Determination of the Existence and Distribution of HIV-I Chemokine Co-Receptor 5 Polymorphism in a Sampled Population from Kenya

Human Immunodeficiency Virus/Acquired Immunodeficiency Syndrome (HIV/AIDS) is a major public health problem, socio-economic burden and a serious threat to development. Entry of human immunodeficiency virus type 1 (HIV-1) into target cells requires the binding of the external envelope glycoprotein gp 120 to both the CD4 molecule and one of several chemokine receptors, recently discovered to function as co-receptors. T –cell line tropic HIV-1 strains utilize the a-chemokine receptor CXCR4, whereas the b-chemokine receptor 5 (CCR5), which is expressed on monocytes/macrophages, T cells and granulocyte precursors, is the key co-factor for macrophage-tropic HIV-1 strains, which predominate during the asymptomatic phase of infection. A thirty two–base pair (bp) deletion mutation (? 32) within the second extra cellular loop-encoding region of the CCR5 gene, which results in a truncated, non-functional protein, has been associated with relative resistance to HIV -1 infection and slower progression to acquired immunodeficiency syndrome (AIDS). Specifically, ?32/?32 homozygotes are protected against acquisition of HIV-1 by the mucosal route despite high risk exposure, whereas disease progression among CCR5/?32 heterozygote occurs more slowly. In this study, the status of the CCR5 gene polymorphism in Kenyan population was investigated in an attempt to explain the differences in HIV prevalence in different parts of the country. To determine this, 200 samples were collected from the 8 provinces of Kenya, that is, 25 samples per province, some of which were positive for HIV-1. Twenty-five samples were randomly selected from a batch of 250 per province, that is, every tenth sample. The samples were collected from HIV screening centers, district and provincial hospitals. Peripheral blood mononuclear cells (PBMC) were extracted from whole blood. Genomic deoxyribonucleic acid (DNA) was then extracted from PBMC. A targeted region of the CCR5 gene flanking the 32bp deletion was amplified by polymerase chain reaction (PCR) using CCR5 specific primers. All the PCR amplicons were then analyzed by gel electrophoresis. The results showed that CCR5-D 32 mutations do not exist in the Kenyan population. Samples were then randomly selected 4 samples per province and sequenced. This was done to determine the genotype of the PCR products that were amplified. After ClustalW analysis of the sequences generated, it was seen that CCR5 gene is not highly conserved in the Kenyan population, as there were amino acid differences between the sequences analyzed suggesting that CCR5 gene in Kenyan population is highly polymorphic. From this study, it was concluded that CCR5-D 32 mutations do not play any role in HIV-1 susceptibility in the Kenyan population. This is because this mutation does not exist in the Kenyan population as per the samples analyzed. The differences in prevalence of HIV in different parts of the country may be due to cultural practices, religious backgrounds, socio-economic status and other intrinsic genetic factors. Keywords: HIV/AIDS, chemokine receptor

Monitoring HIV viral load in resource limited settings: still a matter of debate?

Introduction Consequences of lack of viral monitoring in predicting the effects of development of HIV drug resistance mutations during HAART in resource-limited settings (RLS) is still a matter of debate. Design To assess, among HIV+ patients receiving their first-line HAART, prevalence of virological failure and genotypic resistance mutations pattern in a Médécins Sans Frontières/Ministry of Health programme in Busia District (Kenya). Methods Patients with HAART treatment for ≥12 months were eligible for the study and those with HIV-RNA ≥5000 copies/ml underwent genotypic study. Total HIV-1 RNA from Dried Blood Spots was extracted using Nuclisens method. Results 926 patients were included. Among 274 (29.6%) patients with detectable viral load, 55 (5.9%) experienced treatment failure (viral load >5.000 copies/ml); 61.8% were female and 10 (18.2%) had clinical failure. Median CD4 cell count was 116 cell/mm3 (IQR: 54-189). Median HIV-RNA was 32,000 copies/ml (IQR: 11000-68000). Eighteen out of 55 (33%) samples could be sequenced on PR and RT genes, with resistance associated mutations (RAMs) in 15 out of 18 samples (83%). Among patients carrying RAMs, 12/15 (81%) harboured RAMs associated to thymidine analogues (TAMs). All of them (100%) showed M184V resistance associated mutation to lamivudine as well as NNRTI's RAMS. Conclusions Virological failure rate in resource-limited settings are similar to those observed in developed countries. Resistance mutation patterns were concordant with HAART received by failing patients. Long term detectable viral load confers greater probability of developing resistance and as a consequence, making difficult to find out a cost-effective subsequent treatment regimen

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Monitoring HIV viral load in resource limited settings: still a matter of debate?

INTRODUCTION: Consequences of lack of viral monitoring in predicting the effects of development of HIV drug resistance mutations during HAART in resource-limited settings (RLS) is still a matter of debate. DESIGN: To assess, among HIV+ patients receiving their first-line HAART, prevalence of virological failure and genotypic resistance mutations pattern in a Médécins Sans Frontières/Ministry of Health programme in Busia District (Kenya). METHODS: Patients with HAART treatment for ≥12 months were eligible for the study and those with HIV-RNA ≥5000 copies/ml underwent genotypic study. Total HIV-1 RNA from Dried Blood Spots was extracted using Nuclisens method. RESULTS: 926 patients were included. Among 274 (29.6%) patients with detectable viral load, 55 (5.9%) experienced treatment failure (viral load >5.000 copies/ml); 61.8% were female and 10 (18.2%) had clinical failure. Median CD4 cell count was 116 cell/mm3 (IQR: 54-189). Median HIV-RNA was 32,000 copies/ml (IQR: 11000-68000). Eighteen out of 55 (33%) samples could be sequenced on PR and RT genes, with resistance associated mutations (RAMs) in 15 out of 18 samples (83%). Among patients carrying RAMs, 12/15 (81%) harboured RAMs associated to thymidine analogues (TAMs). All of them (100%) showed M184V resistance associated mutation to lamivudine as well as NNRTI's RAMS. CONCLUSIONS: Virological failure rate in resource-limited settings are similar to those observed in developed countries. Resistance mutation patterns were concordant with HAART received by failing patients. Long term detectable viral load confers greater probability of developing resistance and as a consequence, making difficult to find out a cost-effective subsequent treatment regimen

Monitoring HIV viral load in resource limited settings: still a matter of debate?

Introduction Consequences of lack of viral monitoring in predicting the effects of development of HIV drug resistance mutations during HAART in resource-limited settings (RLS) is still a matter of debate. Design To assess, among HIV+ patients receiving their first-line HAART, prevalence of virological failure and genotypic resistance mutations pattern in a Médécins Sans Frontières/Ministry of Health programme in Busia District (Kenya). Methods Patients with HAART treatment for ≥12 months were eligible for the study and those with HIV-RNA ≥5000 copies/ml underwent genotypic study. Total HIV-1 RNA from Dried Blood Spots was extracted using Nuclisens method. Results 926 patients were included. Among 274 (29.6%) patients with detectable viral load, 55 (5.9%) experienced treatment failure (viral load >5.000 copies/ml); 61.8% were female and 10 (18.2%) had clinical failure. Median CD4 cell count was 116 cell/mm3 (IQR: 54-189). Median HIV-RNA was 32,000 copies/ml (IQR: 11000-68000). Eighteen out of 55 (33%) samples could be sequenced on PR and RT genes, with resistance associated mutations (RAMs) in 15 out of 18 samples (83%). Among patients carrying RAMs, 12/15 (81%) harboured RAMs associated to thymidine analogues (TAMs). All of them (100%) showed M184V resistance associated mutation to lamivudine as well as NNRTI's RAMS. Conclusions Virological failure rate in resource-limited settings are similar to those observed in developed countries. Resistance mutation patterns were concordant with HAART received by failing patients. Long term detectable viral load confers greater probability of developing resistance and as a consequence, making difficult to find out a cost-effective subsequent treatment regimen

RAMS among patients with virological failure (VL>5.000 copies/ml) amplified and evaluated by genotyping.

<p>RAMS among patients with virological failure (VL>5.000 copies/ml) amplified and evaluated by genotyping.</p

Patient's disposition.

<p>Patient's disposition.</p

Patients with VL >5.000 copies/ml (N = 55).

<p>Patients with VL >5.000 copies/ml (N = 55).</p