Immune escape mutations selected by neutralizing antibodies in natural HIV-1 infection can alter coreceptor usage repertoire of the transmitted/founder virus

The ability of HIV-1 to evade neutralizing antibodies (NAbs) in vivo is well demonstrated, but the impact of NAb escape mutations on HIV-1 phenotype other than immune escape itself has rarely been studied. Here, we show that immune escape mutations selected by V3-glycan specific NAbs in vivo can alter the coreceptor usage repertoire of the transmitted/founder (T/F) HIV-1. In a participant developed V3-glycan NAb response, naturally selected mutations at the V3 N301 and N332 glycan sites abrogated CCR8 usage while conferred APJ usage on the cognate T/F strain. Mutations at the N301 glycan also impaired CCR3 usage and partially compromised the efficiency in using CCR5, which could be fully restored by a single escape mutation at the N332 glycan site. Our study demonstrates the link between NAb escape and coreceptor usage alteration in natural HIV-1 infection and indicates that NAb response could drive virus entry tropism evolution in vivo.

Existence of Replication-Competent Minor Variants with Different Coreceptor Usage in Plasma from HIV-1-Infected Individuals

ABSTRACT Cell entry by HIV-1 is mediated by its principal receptor, CD4, and a coreceptor, either CCR5 or CXCR4, with viral envelope glycoprotein gp120. Generally, CCR5-using HIV-1 variants, called R5, predominate over most of the course of infection, while CXCR4-using HIV-1 variants (variants that utilize both CCR5 and CXCR4 [R5X4, or dual] or CXCR4 alone [X4]) emerge at late-stage infection in half of HIV-1-infected individuals and are associated with disease progression. Although X4 variants also appear during acute-phase infection in some cases, these variants apparently fall to undetectable levels thereafter. In this study, replication-competent X4 variants were isolated from plasma of drug treatment-naive individuals infected with HIV-1 strain CRF01_AE, which dominantly carries viral RNA (vRNA) of R5 variants. Next-generation sequencing (NGS) confirmed that sequences of X4 variants were indeed present in plasma vRNA from these individuals as a minor population. On the other hand, in one individual with a mixed infection in which X4 variants were dominant, only R5 replication-competent variants were isolated from plasma. These results indicate the existence of replication-competent variants with different coreceptor usage as minor populations. IMPORTANCE The coreceptor switch of HIV-1 from R5 to CXCR4-using variants (R5X4 or X4) has been observed in about half of HIV-1-infected individuals at late-stage infection with loss of CD4 cell count and disease progression. However, the mechanisms that underlie the emergence of CXCR4-using variants at this stage are unclear. In the present study, CXCR4-using X4 variants were isolated from plasma samples of HIV-1-infected individuals that dominantly carried vRNA of R5 variants. The sequences of the X4 variants were detected as a minor population using next-generation sequencing. Taken together, CXCR4-using variants at late-stage infection are likely to emerge when replication-competent CXCR4-using variants are maintained as a minor population during the course of infection. The present study may support the hypothesis that R5-to-X4 switching is mediated by the expansion of preexisting X4 variants in some cases.

Emergence of HIV-1 C/A1 and C/A1/D circulating recombinant forms, and dominance of subtype C and R5 use from whole genome sequence analysis in Addis Ababa

Background The HIV pandemic in Ethiopia is dominated by subtype C with sporadic A and D epidemiology. The presence of subtypes A and D may result in emergence of recombinant viruses, and increase the genetic diversity that makes monitoring the HIV epidemic, and the development of vaccines and therapeutics difficult. This study is aimed at determining subtypes, circulating recombinant forms (CRFs), and the dominant coreceptor use in Addis Ababa, Ethiopia. Methods Participants with a range of purposely selected CD4+ T-cell counts were included. Chi-square and Mann-Whitney tests were used. Whole genome next-generation sequencing (NGS) of HIV was performed using a PCR amplification method and Illumina MiSeq. Subtyping and scanning of recombination were done by the REGA subtyping tool version 3.0. Prediction of coreceptor usage was performed using Geno2Pheno clonal-model and PhenoSeq. Signature amino acids and positive charges were also used in the tropism prediction. Phylogenetic analyses were conducted with MEGA version 6 using maximum likelihood with the neighbor-joining (N-J) methods. Results Sixty participants were included with a median age of 34.5 [interquartile range (IQR) 30.0-40.0]. Seven (11.7%) of the study participants were at WHO clinical stage 3/4 and 13 (21.7%) were at AIDS stage with CD4+ T cell count <200 cells/uL. Among the total 60 HIV genomes sequenced, 49 were subtype C (81.7%), one was subtype A1 (1.7%), six were recombinant C/A1 (10%), three were recombinant C/A1/D (5.0%), and one was unassigned. From 50 of the sequences where coreceptor usage was determined by PhenoSeq, 44 (88.0%) were CCR5-tropic and six (12.0%) used CXCR4. Conclusion The study confirmed that the dominant subtype in Addis Ababa is HIV-1 subtype C. In addition, HIV-1 subtype A1, CRFs C/A1 and C/A1/D were also identified. The dominance of R5-tropic viruses was detected and these were associated with a higher CD4 T-cell count and lower viral load. Further studies on HIV subtypes and CRFs will be essential to fully understand HIV/AIDS epidemiology. In addition, the tropism information is important in Ethiopia if the use of the co-receptor antagonist maraviroc is planned.

PO 8411 MOLECULAR CHARACTERISATION OF GP41 AND GP120 V3 LOOP IN HIV-1C PATIENTS FAILING SALVAGE THERAPY IN BOTSWANA

BackgroundTriple class drug-resistant HIV-1 infection remains a global challenge in individuals with extensive antiretroviral treatment (ART) experience, in terms of high mortality and probability of onward transmission. New therapeutic options within old and new drug classes are therefore essential. We determined if patients failing salvage therapy in Botswana are eligible for maraviroc (MVC) and enfuvirtide (T20) viral entry inhibitors based on the coreceptor usage and drug-resistant mutations in envelope gp120 and gp41.MethodsA total of 38 deep salvage patients were included in the analysis. We amplified and sequenced gp41 and V3 regions of HIV-1 envelope. Drug resistance mutations were analysed according to the IAS-USA 2017 reference mutation lists. Coreceptor usage was determined using PSSM and Geno2Pheno using a false-positive rate (FPR) of 10%.ResultsAmong 38 participants, 34 (89%) were successfully sequenced and amplified gp41 and 26 (68%) gp120 V3 loop sequences were obtained. Major T20 mutation G36S was obtained in 1/34 samples (5.8%) within the study population. Polymorphisms I169V(97%), I135L(100%), E151A(70.6%) and N42S(70.6%) were detected in HR1 and HR2 of gp41. CXCR4 coreceptor associated use, mutation L34M in gp41 HR1 was detected in 2 samples (5%). Analysis of coreceptor usage showed (17/26) 65.4% use of CCR5, and a (9/26) 34.6% use of the CXCR4 coreceptor.ConclusionA moderately high proportion of treatment-experienced (deep salvage) participants had CXCR4 coreceptor using strains. The use of maraviroc in Botswana would require coreceptor tropism testing. Non-T20 treatment experience in Botswana reduces the prevalence of the major mutations that confer resistance to the drug. T20 is therefore a potential alternative drug for patients failing salvage therapy in Botswana.