Dans certaines situations cliniques, le séquençage de l'ADN VIH est réalisé pour rechercher les mutations de résistances aux antirétroviraux (ARV). De nombreuses disparités dans les pratiques étant observées, un consensus national de type DELPHI a été initié dans l'objectif de faciliter l'homogénéisation des situations de recours et l'interprétation des résultats obtenus. Après analyse de la littérature et partage d'expériences, un Comité de Pilotage (CP) composé de 8 virologues et 1 infectiologue a formulé des assertions soumises au vote électronique, indépendant et anonyme de virologues et de cliniciens en France, sélectionnés pour leur expertise dans le suivi clinique des PVVIH, entre octobre et décembre 2022. Les votes étaient recueillis à l'aide d'une échelle de 1 «Pas du tout d'accord» à 9 «Tout à fait d'accord». Un fort ou un bon consensus était atteint lorsque plus de 75% des scores étaient ≥7 et/ou que le score médian était ≥8. Les assertions n'ayant pas satisfait à ces critères étaient reformulées par le CP puis soumises à un 2nd tour de vote. Le CP a formulé initialement 21 assertions regroupées en 6 catégories: situations cliniques de recours au génotypage sur ADN VIH, techniques de réalisation, prise en considération des mutations APOBEC, rendu des résultats, recyclage des ARV et disponibilité des tests. Vingt et un virologues et 47 cliniciens ont participé à chacun des 2 tours de votes. Au 1er tour, 14/21 assertions ont recueilli un bon ou fort consensus et 7/21 une absence de consensus. Le CP a reformulé 5 assertions avec bon consensus et les 7 avec absence de consensus, dont 2 ont été fusionnées. Au 2nd tour, 2/20 assertions sont restées sans consensus. Au final, 18/20 (90%) assertions ont atteint un consensus fort (après les 2 tours), notamment: la réalisation préalable d'un génotypage sur ADN VIH est utile à la décision clinique en cas de volonté d'allégement/simplification vers un traitement en «long acting» ou par diminution du nombre d'ARV chez un patient en succès virologique pour qui les données ARN à disposition ne sont pas exploitables ou suffisamment informatives; il n'est pas indispensable pour passer au traitement 4 ou 5 jours/7 sans changement de molécule; les mutations de résistance attribuables à APOBEC détectées lors du test doivent être mentionnées dans l'analyse du résultat. En revanche, le panel de virologues n'a pas validé de manière unanime l'intérêt de signaler toute population minoritaire détectée afin de la discuter en réunion de concertation pluridisciplinaire. De même, le panel de cliniciens était partagé sur le risque virologique possible de l'utilisation d'une association INI + XTC chez un patient présentant une charge virale indétectable ≥1 an et en présence d'une mutation M184V documentée <5 ans. Ce consensus de type DELPHI permettra de renforcer et d'harmoniser les bonnes pratiques de réalisation du génotypage sur ADN VIH, notamment vis-à-vis de la réduction du nombre d'ARV et de l'utilisation des «long acting». E. A-G: ViiV, Gilead et MSD

OBJECTIVE:The objective was to evaluate the association between age-related comorbidities (ARCs) and 5-year HIV-related excess mortality in people living with HIV aged ≥60 years.DESIGN:Cohort study using relative survival analysis (Estève's model).SETTING:The French multicentre prospective Dat'AIDS cohort that involves 12 French hospitals.PARTICIPANTS:Inclusion of 1415 HIV-1 infected patients actively followed aged ≥60 years on January 2008, with a 5-year follow-up period in the late combination antiretroviral therapy era.RESULTS:Among 1415 patients included, 154 died. By multivariable analysis, factors predictive of 5-year HIV-related excess mortality were non-AIDS-related cancer (adjusted excess HR (aEHR)=2.94; 95% CI 1.32 to 6.57), cardiovascular disease (aEHR=6.00; 95% CI 2.45 to 14.65), chronic renal disease (aEHR=4.86; 95% CI 2.24 to 10.53), cirrhosis (aEHR=3.58; 95% CI 1.25 to 10.28), hepatitis C co-infection (aEHR=3.63; 95% CI 1.44 to 9.12), body mass index<18.5 kg/m² (aEHR=4.10; 95% CI 1.61 to 10.48) and having a CD4 cell count ≤200/mm3 (aEHR=5.79; 95% CI 2.28 to 14.69).CONCLUSIONS:ARCs, particularly cardiovascular disease and chronic renal disease, are predictive of HIV-related excess mortality, with an increase in hazard similar to that of CD4 cell count.TRIAL REGISTRATION NUMBER:NCT02898987.

OBJECTIVES:We estimated the prevalence of transmitted-drug-resistance-associated mutations (TDRAMs) in antiretroviral-naive chronically HIV-1-infected patients.PATIENTS AND METHODS:TDRAMs were sought in samples from 660 diagnosed HIV-1-infected individuals in 2015/2016 in 33 HIV clinical centres. Weighted analyses, considering the number of patients followed in each centre, were used to derive representative estimates of the percentage of individuals with TDRAMs. Results were compared with those of the 2010/2011 survey (n = 661) using the same methodology.RESULTS:At inclusion, median CD4 cell counts and plasma HIV-1 RNA were 394 and 350/mm3 (P = 0.056) and 4.6 and 4.6 log10 copies/mL (P = 0.360) in the 2010/2011 survey and the 2015/2016 survey, respectively. The frequency of non-B subtypes increased from 42.9% in 2010/2011 to 54.8% in 2015/2016 (P < 0.001), including 23.4% and 30.6% of CRF02_AG (P = 0.004). The prevalence of virus with protease or reverse-transcriptase TDRAMs was 9.0% (95% CI = 6.8-11.2) in 2010/2011 and 10.8% (95% CI = 8.4-13.2) in 2015/2016 (P = 0.269). No significant increase was observed in integrase inhibitor TDRAMs (6.7% versus 9.2%, P = 0.146). Multivariable analysis showed that men infected with the B subtype were the group with the highest risk of being infected with a resistant virus compared with others (adjusted OR = 2.2, 95% CI = 1.3-3.9).CONCLUSIONS:In France in 2015/2016, the overall prevalence of TDRAMs was 10.8% and stable compared with 9.0% in the 2010/2011 survey. Non-B subtypes dramatically increased after 2010. Men infected with B subtype were the group with the highest risk of being infected with a resistant virus, highlighting the need to re-emphasize safe sex messages.

Introduction. - The metabolic pathways of dolutegravir suggest a potential predator effect of nevirapine on dolutegravir pharmacokinetics and switching from a nevirapine- to a dolutegravir-containing regimen could lead to a lower and suboptimal exposure to dolutegravir several weeks after the switch in case of persistent inducer effect. Patients and method. - Prospective, pilot, single-arm, open-label, non-comparative, bicentric study to evaluate the pharmacokinetics, virologic outcomes, safety, and patient satisfaction of switching from abacavir/lamivudine and nevirapine to a single tablet of abacavir/lamivudine/dolutegravir. The primary endpoint was the maintenance of virologic suppression (HIV-1 RNA < 50 copies/mL) at week 12. Secondary endpoints were virologic suppression at week 48, safety and tolerability, patient satisfaction, and pharmacokinetic interaction between nevirapine and dolutegravir. Fifty-three adults on stable abacavir/lamivudine and nevirapine regimen for a median duration of 6 years and virologically suppressed for 9.6 years were included. Results. - Dolutegravir reached steady state by week 4/week 12 when expected by day 5/day 10. All subjects maintained plasma HIV-RNA < 50 copies/mL at week 12 and week 48. Abacavir/lamivudine/dolutegravir was well-tolerated, with two cases of serious adverse events deemed unrelated to study drugs (coronary syndrome in both cases), and one discontinuation for renal impairment at week 24 with a slight improvement after dolutegravir discontinuation. Level of treatment satisfaction remained high after the switch. Conclusion. - The transient predator effect of nevirapine on dolutegravir had no clinical consequences after switching from nevirapine to dolutegravir, neither on safety nor maintenance of virologic suppression. It also had no consequences on patient satisfaction. (C) 2018 Elsevier Masson SAS. All rights reserved.

Switch of antiretroviral therapy in virologically suppressed HIV-infected patients is frequent, to prevent toxicities, for simplification or convenience reasons. Pretherapeutic genotypic resistance testing on RNA can be lacking in some patients, which could enhance the risk of virologic failure, if resistance-associated mutations of the new regimen are not taken into account. Proviral DNA resistance testing in 69 virologically suppressed patients on antiretroviral treatment with no history of virological failure were pair-wised compared with pre-ART plasma RNA resistance testing. The median time between plasma (RNA testing) and whole blood (proviral DNA testing) was 47 months (IQR 29-63). A stop codon was evidenced in 23% (16/69) of proviral DNA sequences; these strains were considered as defective, non-replicative, and not taken into consideration. Within the non defective strains, concordance rate between plasma RNA and non-defective proviral DNA was high both on protease (194/220 concordant resistance-associated mutations=88%) and reverse transcriptase (28/37 concordant resistance-associated mutations=76%) genes. This study supports that proviral DNA testing might be an informative tool before switching antiretrovirals in virologically suppressed patients with no history of virological failure, but the interpretation should be restricted to non-defective viruses.

En 2014 de nouveaux antiviraux d'action directe (AAD) anti-VHC ont progressivement été prescrits chez les patients infectés par le VHC : trois anti-NS3, deux anti-NS5B et cinq anti-NS5A. Ces AAD sont puissants, pour certains pangénotypiques et avec une barrière génétique élevée. Cette étude rétrospective monocentrique évalue, au sein d'une cohorte de patients co-infectés VIH-VHC, le taux de succès virologique et analyse la résistance du VHC à ces nouveaux AAD sur la période du 01/01/2014 au 31/12/2017. Le génotype de la souche ainsi que son polymorphisme est déterminé par l'analyse de la région NS5A. En cas d'échec virologique, le séquençage des gènes des différentes protéines cibles des AAD (NS5A, NS5B et/ou NS3) a été réalisé sur les prélèvements pré-thérapeutiques et post-rechutes. Les données clinicobiologiques ont été collectées à partir du dossier patient (logiciel Nadis). Sur la période analysée, 2387 patients ont eu au moins un recours pour leur suivi VIH. Parmi eux, 245 (10,3 %) avaient une sérologie VHC positive avec une PCR VHC positive pour 105 patients. Cent un (41 %) patients ont reçu un traitement par AAD entre 2014 et 2017 : infectés par le VIH depuis 21 [7 ; 27] ans avec une médiane de 653 [452 ; 823] CD4/mm3, une CV VHC médiane à 6,12 [4,96–6,67] log UI/mL et une CV VIH < 50 copies/mL pour 92 % à l'initiation de l'AAD, 78 % d'hommes. La répartition des génotypes G était : 35 % G1a, 8 % G1b, 2 % G1e, 1 % G2, 17 % G3a et 37 % G4. Les AAD majoritairement prescrits (associés entre eux et ± associés à la ribavirine et au Peg-Interféron) étaient : sofosbuvir (SOF) (n = 93), lédipasvir (LDV) (n = 53), daclatasvir (DCV) (n = 22), velpatasvir (VEL) (n = 8), simeprevir (SMV) (n = 5) et ombitasvir + paritaprévir (OBV + PTV) (n = 4). Le stade de fibrose était ≥ F2 pour 34/95 (36 %) patients. Quatre rechutes sont survenues : deux G4, un G1a et un G3 sous respectivement SOF + PegINF + Riba (F4), SOF + LDV (F4), SOF + DCV (F1), SOF + VEL (F1). Le séquençage de NS5B des souches des patients en rechute ne retrouve aucune mutation décrite et celui de NS5A une mutation Y93N pour une souche G1a. Durant cette même période, il y a eu 25 hépatites C aiguës, 4 patients ont présenté une ou plusieurs réinfection(s) par le VHC et 28 n'avaient pas encore été traités. Dans cette cohorte, 100 % des patients présentaient une CV VHC indétectable à la fin de leur traitement. Les rares échecs virologiques sont des rechutes sans mutation de résistance sur NS5B. À l'ère de la seconde génération d'AAD où le taux de succès est bien > 95 % le problème reste, chez des patients infectés par le VIH, l'exposition au risque d'infection ou de réinfection par le VHC.

Background: The NS5A protein of the hepatitis C virus has been shown to be involved in the development of hepatocellular carcinoma.Objectives: In a French multicenter study, we investigated the clinical and epidemiological features of a new HCV genotype lb strain bearing a wide insertion into the V3 domain.Study Design: We studied NS5A gene sequences in 821 French patients infected with genotype 1b HCV.Results: We identified an uncharacterized V3 insertion without ORF disruption in 3.05% of the HCV sequences. The insertion comprised 31 amino-acids for the majority of patients; 3 patients had 27 aminoacids insertions and 1 had a 12 amino-acids insertion. Sequence identity between the 31 amino-acids insertions and the V3 domain ranged from 48 to 96% with E-values above 4e(-5), thus illustrating sequence homology and a partial gene duplication event that to our knowledge has never been reported in HCV. Moreover we showed the presence of the duplication at the time of infection and its persistence at least during 12 years in the entire quasispecies. No association was found with extrahepatic diseases. Conversely, patients with cirrhosis were two times more likely to have HCV with this genetic characteristic (p = 0.04). Moreover, its prevalence increased with liver disease severity (from 3.0% in patients without cirrhosis to 9.4% in patients with both cirrhosis and HCC, p for trend = 0.045).Conclusions: We identified a duplicated V3 domain in the HCV-1b NS5A protein for the first time. The duplication may be associated with unfavorable evolution of liver disease including a possible involvement in liver carcinogenesis. (C) 2015 Elsevier B.V. All rights reserved.

OBJECTIVES Nevirapine is an inducer of hepatic metabolism. After discontinuation, nevirapine has an inductive effect on cytochrome P450 3A4, which persists for a few weeks and which, after switching to rilpivirine, may reduce rilpivirine exposures and have a negative clinical impact. This study evaluates the virological outcome, pharmacokinetics and safety of switching virologically suppressed, HIV-1-infected patients from nevirapine to rilpivirine. PATIENTS AND METHODS This 24 week open-label single-centre study included HIV-1-infected adults with HIV-1 RNA <50 copies/mL for >6 months on tenofovir/emtricitabine and nevirapine, who were willing to simplify their regimen to tenofovir/emtricitabine/rilpivirine. Virological suppression, safety and nevirapine and rilpivirine pharmacokinetics were assessed. RESULTS At weeks 12 and 24, all 32 subjects remained virologically suppressed. One subject discontinued at week 1 for rilpivirine-associated insomnia and two patients chose to resume tenofovir/emtricitabine and nevirapine after week 12 because of rilpivirine-associated food constraint. There was no grade 3/4 laboratory abnormality. Rilpivirine trough concentrations were above the mean trough concentrations observed in Phase 3 studies by 1 week post-switch. Twenty-seven out of 32 patients had no measurable levels of nevirapine by 2 weeks post-switch. The meal accompanying tenofovir/emtricitabine/rilpivirine intake satisfied food requirements in 81% of cases. Overall general satisfaction was improved in 90% of the subjects despite food constraints. CONCLUSION Nevirapine has a short and limited inductive effect on rilpivirine metabolism, which is not clinically significant. Tenofovir/emtricitabine/rilpivirine is an efficacious and safe option for virologically suppressed HIV-infected patients on nevirapine wishing to simplify their regimen.

4. the dose-dependent neutralization of HCV infection by CD81 specific antibodies and convalescent serum from a recovered HCV genotype 1 patient. Conclusion: This new ex vivo model allows to support HCV replication. It provides a powerful tool for studying the viral life cycle, the dynamics of viral spread in the native tissue and for evaluating the potency and resistance profile of new antiviral therapies.

Évaluer la survenue d’échecs virologiques (EV) chez des patients infectés par le VIH-1 ayant débuté un traitement à partir de 2001 au CHU de Nantes. Étude rétrospective à partir de la base de données NADIS concernant tous les patients ayant débuté un traitement ARV (TARV) entre janvier 2001 et 12/2004. L’EV était défini par 2 CV > 200 c/ml consécutives au moins 6 mois après le début du traitement. Parmi les 277 patients (2 INTI + IP/r : 52 %, 2 INTI + INNTI : 25 %, 3 INTI : 14 %), ≥ 1 EV était survenu au 30/04/2007 chez 57 patients (20 %). Au 1 er EV, 24/52 souches (46 %) étaient sauvages et 28 (54 %) présentaient ≥ 1 mutation de résistance (28 % sous INTI seuls, 13 % sous INTI + INNTI et 3 % sous INTI + IP/r). M184V était la mutation la plus fréquente, retrouvée seule ( n = 7) ou associée à d’autres mutations aux INTI (TAMs n = 5, L74V n = 2, K65R n = 2) ou aux INNTI ( n = 3). Pour 8 virus seules des mutations aux INNTI étaient retrouvées, et dans 1 cas une mutation majeure (M46I) isolée sur la protéase. Au 2 e EV ( n = 24), 11 souches présentaient des mutations et 7/24 une accumulation de résistance par rapport au 1 er échec. Le nombre médian de molécules « perdues » était de 4, majoritairement FTC-3TC, EFV et NVP. Deux virus avaient acquis une résistance aux IP. Chez les patients avec ≥ 3 EV ( n = 8), seuls 2 virus avaient accumulé des mutations et uniquement sur la TI. L’avènement des trithérapies montre que 80 % des patients initiant un traitement sont en succès virologique après un suivi médian de 4 ans. Lors de l’échec près de 50 % des souches ont un génotype sauvage soulignant la fréquence des problèmes d’observance. Les mutations sont plus fréquentes sous trithérapie d’INTI seuls, association non recommandée en 2008. Les EV multiples avec accumulation de résistance sont rares.

Objectives: Determine if a nucleoside reverse transcriptase inhibitors (NRTI)-free regimen affected mitochondrial DNA (mtDNA) levels in peripheral blood mononuclear cells (PBMCs) of patients enrolled in BIKS-2 trial.Methods: Antiretroviral (ARV) naive (N = 13) and NRTI experienced (N = 7) patients, received lopinavir/ritonavir, a boosted protease inhibitor, and efavirenz, a non-nucleoside reverse transcriptase inhibitor from Month (M) 0 to M12 (1-year BIKS trial) and from M12 to M36 (2-year BIKS-2 trial). MtDNA was quantified at M12, M24 and M36 via real-time PCR assay.Results: From M12 to M36, the 20 patients have maintained undetectable plasma HIV-1 RNA, gained CD4 cells and had no side effects attributable to these drugs. Median mtDNA contents were constant: 478.6 at M12, 478.6 at M24 and 324.4 copies/cell at M36 (pM12-M36 = 0.5). Because MO data is missing, these results were compared to those of two groups of age matched individuals: healthy donors and HIV-infected patients before and after exposure to NRTIs. Healthy donors have higher contents (871), followed by patients never treated (602), than by BIKS patients where 7 had toxic NRTIs (478.6) and at last by patients exposed for six months to the most toxic combination (ddl-d4T) (85 copies/cell).Conclusion: Lopinavir/ritonavir + efavirenz did not affect mtDNA contents in PBMCs. (C) 2006 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

Human cytomegalovirus (CMV) has been shown to alter adhesion molecule expression on permissive cells such as endothelial cells. The aim of the present study was to investigate expression of receptors for these molecules on CMV infected polymorphonuclear leukocytes (PMNLs). CMV-induced variations on cellular integrin expression were examined using an in vitro system to obtain infected PMNLs. A triparametric flow cytometry approach was developed, which allows combined detection, in a single experiment, of both viral intranuclear antigen in the selected PMNLs and cellular CD11/CD18 expression. Comparison of infected PMNLs with uninfected cells showed a decrease of up to 50% in the expression of CD11b, CD11c, and CD18. This study thus demonstrates that the presence of CMV in PMNLs, which characterizes active infection, modifies the expression of integrins and may thus affect cell-to-cell interactions and immune functions.

BK virus (BKV) infection during the first year after renal transplantation was studied prospectively in 104 unselected consecutive patients. Viral DNA in urine (DNAuria) and plasma (DNAemia) samples was detected and quantified by real-time PCR. The noncoding control region (NCCR) of BKV isolates was sequenced. DNAuria and DNAemia occurred in 57% and 29% of patients, respectively. Three groups were defined, uninfected patients (group 1, n=45), patients with DNAuria (group 2, n=29) and patients with positive DNAemia (group 3, n=30). Active infection started within the first 3 months in 80% of patients. Cold ischemia duration over 24 h and the administration of tacrolimus were identified as significant risks factors for DNAuria, whereas it remains more frequently negative in patients receiving cyclosporine A. The risk for positive DNAemia was higher in patients with DNAuria (notably for viral load (VL)>4 log/mL) or treated with tacrolimus. No relationship was found with genetic variability in the NCCR sequence. Our data highlight the high frequency of active BKV infection after renal transplantation. Although high VL was detected in some patients, none developed a BKV nephropathy. A prospective follow-up of the whole population during the first year post renal transplantation is thus not useful to predict BKV disease.

Human herpesvirus 6 (HHV-6) replication was evaluated during in vitro expansion of CD34-positive cells that were selected from 11 peripheral blood progenitor cell (PBPC) samples. In order to permit cellular differentiation towards the myeloid lineage, PBPCs were cultured for 14-21 days in a liquid, serum-free medium supplemented with interleukin 1 (IL1), IL3, IL6, granulocyte-macrophage colony-stimulating factor, granulocyte colony-stimulating factor and stem-cell factor. Among the 10 cultures from HHV-6-seropositive patients, the late, alternatively spliced U100 viral mRNA was detected in five of them after PBPC culture for 14 or 21 days. Recovery of infectious virus from one of the expansions, associated with an increase of HHV-6 viral load and detection of the U100 spliced messenger, confirmed the occurrence of a complete replicative cycle. These data thus demonstrate for the first time that haematopoietic differentiation can lead to HHV-6 reactivation.

In order to confirm the occurrence of active Human herpesvirus-6 (HHV-6) infection, two optimal procedures were developed to detect directly replicating virus. MT4 cells and peripheral blood mononuclear cells (PBMCs) infected with two different strains (HST and a patient strain GUI) were used. The first method consisted of a one-step reverse transcription PCR amplifying a part of the late alternatively spliced U100 gene which encode the gp 82–105 viral glycoprotein. Two extraction methods and two RT-PCR kits were evaluated, leading to the selection of TaKaRa mRNA selective PCR kit. The second procedure consisted in a flow cytometry method to analyze the expression of two late viral HHV-6 antigens using 7C7 and 10G6 monoclonal antibodies. Four fixation permeabilization procedures were compared and the preparation of cells with paraformaldehyde (PFA) 4% was found to be optimal. Evaluation of these methods was then realized during a sequential culture of HST strain on MT4 cells. This kinetic study confirmed that Mabs recognized late antigens and demonstrate that the U100 gene splicing starts at a late stage of multiplication whereas unspliced forms are detectable earlier in the cycle.