Maria Casadellà

Gut Microbiota Linked to Sexual Preference and HIV Infection

The precise effects of HIV-1 on the gut microbiome are unclear. Initial cross-sectional studies provided contradictory associations between microbial richness and HIV serostatus and suggested shifts from Bacteroides to Prevotella predominance following HIV-1 infection, which have not been found in animal models or in studies matched for HIV-1 transmission groups. In two independent cohorts of HIV-1-infected subjects and HIV-1-negative controls in Barcelona (n = 156) and Stockholm (n = 84), men who have sex with men (MSM) predominantly belonged to the Prevotella-rich enterotype whereas most non-MSM subjects were enriched in Bacteroides, independently of HIV-1 status, and with only a limited contribution of diet effects. Moreover, MSM had a significantly richer and more diverse fecal microbiota than non-MSM individuals. After stratifying for sexual orientation, there was no solid evidence of an HIV-specific dysbiosis. However, HIV-1 infection remained consistently associated with reduced bacterial richness, the lowest bacterial richness being observed in subjects with a virological-immune discordant response to antiretroviral therapy. Our findings indicate that HIV gut microbiome studies must control for HIV risk factors and suggest interventions on gut bacterial richness as possible novel avenues to improve HIV-1-associated immune dysfunction.

Improved Prediction of Salvage Antiretroviral Therapy Outcomes Using Ultrasensitive HIV-1 Drug Resistance Testing

BACKGROUND The clinical relevance of ultrasensitive human immunodeficiency virus type 1 (HIV-1) genotypic resistance testing in antiretroviral treatment (ART)-experienced individuals remains unknown. METHODS This was a retrospective, multicentre, cohort study in ART-experienced, HIV-1-infected adults who initiated salvage ART including, at least 1 ritonavir-boosted protease inhibitor, raltegravir or etravirine. Presalvage ART Sanger and 454 sequencing of plasma HIV-1 were used to generate separate genotypic sensitivity scores (GSS) using the HIVdb, ANRS, and REGA algorithms. Virological failure (VF) was defined as 2 consecutive HIV-1 RNA levels ≥200 copies/mL at least 12 weeks after salvage ART initiation, whereas subjects remained on the same ART. The ability of Sanger and 454-GSS to predict VF was assessed by receiver operating characteristic (ROC) curves and survival analyses. RESULTS The study included 132 evaluable subjects; 28 (21%) developed VF. Using HIVdb, 454 predicted VF better than Sanger sequencing in the ROC curve analysis (area under the curve: 0.69 vs 0.60, Delong test P = .029). Time to VF was shorter for subjects with 454-GSS < 3 vs 454-GSS ≥ 3 (Log-rank P = .003) but not significantly different between Sanger-GSS < 3 and ≥3. Factors independently associated with increased risk of VF in multivariate Cox regression were a 454-GSS < 3 (HR = 4.6, 95 CI, [1.5, 14.0], P = .007), and the number of previous antiretrovirals received (HR = 1.2 per additional drug, 95 CI, [1.1, 1.3], P = .001). Equivalent findings were obtained with the ANRS and REGA algorithms. CONCLUSIONS Ultrasensitive HIV-1 genotyping improves GSS-based predictions of virological outcomes of salvage ART relative to Sanger sequencing. This may improve the clinical management of ART-experienced subjects living with HIV-1. CLINICAL TRIALS REGISTRATION NCT01346878.

Primary resistance to integrase strand-transfer inhibitors in Europe

OBJECTIVES The objective of this study was to define the natural genotypic variation of the HIV-1 integrase gene across Europe for epidemiological surveillance of integrase strand-transfer inhibitor (InSTI) resistance. METHODS This was a multicentre, cross-sectional study within the European SPREAD HIV resistance surveillance programme. A representative set of 300 samples was selected from 1950 naive HIV-positive subjects newly diagnosed in 2006-07. The prevalence of InSTI resistance was evaluated using quality-controlled baseline population sequencing of integrase. Signature raltegravir, elvitegravir and dolutegravir resistance mutations were defined according to the IAS-USA 2014 list. In addition, all integrase substitutions relative to HXB2 were identified, including those with a Stanford HIVdb score ≥ 10 to at least one InSTI. To rule out circulation of minority InSTI-resistant HIV, 65 samples were selected for 454 integrase sequencing. RESULTS For the population sequencing analysis, 278 samples were retrieved and successfully analysed. No signature resistance mutations to any of the InSTIs were detected. Eleven (4%) subjects had mutations at resistance-associated positions with an HIVdb score ≥ 10. Of the 56 samples successfully analysed with 454 sequencing, no InSTI signature mutations were detected, whereas integrase substitutions with an HIVdb score ≥ 10 were found in 8 (14.3%) individuals. CONCLUSIONS No signature InSTI-resistant variants were circulating in Europe before the introduction of InSTIs. However, polymorphisms contributing to InSTI resistance were not rare. As InSTI use becomes more widespread, continuous surveillance of primary InSTI resistance is warranted. These data will be key to modelling the kinetics of InSTI resistance transmission in Europe in the coming years.

Switching the third drug of antiretroviral therapy to maraviroc in aviraemic subjects: a pilot, prospective, randomized clinical trial

OBJECTIVES To evaluate the safety and efficacy of switching the third drug of antiretroviral treatment to maraviroc in aviraemic subjects infected with R5 HIV. PATIENTS AND METHODS This is a pilot, prospective, randomized clinical trial (ClinicalTrials ID: NCT00966329). Eighty HIV-1-infected aviraemic adults on stable antiretroviral treatment for ≥1 year and no antiretroviral drug resistance were screened for the presence of non-R5 HIV by triplicate proviral V3 population sequencing. From them, 30 subjects with R5 HIV-1 were randomized 1 : 1 to switch the non-nucleoside reverse transcriptase inhibitor or ritonavir-boosted protease inhibitor to maraviroc (n = 15) or to continue the same antiretroviral treatment (controls, n = 15). The principal endpoint was the proportion of subjects with HIV-1 RNA <50 copies/mL at week 48. Ultrasensitive proviral HIV-1 tropism testing (454 sequencing) was performed retrospectively at weeks 0, 4, 12, 24, 36 and 48. RESULTS One subject in the maraviroc arm and one control had non-R5 HIV in proviral DNA by retrospective 454 sequencing. The subject receiving maraviroc was the only individual to develop virological failure. However, plasma HIV at failure was R5. Switching to maraviroc was well tolerated and associated with small, but statistically significant, declines in total, high-density lipoprotein and low-density lipoprotein cholesterol. Median (IQR) triglyceride [1 (0.67-1.22) versus 1.6 (1.4-3.1) mmol/L, P = 0.003] and total cholesterol [4.3 (4.1-4.72) versus 5.4 (4-5.7) mmol/L, P = 0.059] values were lower in the maraviroc arm than in controls at week 48. CONCLUSIONS In this pilot, prospective, randomized clinical trial, switching the third drug to maraviroc was safe, efficacious and improved lipid parameters.

Deep sequencing for HIV-1 clinical management

The emerging HIV-1 resistance epidemic is threatening the impressive global advances in HIV-1 infection treatment and prevention achieved in the last decade. Next-generation sequencing is improving our ability to understand, diagnose and prevent HIV-1 resistance, being increasingly cost-effective and more accessible. However, NGS still faces a number of limitations that need to be addressed to enable its widespread use. Here, we will review the main NGS platforms available for HIV-1 diagnosis, the factors affecting the clinical utility of NGS testing and the evidence supporting -or not- ultrasensitive genotyping over Sanger sequencing for routine HIV-1 diagnosis. Now that global HIV-1 eradication might be within our reach, making NGS accessible also to LMICs has become a priority. Reductions in sequencing costs, particularly in library preparation, and accessibility to low-cost, robust but simplified automated bioinformatic analyses of NGS data will remain essential to end the HIV-1 pandemic.

No detection of the NS5B S282T mutation in treatment-naïve genotype 1 HCV/HIV-1 coinfected patients using deep sequencing

BACKGROUND The S282T mutation is the main variant described associated with resistance to nucleos(t)ide analogues hepatitis C virus (HCV) NS5B polymerase inhibitors. OBJECTIVE We aimed here to investigate whether this substitution pre-existed in treatment naive HCV/HIV-1 coinfected patients. STUDY DESIGN NS5B polymerase deep sequencing was performed at a median coverage per base of 4471 in 16 patient samples. RESULTS No S282T variant was detected in the 16 analyzed samples. CONCLUSION This finding is in agreement with the high genetic barrier of nucleoside analogues NS5B polymerase inhibitors and the clinical efficacy of these compounds.

Treatment options after virological failure of first-line tenofovir-based regimens in South Africa: an analysis by deep sequencing.

In a South African cohort of participants living with HIV developing virological failure on first-line tenofovir disoproxyl fumarate (TDF)-based regimens, at least 70% of participants demonstrated TDF resistance according to combined Sanger and MiSeq genotyping. Sanger sequencing missed the K65R mutation in 30% of samples. Unless HIV genotyping is available to closely monitor epidemiological HIV resistance to TDF, its efficacy as second-line therapy will be greatly compromised.

Clinical value of ultradeep HIV-1 genotyping and tropism testing in late presenters with advanced disease.

Objective:This article aims to investigate if the detection of preexisting drug-resistant minority variant (DRMV) and/or X4 HIV-1 variants could improve the efficacy of first-line combined antiretroviral therapy (ART) in late presenters. Design:Post-hoc, combined analysis of two open-label, prospective, randomized clinical trials comparing first-line ART with efavirenz (EFV) vs. ritonavir-boosted protease inhibitor (PI/r)-based regimens in ART-naive, HIV-1-infected patients, with CD4+ T-cell counts less than 100 cells/&mgr;l and wild-type HIV-1 by bulk sequencing. Methods:Pre-ART samples were reanalyzed for the presence of DRMVs and X4 HIV-1 using 454 sequencing. Kaplan–Meier curves and Cox regression were used to evaluate the association between X4 HIV and DRMVs and risk of virological failure. Results:From 141 evaluable patients, 57 received EFV, and 84 received PI/r, including first-line ART. Median pre-ART CD4+ T-cell counts and HIV-1 RNA levels were 39 cells/&mgr;l and 257 424 copies/ml, respectively; 35.5% of patients had X4 HIV variants. Detection of DRMVs leading to an ART-specific cumulative HIVdb score of at least 10 increased the risk of virological failure in patients initiating EFV [log-rank P = 0.048, hazard ratio = 4.3 (95% confidence interval: 0.8, 25.0), P = 0.074], but not in those starting PI/r. Presence of X4 HIV did not affect virological outcomes, but was associated with impaired CD4+ T-cell count recovery over 2 years (214 vs. 315 cells/&mgr;l with X4 vs. R5 HIV-1 tropism, respectively, P = 0.017). Conclusion:Accounting for preexisting DRMVs may improve the outcomes of first-line nonnucleoside reverse transcriptase inhibitor-based ART in late presenters with advanced immune suppression. Presence of X4 HIV-1 at diagnosis predicts impaired immune restoration under ART.

HLA class I protective alleles in an HIV-1-infected subject homozygous for CCR5-Δ32/Δ32

Homozygosity for a 32 bp deletion in CCR5 (CCR5-Δ32/Δ32) is associated with strong resistance against HIV-1 infection. Several HLA types have been associated to improved viral control and/or delayed progression to AIDS. We report a unique HIV-1 infected individual homozygous for CCR5-Δ32/Δ32 and carrier of HLA-A*2402 and HLA-B*5701. In comparison with earlier data and although a replication competent virus has been isolated, the patient presents better immune status, response to treatment and disease evolution, which may be related to the control exerted by HLA class I restricted T cell immunity. Importantly, the accumulation of protective factors does not warrant a complete protection to HIV infection and the subsequent life-long treatment.

Stable HIV-1 integrase diversity during initial HIV-1 RNA Decay suggests complete blockade of plasma HIV-1 replication by effective raltegravir-containing salvage therapy

BackgroundThere is legitimate concern that minority drug-resistant mutants may be selected during the initial HIV-1 RNA decay phase following antiretroviral therapy initiation, thus undermining efficacy of treatment. The goal of this study was to characterize viral resistance emergence and address viral population evolution during the first phase of viral decay after treatment containing initiation.Findings454 sequencing was used to characterize viral genetic diversity and polymorphism composition of the HIV-1 integrase gene during the first two weeks following initiation of raltegravir-containing HAART in four ART-experienced subjects. No low-prevalence Raltegravir (RAL) drug resistance mutations (DRM) were found at baseline. All patients undergoing treatment received a fully active ART according to GSS values (GSS ≥ 3.5). No emergence of DRM after treatment initiation was detected. Longitudinal analysis showed no evidence of any other polymorphic mutation emergence or variation in viral diversity indexes.ConclusionsThis suggests that fully active salvage antiretroviral therapy including raltegravir achieves a complete blockade of HIV-1 replication in plasma. It is unlikely that raltegravir-resistant HIV-1 may be selected in plasma during the early HIV-1 RNA decay after treatment initiation if the administered therapy is active enough.