HIV-1-specific interferon (IFN)-γ+ CD8+ T cell responses gradually increased over time but did not differ between slow and rapid progressors. However, polyfunctional HIV-1-specific CD8+ T cell responses, as assessed by the expression of four functions (IFN-γ, CD107a, TNF-α, MIP-1β), were higher in slow compared to rapid progressors (p=0.05) early in infection, and was associated with slower subsequent disease progression. These data suggest that the quality of the HIV-specific CD8+ T cell response is associated with the control of disease in children as has been shown in adult infection.
The genetic polymorphism that has the greatest impact on immune control of human immunodeficiency virus (HIV) infection is expression of HLA-B*57. Understanding of the mechanism for this strong effect remains incomplete. HLA-B*57 alleles and the closely related HLA-B*5801 are often grouped together because of their similar peptide-binding motifs and HIV disease outcome associations. However, we show here that the apparently small differences between HLA-B*57 alleles, termed HLA-B*57 micropolymorphisms, have a significant impact on immune control of HIV. In a study cohort of >2,000 HIV C-clade-infected subjects from southern Africa, HLA-B*5703 is associated with a lower viral-load set point than HLA-B*5702 and HLA-B*5801 (medians, 5,980, 15,190, and 19,000 HIV copies/ml plasma; P = 0.24 and P = 0.0005). In order to better understand these observed differences in HLA-B*57/5801-mediated immune control of HIV, we undertook, in a study of >1,000 C-clade-infected subjects, a comprehensive analysis of the epitopes presented by these 3 alleles and of the selection pressure imposed on HIV by each response. In contrast to previous studies, we show that each of these three HLA alleles is characterized both by unique CD8+ T-cell specificities and by clear-cut differences in selection pressure imposed on the virus by those responses. These studies comprehensively define for the first time the CD8+ T-cell responses and immune selection pressures for which these protective alleles are responsible. These findings are consistent with HLA class I alleles mediating effective immune control of HIV through the number of p24 Gag-specific CD8+ T-cell responses generated that can drive significant selection pressure on the virus.
It is unknown whether favorable HLA class II alleles may attenuate HIV-1 through selection pressure in a manner similar to that of protective HLA class I alleles. We investigated the relationship between HLA class II alleles and in vitro replication capacities of recombinant viruses encoding HIV-1 subtype C Gag-protease from chronically infected individuals. No associations were found between individual alleles and lower replication capacity, suggesting no significant HIV-1 attenuation by HLA class II-restricted Gag-specific CD4+ T cell immune pressure.
Monitoring of latent Mycobacterium tuberculosis infection may prevent disease. We tested an ESAT-6 and CFP-10-specific IFN-γ Elispot assay (RD1-Elispot) on 163 HIV-infected individuals living in a TB-endemic setting. An RD1-Elispot was performed every 3 months for a period of 3–21 months. 62% of RD1-Elispot negative individuals were positive by cultured Elispot. Fluctuations in T cell response were observed with rates of change ranging from −150 to +153 spot-forming cells (SFC)/200,000 PBMC in a 3-month period. To validate these responses we used an RD1-specific real time quantitative PCR assay for monokine-induced by IFN-γ (MIG) and IFN-γ inducible protein-10 (IP10) (MIG: r = 0.6527, p = 0.0114; IP-10: r = 0.6967, p = 0.0056; IP-10+MIG: r = 0.7055, p = 0.0048). During follow-up 30 individuals were placed on ARVs and 4 progressed to active TB. Fluctuations in SFC did not correlate with CD4 count, viral load, treatment initiation, or progression to active TB. The RD1-Elispot appears to have limited value in this setting.
We investigated the association of polymorphisms in CCR5, the major human immunodeficiency virus (HIV)–1 coreceptor, and copy number of its potent ligand CCL3L1 with tuberculosis in 298 individuals from Colombia. The CCR5-HHD haplotype, a known genetic determinant of increased susceptibility to HIV-AIDS, and a high copy number of CCL3L1, a known genetic determinant of enhanced CCL3/CCL3L1 chemokine expression, each associated with presence of tuberculosis. Furthermore, CCR5-HHD was associated with higher CCR5 gene and surface expression. These results substantiate the strong link between the pro-inflammatory effects of CCR5 and its ligands with active tuberculosis and suggest that chemokine-chemokine receptor genetic determinants may influence tuberculosis in addition to HIV/AIDS.