Impact of COVID-19 on the Belgian HIV epidemic: slowdown of HIV transmission and testing and adaptation of care.

BACKGROUND: To gain insight into the impact of the COVID-19 pandemic and containment measures on the HIV epidemic and services, this study aims to describe HIV trends in 2020 and compare them with previous years. METHODS: Belgian national HIV surveillance data 2017-2020 were analysed for trends in HIV testing, HIV diagnoses, VL measurements, ART uptake and PrEP purchase. Descriptive statistics from 2020 are compared to annual averages from 2017 to 2019 (proportional difference, %). RESULTS: In 2020, 725 HIV infections were diagnosed in Belgium (- 21.5% compared to 2019). The decline was most pronounced during the first lockdown in April-May but also present in July-December. The number of HIV tests performed decreased by 17.6% in 2020, particularly in March-May and October-December (- 57.5% in April and -25.4% in November 2020 compared to monthly 2017-19 numbers). Diagnosis of acute HIV infections decreased by 47.1% in 2020 (n = 27) compared to 2019 (n = 51). Late HIV diagnoses decreased by 24.7% (95% CI [- 40.7%; -9.7%]) in 2020 compared to 2019. Of patients in care in 2019, 11.8% interrupted HIV care in 2020 compared to 9.1% yearly in the 3 previous years. The number of HIV patients with VL monitoring per month dropped in March-May 2020, whilst proportions of VL suppression and ART coverage remained above 86% and 98.5% respectively in 2020. PrEP purchases, number of purchasers and starters dropped during April-May 2020 (respectively - 45.7%, - 47.4%, - 77.9% in April compared to February 2020). CONCLUSIONS: The significant decrease in HIV diagnoses in Belgium in 2020 coincided with the COVID-19 pandemic and following containment measures, particularly in April-May during the first lockdown. A slowdown of HIV transmission due to reduced HIV risk exposure is suggested by the halving in diagnosis of acute HIV infections in March-December 2020 compared to the previous year, and the adaptive decrease in PrEP use and PrEP initiation from April onwards. Despite a slight increase in HIV care interruptions, the indicators of quality of HIV care remained stable. Access to prevention, testing and care for all people living with HIV and at risk of acquiring HIV is a priority during and after times of 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.

Comparison of Sanger sequencing for hepatitis C virus genotyping with a commercial line probe assay in a tertiary hospital.

BACKGROUND: The technique most frequently used to genotype HCV is quantitative RT-PCR. This technique is unable to provide an accurate genotype/subtype for many samples; we decided to develop an in-house method with the goal of accurately identifying the genotype of all samples. As a Belgium National Centre of reference for hepatitis, we developed in-house sequencing not only for 5'UTR and core regions starting from VERSANT LiPA amplicons but also for NS5B regions. The sequencing of VERSANT LiPA amplicons might be useful for many laboratories worldwide using the VERSANT LiPA assay to overcome undetermined results. METHODS: 100 samples from Hepatitis C virus infected patients analysed by the VERSANT HCV Genotype 2.0 LiPA Assay covering frequent HCV types and subtypes were included in this study. NS5B, 5'UTR and Core home-made sequencing were then performed on these samples. The sequences obtained were compared with the HCV genomic BLAST bank. RESULTS: All the samples were characterised by the VERSANT LiPA assay (8 G1a, 17 G1b, 6 G2, 11 G3, 13 G4, and 10 G6). It was not possible to discriminate between G6 and G1 by the VERSANT LiPA assay for 8 samples and 27 had an undetermined genotype. Forty-one samples were sequenced for the three regions: NS5B, 5'UTR and Core. Twenty-three samples were sequenced for two regions: 5' UTR and Core and 36 samples were sequenced only for NS5B. Of the 100 samples included, 64 samples were analysed for 5'UTR and Core sequencing and 79 samples were analysed for NS5B sequencing. The global agreement between VERSANT LiPA assay and sequencing was greater than 95%. CONCLUSIONS: In this study, we describe a new, original method to confirm HCV genotypes of samples not discriminated by a commercial assay, using amplicons already obtained by the screening method, here the VERSANT LiPA assay. This method thus saves one step if a confirmation assay is needed and might be of usefulness for many laboratories worldwide performing VERSANT LiPA assay in particular.

Piloting a surveillance system for HIV drug resistance in the European Union.

BackgroundA steady increase in HIV drug resistance (HIVDR) has been demonstrated globally in individuals initiating first-line antiretroviral therapy (ART). To support effective use of ART and prevent spread of HIVDR, monitoring is essential.AimWe piloted a surveillance system for transmitted HIVDR to assess the feasibility of implementation at the European level.MethodAll 31 countries in the European Union and European Economic Area were invited to retrospectively submit data on individuals newly diagnosed with HIV in 2015 who were tested for antiviral susceptibility before ART, either as case-based or as aggregate data. We used the Stanford HIV database algorithm to translate genetic sequences into levels of drug resistance.ResultsNine countries participated, with six reporting case-based data on 1,680 individuals and four reporting aggregated data on 1,402 cases. Sequence data were available for 1,417 cases: 14.5% of individuals (n = 244) showed resistance to at least one antiretroviral drug. In case-based surveillance, the highest levels of transmitted HIVDR were observed for non-nucleoside reverse-transcriptase inhibitors (NNRTIs) with resistance detected in 8.6% (n = 145), followed by resistance to nucleoside reverse-transcriptase inhibitors (NRTI) (5.1%; n = 85) and protease inhibitors (2.0%; n = 34).ConclusionWe conclude that standard reporting of HIVDR data was feasible in the participating countries. Legal barriers for data sharing, consensus on definitions and standardisation of interpretation algorithms should be clarified in the process of enhancing European-wide HIV surveillance with drug resistance information.

Influence of chronic azithromycin treatment on the composition of the oropharyngeal microbial community in patients with severe asthma.

BACKGROUND: This study of the oropharyngeal microbiome complements the previously published AZIthromycin in Severe ASThma (AZISAST) clinical trial, where the use of azithromycin was assessed in subjects with exacerbation-prone severe asthma. Here, we determined the composition of the oropharyngeal microbial community by means of deep sequencing of the amplified 16S rRNA gene in oropharyngeal swabs from patients with exacerbation-prone severe asthma, at baseline and during and after 6 months treatment with azithromycin or placebo. RESULTS: A total of 1429 OTUs were observed, of which only 59 were represented by more than 0.02% of the reads. Firmicutes, Bacteroidetes, Fusobacteria, Proteobacteria and Actinobacteria were the most abundant phyla and Streptococcus and Prevotella were the most abundant genera in all the samples. Thirteen species only accounted for two thirds of the reads and two species only, i.e. Prevotella melaninogenica and Streptococcus mitis/pneumoniae, accounted for one fourth of the reads. We found that the overall composition of the oropharyngeal microbiome in patients with severe asthma is comparable to that of the healthy population, confirming the results of previous studies. Long term treatment (6 months) with azithromycin increased the species Streptococcus salivarius approximately 5-fold and decreased the species Leptotrichia wadei approximately 5-fold. This was confirmed by Boruta feature selection, which also indicated a significant decrease of L. buccalis/L. hofstadtii and of Fusobacterium nucleatum. Four of the 8 treated patients regained their initial microbial composition within one month after cessation of treatment. CONCLUSIONS: Despite large diversity of the oropharyngeal microbiome, only a few species predominate. We confirm the absence of significant differences between the oropharyngeal microbiomes of people with and without severe asthma. Possibly, long term azithromycin treatment may have long term effects on the composition of the oropharygeal microbiome in half of the patients.

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.

Deep Sequencing of HIV-1 RNA and DNA in Newly Diagnosed Patients with Baseline Drug Resistance Showed No Indications for Hidden Resistance and Is Biased by Strong Interference of Hypermutation.

Deep sequencing of plasma RNA or proviral DNA may be an interesting alternative to population sequencing for the detection of baseline transmitted HIV-1 drug resistance. Using a Roche 454 GS Junior HIV-1 prototype kit, we performed deep sequencing of the HIV-1 protease and reverse transcriptase genes on paired plasma and buffy coat samples from newly diagnosed HIV-1-positive individuals. Selection was based on the outcome of population sequencing and included 12 patients with either a revertant amino acid at codon 215 of the reverse transcriptase or a singleton resistance mutation, 4 patients with multiple resistance mutations, and 4 patients with wild-type virus. Deep sequencing of RNA and DNA detected 6 and 43 mutations, respectively, that were not identified by population sequencing. A subsequently performed hypermutation analysis, however, revealed hypermutation in 61.19% of 3,188 DNA reads with a resistance mutation. The removal of hypermutated reads dropped the number of additional mutations in DNA from 43 to 17. No hypermutation evidence was found in the RNA reads. Five of the 6 additional RNA mutations and all additional DNA mutations, after full exclusion of hypermutation bias, were observed in the 3 individuals with multiple resistance mutations detected by population sequencing. Despite focused selection of patients with T215 revertants or singleton mutations, deep sequencing failed to identify the resistant T215Y/F or M184V or any other resistance mutation, indicating that in most of these cases there is no hidden resistance and that the virus detected at diagnosis by population sequencing is the original infecting variant.

HIV-1 RNA and HIV-1 DNA persistence during suppressive ART with PI-based or nevirapine-based regimens.

OBJECTIVES: Whether ART regimens differ in their propensity to allow persistent HIV-1 detection remains unclear. To investigate this, we performed a cross-sectional study to characterize HIV-1 persistence in peripheral blood during suppressive therapy with NRTIs plus a PI or nevirapine. METHODS: Residual plasma HIV-1 RNA was quantified by real-time PCR. Cell-associated proviral total HIV-1 DNA, unspliced and multiply spliced HIV-1 RNA and 2-long terminal repeat (2-LTR) circles were quantified by digital PCR. RESULTS: Comparing PI with nevirapine recipients, residual plasma HIV-1 RNA detection rates were 47/80 (58.8%) versus 37/81 (45.7%), with median (IQR) levels of 4 (3-8) versus 4 (3-7) copies/mL (P = 0.207); detection was less likely with longer duration of suppressive ART (P = 0.020), independently of treatment. HIV-1 DNA was detected in all patients, with median levels of 2.3 (IQR 2.0-2.7) versus 2.5 (IQR 2.1-2.7) log10 copies/10(6) PBMCs, respectively; HIV-1 DNA levels were associated with pre-ART viral load (P = 0.004) and with residual HIV-1 RNA (P = 0.034), unspliced HIV-1 RNA (P = 0.001) and 2-LTR circles (P = 0.005), independently of treatment. CONCLUSIONS: No significant differences were revealed in levels of residual plasma HIV-1 RNA, total HIV-1 DNA or intracellular markers of ongoing virus replication (unspliced and multiply spliced HIV-1 RNA and 2-LTR circles) between treatment groups.

Characteristics and spread to the native population of HIV-1 non-B subtypes in two European countries with high migration rate.

BACKGROUND: Non-B subtypes account for at least 50 % of HIV-1 infections diagnosed in Belgium and Luxembourg. They are considered to be acquired through heterosexual contacts and infect primarily individuals of foreign origin. Information on the extent to which non-B subtypes spread to the local population is incomplete. METHODS: Pol and env gene sequences were collected from 410 non-subtype B infections. Profound subtyping was performed using 5 subtyping tools and sequences of both pol and env. Demographic information, disease markers (viral load, CD4 count) and viral characteristics (co-receptor tropism) were compared between subtypes. Maximum likelihood phylogenetic trees were constructed and examined for clustering. RESULTS: The majority of non-B infections were diagnosed in patients originating from Africa (55.8 %), individuals born in Western Europe represented 30.5 %. Heterosexual transmission was the most frequently reported transmission route (79.9 %), MSM transmission accounted for 12.2 % and was significantly more frequently reported for Western Europeans (25.7 % versus 4.3 % for individuals originating from other regions; p < 0.001). Subtypes A and C and the circulating recombinant forms CRF01_AE and CRF02_AG were the most represented and were included in the comparative analysis. Native Western Europeans were underrepresented for subtype A (14.5 %) and overrepresented for CRF01_AE (38.6 %). The frequency of MSM transmission was the highest for CRF01_AE (18.2 %) and the lowest for subtype A (0 %). No differences in age, gender, viral load or CD4 count were observed. Prevalence of CXCR4-use differed between subtypes but largely depended on the tropism prediction algorithm applied. Indications for novel intersubtype recombinants were found in 20 patients (6.3 %). Phylogenetic analysis revealed only few and small clusters of local transmission but could document one cluster of CRF02_AG transmission among Belgian MSM. CONCLUSIONS: The extent to which non-B subtypes spread in the native Belgian-Luxembourg population is higher than expected, with 30.5 % of the non-B infections diagnosed in native Western Europeans. These infections resulted from hetero- as well as homosexual transmission. Introduction of non-B variants in the local high at risk population of MSM may lead to new sub-epidemics and/or increased genetic variability and is an evolution that needs to be closely monitored.

Comparative performances of HIV-1 RNA load assays at low viral load levels: results of an international collaboration.

Low-level viremia during antiretroviral therapy and its accurate measurement are increasingly relevant. Here, we present an international collaboration of 4,221 paired blood plasma viral load (pVL) results from four commercial assays, emphasizing the data with low pVL. The assays compared were the Abbott RealTime assay, the Roche Amplicor assay, and the Roche TaqMan version 1 and version 2 assays. The correlation between the assays was 0.90 to 0.97. However, at a low pVL, the correlation fell to 0.45 to 0.85. The observed interassay concordance was higher when detectability was defined as 200 copies/ml than when it was defined as 50 copies/ml. A pVL of ∼100 to 125 copies/ml by the TaqMan version 1 and version 2 assays corresponded best to a 50-copies/ml threshold with the Amplicor assay. Correlation and concordance between the viral load assays were lower at a low pVL. Clear guidelines are needed on the clinical significance of low-level viremia.

Markers associated with persisting low-level viraemia under antiretroviral therapy in HIV-1 infection.

OBJECTIVES: To identify host and viral characteristics associated with long-term persisting low-level viraemia (PLLV) under antiretroviral therapy (ART). PATIENTS AND METHODS: Seventy-one ART-treated patients with long-term PLLV (20-250 copies/mL) and 102 control patients with systematically undetectable viral load (VL) were selected retrospectively from ART-treated patients followed at the Ghent HIV reference centre. Host and viral characteristics were compared using univariate and multivariate analyses. RESULTS: Higher plasma VL at therapy initiation (OR 3.52; 95% CI 1.86-6.65; P < 0.001), therapy re-initiation after an interruption (OR 3.94; 95% CI 1.70-9.16; P = 0.001), male gender (OR 4.28; 95% CI 1.40-13.00; P = 0.011), a protease inhibitor-based regimen (OR 2.90; 95% CI 1.20-6.97; P = 0.017) and predicted CCR5 co-receptor tropism (OR 2.53; 95% CI 1.05-6.11; P = 0.039) were independently associated with PLLV. CONCLUSIONS: VL at ART initiation, therapy history, gender, ART regimen and co-receptor tropism were independently associated with PLLV. Gender, therapy history, co-receptor tropism and VL at ART initiation could be valuable predictive markers to identify patients at risk for PLLV.

Reverse transcription of plasma-derived HIV-1 RNA generates multiple artifacts through tRNA(Lys-3)-priming.

In vitro reverse transcription of full-length HIV-1 RNA extracted from the blood plasma of people living with HIV-1 remains challenging. Here, we describe the initiation of reverse transcription of plasma-derived viral RNA in the absence of an exogenous primer. Real-time PCR and Sanger sequencing were applied to identify the source and to monitor the outcome of this reaction. Results demonstrated that during purification of viral RNA from plasma, tRNA(Lys-3) is co-extracted in a complex with the viral RNA. In the presence of a reverse transcription enzyme, this tRNA(Lys-3) can induce reverse transcription, a reaction that is not confined to transcription of the 5' end of the viral RNA. A range of cDNA products is generated, most of them indicative for the occurrence of in vitro strand transfer events that involve translocation of cDNA from the 5' end to random positions on the viral RNA. This process results in the formation of cDNAs with large internal deletions. However, near full-length cDNA and cDNA with sequence patterns resembling multiple spliced HIV-1 RNA were also detected. Despite its potential to introduce significant bias in the interpretation of results across various applications, tRNA(Lys-3)-driven reverse transcription has been overlooked thus far. A more in-depth study of this tRNA-driven in vitro reaction may provide new insight into the complex process of in vivo HIV-1 replication.IMPORTANCEThe use of silica-based extraction methods for purifying HIV-1 RNA from viral particles is a common practice, but it involves co-extraction of human tRNA(Lys-3) due to the strong interactions between these molecules. This co-extraction becomes particularly significant when the extracted RNA is used in reverse transcription reactions, as the tRNA(Lys-3) then serves as a primer. Reverse transcription from tRNA(Lys-3) is not confined to cDNA synthesis of the 5' end of the viral RNA but extends across various regions of the viral genome through in vitro strand transfer events. Co-extraction of tRNA(Lys-3) has been overlooked thus far, despite its potential to introduce bias in downstream, reverse transcription-related applications. The observed events in the tRNA(Lys-3)-induced in vitro reverse transcription resemble in vivo replication processes. Therefore, these reactions may offer a unique model to better understand the replication dynamics of HIV-1.

Touchdown digital polymerase chain reaction for quantification of highly conserved sequences in the HIV-1 genome.

Digital polymerase chain reaction (PCR) is an emerging absolute quantification method based on the limiting dilution principle and end-point PCR. This methodology provides high flexibility in assay design without influencing quantitative accuracy. This article describes an assay to quantify HIV DNA that targets a highly conserved region of the HIV-1 genome that hampers optimal probe design. To maintain high specificity and allow probe binding and hydrolysis of a probe with low melting temperature, a two-stage touchdown PCR was designed with a first round of amplification at high temperature and a subsequent round at low temperature to allow accumulation of fluorescence.

Presence of CXCR4-using HIV-1 in patients with recently diagnosed infection: correlates and evidence for transmission.

BACKGROUND: The prevalence and correlates of CXCR4-use in recently diagnosed patients and the impact of X4/DM transmission remain largely unknown. METHOD: Genotypic coreceptor use determination on the baseline sample of 539 recently diagnosed individuals. Correlation of coreceptor use with clinical, viral and epidemiological data and with information on transmission events as obtained through phylogenetic analysis of protease and reverse transcriptase sequences. Results. CXCR4-use was predicted in 12 to 19% of the patients, depending on the interpretative cutoff used. CXCR4-use was correlated with lower CD4(+) T cell counts and subtype 01_AE infection. No association with viral load was observed. Seven (11%) of 63 transmission clusters and 4 (31%) of 13 donor-source pairs resulted from X4/DM transmission. CONCLUSION: The results confirmed the relation between CXCR4-use at diagnosis and low baseline CD4+ T cell counts. Significantly more CXCR4-use was predicted in 01_AE infections, which may impose constraints on the use of CCR5 antagonists in certain regions of the world. Observations from the transmission cluster analysis contradict the hypothesis that R5 viruses are selected at transmission, and support the idea that R5 or X4/DM infections result from a stochastic process.

Towards Novel HIV-1 Serodiagnostic Tests without Vaccine-Induced Seroreactivity.

Vaccine-induced seroreactivity/positivity (VISR/P) poses a significant and common challenge to HIV vaccine implementation, as up to 95% of vaccine recipients may be misclassified as having HIV infection by current HIV screening and confirmatory serological assays. We investigated whether internal HIV proteins could be used to overcome VISR and discovered a set of 4 antigens (gp41 endodomain, p31 integrase, p17 matrix protein, and Nef) that are recognized by antibodies produced in individuals with HIV infection but not in vaccinated individuals. When evaluated in a multiplex double-antigen bridging ELISA, this antigen combination had specificities of 98.1% prevaccination and 97.1% postvaccination, demonstrating the assay is minimally impacted by vaccine-induced antibodies. The sensitivity was 98.5%, further increasing to 99.7% when p24 antigen testing was included. Results were similar across HIV-1 clades. Although more technical advancements will be desired, this research provides the groundwork for the development of new fourth-generation HIV tests unaffected by VISR. IMPORTANCE While the detection of HIV infection is accomplished by several methods, the most common are serological tests that detect host antibodies produced in response to viral infection. However, the use of current serological tests may present a significant challenge to the adoption of an HIV vaccine in the future because the antibodies to HIV antigens detected in currently available tests also tend to be included as antigens in the HIV vaccines in development. The use of these serological tests may thus result in the misclassification of vaccinated HIV-negative individuals, which can have potential for significant harms for individuals and could prevent the widespread adoption and implementation of HIV vaccines. Our study aimed to identify and evaluate target antigens for inclusion in new serological tests that can be used to identify HIV infections without interference from vaccine-induced antibodies but also fit within existing platforms for HIV diagnostics.

Longitudinal patterns of inflammatory mediators after acute HIV infection correlate to intact and total reservoir.

Background: Despite the beneficial effects of antiretroviral therapy (ART) initiation during acute HIV infection (AHI), residual immune activation remains a hallmark of treated HIV infection. Methods: Plasma concentrations of 40 mediators were measured longitudinally in 39 early treated participants of a Belgian AHI cohort (HIV+) and in 21 HIV-negative controls (HIV-). We investigated the association of the inflammatory profile with clinical presentation, plasma viral load, immunological parameters, and in-depth characterization of the HIV reservoir. Results: While levels of most soluble mediators normalized with suppressive ART, we demonstrated the persistence of a pro-inflammatory signature in early treated HIV+ participants in comparison to HIV- controls. Examination of these mediators demonstrated a correlation with their levels during AHI, which seemed to be viremia-driven, and suggested involvement of an activated myeloid compartment, IFN-γ-signaling, and inflammasome-related pathways. Interestingly, some of these pro-inflammatory mediators correlated with a larger reservoir size and slower reservoir decay. In contrast, we also identified soluble mediators which were associated with favorable effects on immunovirological outcomes and reservoir, both during and after AHI. Conclusion: These data highlight how the persistent pro-inflammatory profile observed in early ART treated individuals is shaped during AHI and is intertwined with viral dynamics.

Benefits of antiretroviral therapy initiation during acute HIV infection.

Objectives: In the last decade, there has been increasing scientific and legislative focus on antiretroviral treatment (ART) for all people living with HIV. Especially early ART initiation, preferably during acute HIV infection, has been named as a promising strategy, both for the individual and for the society. This article will review the benefits and possible future applications of immediate ART initiation during acute HIV infection and explore the remaining hurdles towards this strategy.Results: On an individual level, initiation of ART during acute HIV infection limits the viral reservoir, preserves immune function, and decreases systemic inflammation. In addition, obtaining viral suppression soon after infection can be beneficial for the society by decreasing the chance of onward HIV transmission. Reducing the transmission will reduce HIV incidence and can curtail HIV-related health expenditure. Furthermore, the favorable immunological and virological profile obtained by treating during acute HIV infection will form an ideal starting point for several HIV cure strategies.Conclusions: Initiation of ART during acute HIV infection has shown distinct benefits for the individual, for the society, and for future research on HIV cure. In order to implement this strategy, equal focus should be placed on early diagnosis.

Screening algorithms for HBV, HCV, HIV and syphilis in an anatomical donation program.

BACKGROUND: Users of anatomical donors are at risk of exposure to bloodborne pathogens. This study evaluated screening algorithms for hepatitis B and C virus, human immunodeficiency virus and Treponema pallidum during donor allocation and assessed the impact of postmortem time on hemolysis and how hemolysis affects test results and donor discard rate. METHODS: From 2011 to 2018, demographic data of anatomical donors, time of postmortem blood sampling, presence of sample hemolysis, serological test results (negative; active infection; false reactive screening test; historic infection; inconclusive; technically impracticable) and the actual donor allocation were collected. RESULTS: Donors (n = 537) had a mean age of 77.53 ± 13.67 (24-103) year. Nine (1.68%) had laboratory test results indicative for active infection for hepatitis B (n = 1) and C virus (n = 2), human immunodeficiency virus (n = 5) and T. pallidum (n = 1). Negative screenings ranged from 74.67 to 97.58%, depending on the pathogen. According to the original screening algorithms, 479 (89.20%) donors should have been accepted. In practice, a donor acceptance rate of 91.20% was found. Analysis of potential donor allocation interpretation obstacles resulted in simplification of the in-house laboratory testing algorithms and addition of a nucleic acid test to increase the reliability for identification of active (acute) human immunodeficiency virus infection. Hemolysis was more common when sampling was performed more than 24 h after death (p < 0.001). Hemolytic samples more frequently showed a reactive or indeterminate human immunodeficiency virus test result (p < 0.001). Screening for human immunodeficiency virus and T. pallidum was technically more impracticable when hemolysis was present (p = 0.042 and p = 0.003, respectively). Donors with hemolytic blood samples were more often discarded (46.88%) compared to bodies with non-hemolytic samples (6.32%) (p < 0.001). CONCLUSIONS: Despite the implementation of donor screening algorithms, a significant number of bodies have an inconsistent allocation. New algorithms, to be evaluated in future research, were suggested. Early postmortem blood sampling is key as hemolysis can influence certain test results and donor allocation.

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.

Comparison of phenotypic and genotypic tropism determination in triple-class-experienced HIV patients eligible for maraviroc treatment.

BACKGROUND: Determination of HIV-1 tropism is a pre-requisite to the use of CCR5 antagonists. This study evaluated the potential of population genotypic tropism tests (GTTs) in clinical practice, and the correlation with phenotypic tropism tests (PTTs) in patients accessing routine HIV care. METHODS: Forty-nine consecutive plasma samples for which an original Trofile(TM) assay was performed were obtained from triple-class-experienced patients in need of a therapy change. Viral tropism was defined as the consensus of three or more tropism calls obtained from the combination of two independent population PTT assays (Trofile Biosciences, San Francisco, CA, USA, and Virco, Beerse, Belgium), population GTTs and GTTs based on ultra-deep sequencing. If no consensus was reached, a clonal PTT was performed in order to finalize the tropism call. This two-step approach allowed the definition of a reference tropism call. RESULTS: According to the reference tropism result, 35/49 samples were CCR5 tropic (R5) (patients eligible for maraviroc treatment) and 14/49 were assigned as non-R5 tropic. The non-R5 samples [patients not eligible for maraviroc treatment according to the FDA/European Medicines Agency (EMEA) label] group included both the CXCR4 (X4) samples and the dual and mixed CCR5/CXCR4 (R5/X4) samples. Compared with Trofile(TM) population PTTs, population GTTs showed a higher sensitivity (97%) and a higher negative predictive value (91%), but almost equal specificity and an equal positive predictive value. CONCLUSIONS: In line with recent reports from clinical trial data, our data support the use of population genotypic tropism testing as a tool for tropism determination before the start of maraviroc.