Marcel Tongo

Inaccurate Diagnosis of HIV-1 Group M and O Is a Key Challenge for Ongoing Universal Access to Antiretroviral Treatment and HIV Prevention in Cameroon

Background Increased access to HIV testing is essential in working towards universal access to HIV prevention and treatment in resource-limited countries. We here evaluated currently used HIV diagnostic tests and algorithms in Cameroon for their ability to correctly identify HIV infections. Methods We estimated sensitivity, specificity, and positive and negative predictive values of 5 rapid/simple tests, of which 3 were used by the national program, and 2 fourth generation ELISAs. The reference panel included 500 locally collected samples; 187 HIV -1 M, 10 HIV-1 O, 259 HIV negative and 44 HIV indeterminate plasmas. Results None of the 5 rapid assays and only 1 ELISA reached the current WHO/UNAIDS recommendations on performance of HIV tests of at least 99% sensitivity and 98% specificity. Overall, sensitivities ranged between 94.1% and 100%, while specificities were 88.0% to 98.8%. The combination of all assays generated up to 9% of samples with indeterminate HIV status, because they reacted discordantly with at least one of the different tests. Including HIV indeterminate samples in test efficiency calculations significantly decreased specificities to a range from 77.9% to 98.0%. Finally, two rapid assays failed to detect all HIV-1 group O variants tested, with one rapid test detecting only 2 out of 10 group O specimens. Conclusion In the era of ART scaling-up in Africa, significant proportions of false positive but also false negative results are still observed with HIV screening tests commonly used in Africa, resulting in inadequate treatment and prevention strategies. Depending on tests or algorithms used, up to 6% of HIV-1 M and 80% of HIV-1 O infected patients in Cameroon do not receive ART and adequate counseling to prevent further transmission due to low sensitivities. Also, the use of tests with low specificities could imply inclusion of up to 12% HIV negative people in ART programs and increase budgets in addition to inconveniences caused to patients.

Characterization of HIV-1 gag and nef in Cameroon: further evidence of extreme diversity at the origin of the HIV-1 group M epidemic

BackgroundCameroon, in west central Africa, has an extraordinary degree of HIV diversity, presenting a major challenge for the development of an effective HIV vaccine. Given the continuing need to closely monitor the emergence of new HIV variants in the country, we analyzed HIV-1 genetic diversity in 59 plasma samples from HIV-infected Cameroonian blood donors. Full length HIV gag and nef sequences were generated and phylogenetic analyses were performed.FindingsAll gag and nef sequences clustered within HIV-1M. Circulating recombinant form CRF02_AG predominated, accounting for 50% of the studied infections, followed by clade G (11%), clade D and CRF37_cpx (4% each), and clades A, F, CRF01_AE and CRF36_cpx (2% each). In addition, 22% of the studied viruses apparently had nef and gag genes from viruses belonging to different clades, with the majority (8/10) having either a nef or gag gene derived from CRF02_AG. Interestingly, five gag sequences (10%) and three (5%) nef sequences were neither obviously recombinant nor easily classifiable into any of the known HIV-1M clades.ConclusionThis suggests the widespread existence of highly divergent HIV lineages in Cameroon. While the genetic complexity of the Cameroonian HIV-1 epidemic has potentially serious implications for the design of biomedical interventions, detailed analyses of divergent Cameroonian HIV-1M lineages could be crucial for dissecting the earliest evolutionary steps in the emergence of HIV-1M.

Challenges in the Design of a T Cell Vaccine in the Context of HIV-1 Diversity

The extraordinary variability of HIV-1 poses a major obstacle to vaccine development. The effectiveness of a vaccine is likely to vary dramatically in different populations infected with different HIV-1 subtypes, unless innovative vaccine immunogens are developed to protect against the range of HIV-1 diversity. Immunogen design for stimulating neutralizing antibody responses focuses on “breadth” – the targeting of a handful of highly conserved neutralizing determinants on the HIV-1 Envelope protein that can recognize the majority of viruses across all HIV-1 subtypes. An effective vaccine will likely require the generation of both broadly cross-neutralizing antibodies and non-neutralizing antibodies, as well as broadly cross-reactive T cells. Several approaches have been taken to design such broadly-reactive and cross-protective T cell immunogens. Artificial sequences have been designed that reduce the genetic distance between a vaccine strain and contemporary circulating viruses; “mosaic” immunogens extend this concept to contain multiple potential T cell epitope (PTE) variants; and further efforts attempt to focus T cell immunity on highly conserved regions of the HIV-1 genome. Thus far, a number of pre-clinical and early clinical studies have been performed assessing these new immunogens. In this review, the potential use of these new immunogens is explored.

High Degree of HIV-1 group M Genetic Diversity within Circulating Recombinant Forms: Insight into the Early Events of HIV-1M Evolution.

ABSTRACT The existence of various highly divergent HIV-1 lineages and of recombination-derived sequence tracts of indeterminate origin within established circulating recombinant forms (CRFs) strongly suggests that HIV-1 group M (HIV-1M) diversity is not fully represented under the current classification system. Here we used a fully exploratory screen for recombination on a set of 480 near-full-length genomes representing the full known diversity of HIV-1M. We decomposed recombinant sequences into their constituent parts and then used maximum-likelihood phylogenetic analyses of this mostly recombination-free data set to identify rare divergent sequence lineages that fall outside the major named HIV-1M taxonomic groupings. We found that many of the sequence fragments occurring within CRFs (including CRF04_cpx, CRF06_cpx, CRF11_cpx, CRF18_cpx, CRF25_cpx, CRF27_cpx, and CRF49_cpx) are in fact likely derived from divergent unclassified parental lineages that may predate the current subtypes, even though they are presently identified as derived from currently defined HIV-1M subtypes. Our evidence suggests that some of these CRFs are descended predominantly from what were or are major previously unidentified HIV-1M lineages that were likely epidemiologically relevant during the early stages of the HIV-1M epidemic. The restriction of these divergent lineages to the Congo basin suggests that they were less infectious and/or simply not present at the time and place of the initial migratory wave that triggered the global epidemic. IMPORTANCE HIV-1 group M (HIV-1M) likely spread to the rest of the world from the Congo basin in the mid-1900s (N. R. Faria et al., Science 346:56–61, 2014, http://dx.doi.org/10.1126/science.1256739) and is today the principal cause of the AIDS pandemic. Here, we show that large sequence fragments from several HIV-1M circulating recombinant forms (CRFs) are derived from divergent parental lineages that cannot reasonably be classified within the nine established HIV-1M subtypes. These lineages are likely to have been epidemiologically relevant in the Congo basin at the onset of the epidemic. Nonetheless, they appear not to have undergone the same explosive global spread as other HIV-1M subtypes, perhaps because they were less transmissible. Concerted efforts to characterize more of these divergent lineages could allow the accurate inference and chemical synthesis of epidemiologically key ancestral HIV-1M variants so as to directly test competing hypotheses relating to the viral genetic factors that enabled the present pandemic.

Refined Identification of Neutralization-Resistant HIV-1 CRF02_AG Viruses

ABSTRACT We studied neutralization of CRF02_AG HIV-1-infected plasma samples. In contrast to previous reports, these samples neutralized CRF02_AG viruses better than other viruses. This included six of eight CRF02_AG viruses previously designated resistant (tier 2/3 or 3). Only viruses 253-11 and 278-50 remained highly resistant, but they were sensitive to membrane-proximal external region (MPER)-specific monoclonal antibodies, suggesting neutralization targets for even these viruses. We propose using high-neutralizing-within-subtype samples for evaluation of neutralization resistance of viruses.

Near full-length HIV type 1M genomic sequences from Cameroon Evidence of early diverging under-sampled lineages in the country

Background: Cameroon is the country in which HIV-1 group M (HIV-1M) likely originated and is today a major hotspot of HIV-1M genetic diversity. It remains unclear, however, whether the highly divergent HIV-1M lineages found in this country arose during the earliest phases of the global HIV-1M epidemic, or whether they arose more recently as a result of recombination events between globally circulating HIV-1M lineages. Methodology: To differentiate between these two possibilities, we performed phylogenetic analyses of the near full genome sequences of nine newly sequenced divergent HIV-1M isolates and 15 previously identified, apparently unique recombinant forms (URFs) from Cameroon. Results: Although two of the new genome sequences were clearly classifiable within subtype G, the remaining seven were highly divergent and phylogenetically branched either outside of, or very near the bases of clades containing the well characterised globally circulating viral lineages that they were most closely related to. Recombination analyses further revealed that these divergent viruses were likely complex URFs. We show, however that substantial portions (>1 Kb) of three of the new genome sequences and 15 of the previously characterised Cameroonian URFs have apparently been derived from divergent parental viruses that branch phylogenetically near the bases of the major HIV-1M clades. Conclusions and implications: Our analyses indicate the presence in Cameroon of contemporary descendants of numerous early-diverging HIV-1M lineages. Further efforts to sample and sequence viruses from such lineages could be crucial both for retracing the earliest evolutionary steps during the emergence of HIV-1M in humans, and accurately reconstructing the ancestral sequences of the major globally circulating HIV-1M lineages.

Striking lack of T cell immunodominance in both a multiclade and monoclade HIV-1 epidemic: Implications for vaccine development

Understanding the impact of HIV diversity on immunological responses to candidate immunogens is critical for HIV vaccine development. We investigated the reactivity and immunodominance patterns of HIV-1 consensus group M Gag and Nef in (i) Cameroon, where individuals infected with the predominant CRF02_AG clade were compared with those infected with diverse non-CRF02_AG clades; and (ii) in a multiclade epidemic, namely Cameroon, compared with a monoclade C epidemic, South Africa. We analyzed 57 HIV-infected individuals from Cameroon and 44 HIV-infected individuals from South Africa for differences in detecting HIV-1 consensus M Gag and Nef T cell responses using the IFN-γ ELISpot assay. We found no difference in the predicted epitope coverage between CRF02_AG and non-CRF02_AG viruses for either Gag or Nef. There were no differences in the magnitude and breadth of responses for CRF02_AG and non-CRF02_AG-infected individuals. In contrast, the specificity of epitope targeting was markedly different between the two groups, with fewer than one third (11/38) of peptides commonly recognized in Gag. Furthermore, only one peptide was commonly recognized by at least three individuals from both AG and non-AG groups, indicating poor immunodominance. For Nef, more than half of all targeted peptides (14/27) were recognized by both groups, and four peptides were commonly targeted by at least three individuals. Three times more peptides were exclusively targeted in the diverse non-CRF02_AG group compared to the CRF02_AG group (10 vs. 3). Of note, similar results were obtained when South Africa, a monoclade C epidemic, and Cameroon, a multiclade epidemic, were compared. The central nature of HIV-1 consensus M sequences resulted in their broad recognition, but failed to identify highly immunodominant peptides between homogeneous and diverse HIV epidemics.

Phylogenetics of HIV-1 subtype G env: Greater complexity and older origins than previously reported.

HIV-1 subtype G has played an early and central role in the emergent complexity of the HIV-1 group M (HIV-1M) epidemic in central/west Africa. Here, we analysed new subtype G env sequences sampled from 8 individuals in Yaoundé, Cameroon during 2007-2010, together with all publically available subtype G-attributed full-length env sequences with known sampling dates and locations. We inferred that the most recent common ancestor (MRCA) of the analysed subtype G env sequences most likely occurred in ∼1953 (95% Highest Posterior Density interval [HPD] 1939-1963): about 15 years earlier than previous estimates. We found that the subtype G env phylogeny has a complex structure including seven distinct lineages, each likely dating back to the late 1960s or early 1970s. Sequences from Angola, Gabon and the Democratic Republic of Congo failed to group consistently in these lineages, possibly because they are related to more ancient sequences that are poorly sampled. The circulating recombinant form (CRF), CRF06_cpx env sequences but not CRF25_cpx env sequences are phylogenetically nested within the subtype G clade. This confirms that the CRF06_cpx env plausibly was derived through recombination from a subtype G parent, and suggests that the CRF25_cpx env was likely derived from an HIV-1M lineage related to the MRCA of subtype G that has remained undiscovered and may be extinct. Overall, this fills important gaps in our knowledge of the early events in the spread of HIV-1M.

Evaluating potential T-cell epitope peptides for detecting HIV-specific T cell responses in a highly diverse HIV-1 epidemic from Cameroon.

HIV genetic diversity is a major obstacle for vaccine development. To define whether potential T-cell epitope (PTE) peptide usage improves the detection of T cell responses in a highly diverse HIV-1 epidemic, we compared the magnitude, breadth and depth of group M PTE peptide responses to consensus M peptides in Gag and Nef proteins. Gag PTE responses were detected at a higher magnitude, more Nef PTE responses were detected at a cohort (but not individual) level and depth was detected in both Gag and Nef responses.

Unravelling the complicated evolutionary and dissemination history of HIV-1M subtype A lineages

Abstract Subtype A is one of the rare HIV-1 group M (HIV-1M) lineages that is both widely distributed throughout the world and persists at high frequencies in the Congo Basin (CB), the site where HIV-1M likely originated. This, together with its high degree of diversity suggests that subtype A is amongst the fittest HIV-1M lineages. Here we use a comprehensive set of published near full-length subtype A sequences and A-derived genome fragments from both circulating and unique recombinant forms (CRFs/URFs) to obtain some insights into how frequently these lineages have independently seeded HIV-1M sub-epidemics in different parts of the world. We do this by inferring when and where the major subtype A lineages and subtype A-derived CRFs originated. Following its origin in the CB during the 1940s, we track the diversification and recombination history of subtype A sequences before and during its dissemination throughout much of the world between the 1950s and 1970s. Collectively, the timings and numbers of detectable subtype A recombination and dissemination events, the present broad global distribution of the sub-epidemics that were seeded by these events, and the high prevalence of subtype A sequences within the regions where these sub-epidemics occurred, suggest that ancestral subtype A viruses (and particularly sub-subtype A1 ancestral viruses) may have been genetically predisposed to become major components of the present epidemic.