[Show abstract][Hide abstract] ABSTRACT: Molecular surveillance is essential to monitor HIV diversity and track emerging strains. We have developed a universal library
preparation method (HIV-SMART) for next-generation sequencing that harnesses the specificity of HIV-directed priming to enable
full genome characterization of all HIV-1 groups (M, N, O, and P) and HIV-2. Broad application of the HIV-SMART approach was
demonstrated using a panel of diverse cell-cultured virus isolates. HIV-1 non-subtype B infected clinical specimens from Cameroon
were then used to optimize the protocol to sequence directly from plasma. When multiplexing 8 or more libraries per MiSeq
run, full genome coverage at a median ∼2000 fold depth was routinely obtained for either sample type. The method reproducibly
generated the same consensus sequence, consistently identified viral sequence heterogeneity present in specimens, and at viral
loads ≤ 4.5 log copies/ml yielded sufficient coverage to permit strain classification. HIV-SMART provides an unparalleled
opportunity to identify diverse HIV strains in patient specimens and to determine phylogenetic classification based on the
entire viral genome. Easily adapted to sequence any RNA virus, this technology illustrates the utility of NGS for viral characterization
Preview · Article · Dec 2015 · Journal of clinical microbiology
[Show abstract][Hide abstract] ABSTRACT: Background:
Highly potent broadly neutralizing monoclonal antibodies (bNabs) have been obtained from individuals infected by HIV-1 group M variants. We analyzed the cross-group neutralization potency of these bNabs towards non-M primary isolates (PI).
Material & methods:
The sensitivity to neutralization was analyzed in a neutralization assay using TZM-bl cells. Twenty three bNabs were used, including reagents targeting the CD4 binding site (CD4bs), the N160 glycan-V1V2 site, the N332 glycan-V3 site, the membrane proximal external region of gp41, and complex epitopes spanning both Env subunits. Two bispecific antibodies that combine the inhibitory activity of an anti-CD4 with that of PG9 or PG16 (BibNabs) were included in the study (PG9-iMab and PG16-iMab).
Cross-group neutralization was observed only with the bNabs targeting the N160 glycan-V1V2 site. Four group O PIs, one group N PI and the group P PI were neutralized by PG9 and/or PG16 or PGT145 at low concentrations (0.04-9.39 µg/mL). None of the non-M PIs was neutralized by the bNabs targeting other regions at the highest concentration tested, except 10E8 that neutralized weakly two group N PIs and 35O22 that neutralized one group O PI. The BibNabs neutralized very efficiently all the non-M PIs with IC50 below 1 µg/mL, except two group O strains.
The N160 glycan-V1V2 site is the most conserved neutralizing site within the four groups of HIV-1. This makes it an interesting target for the development of HIV vaccine immunogens. The corresponding bNabs may be useful for immunotherapeutic strategies in patients infected by non-M variants.
No preview · Article · Sep 2015 · JAIDS Journal of Acquired Immune Deficiency Syndromes
[Show abstract][Hide abstract] ABSTRACT: We recently published a paper in the Journal of Medical Virology titled "Performance of rapid tests for discrimination between HIV-1 and/or HIV-2 infections" [Gautheret-Dejean et al., 2015]. Hønge et al. followed up with a commentary on our findings, which we have read with great interest. We are grateful for this opportunity to respond to their comments and give some precisions/clarifications in this letter. This article is protected by copyright. All rights reserved.
No preview · Article · Sep 2015 · Journal of Medical Virology
[Show abstract][Hide abstract] ABSTRACT: Objective:
Despite the genetic divergence between HIV-1 groups M and O, HIV-1 M/O intergroup recombinants were reported. Actually, there is no data on the transmissibility of such recombinant forms. During a surveillance of HIV genetic diversity in Cameroon, we investigated the possible direct transmission of an HIV-1 M/O recombinant virus in an HIV-infected couple.
Consecutive samples obtained from the couple were analysed for detection of dual HIV-1 groups M and O infections, and HIV-1 M/O recombinant forms. Analyses were performed using a serological and molecular algorithm based on HIV serotyping and group-specific PCRs targeting the polymerase and envelope genes. Pattern characterization of the strains found in both patients was based on complete genome sequencing. Phylogenetic and similarity profile analyses were performed to investigate the genetic relationship between viruses from both spouses and the previously described recombinant forms.
The sero-molecular algorithm data showed a group O serotype confirmed by molecular analysis in the envelope regions, whereas molecular tests identified HIV-1 group M in the polymerase. Phylogenetic analyses and similarity profiles of the full-length genome sequences showed that both spouses were infected with a unique recombinant virus having two recombination breakpoints in the vpr gene and LTR region. No phylogenetic link was found with the previous M/O recombinants.
We provide, for the first time, molecular evidence of direct transmission of an HIV-1 M/O recombinant, highlighting the potential spread of these divergent viruses. The importance of HIV-1 recombination on genetic evolution and public health when implying divergent strains as group O has to be carefully considered.
[Show abstract][Hide abstract] ABSTRACT: Author Summary
HIV-1 group O is one of the causal agents of AIDS, together with HIV-1 groups M (responsible for the pandemic), N and P (15 and 2 cases detected respectively, from Cameroon) and HIV-2 groups A to I (mostly found in West Africa), each group resulting from a distinct cross species transmission event from non-human primates. Even though mostly restricted to Cameroon, group O infections have been found in other African countries as well as in Europe and in the US. Due to their genetic distance from the pandemic HIV-1 group M, group O viruses still impact diagnosis, virological and therapeutic monitoring. Moreover, very few data are available on the natural history and epidemiology of these infections, as well as their genetic diversity and evolution. In particular, there is currently no explanation of the lack of spread of these variants, compared to the pandemic viruses from group M. Analysis of HIV-1 group O molecular evolution, from sequences spanning more than 2 decades, is an opportunity to better understand the phylodynamics of group O infection. We investigate it further by producing the largest set of group O sequences described. We show that the previous classifications proposed do not agree with each other and do not fit with the extensive genetic diversity of this group. We also estimate that group O MRCA existed in the 1930s (95% Higher Posterior Density: 1914–1944), and show that group O has diversified during two successive phases that could be linked to the specific historical context of Cameroon. These results contribute to a better understanding of the factors influencing HIV evolution, especially in the local context of west central Africa and lead to new hypotheses on the limited diffusion of such variants.
[Show abstract][Hide abstract] ABSTRACT: Background
Plasma HIV-1 RNA monitoring is one of the standard tests for the management of HIV-1 infection. While HIV-1 RNA can be quantified using several commercial tests, no test has been commercialized for HIV-2 RNA quantification. We studied the relationship between plasma HIV-2 viral load (VL) and CD4 count in West African patients who were either receiving antiretroviral therapy (ART) or treatment-naïve.
A cross sectional survey was conducted among HIV-2-infected individuals followed in three countries in West Africa from March to December 2012. All HIV-2 infected-patients who attended one of the participating clinics were proposed a plasma HIV-2 viral load measurement. HIV-2 RNA was quantified using the new ultrasensitive in-house real-time PCR assay with a detection threshold of 10 copies/ mL (cps/mL).
A total of 351 HIV-2-infected individuals participated in this study, of whom 131 (37.3%) were treatment naïve and 220 (62.7%) had initiated ART. Among treatment-naïve patients, 60 (46.5%) had undetectable plasma HIV-2 viral load (<10 cps/mL), it was detectable between 10-100 cps/mL in 35.8%, between 100-1000 cps/mL in 11.7% and >1000 cps/mL in 6.0% of the patients. Most of the treatment-naïve patients (70.2%) had CD4-T cell count ≥500 cells/mm3 and 43 (46.7%) of these patients had a detectable VL (≥10 cps/mL). Among the 220 patients receiving ART, the median CD4-T cell count rose from 231 to 393 cells/mm3 (IQR [259-561]) after a median follow-up duration of 38 months and 145 (66.0%) patients had CD4-T cell count ≤ 500 cells/mm3 with a median viral load of 10 cps/mL (IQR [10-33]). Seventy five (34.0%) patients had CD4-T cell count ≥ 500 cells/mm3, among them 14 (18.7%) had a VL between 10-100 cps/mL and 2 (2.6%) had VL >100 cps/mL.
This study suggests that the combination of CD4-T cell count and ultrasensitive HIV-2 viral load quantification with a threshold of 10 cps/mL, could improve ART initiation among treatment naïve HIV-2-infected patients and the monitoring of ART response among patients receiving treatment.
[Show abstract][Hide abstract] ABSTRACT: HIV-1 group O (HIV-O) is a rare variant that is characterized by a high number of natural polymorphisms in the integrase coding region that may impact on susceptibility to integrase strand transfer inhibitors (INSTIs) and on the emergence of resistance substitutions. We previously reported that HIV-O is more susceptible to RAL than HIV-1 group M (HIV-M).
The aim of the present study was to assess pathways of resistance to INSTIs in group 0 variants. Accordingly, we selected for resistance to each of raltegravir (RAL), elvitegravir (EVG), and dolutegravir (DTG) in cord blood mononuclear cells using HIV group O subtypes A and B, a HIV-O divergent isolate, and HIV-1 group M (subtype B, which served as a reference). Site directed mutagenesis was performed on the pCOM2.5 HIV group 0 infectious clone to ascertain the impact of INSTI resistance substitutions at positions Q148R, N155H, and R263K within integrase on susceptibility to INSTIs and infectiousness.
Cell culture selections of group O variants yielded similar patterns of resistance to RAL, EVG, and DTG as observed for subtype B. In the DTG selections, subtype B yielded S153Y whereas a natural S153A polymorphism sometimes led to A153V in group O. The pCMO2.5/Q148R and pCMO2.5/N155H variants displayed far higher levels of resistance to DTG (> 1000 FC) than was seen for group M viruses.
HIV-O harboring Q148R and N155H show higher resistance to DTG compared to HIV-M subtype B.
Full-text · Article · May 2015 · JAIDS Journal of Acquired Immune Deficiency Syndromes
[Show abstract][Hide abstract] ABSTRACT: Background
Acute respiratory infections (ARIs) are a major cause of morbidity and mortality in children in Africa. The circulation of viruses classically implicated in ARIs is poorly known in Burkina Faso. The aim of this study was to identify the respiratory viruses present in children admitted to or consulting at the pediatric hospital in Ouagadougou.
From July 2010 to July 2011, we tested nasal aspirates of 209 children with upper or lower respiratory infection for main respiratory viruses (respiratory syncytial virus (RSV), metapneumovirus, adenovirus, parainfluenza viruses 1, 2 and 3, influenza A, B and C, rhinovirus/enterovirus), by immunofluorescence locally in Ouagadougou, and by PCR in France. Bacteria have also been investigated in 97 samples.
153 children (73.2%) carried at least one virus and 175 viruses were detected. Rhinoviruses/enteroviruses were most frequently detected (rhinovirus n = 88; enterovirus n = 38) and were found to circulate throughout the year. An epidemic of RSV infections (n = 25) was identified in September/October, followed by an epidemic of influenza virus (n = 13), mostly H1N1pdm09. This epidemic occurred during the period of the year in which nighttime temperatures and humidity were at their lowest. Other viruses tested were detected only sporadically. Twenty-two viral co-infections were observed. Bacteria were detected in 29/97 samples with 22 viral/bacterial co-infections.
This study, the first of its type in Burkina Faso, warrants further investigation to confirm the seasonality of RSV infection and to improve local diagnosis of influenza. The long-term objective is to optimize therapeutic management of infected children.
[Show abstract][Hide abstract] ABSTRACT: The presence of HIV-1 non-B subtypes in Western Europe is commonly attributed to migration of individuals from non-European
countries, but the possible role of domestic infections with non-B subtypes is not well investigated. The French mandatory
anonymous reporting system for HIV is linked to a virological surveillance using assays for recent infection (<6 months) and
serotyping. During the first semester of years 2007 to 2010, any sample corresponding to a non-B recent infection was analyzed
by sequencing a 415-bp env region, followed by phylogenetic analysis and search for transmission clusters. Two hundred thirty-three recent HIV-1 infections
with non-B variants were identified. They involved 5 subtypes and 7 circulating recombinant forms (CRFs). Ninety-two cases
(39.5%) were due to heterosexual transmissions, of which 39 occurred in patients born in France. Eighty-five cases (36.5%)
were identified in men having sex with men (MSM). Forty-three recent non-B infections (18.5%) segregated into 14 clusters,
MSM being involved in 11 of them. Clustered transmission events included 2 to 7 cases per cluster. The largest cluster involved
MSM infected by a CRF02_AG variant. In conclusion, we found that the spread of non-B subtypes in France occurs in individuals
of French origin and that MSM are particularly involved in this dynamic.
[Show abstract][Hide abstract] ABSTRACT: We report the case of a multi-experienced patient, infected with an HIV-1 strain, which selected multiple resistance mutations. We designed a novel well-tolerated and effective rescue treatment including dolutegravir, rilpivirine, and foscarnet, allowing a 60 week sustained virological response for the first time in 23 years of HIV infection.
Full-text · Article · Aug 2014 · Journal of Clinical Virology
[Show abstract][Hide abstract] ABSTRACT: The Agence Nationale de Recherche sur le Sida et les hépatites virales (ANRS) previously developed a widely used method for
HIV-1 RNA quantification (Biocentric). Here, we report the development of a new specific and sensitive method for HIV-2 RNA
quantification, based on an adaptation of the existing HIV-1 protocol. The new test is based on TaqMan one-step reverse transcription-quantitative
PCR (qRT-PCR) targeting two conserved consensus regions of HIV-2 (long terminal repeat [LTR] and gag). Analytic performances were determined in three laboratories. Clinical performances were evaluated on 100 plasma samples
from HIV-2-infected patients (groups A, B, and H) by comparison with the assay currently used for the ANRS HIV-2 cohort. The
specificity was 100%. Sensitivity was 50 copies/ml (cp/ml) and was optimized to 10 cp/ml. The within-run coefficients of variation
in the three laboratories varied from 0.54% to 1.61% at 4 log10 copies/ml and from 7.24% to 14.32% at 2 log10 cp/ml. The between-run coefficients of variation varied from 2.28% to 6.43%. Of the 39 clinical samples below 2 log10 in the current assay, the new test improved the detection or quantification of 17 samples, including eight group B samples.
For quantifiable samples, similar loads were obtained with the two assays for group A samples. The median difference between
the two assays for group B samples was +0.18 but with greater heterogeneity than for group A. The HIV-2 group H sample had
similar results with the two assays. This new assay is highly sensitive and accurately quantifies the most prevalent HIV-2
groups. This test will be useful for monitoring low viral loads in HIV-2-infected patients.
Preview · Article · Jun 2014 · Journal of Clinical Microbiology
[Show abstract][Hide abstract] ABSTRACT: To assess the virological response, genotypic resistance profiles, and antiretroviral plasma concentrations in HIV-2 antiretroviral-treated (antiretroviral therapy, ART) patients in Côte d'Ivoire.
A cross-sectional survey was conducted among HIV-2 patients receiving ART. Plasma HIV-2 viral load was performed using the ANRS assay. Protease and reverse transcriptase sequencing was performed using in-house methods and antiretroviral plasma concentrations were assessed using ultra performance liquid chromatography combined with tandem mass spectrometry.
One hundred forty five HIV-2-treated patients were enrolled with a median CD4 cell count of 360/μl (interquartile range, IQR = 215-528). Median duration of ART was 4 years (IQR = 2-7) and 74% of patients displayed viral load less than 50 copies/ml. Median plasma HIV-2 RNA among patients with viral load more than 50 copies/ml was 3016 copies/ml (IQR = 436-5156). Most patients (84%) received a lopinavir/ritonavir-based regimen. HIV-2 resistance mutations to nucleoside reverse transcriptase inhibitors and protease inhibitors were detected in 21 of 25 (84%) and 20 of 29 (69%) samples, respectively. The most prevalent nucleoside reverse transcriptase inhibitor resistance mutations were M184I/V (90%), Q151M (24%), and S215F/Y (24%). The most prevalent protease inhibitor resistance mutations were V47A (60%) and I54M (30%). Median CD4 cell counts were 434/μl (292-573) and 204/μl (122-281) in patients with viral load less than 50 copies/ml and those exhibiting virological failure (P < 0.0001), respectively. The proportions of patients with adequate antiretroviral plasma concentrations were 81 and 93% in patients displaying virological failure and in those with viral load less than 50 copies/ml, respectively (P = 0.046), suggesting good treatment adherence.
We observed adequate drug plasma concentrations and virological suppression in a high proportion of HIV-2-infected patients. However, in cases of virological failure, the limited HIV-2 therapeutic arsenal and cross-resistance dramatically reduced treatment options.
No preview · Article · Feb 2014 · AIDS (London, England)