Performance characteristics of Allele-Specific PCR (ASPCR) in detecting drug resistance mutations among non-B HIV-1 Variants.

Allele-Specific Polymerase Chain Reaction (ASPCR) is an affordable point-mutation assay whose validation could improve the detection of HIV-1 drug resistance mutations (DRMs) in resource-limited settings (RLS). We assessed the performance of ASPCR onforty-four non-B HIV-1 plasma samples from patients who were ARV treated in failure in N'Djamena-Chad. Viral RNA was reverse-transcribed and amplified using LightCycler® FastStart DNA MasterPLUS SYBR Green I. Detection of six major DRMs (K70R, K103N, Y181C, M184V, T215F, T215Y) was evaluated on Roche LightCycler®480 automated system (with dilutions 0.01-100%). ASPCR-results were compared to Sanger-sequencing (gold-standard). Correlations of mutation curves were excellent (R2 >0.97); all DRMs were detected with desirable mutant/wild-type threshold differences (ΔCt≥9) except K70R(ΔCtK70R=6; ΔCtK103N=13; ΔCtM184V=9; ΔCtT215F=12; ΔCtT215Y=12; ΔCtY181C=9) and positive controls were below required thresholds. Also, ASPCR reproducibility on DRMs was assessed by using dilutions of intra-assay and inter-assay coefficient of variations respectively with a threshold of less than 50(i.e.<0.50 variation) which are;: K70R (0.02-0.28 vs. 0.12-0.37), K103N (0.08-0.42 vs. 0.12-0.37), Y181C (0.12-0.39 vs. 0.31-0.37), M184V (0.13-0.39 vs. 0.23-0.42), T215F (0.05-0.43 vs. 0.04-0.45) and T215Y (0.13-0.41 vs. 0.19-0.41). DRM detection-rate by ASPCR vs Sanger was respectively: M184V (63.6% vs. 38.6%); T215F (18.1% vs. 9.1%); T215Y (6.8% vs. 2.3%); K70R (4.5% vs. 2.3%). K103N (22.7% vs. 13.6%); Y181C (13.6% vs. 11.4%). Correlations of mutation curves were excellent (R2 >0.97); all DRMs were detected with desirable mutant/wild-type threshold differences (ΔCt≥9) except K70R(ΔCtK70R=6; ΔCtK103N=13; ΔCtM184V=9; ΔCtT215F=12; ΔCtT215Y=12; ΔCtY181C=9) and positive controls were below required thresholds. Also, ASPCR reproducibility on DRMs was assessed by using dilutions of intra-assay and inter-assay coefficient of variations respectively with a threshold of less than 50(i.e.<0.50 variation) which are;: K70R (0.02-0.28 vs. 0.12-0.37), K103N (0.08-0.42 vs. 0.12-0.37), Y181C (0.12-0.39 vs. 0.31-0.37), M184V (0.13-0.39 vs. 0.23-0.42), T215F (0.05-0.43 vs. 0.04-0.45) and T215Y (0.13-0.41 vs. 0.19-0.41). DRM detection-rate by ASPCR vs Sanger was respectively: M184V (63.6% vs. 38.6%); T215F (18.1% vs. 9.1%); T215Y (6.8% vs. 2.3%); K70R (4.5% vs. 2.3%). K103N (22.7% vs. 13.6%); Y181C (13.6% vs. 11.4%). ASPCR appears more efficient for detecting DRMs on diverse HIV-1 non-B circulating in RLS like Chad.

Prevalence and Evolution of Transmitted Human Immunodeficiency Virus Drug Resistance in Belgium Between 2013 and 2019.

Background: To assess the prevalence and evolution of transmitted drug resistance (TDR) in Belgium, a total of 3708 baseline human immunodeficiency virus (HIV)-1 polymerase sequences from patients diagnosed between 2013 and 2019 were analyzed. Methods: Protease and reverse-transcriptase HIV-1 sequences were collected from the 7 national Aids Reference Laboratories. Subtype determination and drug resistance scoring were performed using the Stanford HIV Drug Resistance Database. Trends over time were assessed using linear regression, and the maximum likelihood approach was used for phylogenetic analysis. Results: A total of 17.9% of the patients showed evidence of TDR resulting in at least low-level resistance to 1 drug (Stanford score ≥15). If only the high-level mutations (Stanford score ≥60) were considered, TDR prevalence dropped to 6.3%. The majority of observed resistance mutations impacted the sensitivity for nonnucleoside reverse-transcriptase inhibitors (NNRTIs) (11.4%), followed by nucleoside reverse-transcriptase inhibitors (6.2%) and protease inhibitors (2.4%). Multiclass resistance was observed in 2.4%. Clustered onward transmission was evidenced for 257 of 635 patients (40.5%), spread over 25 phylogenetic clusters. Conclusions: The TDR prevalence remained stable between 2013 and 2019 and is comparable to the prevalence in other Western European countries. The high frequency of NNRTI mutations requires special attention and follow-up. Phylogenetic analysis provided evidence for local clustered onward transmission of some frequently detected mutations.

Impact of switch from tenofovir disoproxil fumarate-based regimens to tenofovir alafenamide-based regimens on lipid profile, weight gain and cardiovascular risk score in people living with HIV.

BACKGROUND: As cardiovascular diseases represent the main cause of non-AIDS related death in people living with HIV (PLWH) with undetectable viral load, we evaluated lipid profile, weight gain and calculated cardiovascular risk change after switching from tenofovir disoproxil fumarate (TDF)-based regimens to tenofovir alafenamide (TAF)-based regimens. METHODS: For this retrospective study, we selected HIV-infected patients with suppressed viral load who fitted in one of the two groups below: First group (TDF/TDF): Patients treated continuously with TDF-based regimens. Second group (TDF/TAF): Patients treated with TDF-regimens during at least 6 months then switched to TAF-regimens while maintaining other drugs unchanged. Available data included date of birth, gender, ethnicity, lymphocyte T CD4+ count, weight, height, blood pressure, current/ex/non-smoker, diabetes mellitus, familial cardiovascular event, lipid profile, duration and nature of antiretroviral therapy. Lipid parameters, weight and calculated cardiovascular risk using 5-year reduced DAD score algorithm [Friis-Møller et al. in Eur J Cardiovasc Prev Rehabil 17:491-501, 2010] were analyzed in each groups. RESULTS: Switching from TDF to TAF resulted in a significant increase in triglycerides levels, total cholesterol and HDL cholesterol. LDL cholesterol and total cholesterol/HDL ratio did not show significant changes. Calculated cardiovascular risk increased after switch from TDF- to TAF-based therapy. CONCLUSIONS: Together with favorable outcomes at the bone and kidney levels, potential negative impact of TAF on lipid profile should be included in the reflection to propose the most appropriate and tailored ARV treatment.

Impact of the COVID-19 pandemic situation on HIV care in Liège, Belgium.

BACKGROUND: Background: The COVID-19 pandemic and associated containment measures dramatically affected the health care systems including the screening of human immunodeficiency virus and the management people living with HIV around the world by making the access to preventive care services and specific medical monitoring more difficult. OBJECTIVE: Objective: To study the impact of the COVID-19 pandemic on the holistic care of people living with HIV in Liège (Belgium). METHODS: Methods: In this retrospective observational study conducted in Liège University Hospital, we compared the out-patient follow-up of HIV-infected individuals as well as the number of new HIV diagnoses between 2019 and 2020 and between the different waves of the COVID-19 pandemic in 2020. RESULTS: Results: In 2020, when compared to 2019, we observed a significant decrease in the number of new HIV diagnoses, especially during the first wave of the pandemic, and in the number of consultations undertaken by sexual health services, psychologists and specialists in infectious diseases at our HIV clinic. We also observed a decrease in the number of viral load assays and blood CD4 + T-cells count analyses performed, although we found less patients with HIV plasma viral load above 400 copies per mL in 2020. Finally, we noted a significant reduction in terms of screening of our HIV-infected patients for hepatitis C, syphilis, colorectal and anal cancers and hypercholesterolemia. CONCLUSIONS: Conclusions: Our experience exhibits the deleterious impact of the COVID-19 pandemic on the HIV care and the need to implement new strategies to guarantee its continuum.

Evolution of Drug Interactions With Antiretroviral Medication in People With HIV.

BACKGROUND: Polypharmacy and drug interactions are important issues for HIV-infected individuals. The number and nature of those interactions are continuously evolving with the use of new antiretroviral drugs and the aging of HIV-infected individuals. We aimed to analyze this evolution over time. METHODS: This retrospective cohort study was conducted in the University Hospital of Liège (Belgium). Treatments of HIV-infected outpatients attending Liège University Hospital were collected and analyzed in 2012 and 2016. The University of Liverpool HIV drug interactions database was used to determine drug interactions. RESULTS: We included 1038 patients in 2016, of whom 78% had 1 comedication. Polypharmacy was seen in 20% of the cohort. Four percent of the patients presented red flag interactions, and 38% had orange flag interactions. Nonantiretroviral (non-ARV) therapeutic classes involved in drug interactions were mostly cardiovascular and central nervous system drugs. They were followed by hormone drugs and dietary supplements for orange flag interactions. Two factors significantly contributed to both red and orange flag interactions: the number of non-ARV comedications and protease inhibitor-based ARV regimens. The proportion of patients with red or orange flag interactions remained stable from 2012 to 2016. CONCLUSIONS: This study highlights the persistence of an alarming number of contraindicated drug interactions and a high prevalence of potential drug interactions over time. Identification, prevention, and management of drug interactions remain a key priority in HIV care.

Impact of coronavirus pandemic and containment measures on HIV diagnosis.

During the last months and following the implementation of containment measures in the context of coronavirus disease 2019 (COVID-19) pandemic, the number of new human immunodeficiency virus (HIV) diagnoses radically decreased in Liege AIDS Reference Center, Belgium. The number of HIV screening tests has also dramatically dropped down to an unprecedented level. This decline of HIV diagnosis is caused by missed diagnoses of individuals infected before the establishment of such measures and to the reduction of high-risk sexual behaviours during the COVID-19 pandemic.

Chronic and Early Antiretroviral Therapy Impact Human Immunodeficiency Virus (HIV) Serological Assay Sensitivity, Leading to More False-Negative Test Results in HIV Diagnosis.

This retrospective study evaluated the reactivity of 3 human immunodeficiency virus (HIV) confirmatory assays (INNO-LIA, Geenius, and MP) and 7 HIV rapid tests on samples from 2 different study populations in Belgium. For the early-treated cohort (83 HIV-1 adult patients treated within 3 months after infection), HIV-1 diagnosis was not obtained in at least 1 confirmatory assay in 12.0% (10/83) and in an HIV rapid test in 31.3% (26/83). Confirmation assay sensitivities ranged from 87.5% to 95.2%, whereas rapid test assay sensitivities ranged from 75.9% to 100%. The time to treatment initiation or the length of time on treatment did not have a statistical influence on the probability to obtain a false-negative test result. The fastest reversion was demonstrated after 4 months of treatment. Among the long-term treated cohort (390 HIV-1 patients with ≥ 9 years of undetectable viral load), false-negative test results were found in at least 1 HIV confirmatory assay for 2.1% (8/390) of the patients and in a HIV rapid test for 4.9% (19/390). Confirmation assay sensitivities ranged from 98.1% to 99.5%, whereas rapid test sensitivities ranged from 96.2% to 100%. Longer treatment increased nonreactivity of the HIV rapid tests (P = .033). Undetectable viral load decreases the sensitivities of HIV diagnostic tests, and further monitoring of the performance of serological assays is advised.

High frequency of new recombinant forms in HIV-1 transmission networks demonstrated by full genome sequencing.

The HIV-1 epidemic in Belgium is primarily driven by MSM. In this patient population subtype B predominates but an increasing presence of non-B subtypes has been reported. We aimed to define to what extent the increasing subtype heterogeneity in a high at risk population induces the formation and spread of new recombinant forms. The study focused on transmission networks that reflect the local transmission to an important extent. One hundred and five HIV-1 transmission clusters were identified after phylogenetic analysis of 2849 HIV-1 pol sequences generated for the purpose of baseline drug resistance testing between 2013 and 2017. Of these 105 clusters, 62 extended in size during the last two years and were therefore considered as representing ongoing transmission. These 62 clusters included 774 patients in total. From each cluster between 1 and 3 representative patients were selected for near full-length viral genome sequencing. In total, the full genome sequence of 101 patients was generated. Indications for the presence of a new recombinant form were found for 10 clusters. These 10 clusters represented 105 patients or 13.6% of the patients covered by the study. The findings clearly show that new recombinant strains highly contribute to local transmission, even in an epidemic that is largely MSM and subtype B driven. This is an evolution that needs to be monitored as reshuffling of genome fragments through recombination may influence the transmissibility of the virus and the pathology of the infection. In addition, important changes in the sequence of the viral genome may challenge the performance of tests used for diagnosis, patient monitoring and drug resistance analysis.

Detectability of HIV Residual Viremia despite Therapy Is Highly Associated with Treatment with a Protease Inhibitor-Based Combination Antiretroviral Therapy.

HIV persistence despite therapy contributes to chronic immune activation and inflammation, increasing the risk of aging-associated events in HIV-infected individuals. We sought here to better understand the complex link between clinical and treatment features and HIV persistence despite therapy. A total of 11,045 samples from 1,160 individuals under combination antiretroviral therapy (cART) with an unquantifiable viral load (VL; limit of quantification, 20 copies/ml) were categorized as detectable or undetectable depending on the detection of a PCR signal using a commercially available assay. Generalized estimating equation (GEE) regression was used to model viral load detectability and to assess the determinants of residual viremia (RV; VL detected below 20 copies/ml) despite therapy. A high VL zenith was associated with a higher probability to have a detectable viremia under cART. Conversely, the probability to have a detectable viral load below 20 copies/ml decreased with time under therapy. Of therapy regimens, protease inhibitor (PI)-based cART was associated with a significantly higher probability of detectable RV compared to nonnucleoside transcriptase inhibitor- or integrase inhibitor-based cART. We found that a PI-based treatment regimen is highly associated with an increased frequency of RV, supporting previous evidence suggesting that PI-based cART regimens could favor ongoing viral replication in some individuals.

Exploring HIV-1 Transmission Dynamics by Combining Phylogenetic Analysis and Infection Timing.

HIV-1 pol sequences obtained through baseline drug resistance testing of patients newly diagnosed between 2013 and 2017 were analyzed for genetic similarity. For 927 patients the information on genetic similarity was combined with demographic data and with information on the recency of infection. Overall, 48.3% of the patients were genetically linked with 11.4% belonging to a pair and 36.9% involved in a cluster of ≥3 members. The percentage of early diagnosed (≤4 months after infection) was 28.6%. Patients of Belgian origin were more frequently involved in transmission clusters (49.7% compared to 15.3%) and diagnosed earlier (37.4% compared to 12.2%) than patients of Sub-Saharan African origin. Of the infections reported to be locally acquired, 69.5% were linked (14.1% paired and 55.4% in a cluster). Equal parts of early and late diagnosed individuals (59.9% and 52.4%, respectively) were involved in clusters. The identification of a genetically linked individual for the majority of locally infected patients suggests a high rate of diagnosis in this population. Diagnosis however is often delayed for >4 months after infection increasing the opportunities for onward transmission. Prevention of local infection should focus on earlier diagnosis and protection of the still uninfected members of sexual networks with human immunodeficiency virus (HIV)-infected members.

Distribution of HCV genotypes in Belgium from 2008 to 2015.

BACKGROUND: The knowledge of circulating HCV genotypes and subtypes in a country is crucial to guide antiviral therapy and to understand local epidemiology. Studies investigating circulating HCV genotypes and their trends have been conducted in Belgium. However they are outdated, lack nationwide representativeness or were not conducted in the general population. METHODS: In order to determine the distribution of different circulating HCV genotypes in Belgium, we conducted a multicentre study with all the 19 Belgian laboratories performing reimbursed HCV genotyping assays. Available genotype and subtype data were collected for the period from 2008 till 2015. Furthermore, a limited number of other variables were collected: some demographic characteristics from the patients and the laboratory technique used for the determination of the HCV genotype. RESULTS: For the study period, 11,033 unique records collected by the participating laboratories were used for further investigation. HCV genotype 1 was the most prevalent (53.6%) genotype in Belgium, with G1a and G1b representing 19.7% and 31.6%, respectively. Genotype 3 was the next most prevalent (22.0%). Further, genotype 4, 2, and 5 were responsible for respectively 16.1%, 6.2%, and 1.9% of HCV infections. Genotype 6 and 7 comprise the remaining <1%. Throughout the years, a stable distribution was observed for most genotypes. Only for genotype 5, a decrease as a function of the year of analysis was observed, with respectively 3.6% for 2008, 2.3% for 2009 and 1.6% for the remaining years. The overall M:F ratio was 1.59 and was mainly driven by the high M:F ratio of 3.03 for patients infected with genotype 3. Patients infected with genotype 3 are also younger (mean age 41.7 years) than patients infected with other genotypes (mean age above 50 years for all genotypes). The patients for whom a genotyping assay was performed in 2008 were younger than those from 2015. Geographical distribution demonstrates that an important number of genotyped HCV patients live outside the Belgian metropolitan cities. CONCLUSION: This national monitoring study allowed a clear and objective view of the circulating HCV genotypes in Belgium and will help health authorities in the establishment of cost effectiveness determinations before implementation of new treatment strategies. This baseline characterization of the circulating genotypes is indispensable for a continuous surveillance, especially for the investigation of the possible impact of migration from endemic regions and prior to the increasing use of highly potent direct-acting antiviral (DAA) agents.

Factors associated with late presentation for HIV care in a single Belgian reference center: 2006-2017.

Late presentation for HIV care is a major issue and the cause of higher morbidity, mortality and transmission. In this regard, we analyzed the characteristics of patients presenting for care at our center from January 2006 to July 2017 (n = 687). The majority of the studied population was of African origin (54.3%) with heterosexual women representing the main group (n = 292; 42.5%). 44% of the patients were late presenters (LP) (presenting for care with CD4 T cells <350/mm3 or an AIDS defining event) and 24% were late presenters with advanced disease (LP-AD) (presenting for care with CD4 T cells <200/mm3 or an AIDS defining event). A very high risk of being LP and LP-AD was associated with Sub-Saharan origin (OR 3.4 and 2.6 respectively). Other factors independently associated with LP or LP-AD were age (OR 1.3), male gender (OR 2.0 and 1.5 respectively) and heterosexual route of transmission (OR 2.4 and 2.3 respectively). A significant increase in HIV screening without forgetting those groups would contribute to earlier HIV diagnosis, a key element to end the HIV epidemic. To achieve this goal, addressing the specific hurdles to HIV testing in the migrant population is critical.

Phylogenetic analysis of the Belgian HIV-1 epidemic reveals that local transmission is almost exclusively driven by men having sex with men despite presence of large African migrant communities.

To improve insight in the drivers of local HIV-1 transmission in Belgium, phylogenetic, demographic, epidemiological and laboratory data from patients newly diagnosed between 2013 and 2015 were combined and analyzed. Characteristics of clustered patients, paired patients and patients on isolated branches in the phylogenetic tree were compared. The results revealed an overall high level of clustering despite the short time frame of sampling, with 47.6% of all patients having at least one close genetic counterpart and 36.6% belonging to a cluster of 3 or more individuals. Compared to patients on isolated branches, patients in clusters more frequently reported being infected in Belgium (95.1% vs. 47.6%; p < 0.001), were more frequently men having sex with men (MSM) (77.9% vs. 42.8%; p < 0.001), of Belgian origin (68.2% vs. 32.9%; p < 0.001), male gender (92.6% vs. 65.8%; p < 0.001), infected with subtype B or F (87.8% vs. 43.4%; p < 0.001) and diagnosed early after infection (55.4% vs. 29.0%; p < 0.001). Strikingly, Sub-Saharan Africans (SSA), overall representing 27.1% of the population were significantly less frequently found in clusters than on individual branches (6.0% vs. 41.8%; p < 0.001). Of the SSA that participated in clustered transmission, 66.7% were MSM and this contrasts sharply with the overall 12.0% of SSA reporting MSM. Transmission clusters with SSA were more frequently non-B clusters than transmission clusters without SSA (44.4% versus 18.2%). MSM-driven clusters with patients of mixed origin may account, at least in part, for the increasing spread of non-B subtypes to the native MSM population, a cross-over that has been particularly successful for subtype F and CRF02_AG. The main conclusions from this study are that clustered transmission in Belgium remains almost exclusively MSM-driven with very limited contribution of SSA. There were no indications for local ongoing clustered transmission of HIV-1 among SSA.

Decision tree for accurate infection timing in individuals newly diagnosed with HIV-1 infection.

BACKGROUND: There is today no gold standard method to accurately define the time passed since infection at HIV diagnosis. Infection timing and incidence measurement is however essential to better monitor the dynamics of local epidemics and the effect of prevention initiatives. METHODS: Three methods for infection timing were evaluated using 237 serial samples from documented seroconversions and 566 cross sectional samples from newly diagnosed patients: identification of antibodies against the HIV p31 protein in INNO-LIA, SediaTM BED CEIA and SediaTM LAg-Avidity EIA. A multi-assay decision tree for infection timing was developed. RESULTS: Clear differences in recency window between BED CEIA, LAg-Avidity EIA and p31 antibody presence were observed with a switch from recent to long term infection a median of 169.5, 108.0 and 64.5 days after collection of the pre-seroconversion sample respectively. BED showed high reliability for identification of long term infections while LAg-Avidity is highly accurate for identification of recent infections. Using BED as initial assay to identify the long term infections and LAg-Avidity as a confirmatory assay for those classified as recent infection by BED, explores the strengths of both while reduces the workload. The short recency window of p31 antibodies allows to discriminate very early from early infections based on this marker. BED recent infection results not confirmed by LAg-Avidity are considered to reflect a period more distant from the infection time. False recency predictions in this group can be minimized by elimination of patients with a CD4 count of less than 100 cells/mm3 or without no p31 antibodies. For 566 cross sectional sample the outcome of the decision tree confirmed the infection timing based on the results of all 3 markers but reduced the overall cost from 13.2 USD to 5.2 USD per sample. CONCLUSIONS: A step-wise multi assay decision tree allows accurate timing of the HIV infection at diagnosis at affordable effort and cost and can be an important new tool in studies analyzing the dynamics of local epidemics or the effects of prevention strategies.

Virological response, HIV-1 drug resistance mutations and genetic diversity among patients on first-line antiretroviral therapy in N'Djamena, Chad: findings from a cross-sectional study.

BACKGROUND: The national antiretroviral therapy in the Republic of Chad provides free of charge antiretroviral regimens and therapeutic monitoring for patients receiving antiretroviral therapy nationwide. For a successful programmatic uptake, these efforts merit to be supported by thorough assessments of antiretroviral therapy response and HIV-1 drug resistance surveillance, especially with risks of cross-resistance due to the gradual stavudine phasing out in such national settings. We therefore evaluated the virological response to antiretroviral therapy, HIV-1 drug resistance emergence and circulating HIV-1 clades in a Chad context. A cross-sectional and prospective study was conducted among 116 patients (41 [δ ± 6.87] years, 59% female) receiving first-line antiretroviral therapy for ≥ 6 months in Ndjamena, Chad, in 2011-2012, enrolled consecutively. To ensure accuracy, plasma viral load was concomitantly measured using Abbott Real-Time and Cobas AmpliPrep/TaqMan (v2.0), and virological failure defined as ≥ 1000 HIV-1 RNA copies/ml. Plasma from patients experiencing virological failure were processed for sequencing of HIV-1 protease-reverse transcriptase using the ANRS-AC.11 resistance testing protocol; drug resistant mutations were interpreted using the ANRS-AC11 algorithm; and phylogenetic analysis was performed using MEGA.v.6. RESULTS: Majority of patients was receiving zidovudine plus lamivudine plus nevirapine (46%), stavudine plus lamivudine plus nevirapine (41%) and tenofovir plus emtricitabine plus efavirenz (11%), for a median time-on-treatment of 5 [IQR 4-7] years. The rate of virological failure was 43% (50/116), with 86% (43/50) sequencing performance. Overall, 32% (37/116) patients presented ≥ one major drug resistant mutation(s), with 29% (34/116) to nucleos(t)ide reverse transcriptase inhibitors (67% [29/43] M184V/I, 30% [13/43] T215Y/F, 19% [8/43] V75A/F/I/L/M, 9% [4/43] K70P/R/W, 9% [4/43] K219E/N/Q and 5% [2/43] A62V); 86% (37/43) to non-nulceos(t)ide reverse transcriptase inhibitors (30% [13/43] K103N/S/E, 26% [11/43] Y181C/V/F/L, 2% [1/43] L100I, 2% [1/43] F227L, 2% [1/43] P225H); and 2% (1/43) to protease inhibitors (M46I, I54V, V82S). Six HIV-1 subtypes were found: 30% circulating recombinant form (CRF02_AG), 30% J, 16% G, 9% A, 9% D, 5% F. CONCLUSIONS: In Chad, almost half of patients are failing first-line antiretroviral therapy after 5 years, with considerable drug resistant mutations at failure. Absence of K65R supports the use of tenofovir-containing regimens as preferred first-line and as suitable drug for second-line combinations, in this setting with significant HIV-1 genetic diversity.

Exploring the link between innate immune activation and thymic function by measuring sCD14 and TRECs in HIV patients living in Belgium.

Microbial translocation is now viewed as a central event in the pathogenesis of chronic inflammation during HIV infection. Thymic function failure is another crucial factor involved in HIV disease progression. The goal of this study was to explore the hypothesis of potential links between microbial translocation and thymic function in HIV-1 patients living in Belgium. The extent of microbial translocation was assessed through the measurement of soluble CD14 (sCD14). T-cell receptor excision circles (sjTRECs and dßTRECs) were used as a measure of thymic function. Data were collected from 75 HIV-infected patients. Simple and complex linear regressions were done to analyze the link between these two processes. We found a statistically relevant negative correlation between thymopoiesis (sjTREC) and sCD14 level (p = 0.004). These results suggest a link between thymic function failure, microbial translocation and innate immune activation.

Factors associated with the continuum of care of HIV-infected patients in Belgium.

INTRODUCTION: We studied factors associated with the continuum of HIV care in Belgium. METHODS: Data of the national registration of new HIV diagnosis and of the national cohort of HIV-infected patients in care were combined to obtain estimates of and factors related with proportions of HIV-infected patients in each step of the continuum of care from diagnosis to suppressed viral load (VL). Factors associated with ignorance of HIV seropositivity were analyzed among patients co-infected with HIV and STI in the Belgian STI sentinel surveillance network. Associated factors were identified by multivariate logistic regression. RESULTS: Among 4038 individuals diagnosed with HIV between 2007 and 2010, 90.3% were linked to care. Of 11684 patients in care in 2010, 90.8% were retained in care up to the following year, 88.3% of those were on ART, of whom 95.3% had suppressed VL (<500 cp/ml) (Figure 1). In multivariate analyses, factors associated with ignoring HIV+ status were being younger (p<0.001), being heterosexual compared to MSM, and of a region of origin other than Belgium, Sub-Saharan Africa and Europe. Non-Belgian regions of origin were associated with lower entry and retention in care (p<0.001 for both). Preoperative HIV testing was associated with lower entry in care (p=0.003). MSM had a higher retention in care (p<0.001), whilst IDU had lower retention (p=0.004). Low CD4 at first clinical contact and clinical reasons for HIV testing were independently associated with being on ART (p<0.001 for both); whilst prenatal HIV diagnosis was associated with lower proportion on ART (p=0.016) and lower proportion with suppressed VL among those on ART (p=0.005). Older age was associated with both being on ART and having suppressed VL among those on ART (p=0.007 and p<0.001 respectively), independently of time since HIV diagnosis (Table 1). CONCLUSIONS: Regions of origin and risk groups (MSM/heterosexual/IDU) are the main factors associated with ignorance of HIV seropositivity, entry and retention in care, but once the HIV patient is retained in care, no effect of these factors on the proportions on ART and with suppressed VL are observed. The association of prenatal HIV diagnosis and proportions on ART and with suppressed VL could be biased by transitory CD4 disturbances during pregnancy and ART discontinuation after pregnancy. The higher probabilities of older patients to be on ART and have suppressed VL once retained in care could be influenced by factors not studied here like comorbidities, adherence or duration on ART.

First-line antiretroviral therapy with nevirapine versus lopinavir-ritonavir based regimens in a resource-limited setting.

OBJECTIVE: To compare WHO first-line antiretroviral therapy (ART) with nonnucleoside reverse transcriptase inhibitors (NNRTI)-based regimen with a boosted protease inhibitor (bPI) regimen in a resource-limited setting regarding treatment outcome and emergence of drug resistance mutations (DRMs). METHODS: Treatment-naive adults were randomized to nevirapine (NVP) or ritonavir-boosted lopinavir (LPV/r) regimens each in combination with tenofovir (TDF)/emtricitabine (FTC) or zidovudine (ZDV)/lamivudine (3TC). Primary endpoint was the incidence of therapeutical (clinical and/or virologic) failure at week 48 with follow-up till week 96. RESULTS: Four hundred and twenty-five patients (120 men; 305 women) received at least one dose of the study drug. mITT analysis showed no difference in proportion of therapeutical failure between treatment arms [67/209 (32%) in NVP vs. 63/216 (29%) LPV/r at week 48 (P = 0.53); 88/209 (42%) in NVP vs. 83/216 (38%) in LPV/r at week 96 (P = 0.49)]. Per-protocol analysis demonstrated significantly more virologic failure with NVP than with LPV/r regimens [at week 48: 19/167 (11%) vs. 7/166 (4%), P = 0.014; at week 96: 27/158 (17%) vs. 13/159 (8%), P =  0.019)]. Drug resistance mutations to NNRTI were detected in 19 out of 22 (86.3%) and dual-class resistance to nucleoside reverse transcriptase inhibitor (NRTI) and NNRTI in 15 out of 27 (68.2%) of NVP failing patients. K65R mutation was present in seven out of 14 patients failing NVP-TDF/FTC regimen. No major protease inhibitor-DRM was detected among LPV/r failing patients. Discontinuation for adverse events was similar between treatment groups. CONCLUSION: In resource-limited settings, first-line NNRTI-NRTI regimen as compared with bPI-based regimen provides similar outcome but is associated with a significantly higher number of virologic failure and resistance mutations in both classes that jeopardize future options for second-line therapy.

Horizontal gene transfer from human host to HIV-1 reverse transcriptase confers drug resistance and partly compensates for replication deficits.

We investigated the origin and the effect of insertion D67D-THGERDLGPA within HIV-1 RT from a patient failing antiviral therapy. The insertion developed within the context of pre-existing NRTI and NNRTI mutations (M41L, L210W, T215Y and N348I). Concurrently, the NRTI mutations T69I and V118I and the NNRTI mutations K103N and Y181C were detected for the first time. High-level drug resistance (fold-changes≥50) and a good replication capacity (87% of wild-type) were observed, significantly higher than for the previous virus without insertion. The insertion was very similar to a region within human chromosome 17 (31/34 nucleotide identity), and had already been detected independently in a Japanese HIV-1 isolate. These results suggest that a particular sequence within human chromosome 17 is prone to horizontal gene transfer into the HIV-1 RT finger subdomain. This insertion confers selective advantage to HIV-1 by its contribution to multi-drug resistance and restoration of impaired replication capacity.

Minocycline attenuates HIV-1 infection and suppresses chronic immune activation in humanized NOD/LtsZ-scidIL-2Rγ(null) mice.

More than a quarter of a century of research has established chronic immune activation and dysfunctional T cells as central features of chronic HIV infection and subsequent immunodeficiency. Consequently, the search for a new immunomodulatory therapy that could reduce immune activation and improve T-cell function has been increased. However, the lack of small animal models for in vivo HIV study has hampered progress. In the current study, we have investigated a model of cord blood haematopoietic progenitor cells (CB-HPCs) -transplanted humanized NOD/LtsZ-scidIL-2Rγ(null) mice in which progression of HIV infection is associated with widespread chronic immune activation and inflammation. Indeed, HIV infection in humanized NSG mice caused up-regulation of several T-cell immune activation markers such as CD38, HLA-DR, CD69 and co-receptor CCR5. T-cell exhaustion markers PD-1 and CTLA-4 were found to be significantly up-regulated on T cells. Moreover, increased plasmatic levels of lipopolysaccharide, sCD14 and interleukin-10 were also observed in infected mice. Treatment with minocycline resulted in a significant decrease of expression of cellular and plasma immune activation markers, inhibition of HIV replication and improved T-cell counts in HIV-infected humanized NSG mice. The study demonstrates that minocycline could be an effective, low-cost adjunctive treatment to regulate chronic immune activation and replication of HIV.