Molecular epidemiology of hepatitis C virus in Cambodia during 2016-2017.

In Cambodia, little epidemiological data of hepatitis C virus (HCV) is available. All previous studies were limited to only small or specific populations. In the present study, we performed a characterization of HCV genetic diversity based on demography, clinical data, and phylogenetic analysis of HCV non-structural 5B (NS5B) sequences belonging to a large cohort of patients (n = 3,133) coming from majority part of Cambodia between September 2016 and December 2017. The phylogenetic analysis revealed that HCV genotype 1 and 6 were the most predominant and sharing equal proportions (46%). The remaining genotypes were genotype 2 (4.3%) and unclassified variants (3.6%). Among genotype 1, subtype 1b was the most prevalent subtype accounting for 94%. Within genotype 6, we observed a high degree of diversity and the most common viral subtypes were 6e (44%) and 6r (23%). This characteristic points to the longstanding history of HCV in Cambodia. Geographic specificity of viral genotype was not observed. Risks of HCV infection were mainly associated with experience of an invasive medical procedure (64.7%), having partner with HCV (19.5%), and blood transfusion (9.9%). In addition, all of these factors were comparable among different HCV genotypes. All these features define the specificity of HCV epidemiology in Cambodia.

Field evaluation of GeneXpert® (Cepheid) HCV performance for RNA quantification in a genotype 1 and 6 predominant patient population in Cambodia.

GeneXpert® (Cepheid) is the only WHO prequalified platform for hepatitis C virus (HCV) nucleic acid amplification testing that is suitable for point-of-care use in resource-limited contexts. However, its application is constrained by the lack of evidence on genotype 6 (GT6) HCV. We evaluated its field performance among a patient population in Cambodia predominantly infected with GT6. Between August and September 2017, we tested plasma samples obtained from consenting patients at Médecins Sans Frontières' HCV clinic at Preah Kossamak Hospital for HCV viral load (VL) using GeneXpert® and compared its results to those obtained using COBAS® AmpliPrep/Cobas® TaqMan® HCV Quantitative Test, v2.0 (Roche) at the Institut Pasteur du Cambodge. Among 769 patients, 77% of the seropositive patients (n = 454/590) had detectable and quantifiable VL using Roche and 43% (n = 195/454) were GT6. The sensitivity and specificity of GeneXpert® against Roche were 100% (95% CI 99.2, 100.0) and 98.5% (95% CI 94.8, 99.8). The mean VL difference was -0.01 (95% CI -0.05, 0.02) log10  IU/mL for 454 samples quantifiable on Roche and -0.07 (95% CI -0.12, -0.02) log10  IU/mL for GT6 (n = 195). The limit of agreement (LOA) was -0.76 to 0.73 log10  IU/mL for all GTs and -0.76 to 0.62 log10 IU/mL for GT6. Twenty-nine GeneXpert® results were outside the LOA. Frequency of error and the median turnaround time (TAT) for GeneXpert® were 1% and 0 days (4 days using Roche). We demonstrated that the GeneXpert® HCV assay has good sensitivity, specificity, quantitative agreement, and TAT in a real-world, resource-limited clinical setting among GT6 HCV patients.

Usefulness of a serial algorithm of HBsAg and HBeAg rapid diagnosis tests to detect pregnant women at risk of HBV mother-to-child transmission in Cambodia, the ANRS 12328 pilot study.

BACKGROUND: In Cambodia, access to hepatitis B surface antigen (HBsAg) screening is low for pregnant women and Hepatitis B Virus (HBV) DNA quantification is poorly accessible. OBJECTIVES: To evaluate the performance of a serial algorithm using two HBV rapid diagnostic tests (RDTs), in which samples positive for HBsAg were further tested for HBeAg as a surrogate marker for HBV DNA quantification. STUDY DESIGN: In 2015, we prospectively collected plasma samples from 250 pregnant women consulting for antenatal care in one hospital in Phnom Penh including 128 with a known positive HBsAg status. All specimens were tested with the SD BIOLINE HBsAg RDT and HBsAg ELISA assay. In ELISA-positive samples, HBeAg status was determined using the SD BIOLINE HBeAg RDT and HBV DNA quantification was assessed. RESULTS: Sensitivity and specificity of HBsAg RDT were 99.2% (97.7-99.9) and 100% (97.0-100), respectively. Among the 128 ELISA-positive samples, 29 (23%) tested HBeAg positive and 34 (26.5%) had HBV DNA > 5.3 Log10 IU/mL. Sensitivity and specificity of HBeAg RDT in identifying viremic samples were 76.5% (62.2.0-90.7) and 96.8% (93.3-100) for HBV DNA > 5.3 Log10 IU/mL and 89.3% (77.8-100) and 96.0% (92.2-99.8) for HBV DNA > 7.3 Log10IU/mL. Among the 99 negative HBeAg RDT women, 8 had HBV DNA > 5.3 Log10 IU/mL and 7 of them harbored BCP/PC HBV mutants. CONCLUSIONS: A combination of HBsAg and HBeAg RDTs could be a low-cost strategy to identify HBV-infected pregnant women at risk of perinatal transmission in a country were HBV DNA quantification is not routinely available.

Massive Iatrogenic Outbreak of Human Immunodeficiency Virus Type 1 in Rural Cambodia, 2014-2015.

Background: In 2014-2015, 242 individuals aged 2-89 years were newly diagnosed with human immunodeficiency virus type 1 (HIV-1) in Roka, a rural commune in Cambodia. A case-control study attributed the outbreak to unsafe injections. We aimed to reconstruct the likely transmission history of the outbreak. Methods: We assessed in 209 (86.4%) HIV-infected cases the presence of hepatitis C virus (HCV) and hepatitis B virus (HBV). We identified recent infections using antibody (Ab) avidity testing for HIV and HCV. We performed amplification, sequencing, and evolutionary phylogenetic analyses of viral strains. Geographical coordinates and parenteral exposure through medical services provided by an unlicensed healthcare practitioner were obtained from 193 cases and 1499 controls during interviews. Results: Cases were coinfected with HCV (78.5%) and HBV (12.9%). We identified 79 (37.8%) recent (<130 days) HIV infections. Phylogeny of 202 HIV env C2V3 sequences showed a 198-sample CRF01_AE strains cluster, with time to most recent common ancestor (tMRCA) in September 2013 (95% highest posterior density, August 2012-July 2014), and a peak of 15 infections/day in September 2014. Three geospatial HIV hotspots were discernible in Roka and correlated with high exposure to the practitioner (P = .04). Fifty-nine of 153 (38.6%) tested cases showed recent (<180 days) HCV infections. Ninety HCV NS5B sequences formed 3 main clades, 1 containing 34 subtypes 1b with tMRCA in 2012, and 2 with 51 subtypes 6e and tMRCAs in 2002-2003. Conclusions: Unsafe injections in Cambodia most likely led to an explosive iatrogenic spreading of HIV, associated with a long-standing and more genetically diverse HCV propagation.

Phyloepidemiological Analysis Reveals that Viral Divergence Led to the Paucity of Simian Immunodeficiency Virus SIVmus/gsn/mon Infections in Wild Populations.

Human immunodeficiency virus type 1 (HIV-1) is the result of cross-species transmission of simian immunodeficiency virus from chimpanzees (SIVcpz). SIVcpz is a chimeric virus which shares common ancestors with viruses infecting red-capped mangabeys and a subset of guenon species. The epidemiology of SIV infection in hominoids is characterized by low prevalences and an uneven geographic distribution. Surveys in Cameroon indicated that two closely related members of the guenon species subset, mustached guenons and greater spot-nosed guenons, infected with SIVmus and SIVgsn, respectively, also have low rates of SIV infections in their populations. Compared to that for other monkeys, including red-capped mangabeys and closely related guenon species, such an epidemiology is unusual. By intensifying sampling of geographically distinct populations of mustached and greater spot-nosed guenons in Gabon and including large sample sets of mona guenons from Cameroon, we add strong support to the hypothesis that the paucity of SIV infections in wild populations is a general feature of this monophyletic group of viruses. Furthermore, comparative phylogenetic analysis reveals that this phenotype is a feature of this group of viruses infecting phylogenetically disparate hosts, suggesting that this epidemiological phenotype results from infection with these HIV-1-related viruses rather than from a common host factor. Thus, these HIV-1-related viruses, i.e., SIVcpz and the guenon viruses which share an ancestor with part of the SIVcpz genome, have an epidemiology distinct from that found for SIVs in other African primate species.IMPORTANCE Stable virus-host relationships are established over multiple generations. The prevalence of viral infections in any given host is determined by various factors. Stable virus-host relationships of viruses that are able to cause persistent infections and exist with high incidences of infection are generally characterized by a lack of morbidity prior to host reproduction. Such is the case for cytomegalovirus (CMV) and Epstein-Barr virus (EBV) infections of humans. SIV infections of most African primate species also satisfy these criteria, with these infections found at a high prevalence and with rare cases of clinical disease. In contrast, SIVcpz, the ancestor of HIV-1, has a different epidemiology, and it has been reported that infected animals suffer from an AIDS-like disease in the wild. Here we conclusively demonstrate that viruses which are closely related to SIVcpz and infect a subset of guenon monkeys show an epidemiology resembling that of SIVcpz.

Hepatitis E virus antibody prevalence, RNA frequency, and genotype among blood donors in Cambodia (Southeast Asia).

BACKGROUND: Recent studies conducted in developed countries described hepatitis E virus (HEV) as an emerging infectious threat to blood safety. However, data on HEV among blood donors from southeast Asia are lacking. STUDY DESIGN AND METHODS: Between July and August 2014, we assessed the presence of HEV immunoglobulin (Ig)G and IgM in 301 Cambodian blood donors. All samples were further tested for the presence of HEV RNA using an in-house reverse transcription-polymerase chain reaction. ORF2/ORF3 phylogenetic analysis was performed on positive HEV RNA specimens. RESULTS: We found HEV IgG in 28.2% of blood donors from Cambodia. Three blood donors tested positive for HEV IgM with three distinct patterns: IgM(+)/IgG(-)/RNA(-) (n = 1), IgM(+)/IgG(+)/RNA(-) (n = 1), and IgM(+)/IgG(+)/RNA(+) (n = 1). Thus, the prevalence rates of HEV IgM and HEV RNA were 1.0 and 0.3%. Interestingly, the viremic blood donor harbored a HEV strain that belonged to Genotype 3 (HEV-3) and clustered with a Cambodian riverine HEV-3 isolate. CONCLUSION: Due to the high frequency of Cambodian blood donors with positive HEV IgG, we conclude that HEV is endemic in this country. Large-scale studies must be considered to determine whether Cambodian blood donation screening is warranted to enhance blood safety in regard to HEV. In addition, our findings suggest that river water may be a significant source of exposure to HEV-3.

Estimation of HIV-1 DNA Level Interfering with Reliability of HIV-1 RNA Quantification Performed on Dried Blood Spots Collected from Successfully Treated Patients.

The impact of HIV-1 DNA coamplification during HIV-1 RNA quantification on dried blood spots (DBS) was explored. False-positive HIV RNA detection (22/62, 35%) was associated with high HIV-1 DNA levels. Specificity of HIV-1 RNA assays on DBS should be evaluated following manufacturer protocols on samples with HIV-1 DNA levels of ≥1,000 copies/10(6) peripheral blood mononuclear cells.

Cluster of HIV Infections Attributed to Unsafe Injection Practices--Cambodia, December 1, 2014-February 28, 2015.

In December 2014, local health authorities in Battambang province in northwest Cambodia reported 30 cases of human immunodeficiency virus (HIV) infection in a rural commune (district subdivision) where only four cases had been reported during the preceding year. The majority of cases occurred in residents of Roka commune. The Cambodian National Center for HIV/AIDS (acquired immunodeficiency syndrome), Dermatology and Sexually Transmitted Diseases (NCHADS) investigated the outbreak in collaboration with the University of Health Sciences in Phnom Penh and members of the Roka Cluster Investigation Team. By February 28, 2015, NCHADS had confirmed 242 cases of HIV infection among the 8,893 commune residents, an infection rate of 2.7%. Molecular investigation of the HIV strains present in this outbreak indicated that the majority of cases were linked to a single HIV strain that spread quickly within this community. An NCHADS case-control study identified medical injections and infusions as the most likely modes of transmission. In response to this outbreak, the Government of Cambodia has taken measures to encourage safe injection practices by licensed medical professionals, ban unlicensed medical practitioners, increase local capacity for HIV testing and counseling, and expand access to HIV treatment in Battambang province. Measures to reduce the demand for unnecessary medical injections and the provision of unsafe injections are needed. Estimates of national HIV incidence and prevalence might need to be adjusted to account for unsafe injection as a risk exposure.

Factors associated with the failure of first and second-line antiretroviral therapies therapy, a case control study in Cambodian HIV-1 infected children.

BACKGROUND: Little is known about the efficacy of first and and second-line antiretroviral therapies (ART) for HIV-1 infected children in resource limited Southeast Asian settings. Previous studies have shown that orphans are at a higher risk for virological failure (VF) in Cambodia. Consequently most of them required transfer to second-line ART. We assessed the factors associated with VF among HIV-1 infected children who were either under first-line (mostly 3TC + D4T + NVP) or under second-line (mostly ABC + DDI + LPV) therapies at a referral hospital in Cambodia. METHODS: A case-control study was conducted from February to July 2013 at the National Pediatric Hospital among HIV-1 infected children (aged 1-15 years) under second-line ART (cases) or first-line (matched controls at a ratio of 1:3) regimens. Children were included if a HIV-1 RNA plasma viral load (VL) result was available for the preceding 12 months. A standardized questionnaire explored family sociodemographics, HIV history, and adherence to ART. Associations between VF (HIV-1 RNA levels ≥1000 copies/ml) and the children's characteristics were assessed using bivariate and multivariate analyses. RESULTS: A total of 232 children, 175 (75.4 %) under first-line and 57 (24.6 %) under second-line ART, for a median of 72.0 (IQR: 68.0-76.0) months, were enrolled. Of them, 94 (40.5 %) were double orphans and 51 (22.0 %) single orphans, and 77 (33.2 %) were living in orphanages. A total of 222 children (95.6 %) were deemed adherent to ART. Overall, 18 (7.7 %; 95 % CI 4.6-11.9) showed a VF, 14 (8.6 %; 95 % CI 4.8-14.0) under first-line and 4 (7.0 %; 95 % CI 1.9-17.0) under second-line ART (p = 0.5). Their median CD4 percentage was 8 % (IQR 2.9-12.9) at ART initiation. Children under second-line ART were older; more often double orphans, and had lower CD4 cell counts at the last control. In the multivariate analysis, having the last CD4 percentage below 15 % was the only factor associated with VF for ART regimen separately or when combined (OR 40.4; 95 % CI 11-134). CONCLUSIONS: The pattern of risk factors for VF in children is changing in Cambodia. Improved adherence evaluation and intensified monitoring of children with low CD4 counts is needed to decrease the risk of VF.

Broad Range of Hepatitis B Virus (HBV) Patterns, Dual Circulation of Quasi-Subgenotype A3 and HBV/E and Heterogeneous HBV Mutations in HIV-Positive Patients in Gabon.

Integrated data on hepatitis B virus (HBV) patterns, HBV genotypes and mutations are lacking in human immunodeficiency virus type 1 (HIV-1) co-infected patients from Africa. This survey was conducted in 2010-2013 among 762 HIV-1-positive adults from Gabon who were predominantly treated with 3TC-based antiretroviral treatment. HBV patterns were identified using immunoassays detecting total antibody to hepatitis B core antigen (HBcAb), hepatitis B surface antigen (HBsAg), IgM HBcAb, hepatitis B e antigen (HBeAg), antibody to HBsAg (HBsAb) and an in-house real-time PCR test for HBV DNA quantification. Occult hepatitis B (OBI) was defined by the presence of isolated anti-HBc with detectable serum HBV DNA. HBV genotypes and HBV mutations were analyzed by PCR-direct sequencing method. Seventy-one (9.3%) patients tested positive for HBsAg, including one with acute hepatitis B (0.1%; 95% CI, 0.0%-0.2%), nine with HBeAg-positive chronic hepatitis B (CHB) (1.2%; 95% CI, 0.6%-2.2%), 16 with HBeAg-negative CHB (2.1%; 95% CI, 1.2%-3.3%) and 45 inactive HBV carriers (5.9%; 95% CI, 4.4%-7.8%). Sixty-one (8.0%; 95% CI, 6.2%-10.1%) patients showed OBI. Treated patients showed similar HBV DNA levels to those obtained in untreated patients, regardless of HBV patterns. Around 15.0% of OBI patients showed high (>1,000 UI/mL) viremia. The mutation M204V/I conferring resistance to 3TC was more common in HBV/A (47.4%) than in HBV/E isolates (0%) (P = .04). Our findings encouraged clinicians to promote HBV vaccination in patients with no exposure to HBV and to switch 3TC to universal TDF in those with CHB.

High Rate of Simian Immunodeficiency Virus (SIV) Infections in Wild Chimpanzees in Northeastern Gabon.

The emergence of HIV-1 groups M, N, O, and P is the result of four independent cross-species transmissions between chimpanzees (cpz) and gorillas (gor) from central/south Cameroon and humans respectively. Although the first two SIVcpz were identified in wild-born captive chimpanzees in Gabon in 1989, no study has been conducted so far in wild chimpanzees in Gabon. To document the SIVcpz infection rate, genetic diversity, and routes of virus transmission, we analyzed 1458 faecal samples collected in 16 different locations across the country, and we conducted follow-up missions in two of them. We found 380 SIV antibody positive samples in 6 different locations in the north and northeast. We determined the number of individuals collected by microsatellite analysis and obtained an adjusted SIV prevalence of 39.45%. We performed parental analysis to investigate viral spread between and within communities and found that SIVs were epidemiologically linked and were transmitted by both horizontal and vertical routes. We amplified pol and gp41 fragments and obtained 57 new SIVcpzPtt strains from three sites. All strains, but one, clustered together within a specific phylogeographic clade. Given that these SIV positive samples have been collected nearby villages and that humans continue to encroach in ape's territories, the emergence of a new HIV in this area needs to be considered.

Low frequency of acute hepatitis E virus (HEV) infections but high past HEV exposure in subjects from Cambodia with mild liver enzyme elevations, unexplained fever or immunodeficiency due to HIV-1 infection.

BACKGROUND: In Cambodia, previous studies conducted on hepatitis E virus (HEV) infection are scant, sometimes old, and showed inconsistent results. Moreover, there is no data about HEV infection in Cambodian HIV-1-infected patients. OBJECTIVES: To assess the occurrence of acute HEV infections and the level of past HEV exposure in one Mekong country. STUDY DESIGN: Using anti-HEV IgM and HEV RNA detection, we retrospectively investigated the presence of acute HEV infection in 825 individuals, including 350 subjects with or without fever, 300 subjects with or without liver enzyme elevations (LEE) and 175 antiretroviral treatment (ART)-naïve, severely immunocompromised HIV-1-infected patients. The detection of anti-HEV IgG was also performed to assess ancient HEV exposure. RESULTS: Nine individuals tested positive for anti-HEV IgM yielding an overall rate of 1.1% (95% confidence interval (CI), 0.5-2.0). We did not find significant differences for anti-HEV IgM rates between subjects with unexplained fevers (1.5%) and those with malaria or dengue-associated fever (1.7%) or non-febrile individuals (0%) (P=0.49), and between subjects with (1.5%) and without (2.0%) LEE (P=0.87). No HIV-infected patient tested positive for anti-HEV IgM. HEV RNA was not detected in all tested plasma specimens (n=578). Overall, the anti-HEV IgG prevalence rate was 30.1% (95% CI, 27.0-33.2). CONCLUSIONS: The scarcity of recent HEV infection contrasted with the high level of past HEV exposure. The role of HEV in liver disease is likely minor in Cambodia since no HEV RNA was detected in our studied populations, including HIV-positive patients with severe immunodepression.

Simian Immunodeficiency Virus Infection of Chimpanzees (Pan troglodytes) Shares Features of Both Pathogenic and Non-pathogenic Lentiviral Infections.

The virus-host relationship in simian immunodeficiency virus (SIV) infected chimpanzees is thought to be different from that found in other SIV infected African primates. However, studies of captive SIVcpz infected chimpanzees are limited. Previously, the natural SIVcpz infection of one chimpanzee, and the experimental infection of six chimpanzees was reported, with limited follow-up. Here, we present a long-term study of these seven animals, with a retrospective re-examination of the early stages of infection. The only clinical signs consistent with AIDS or AIDS associated disease was thrombocytopenia in two cases, associated with the development of anti-platelet antibodies. However, compared to uninfected and HIV-1 infected animals, SIVcpz infected animals had significantly lower levels of peripheral blood CD4+ T-cells. Despite this, levels of T-cell activation in chronic infection were not significantly elevated. In addition, while plasma levels of ß2 microglobulin, neopterin and soluble TNF-related apoptosis inducing ligand (sTRAIL) were elevated in acute infection, these markers returned to near-normal levels in chronic infection, reminiscent of immune activation patterns in 'natural host' species. Furthermore, plasma soluble CD14 was not elevated in chronic infection. However, examination of the secondary lymphoid environment revealed persistent changes to the lymphoid structure, including follicular hyperplasia in SIVcpz infected animals. In addition, both SIV and HIV-1 infected chimpanzees showed increased levels of deposition of collagen and increased levels of Mx1 expression in the T-cell zones of the lymph node. The outcome of SIVcpz infection of captive chimpanzees therefore shares features of both non-pathogenic and pathogenic lentivirus infections.

Usefulness of a fourth generation ELISA assay for the reliable identification of HCV infection in HIV-positive adults from Gabon (Central Africa).

BACKGROUND/OBJECTIVES: Guidelines for optimized HCV screening are urgently required in Africa, especially for patients infected with HIV, who sometimes show false positive or false negative reactivity in anti-HCV antibody assays. Here, we assessed the usefulness of a fourth-generation HCV Ag-Ab ELISA for the identification of active HCV infection in HIV-positive patients. METHODS: This cross-sectional study was conducted between 03/2010 and 01/2013 and included 762 Gabonese HIV-positive adult patients. The results of ELISA (Monolisa HCV Ag-Ab ULTRA, Bio-Rad) were compared with those obtained by RT-PCR (gold standard). The optimal ELISA signal-to-cutoff (S/CO) ratio to identify patients with active hepatitis C (positive HCV RNA) was determined. Specimens were further tested by the INNO-LIA HCV Score assay (Innogenetics) and the Architect HCV Ag kit (Abbott) to define the best diagnostic strategy. RESULTS: Sixty-seven patients tested positive for HCV (S/CO ratio ≥ 1) by ELISA. Of these, 47 (70.1%) tested positive for HCV RNA. The optimal S/CO associated with active HCV infection was 1.7. At this threshold, the sensitivity of ELISA was 97.9% (95% confidence interval (CI) 90.0-99.9%), its specificity was 91.3% (95% CI 85.0-95.5%), and HCV seroprevalence rate was 7.3% (56/762) (95% CI 5.6-9.4%). Among 57 HCV-seropositive patients with available INNO-LIA results, false reactivity was identified in 14 (24.6%), resolved HCV infection in two (3.5%), possible acute HCV infections in nine (15.8%) and likely chronic HCV infections in 32 (56.1%) patients. HCV core Ag was undetectable in 14/15 (93.3%) specimens that tested negative for HCV RNA whereas it was quantified in 34 (out of 39, 87.2%) samples that tested positive for HCV RNA. CONCLUSIONS: Our study provides comprehensive guidance for HCV testing in Gabon, and will help greatly clinicians to improve case definitions for both the notification and surveillance of HCV in patients co-infected with HIV.

Full-length genome analyses of two new simian immunodeficiency virus (SIV) strains from mustached monkeys (C. Cephus) in Gabon illustrate a complex evolutionary history among the SIVmus/mon/gsn lineage.

The Simian Immunodeficiency Virus (SIV) mus/mon/gsn lineage is a descendant of one of the precursor viruses to the HIV-1/SIVcpz/gor viral lineage. SIVmus and SIVgsn were sequenced from mustached and greater spot nosed monkeys in Cameroon and SIVmon from mona monkeys in Cameroon and Nigeria. In order to further document the genetic diversity of SIVmus, we analyzed two full-length genomes of new strains identified in Gabon. The whole genomes obtained showed the expected reading frames for gag, pol, vif, vpr, tat, rev, env, nef, and also for a vpu gene. Analyses showed that the Gabonese SIVmus strains were closely related and formed a monophyletic clade within the SIVmus/mon/gsn lineage. Nonetheless, within this lineage, the position of both new SIVmus differed according to the gene analyzed. In pol and nef gene, phylogenetic topologies suggested different evolutions for each of the two new SIVmus strains whereas in the other nucleic fragments studied, their positions fluctuated between SIVmon, SIVmus-1, and SIVgsn. In addition, in C1 domain of env, we identified an insertion of seven amino acids characteristic for the SIVmus/mon/gsn and HIV­1/SIVcpz/SIVgor lineages. Our results show a high genetic diversity of SIVmus in mustached monkeys and suggest cross-species transmission events and recombination within SIVmus/mon/gsn lineage. Additionally, in Central Africa, hunters continue to be exposed to these simian viruses, and this represents a potential threat to humans.

HIV-1 DNA concentrations and evolution among African HIV-1-infected children under antiretroviral treatment (ANRS 1244/1278).

OBJECTIVES: The objectives of this study were to describe the pretreatment HIV-1 DNA concentrations in children infected with HIV and to evaluate the impact of antiretroviral therapy (ART) on HIV-DNA concentrations. METHODS: This was a retrospective analysis of all children followed up in the 'Programme Enfant Yopougon' cohort, Abidjan, Côte d'Ivoire, from 2000 to 2004, who had cryopreserved peripheral blood mononuclear cells (PBMCs) and plasma samples. HIV-DNA was measured using a real-time PCR assay. Mixed-model analysis was used to analyse the factors associated with change in HIV-DNA concentration. RESULTS: The study included 121 children infected with HIV-1. The median age at inclusion was 6 years (IQR: 3.5-9) and children were at an advanced stage of HIV disease (46.6% and 20.3% presenting CDC stage B and CDC stage C, respectively). At baseline, the median HIV-DNA concentration was 3.4 log10 copies/10(6) PBMCs (IQR: 3.1-3.6). Fifty-four children were initiated on ART during follow-up. After 24 months of ART, HIV-DNA load decreased by 0.32 (IQR: 0.08-0.57) log10 copies/10(6) PBMCs. The only factor associated with the HIV-DNA decrease was a concomitant low HIV-RNA viral load result. Children with efficient ART had a 0.51 log10 copies (IQR: 0.40-0.86) HIV-DNA decrease per million PBMCs. CONCLUSIONS: HIV-DNA concentrations decreased following ART initiation in a large African paediatric cohort. This decline was exclusively associated with the decrease in ongoing replication level achieved. Our study points out that a strong adherence is needed for ART to be efficient on the viral reservoirs, and further reinforces that adherence support is also essential to diminish the reservoir.

Significant impact of non-B HIV-1 variants genetic diversity in Gabon on plasma HIV-1 RNA quantitation.

Evaluations of HIV-1 RNA viral load assays are lacking in Central Africa. The main objective of our study was to assess the reliability of HIV-1 RNA results obtained with three different assays for samples collected in Gabon. A total of 137 plasma specimens were assessed for HIV-1 RNA using the Abbott RealTime HIV-1® and Nuclisens HIV-1 EasyQ® version 2.0 assays. It included HIV-1 non-B samples (n = 113) representing six subtypes, 10 CRFs and 18 URFs from patients infected with HIV-1 and treated with antiretrovirals that were found HIV-1 RNA positive (≥300 copies/ml) with the Generic HIV viral load® assay; and samples (n = 24) from untreated individuals infected with HIV-1 but showing undetectable (<300 copies/ml) results with the Biocentric kit. For samples found positive with the Generic HIV viral load® test, correlation coefficients obtained between the three techniques were relatively low (R = 0.65 between Generic HIV viral load® and Abbott RealTime HIV-1®, 0.50 between Generic HIV viral load® and Nuclisens HIV-1 EasyQ®, and 0.66 between Abbott RealTime HIV-1® and Nuclisens HIV-1 EasyQ®). Discrepancies by at least one log10 were obtained for 19.6%, 33.7%, and 20% of samples, respectively, irrespective of genotype. Most of samples (22/24) from untreated study patients, found negative with the Biocentric kit, were also found negative with the two other techniques. In Central Africa, HIV-1 genetic diversity remains challenging for viral load testing. Further studies are required in the same area to confirm the presence of HIV-1 strains that are not amplified with at least two different viral load assays.