Impact of Cytotoxic T Lymphocyte (CTL) escape mutations in acute/Early HIV-1 Subtype C Infection on Disease Progression

Includes abstract. Includes bibliographical references (leaves 139-162).Most HIV vaccines currently in development aim to protect people from infection or disease by eliciting strong anti-HIV cytotoxic T lymphocyte (CTL) responses. Evolved evasion mutations that undermine host immune responses pose a major challenge to the development of such vaccines. Understanding the mechanisms that selectively favour the emergence of CTL evasion mutations in vivo and the impact of these mutations on both disease progression and long-term HIV evolution will not only contribute to our understanding of HIV pathogenesis, but will also inform vaccine design strategies. This study aimed at investigating CTL escape mutations in HIV-1 Gag and Nef, during the acute and early phases of infection and the impact of these mutations on subsequent disease progression in a cohort of recently HIV-1 subtype C infected females. Of 36 women recruited into the study within 12 weeks of infection (median 6 weeks) and followed for six months, 32 were infected with single viruses. Two participants were infected with epidemiologically unlinked viruses (dual infection), and in a further two individuals the viruses were highly divergent suggestive of dual infection and/or recombination. These individuals were excluded from further analysis as it was difficult to predict CTL escape due to high degrees of diversity between sequences. In the remaining 32 study participants, there was a high frequency of CTL escape with putative escape mutations identified in 21 of 32 individuals (66%). Twelve of these 21 (33%) harboured viruses which developed escape mutations in Gag, and 17 (53%) developed escape mutations in Nef. In the conserved structural protein, p24, potential reversion mutations were more frequent than potential escape mutations. During the first six months of infection whereas potential reversion mutations occurred at low entropy sites, potential escape mutations occurred at high entropy sites. Although there was no detectable association between the timing of escape mutations and disease progression, there was an association between the degree of deviation of the p24 sequence from the subtype-C population consensus (a measure of escape mutation load) and CD4+ counts. Analysis of the earliest sampled viruses from HLA-B*57/B*5801 negative study participants for viral genetic markers associated with disease progression identified two iv polymorphisms, A146X (n = 9) and T242N (n =6), that were associated with improved viral control. The polymorphisms are well-known escape mutations in HLAB* 57/B*5801 restricted epitopes. This suggested transmission of these variants from individuals carrying these alleles. Further evidence that viruses carrying the T242N and/or A146X mutations had been previously passaged through B*57/B*5801 positive individuals came from the fact that the observed T242N mutations reverted to wild type during follow-up. There was no significant change in viral load and CD4+ counts upon reversion of the T242N mutations. In vitro replication assays using chimeric viruses containing gag sequences from one of participants showed that the virus harbouring the T242N mutation was fitter than that carrying the reversion mutation. These viruses harboured other T242N associated compensatory mutations suggesting that these compensatory mutations may themselves carry a fitness cost in the absence of the T242N mutation. This suggests that there possibly exist networks of B*57/B*5801 associated mutations and that reversion of some of these mutations in isolation does not necessarily restore viral fitness. Lastly, the kinetics of CTL escape in HLA-B*5801 positive participants (n = 6) and the impact of escape on disease progression was investigated. CTL escape within B *5801 positive individuals was found to predominantly occur within the TW10 in Gag (n = 4) and KAF9 in Nef (n = 6) epitopes. The emergence of the T242N mutation in TW10 was always preceded by mutations elsewhere in the epitope and was associated with the occurrence of previously described compensatory mutation upstream of the epitope. The targeting of TW10 and the emergence of T242N escape mutations were associated with higher CD4+ counts at 12 months postinfection in the B*5801 positive individuals (p = 0.0231 and p = 0.0282, respectively). Independent of host HLA genotypes, the presence of the A146X and T242X mutations was associated with higher CD4+ counts (p = 0.0495). This study provides some useful insights into HIV-1 subtype C pathogenesis. The notion that CTL escape mutations do not invariably result in less fit viruses is evidenced by the observation that escape was not obviously associated with disease progression in this cohort, while escape mutations in the Gag p24 region within B*5801 positive individuals v in particular, was associated with improved viral control. There is therefore evidently a complex interaction between escape and compensatory mutations and further work is required to identify the impact of compensatory mutations on viral fitness. Overall, this study provides further evidence that vaccines need to elicit responses that specifically target the functionally constrained regions of the HIV proteome

Molecular characterization of XvlNO1, a myo-inositol 1-phosphate synthase gene from Xerophyta viscosa

Includes bibliographical references.Myo-inositol I-phosphate synthase (INO 1) catalyses the conversion of glucose-6-phosphate to myo-inositol I-phosphate, which is subsequently dephosphorylated to myo-inositol. Myo-inositol is a precursor for a number of important metabolites that include membrane components, storage molecules, phytohormones and a variety of osmoprotectants. Xerophyta viscosa Baker (Family Velloziaceae) is a monocotyledonous angiosperm which has the ability to resume full physiological function after desiccation. The full-length cDNA for INO1 from X viscosa was isolated using the RACE technique

Exploring the trends in prevalence of human immunodeficiency virus drug resistance in South Africa over the course of the HIV epidemic

Magister Public Health - MPHBackground: Antiretroviral therapy (ART) was rolled out in South Africa in the public sector in 2004 and the treatment coverage has increased over the years to 56% in 2016. The increased treatment coverage has the potential to increase the level of HIV drug resistance. Drug resistance presents a major challenge to the management of HIV infection through antiretroviral therapy at the population level. The aim of this study was to determine the impact of the public sector antiretroviral therapy rollout on the prevalence of HIV drug resistance in South Africa and the factors associated with drug resistance. Methodology: A cross-sectional analytical study was used to determine the prevalence of drug resistance before and after ART rollout. The study population was HIV infected South Africans (infected between 1996 and 2011) who were not on antiretroviral therapy. The study sample was therapy naïve HIV infected South Africans who participated in published studies conducted between 1996 and 2011. HIV DNA sequences and associated data (participants’ age, gender, geographic location and estimated year of HIV infection) were accessed through the Los Alamos HIV Database. The database contains all HIV DNA sequences and associated data from all published studies and the data was freely accessible. A descriptive analysis was carried out on the data to determine characteristics of the study sample. Drug resistance mutations were detected using Calibrated Population Resistance Program on the Stanford University HIV Drug Resistance database. The output from the Calibrated Population Resistance Program analysis were used to determine the prevalence of drug resistance mutations. Results: There were 1701 DNA sequences obtained from the Los Alamos HIV Database for the three gene regions targeted by ART (reverse transcriptase, protease and integrase). Of these, 604 (35,5%) were for reverse transcriptase, 794 (46,7%) were for protease and 303 (17,8%) were for integrase. There was overrepresentation of DNA sequences from female participants (91%). There was no significant difference in the prevalence of drug resistance mutations between 1996-2004 (before ART rollout) and 2005-2011 (after ART rollout) in all the drug classes. There was also no association between drug resistance and age as well as gender. Conclusion: The data from this study suggest that the public sector rollout of ART did not result in an increase in the prevalence of drug resistance mutations in therapy naïve HIVinfected South Africans. There is need for further studies, which have a wider coverage of the South African population

Early evolution of HLA-associated escape mutations in variable Gag proteins predicts CD4+ decline in HIV-1 subtype C infected women.

CAPRISA, 2017.Abstract available in pdf

Transmission of HIV-1 CTL escape variants provides HLA - mismatched recipients with a survival advantage.

One of the most important genetic factors known to affect the rate of disease progression in HIV-infected individuals is the genotype at the Class I Human Leukocyte Antigen (HLA) locus, which determines the HIV peptides targeted by cytotoxic T-lymphocytes (CTLs). Individuals with HLA-B*57 or B*5801 alleles, for example, target functionally important parts of the Gag protein. Mutants that escape these CTL responses may have lower fitness than the wild-type and can be associated with slower disease progression. Transmission of the escape variant to individuals without these HLA alleles is associated with rapid reversion to wild-type. However, the question of whether infection with an escape mutant offers an advantage to newly infected hosts has not been addressed. Here we investigate the relationship between the genotypes of transmitted viruses and prognostic markers of disease progression and show that infection with HLA-B*57/B*5801 escape mutants is associated with lower viral load and higher CD4+ counts

Killer-cell Immunoglobulin-like Receptor (KIR) gene profiles modify HIV disease course, not HIV acquisition in South African women.

CAPRISA, 2016.Abstract available in PDF file

Brief report: selection of HIV-1 variants with higher transmission potential by 1% tenofovir gel microbicide.

CAPRISA, 2017.Abstract available in pdf