Search

RATIONALE: Tuberculosis (TB) remains a major cause of mortality. A better understanding of the immune responses to mycobacterial antigens may be helpful to develop improved vaccines and diagnostics. OBJECTIVES: The mycobacterial antigen heparin-binding hemagglutinin (HBHA) induces strong IFN-[gamma] responses by circulating lymphocytes from subjects latently infected with Mycobacterium tuberculosis, and low responses associated with CD4(+) regulatory T (Treg) cells in patients with TB. Here, we investigated HBHA-specific IFN-[gamma] responses at the site of the TB disease. METHODS: Bronchoalveolar lavages, pleural fluids, and blood were prospectively collected from 61 patients with a possible diagnosis of pulmonary or pleural TB. HBHA-specific IFN-[gamma] production was analyzed by flow cytometry and ELISA. The suppressive effect of pleural Treg cells was investigated by depletion experiments. MEASUREMENTS AND MAIN RESULTS: The percentages of HBHA-induced IFN-[gamma](+) alveolar and pleural lymphocytes were higher for pulmonary (P < 0.0001) and for pleural (P < 0.01) TB than for non-TB controls. Local CD4(+) and CD8(+) T cells produced the HBHA-specific IFN-[gamma]. This local secretion was not suppressed by Treg lymphocytes, contrasting with previously reported data on circulating lymphocytes. CONCLUSIONS: Patients with TB display differential effector and regulatory T-cell responses to HBHA in local and circulating lymphocytes with a predominant effector CD4(+) and CD8(+) response locally, compared with a predominant Treg response among circulating lymphocytes. These findings may be helpful for the design of new vaccines against TB, and the detection of HBHA-specific T cells at the site of the infection may be a promising tool for the rapid diagnosis of active TB. [ABSTRACT FROM AUTHOR]

Background: Verona integron-encoded metallo-β-lactamase-producing Pseudomonas aeruginosa (VIM-PA) outbreaks are frequently linked to contaminated sink-drains in the intensive care unit (ICU). This study aims to investigate a VIM-PA outbreak occurring at 4 ICUs in a Belgian university center., Methods: Between 01/01/2019 and 30/07/2023, data were retrospectively retrieved. Whole-genome sequencing of VIM-PA was carried out for available isolates and the core genome multilocus sequencing typing (cgMLST) was used to confirm clonality. New case incidence was estimated by analyzing the weekly data of at-risk and VIM-PA-colonized patients, fitting a regression model., Results: Fifty-one patients were colonized, among them, 32 (63%) were infected by VIM-PA, which contributed to 7 deaths. The outbreak investigation showed that 19 (47%) of the examined sink-drains grew at least once a VIM-PA. Two major clusters were observed by cgMLST: ST111 (59 clones with 40 clinical isolates), and ST17 (8 clones with 6 clinical isolates). The estimated incidence rate of new cases was significantly higher in one unit., Conclusions: A 5-year prolonged outbreak at the UZ Brussel ICUs was caused by only 2 VIM-PA clones, both linked to sink-drains, with minimal mutations occurring throughout the years. Statistical modeling found different incidence rates between units. Tailored interventions were hence prioritized., (Copyright © 2024 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.)

Background: The events during acute HIV infection (AHI) set the stage for the subsequent course of the disease. Early initiation of antiretroviral therapy (ART) has been associated with favorable immunovirological outcomes, yet the precise impact of ART timing during AHI remains unclear, particularly on lymphoid tissues., Materials and Methods: The ACS cohort is a prospective cohort study in Belgium, collecting longitudinal clinical data and human bodily material (HBM) from people diagnosed and treated during AHI. The aim of the cohort is to study the impact of ART initiation during AHI on HIV reservoir and immune dysfunction in peripheral blood and anatomical sanctuary sites, as well as its effect on the gut microbiome. The cohort consists of two HBM sampling trajectories: one limited (blood, stool and leukapheresis) and a more extensive one (blood, stool, leukapheresis, colonoscopy, inguinal lymph node excision and lumbar puncture). Here we describe the baseline characteristics, immunovirological outcomes, safety and tolerability of HBM sampling., Results: Between March 2016 and April 2024, 47 participants were enrolled, predominantly men who have sex with men (MSM), with a median age of 36 years [IQR 30-43.5]. Almost 90 % of participants initiated ART within 72 h after study inclusion, irrespective of HBM sampling trajectory. The timing of ART initiation according to the Fiebig stage did not significantly impact immune recovery (CD4/CD8 ratio ≥1) or the time to viral suppression. Approximately 40 % of participants opted for the extensive HBM sampling trajectory during AHI. However, the participation rate for the extensive trajectory decreased by nearly half at the longitudinal follow-up timepoint. In general, study-related procedures were safe and well-tolerated, with limited procedure-related adverse events (AEs). Inguinal lymph node excision was associated with the highest AE rate, in line with previous reports., Conclusions: Our findings reaffirm the beneficial effect of ART initiation during AHI on long term immunovirological outcomes, regardless of Fiebig stage at treatment initiation. Additionally, we demonstrate that the collection of HBM during and longitudinally after AHI is safe and feasible, without compromising time to ART initiation. Cohorts that integrate comprehensive clinical data with high-quality HBM samples are essential to longitudinally study the impact of early ART on reservoir dynamics and immune responses across various anatomical sites after AHI., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Linos Vandekerckhove reports financial support and travel were provided by ViiV Healthcare. Jozefien De Clercq reports travel was provided by ViiV Healthcare. Sarah Gerlo reports travel was provided by ViiV Healthcare. Linos Vandekerckhove reports financial support was provided by Collen-Francqui Fund. Linos Vandekerckhove reports financial support provided by Research Foundation Flanders. Linos Vandekerckhove reports financial support provided by the Belgium Research on AIDS and HIV Consortium (BREACH). Linos Vandekerckhove reports a relationship with Gilead Sciences Inc that includes: consulting or advisory. Linos Vandekerckhove, co-author serves as an Editorial Board Member for Journal of Virus Eradication. Other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors. Published by Elsevier Ltd.)

Background: Limited data are available on Mpox breakthrough infections., Purpose: The purpose of this study is to investigate a Mpox breakthrough outbreak in 3 vaccinated individuals., Methods: Study participants provided informed consent. Serology testing was performed in one involved individual (ID-1) using an in-house assay detecting anti-orthopoxvirus IgG. Whole genome sequencing (WGS) was carried out and compared with the reference sequence ON563414.3 ( https://www.ncbi.nlm.nih.gov/nuccore/ON563414.3/ )., Results: Three individuals vaccinated with modified vaccinia Ankara-Bavaria Nordic contracted Mpox following one sexual intercourse event. One of them (ID-1) had received only one vaccine dose, while the other two were fully vaccinated. ID-1 presented to the sexual health clinic of the Universitair Ziekenhuis Brussel with proctitis related to Mpox. Despite one vaccination, serology testing Three months post vaccine showed absence of Mpox virus (MPXV) specific antibodies in ID-1. In contrast, 2 weeks after the sexual intercourse, seroconversion occurred. Whole genome sequencing of the isolated MPXV showed, compared with the reference sequence, a total of seven single nucleotide variants with four of them indicating protein amino-acid changes., Conclusion: Incomplete MPXV vaccination as well as MPXV variants might result in breakthrough infections. Preventive measures, such as MPVX vaccination, could maintain immunity in individuals with higher risk of MPXV infection, and might lower disease severity., (Copyright © 2024 American Sexually Transmitted Diseases Association. All rights reserved.)

Introduction: In Belgium, oral HIV pre-exposure prophylaxis (PrEP) is primarily provided in specialized clinical settings. Optimal implementation of PrEP services can help to substantially reduce HIV transmission. However, insights into implementation processes, and their complex interactions with local context, are limited. This study examined factors that influence providers' adaptive responses in the implementation of PrEP services in Belgian HIV clinics., Methods: We conducted a qualitative multiple case study on PrEP care implementation in eight HIV clinics. Thirty-six semi-structured interviews were conducted between January 2021 and May 2022 with a purposive sample of PrEP care providers (e.g. physicians, nurses, psychologists), supplemented by 50 hours of observations of healthcare settings and clinical interactions. Field notes from observations and verbatim interview transcripts were thematically analysed guided by a refined iteration of extended Normalisation Process Theory., Results: Implementing PrEP care in a centralized service delivery system required considerable adaptive capacity of providers to balance the increasing workload with an adequate response to PrEP users' individual care needs. As a result, clinic structures were re-organized to allow for more efficient PrEP care processes, compatible with other clinic-level priorities. Providers adapted clinical and policy norms on PrEP care (e.g. related to PrEP prescribing practices and which providers can deliver PrEP services), to flexibly tailor care to individual clients' situations. Interprofessional relationships were reconfigured in line with organizational and clinical adaptations; these included task-shifting from physicians to nurses, leading them to become increasingly trained and specialized in PrEP care. As nurse involvement grew, they adopted a crucial role in responding to PrEP users' non-medical needs (e.g. providing psychosocial support). Moreover, clinicians' growing collaboration with sexologists and psychologists, and interactions with PrEP users' family physician, became crucial in addressing complex psychosocial needs of PrEP clients, while also alleviating the burden of care on busy HIV clinics., Conclusions: Our study in Belgian HIV clinics reveals that the implementation of PrEP care presents a complex-multifaceted-undertaking that requires substantial adaptive work to ensure seamless integration within existing health services. To optimize integration in different settings, policies and guidelines governing PrEP care implementation should allow for sufficient flexibility and tailoring according to respective local health systems., (© 2024 The Author(s). Journal of the International AIDS Society published by John Wiley & Sons Ltd on behalf of International AIDS Society.)

Background: Infection with SARS-CoV-2 in high-risk groups such as kidney transplant and dialysis patients is shown to be associated with a more serious course of the disease. Four years after the start of the COVID-19 pandemic, crucial knowledge on the immune responses in these patient groups is still lacking. Therefore, this study aimed at investigating the humoral immune response after a SARS-CoV-2 infection compared to vaccination as well as the evolution of immunoglobulins over time., Methods: Kidney transplant recipients, patients on haemodialysis or on peritoneal dialysis and healthy controls were included in this longitudinal multicenter study. SARS-CoV-2 anti-RBD, anti-NP and anti-S1S2 immunoglobulin G (IgG) and A (IgA) as well as the neutralizing antibody capacity were measured., Results: Kidney transplant recipients had a significantly better humoral response to SARS-CoV-2 after infection (86.4%) than after a two-dose mRNA vaccination (55.8%) while seroconversion was comparable in patients on haemodialysis after infection (95.8%) versus vaccination (89.4%). In individuals without prior COVID-19, the IgG levels after vaccination were significantly lower in kidney transplant recipients when compared to all other groups. However, the IgA titres remained the highest in this patient group at each time point, both after infection and vaccination. A history COVID-19 was associated with higher antibody levels after double-dose vaccination in all patient categories and, while decreasing, titres remained high six months after double-dose vaccination., Conclusion: Kidney transplant recipients had a more robust humoral response to SARS-CoV-2 following infection compared to a two-dose mRNA vaccination, while patients on haemodialysis exhibited comparable seroconversion rates. Notably, individuals with prior COVID-19 exhibited higher IgG levels in response to vaccination. Hybrid immunity is thus the best possible defence against severe COVID-19 disease and seems also to hold up for these populations. Next, it is not clear whether the higher IgA levels in the kidney transplant recipients is beneficial for neutralizing SARS-CoV-2 or if it is a sign of disease severity., (© 2024. The Author(s).)

Objective: Describe the prevalence and characteristics of people living with HIV (PLWH) in Belgium with limited/exhausted treatment options., Methods: A cross-sectional, multicenter study involving adult treatment-experienced individuals with limited/exhausted treatment options defined as having a multi-drug resistant HIV-1 or a history of multiple treatment changes. The primary outcome was to determine the prevalence of these individuals and classify them based on their two most recent consecutive HIV-1 viral loads (VLs): suppressed (2 VLs < 50 copies/mL), intermediate (≥1 VL between 50-200 copies/mL), or unsuppressed (2 VLs > 200 copies/mL). Secondary outcome was to characterize the participants included in this analysis., Results: There were 119 individuals included (prevalence of 0.97%; 119 of 12 282 in care). The majority were aged > 50 years (88.2%), women represented 35.3%, and individuals were primarily White (54.7%). Median (IQR) CD4 + T-cell count was 635 (400-875) cells/µL and most (42%) were on a 3-drug ART regimen. Overall, 87.4% were classified as suppressed, 9.2% as intermediate, and 3.4% as unsuppressed. On multivariable analysis, CD4 + T-cell count < 200 cells/µL was associated with being classified as intermediate or unsuppressed ( p  = 0.004)., Conclusion: In this analysis of PLWH in Belgium, individuals with limited/exhausted treatment options represented a small fraction. Most were on a 3-drug ART regimen, were virologically suppressed, and had a CD4 + T-cell count within normal range. A small proportion were not virologically suppressed while others, despite being suppressed, were on ≥ 4-drug ART regimens. As such, new therapeutic options are needed to achieve and maintain virologic suppression in such individuals while decreasing their pill burden.

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., Competing Interests: LV has received consulting fees and travel grants from Gilead Sciences and ViiV Healthcare, paid to his institution. JDC and SG have received travel grants from ViiV Healthcare. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 De Clercq, De Scheerder, Mortier, Verhofstede, Vandecasteele, Allard, Necsoi, De Wit, Gerlo and Vandekerckhove.)

Background: Adequate diagnosis of bacterial respiratory tract co-/superinfection (bRTI) in coronavirus disease (COVID-19) patients is challenging, as there is insufficient knowledge about the role of risk factors and (para)clinical parameters in the identification of bacterial co-/superinfection in the COVID-19 setting. Empirical antibiotic therapy is mainly based on COVID-19 severity and expert opinion, rather than on scientific evidence generated since the start of the pandemic., Purpose: We report the best available evidence regarding the predictive value of risk factors and (para)clinical markers in the diagnosis of bRTI in COVID-19 patients., Methods: A multidisciplinary team identified different potential risk factors and (para)clinical predictors of bRTI in COVID-19 and formulated one or two research questions per topic. After a thorough literature search, research gaps were identified, and suggestions concerning further research were formulated. The quality of this narrative review was ensured by following the Scale for the Assessment of Narrative Review Articles., Results: Taking into account the scarcity of scientific evidence for markers and risk factors of bRTI in COVID-19 patients, to date, COVID-19 severity is the only parameter which can be associated with higher risk of developing bRTI., Conclusions: Evidence on the usefulness of risk factors and (para)clinical factors as predictors of bRTI in COVID-19 patients is scarce. Robust studies are needed to optimise antibiotic prescribing and stewardship activities in the context of COVID-19.

COVID-19 can induce neurological sequelae, negatively affecting the quality of life. Unravelling this illness's impact on structural brain connectivity, white-matter microstructure (WMM), and cognitive performance may help elucidate its implications. This cross-sectional study aimed to investigate differences in these factors between former hospitalised COVID-19 patients (COV) and healthy controls. Group differences in structural brain connectivity were explored using Welch-two sample t-tests and two-sample Mann-Whitney U tests. Multivariate linear models were constructed (one per region) to examine fixel-based group differences. Differences in cognitive performance between groups were investigated using Wilcoxon Rank Sum tests. Possible effects of bundle-specific FD measures on cognitive performance were explored using a two-group path model. No differences in whole-brain structural organisation were found. Bundle-specific metrics showed reduced fiber density (p = 0.012, Hedges' g = 0.884) and fiber density cross-section (p = 0.007, Hedges' g = 0.945) in the motor segment of the corpus callosum in COV compared to healthy controls. Cognitive performance on the motor praxis and digit symbol substitution tests was worse in COV than healthy controls (p < 0.001, r = 0.688; p = 0.013, r = 422, respectively). Associations between the cognitive performance and bundle-specific FD measures differed significantly between groups. WMM and cognitive performance differences were observed between COV and healthy controls., (© 2023. Springer Nature Limited.)

Objectives: Our objective was to evaluate the efficacy, durability, and tolerability of bictegravir/emtricitabine/tenofovir alafenamide (BIC/FTC/TAF) in a real-world setting in Belgium., Methods: This was a retrospective, multicentre cohort study involving adult treatment-naïve (TN) and treatment-experienced (TE) people living with HIV receiving BIC/FTC/TAF between 1 January 2019 and 30 September 2020. The primary outcome was rate of virological suppression (plasma HIV-1 viral load <50 copies/mL; on-treatment analysis) at weeks 24 and 48. The main secondary outcomes included loss of virological suppression (LVS; two consecutive viral loads of >200 copies/mL after being virologically suppressed) by week 48 and analysis of resistance-associated mutations at time of LVS; tolerability of BIC/FTC/TAF over the 48-week study period; and change in weight and proportion of participants reporting a >10% weight gain at week 48., Results: Overall, 2001 participants were included. Through 48 weeks, overall rate of virological suppression was 93.5%, with similar results observed in the following subgroups: age ≥50 years (92.7%), women (92.8%), Black sub-Saharan African (91%), TN (94%), TE (93.2%), and non-suppressed at baseline (86.6%). LVS was observed in 0.7% (n = 14) of participants, with one participant developing resistance-associated mutations to nucleoside reverse transcriptase inhibitors (184 V) and integrase strand transfer inhibitors (263KR). Of the 131 (6.5%) treatment discontinuations, the most common reason was an adverse event (2.4%), with the most frequent being central nervous system/psychiatric (0.4%) and gastrointestinal (0.4%) toxicity. Median weight gain at week 48 was 2 kg (interquartile range -1 to 5), and a >10% weight increase was observed in 11.6% of participants., Conclusion: In this large real-world cohort, BIC/FTC/TAF showed excellent virological efficacy in a diverse population of patients with HIV. Rare occurrence of emergent drug resistance was observed, and treatment was well tolerated., (© 2023 British HIV Association.)

Although natural killer (NK) cells have been studied in connection with dendritic cell (DC)-based vaccination in the field of cancer immunology, their role has barely been addressed in the context of therapeutic vaccination against HIV-1. In this study, we evaluated whether a therapeutic DC-based vaccine consisting of monocyte-derived DCs electroporated with Tat, Rev and Nef encoding mRNA affects NK cell frequency, phenotype and functionality in HIV-1-infected individuals. Although the frequency of total NK cells did not change, we observed a significant increase in cytotoxic NK cells following immunisation. In addition, significant changes in the NK cell phenotype associated with migration and exhaustion were observed together with increased NK cell-mediated killing and (poly)functionality. Our results show that DC-based vaccination has profound effects on NK cells, which highlights the importance of evaluating NK cells in future clinical trials looking at DC-based immunotherapy in the context of HIV-1 infection., (© 2023. The Author(s).)

Objectives: A paradigm shift from three-drug regimens to two-drug regimens (2DRs) is currently taking place in real-world clinical practice. This study aimed to describe the efficacy, durability, and tolerability of dolutegravir (DTG)/lamivudine (3TC) and DTG/rilpivirine (RPV) in a real-world setting., Methods: This was a retrospective, observational, multicentre (ten centres in Belgium) study involving adult treatment-naïve and treatment-experienced people living with HIV on DTG/3TC or DTG/RPV between 1 January 2019 and 30 September 2020. The primary endpoint was rate of virological suppression (VS; plasma HIV-1 viral load [VL] <50 copies/ml) using an on-treatment analysis. Main secondary endpoints included the proportion of people that experienced loss of VS (LVS; defined as two consecutive HIV-1 VLs of >200 copies/ml after initially achieving VS) and a resistance analysis at the time of LVS; rate, incidence, and reasons for discontinuation of treatment (stopping treatment or changing any component of the 2DR); and change in weight, along with the proportion of people reporting a >10% weight gain. Ordinal logistic regression analysis examined associations between baseline variables and >10% on-treatment weight gain., Results: Overall, 948 people were included, of whom 734 (77%) were on DTG/3TC and 214 (23%) were on DTG/RPV. Baseline characteristics included 54% aged ≥50 years, 31% female, 31% Black sub-Saharan African, 95% treatment-experienced, and 8% with HIV-1 VL ≥50 copies/ml. Through 48 weeks, the rate of VS for the overall cohort was 98.3% (99.1% with 3TC; 96.2% with RPV). LVS was observed in 0.5% (n = 5) of the overall population (n = 1 [3TC group], n = 4 [RPV group]). There were 40 treatment discontinuations (4.2%, n = 27 [3TC group]; n = 13 [RPV group]), corresponding to an incidence of 4.7 per 100 patient-years. The most common reason for discontinuation was an adverse event (1.4%), with neurotoxicity the most frequent (0.5%). Median on-treatment weight gain at week 48 was 1 kg (interquartile range [IQR] -1-3) overall, 1 kg (IQR -1-3) in the 3TC group, and 2 kg (IQR 0-4) in the RPV group. A >10% weight increase was observed in 6.3% of people. Regression analysis showed that being on a tenofovir disoproxil fumarate-based regimen prior to 2DR initiation was the only variable associated with a >10% increase in weight from baseline (odds ratio 3.48; 95% confidence interval 1.13-10.68; p = 0.038)., Conclusion: In this real-world analysis, the 2DRs analysed were effective, durable, and safe for those who were treatment-naive and treatment-experienced. A slight increase in weight was associated with these regimens., (© 2022 British HIV Association.)

Background: Long-term sequelae of COVID-19 can result in reduced functionality of the central nervous system and substandard quality of life. Gaining insight into the recovery trajectory of admitted COVID-19 patients on their cognitive performance and global structural brain connectivity may allow a better understanding of the diseases' relevance., Objectives: To assess whole-brain structural connectivity in former non-intensive-care unit (ICU)- and ICU-admitted COVID-19 survivors over 2 months following hospital discharge and correlate structural connectivity measures to cognitive performance., Methods: Participants underwent Magnetic Resonance Imaging brain scans and a cognitive test battery after hospital discharge to evaluate structural connectivity and cognitive performance. Multilevel models were constructed for each graph measure and cognitive test, assessing the groups' influence, time since discharge, and interactions. Linear regression models estimated whether the graph measurements affected cognitive measures and whether they differed between ICU and non-ICU patients., Results: Six former ICU and six non-ICU patients completed the study. Across the various graph measures, the characteristic path length decreased over time (β = 0.97, p = 0.006). We detected no group-level effects (β = 1.07, p = 0.442) nor interaction effects (β = 1.02, p = 0.220). Cognitive performance improved for both non-ICU and ICU COVID-19 survivors on four out of seven cognitive tests 2 months later (p < 0.05)., Conclusion: Adverse effects of COVID-19 on brain functioning and structure abate over time. These results should be supported by future research including larger sample sizes, matched control groups of healthy non-infected individuals, and more extended follow-up periods., (© 2023. The Author(s).)

The unprecedented COVID-19 pandemic took the form of successive variant waves, spreading across the globe. We wanted to investigate any shift in hospitalised patients' profiles throughout the pandemic. For this study, we used a registry that collected data automatically from electronic patient health records. We compared clinical data and severity scores, using the National Institute of Health (NIH) severity scores, from all patients admitted for COVID-19 during four SARS-CoV-2 variant waves. Our study concluded that patients hospitalised for COVID-19 showed very different profiles across the four variant waves in Belgium. Patients were younger during the Alpha and Delta waves and frailer during the Omicron period. 'Critical' patients according to the NIH criteria formed the largest fraction among the Alpha wave patients (47.7%), while 'severe' patients formed the largest fraction among Omicron patients (61.6%). We discussed host factors, vaccination status, and other confounders to put this into perspective. High-quality real-life data remain crucial to inform stakeholders and policymakers that shifts in patients' clinical profiles have an impact on clinical practice.

Objectives: Physicians could request compassionate use of oral and long-acting (LA) cabotegravir + rilpivirine for people living with HIV-1 under a single-patient request programme supported by ViiV Healthcare and Janssen. Outcomes are reported., Methods: Eligibility criteria included need for parenteral therapy, no primary resistance mutations to cabotegravir or rilpivirine, and established retention in care. Demographic, efficacy, and safety data were obtained from standardized programme applications and quarterly clinical updates. Individuals received a loading dose of LA cabotegravir 600 mg + rilpivirine 900 mg, followed by LA maintenance doses of 400 mg/600 mg every 4 weeks; some received lead-in oral cabotegravir and rilpivirine., Results: Through July 2020, 35 people living with HIV-1 had data available. The most frequent reason for compassionate use request was chronic non-adherence due to psychological conditions (n = 15). Of 35 people living with HIV-1, 28 had detectable viremia (median viral load 60 300 copies/mL) and seven were virologically suppressed at programme entry; 16/28 and 6/7 achieved or maintained virological suppression at data cutoff, respectively. Seven people living with HIV-1 discontinued for incomplete virological response, six with detectable viremia at initiation; six and four had new reverse transcriptase and integrase mutations at discontinuation, respectively. Six non-fatal serious adverse events were reported, two considered possibly treatment related. Four deaths were reported; none were treatment related. One individual reported two pregnancies and continued LA dosing., Conclusions: Most people living with HIV-1 had advanced disease and achieved (16/28) or maintained (6/7) virological suppression with LA therapy. Cabotegravir LA + rilpivirine LA as compassionate use provided a valuable treatment option for individuals with adherence issues with oral therapy and advanced disease., (© 2022 ViiV Healthcare and The Author. HIV Medicine published by John Wiley & Sons Ltd on behalf of British HIV Association.)

Objectives: Suppression of viral replication in patients on antiretroviral therapy (ART) is determined by plasma viral load (pVL) measurement. Whenever pVL reaches values below the limit of quantification, the qualitative parameter 'target detected' or 'target not detected' is available but often not reported to the clinician. We investigated whether qualitative pVL measurements can be used to estimate the viral reservoir size., Design: The study recruited 114 people with HIV (PWH) who are stable on ART between 2016 and 2018. The percentage of pVL measurements qualitatively reported as 'target detected' (PTD) within a 2-year period was calculated., Methods: t-DNA and US-RNA were used to estimate viral reservoir size and were quantified on peripheral blood mononuclear cells (PBMCs) using droplet digital PCR., Results: A median of 6.5 pVL measurements over a 2-year period was evaluated for each participant to calculate PTD. A positive correlation was found between t-DNA and PTD (r = 0.24; P = 0.011) but not between US-RNA and PTD (r = 0.1; P = 0.3). A significantly lower PTD was observed in PWH with a small viral reservoir, as estimated by t-DNA less than 66 copies/106 PBMCs and US-RNA less than 10 copies/106 PBMCs, compared with PWH with a larger viral reservoir (P = 0.001). We also show that t-DNA is detectable whenever PTD is higher than 56% and that ART regimen does not affect PTD., Conclusion: Our study shows that PTD provides an efficient parameter to preselect participants with a small viral reservoir based on already available pVL data for future HIV cure trials., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Background: In the period following the declaration of the COVID-19 pandemic, more evidence became available on the epidemiology of bacterial co-/superinfections (bCSs) in hospitalized COVID-19 patients. Various European therapeutic guidelines were published, including guidance on rational antibiotic use., Methods: In this letter to the editor, we provide an overview of the largest meta-analyses or prospective studies reporting on bCS rates in COVID-19 patients and discuss why the reader should interpret the results of those reports with care. Moreover, we compare different national and international COVID-19 therapeutic guidelines from countries of the European Union. Specific attention is paid to guidance dedicated to rational antibiotic use., Results: We found a significant heterogeneity in studies reporting on the epidemiology of bCSs in COVID-19 patients. Moreover, European national and international guidelines differ strongly from each other, especially with regard to the content and extent of antibiotic guidance in hospitalized COVID-19 patients., Conclusion: A standardized way of reporting on bCSs and uniform European guidelines on rational antibiotic use in COVID-19 patients are crucial for antimicrobial stewardship teams to halt unnecessary antibiotic use in the COVID-19 setting., Competing Interests: The authors declare no conflict of interest.

Background: Healthcare-associated SARS-CoV-2 infections need to be explored further. Our study is an analysis of hospital-acquired infections (HAIs) and ambulatory healthcare workers (aHCWs) with SARS-CoV-2 across the pandemic in a Belgian university hospital., Methods: We compared HAIs with community-associated infections (CAIs) to identify the factors associated with having an HAI. We then performed a genomic cluster analysis of HAIs and aHCWs. We used this alongside the European Centre for Disease Control (ECDC) case source classifications of an HAI., Results: Between March 2020 and March 2022, 269 patients had an HAI. A lower BMI, a worse frailty index, lower C-reactive protein (CRP), and a higher thrombocyte count as well as death and length of stay were significantly associated with having an HAI. Using those variables to predict HAIs versus CAIs, we obtained a positive predictive value (PPV) of 83.6% and a negative predictive value (NPV) of 82.2%; the area under the ROC was 0.89. Genomic cluster analyses and representations on epicurves and minimal spanning trees delivered further insights into HAI dynamics across different pandemic waves. The genomic data were also compared with the clinical ECDC definitions for HAIs; we found that 90.0% of the 'definite', 87.8% of the 'probable', and 70.3% of the 'indeterminate' HAIs belonged to one of the twenty-two COVID-19 genomic clusters we identified., Conclusions: We propose a novel prediction model for HAIs. In addition, we show that the management of nosocomial outbreaks will benefit from genome sequencing analyses.

Introduction: Legionnaires' disease (LD) is a recognised cause of community-acquired pneumonia. However, Legionella is an overlooked pathogen in hospital-acquired pneumonia. The European Surveillance System 2008-2017 found 23% of the Belgian LD reported cases being healthcare-related, with a higher death-rate than in community-acquired patients. This study aims to describe patients admitted for community-acquired LD or affected by hospital-acquired LD and investigate discriminants associated with lethality., Methods: Medical records were retrospectively reviewed at three Belgian University Hospitals, between 1 January 2016 up to 31 January 2019. Hospital-acquired LD was defined as symptom onset at 10 days or more after admission, according to the Centres for Disease Control and prevention. Community-acquired LD was defined as diagnosis at admission or within 10 days after admission., Results: Fifty patients were included in the study, among them 26% were diagnosed with hospital-acquired LD. The case-fatality rate was 22%, with eight of the eleven deceased patients (73%) being in the hospital-acquired LD group. Medical history of asthma or chronic obstructive pulmonary disease and higher sequential organ failure assessment (SOFA) score at diagnosis were more frequently observed in the hospital-acquired LD group. Furthermore, significantly lower SOFA score at diagnosis of LD and higher rates of treatment with levofloxacin or moxifloxacin were observed in survivors., Conclusion: In the current cohort, LD death-rate was mainly driven by hospital-acquired LD patients. Hospital-acquired LD might especially affect patients with chronic respiratory disease. Respiratory fluoroquinolones treatment and lower SOFA score at diagnosis may be associated with favourable outcomes.

A major determinant for the success of mRNA-based vaccines is the composition of the nanoparticles (NPs) used for formulation and delivery. Cationic peptides represent interesting candidate carriers for mRNA, since they have been shown to efficiently deliver nucleic acids to eukaryotic cells. mRNA NPs based on arginine-rich peptides have previously been demonstrated to induce potent antigen-specific CD8 + T-cell responses. We therefore compared the histidine-rich amphipathic peptide LAH4-L1 (KKALLAHALHLLALLALHLAHALKKA) to the fully substituted arginine variant (LAH4-L1R) for their capacity to formulate mRNA and transfect dendritic cells (DCs). Although both peptides encapsulated mRNA to the same extent, and showed excellent uptake in DCs, the gene expression level was significantly higher for LAH4-L1. The LAH4-L1-mRNA NPs also resulted in enhanced antigen presentation in the context of MHC I compared to LAH4-L1R in primary murine CD103 + DCs. Both peptides induced DC maturation and inflammasome activation. Subsequent ex vivo stimulation of OT-I splenocytes with transfected CD103 + DCs resulted in a high proportion of polyfunctional CD8 + T cells for both peptides. In addition, in vivo immunization with LAH4-L1 or LAH4-L1R-mRNA NPs resulted in proliferation of antigen-specific T cells. In conclusion, although LAH4-L1 outperformed LAH4-L1R in terms of transfection efficiency, the immune stimulation ex vivo and in vivo was equally efficient.

Background: Health care workers (HCW) are facing the Coronavirus disease 2019 (CoViD-19) epidemic. Consequently, psychological impairments have been reported. However, literature showed controversial results on the relationship between gender, frontline HCW, and psychological impairments. This study aims to investigate CoViD-19 fear and reluctance to work in HCW., Methods: Employees who worked between April and October 2020 at the UZ Brussel were included. Data were prospectively collected in 2 phases through a survey together with serological tests. Sampling strategy was convenience sampling., Results: About 2,336 employees completed the study and response rate was 70%. The prevalence of severe CoViD-19 fear in participants increased from 9% to 15%. Employees showing way less motivation rose from 9% to 14%. The seroprevalence was 7.4% and 7.9%. Multivariable analysis found a relation between reluctance to work, study phase, female gender, shortage of personal protective equipment, and poor education on CoViD-19. Furthermore, CoViD-19 fear was related to the study phase, older age, female gender, being second-line HCW, reported exposure to CoViD-19 during work, and insufficient education on CoViD-19., Discussion: Seroprevalence remained rather stable, but fear and reluctance to work significantly increased. Differences in time of data collection together with epidemiological setting might be responsible for conflicting data reported in literature., Conclusions: The evolution of the epidemiological setting might influence the results of studies investigating psychological impairments in HCW., (Copyright © 2021 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.)

Despite the low rates of bacterial co-/superinfections in COVID-19 patients, antimicrobial drug use has been liberal since the start of the COVID-19 pandemic. Due to the low specificity of markers of bacterial co-/superinfection in the COVID-19 setting, overdiagnosis and antimicrobial overprescription have become widespread. A quantitative and qualitative evaluation of urinary tract infection (UTI) diagnoses and antimicrobial drug prescriptions for UTI diagnoses was performed in patients admitted to the COVID-19 ward of a university hospital between 17 March and 2 November 2020. A team of infectious disease specialists performed an appropriateness evaluation for every diagnosis of UTI and every antimicrobial drug prescription covering a UTI. A driver analysis was performed to identify factors increasing the odds of UTI (over)diagnosis. A total of 622 patients were included. UTI was present in 13% of included admissions, and in 12%, antimicrobials were initiated for a UTI diagnosis (0.71 daily defined doses (DDDs)/admission; 22% were scored as 'appropriate'). An evaluation of UTI diagnoses by ID specialists revealed that of the 79 UTI diagnoses, 61% were classified as probable overdiagnosis related to the COVID-19 hospitalization. The following factors were associated with UTI overdiagnosis: physicians who are unfamiliar working in an internal medicine ward, urinary incontinence, mechanical ventilation and female sex. Antimicrobial stewardship teams should focus on diagnostic stewardship of UTIs, as UTI overdiagnosis seems to be highly prevalent in admitted COVID-19 patients.

Introduction: Although bacterial co- and superinfections are rarely present in patients with COVID-19, overall antibiotic prescribing in admitted patients is high. In order to counter antibiotic overprescribing, antibiotic stewardship teams need reliable data concerning antibiotic prescribing in admitted patients with COVID-19., Methods: In this prospective observational cohort study, we performed a quantitative and qualitative evaluation of antibiotic prescriptions in patients admitted to the COVID-19 ward of a 721-bed Belgian university hospital between 1 May and 2 November 2020. Data on demographics, clinical and microbiological parameters and antibiotic consumption were collected. Defined daily doses (DDD) were calculated for antibiotics prescribed in the context of a (presumed) bacterial respiratory tract infection and converted into two indicators: DDD/admission and DDD/100 hospital bed days. A team of infectious disease specialists performed an appropriateness evaluation for every prescription. A driver analysis was performed to identify factors increasing the odds of an antibiotic prescription in patients with a confirmed COVID-19 diagnosis., Results: Of 403 eligible participants with a suspected COVID-19 infection, 281 were included. In 13.8% of the 203 admissions with a COVID-19 confirmed diagnosis, antibiotics were initiated for a (presumed) bacterial respiratory tract co-/superinfection (0.86 DDD/admission; 8.92 DDD/100 bed days; 39.4% were scored as 'appropriate'). Five drivers of antibiotic prescribing were identified: history of cerebrovascular disease, high neutrophil/lymphocyte ratio in male patients, age, elevated ferritin levels and the collection of respiratory samples for bacteriological analysis., Conclusion: In the studied population, the antibiotic consumption for a (presumed) bacterial respiratory tract co-/superinfection was low. In particular, the small total number of DDDs in patients with confirmed COVID-19 diagnosis suggests thoughtful antibiotic use. However, antibiotic stewardship programmes remain crucial to counter unnecessary and inappropriate antibiotic use in hospitalized patients with COVID-19., Trial Registration: The study is registered at ClinicalTrials.gov (NCT04544072)., (© 2021. The Author(s).)

Background: The diagnostic gold standard for Coronavirus-2019 disease (CoViD-19) is reverse transcriptase-polymerase chain reaction (RT-PCR). However, its sensitivity might be suboptimal. The current study aims to investigate predictive factors for false-negative nasopharyngeal RT-PCR in CoViD-19 patients. Additionally, the specificity and sensitivity of RT-PCR on the nasopharyngeal swab, serology and chest computerized-tomography (CCT) as a screening tool for the diagnosis of CoViD-19 were investigated., Methods: Medical records of patients admitted at the university hospital UZ Brussel during the CoViD-19 epidemic were reviewed. A group of CoViD-19 patients with false-negative RT-PCR was identified through scrupulous examination of medical records. Serological testing was performed through chemiluminescent microparticle assay., Results: Eighteen CoViD-19 patients with 'false negative' RT-PCR were identified and compared to 51 'true positives'. Logistic regression for prediction of 'false negative' RT-PCR found significantly higher serology results at hospitalization and more intensive care unit admission in the group with false-negative testing. In a cohort of 228 patients, the sensitivity of RT-PCR for the diagnosis of CoViD-19 was 85%. The sensitivity of serology was 86% and its specificity 92%. Chest computerized-tomography (CCT) showed a sensitivity of 93%, its specificity was 62%. By combining RT-PCR and serology results any 'false negative' could be excluded., Conclusions: In this cohort, the sensitivity and specificity of RT-PCR and serology for the diagnosis of CoViD-19 were high and comparable. CCT had the highest sensitivity and confirmed its efficacy as a screening tool. CoViD-19 patients, who have a more severe presentation, might have negative RT-PCR and positive serology results.

Introduction : Lemierre's syndrome is a septic thromboembolic complication of an oropharyngeal or neck infection, primarily caused by Fusobacterium species. Although it usually affects young healthy patients, some case reports describe this syndrome in older population. Methods : A case report and a systematic review of the literature were conducted to investigate the late onset of Lemierre's syndrome. Forty-one articles were selected for the qualitative analysis, 39 for the quantitative analysis. Results : The average age of the study population was 52 years old. Diabetes mellitus and upper gastro-intestinal malignancy, common comorbidities in the study population, might play a role in the development of late-onset Lemierre's syndrome. Empiric antibiotic treatment should cover Fusobacterium and Streptococcus species both, which may cooperate to induce purulent disease. Reported unfavourable outcome was more than expected. Conclusion : Lemierre's syndrome in adulthood may differ from the usual version. This disease may further pass unrecognized, if presented out of the expected age range. Nevertheless, early diagnosis and prompt treatment are a requisite to prevent morbidity and mortality, which may be higher in this older population.

Background/aims: SARS-CoV-2 is highly contagious. More evidence concerning extrapulmonary transmission routes such as the eyes is urgently needed. Although the humoral immune response is important in the viral containment, the local response in tears has not yet been studied. The aim of our study was twofold: to assess the prevalence of both SARS-CoV-2 RNA and antibodies in tear fluid., Methods: In a first series, nasopharyngeal sampling and tear sampling by Schirmer test strips were performed in 26 acutely ill patients with COVID-19 to assess the presence of SARS-CoV-2 RNA by reverse transcription PCR. In a second series, IgG and IgA responses to SARS-CoV-2 spike protein in serum and tear fluid of convalescent individuals (n=22) were compared with control individuals (n=15) by ELISA., Results: SARS-CoV-2 RNA was detected in tears of 7/26 (26.9%) patients with COVID-19. None of them had ocular symptoms. Convalescent individuals displayed a significant higher ratio of IgG (p<0.0001) and IgA (p=0.0068) in tears compared with control individuals. A sensitivity of 77.3% and specificity of 93.3% was observed for IgG, and 59.1% and 100% for IgA., Conclusions: Our results demonstrate the presence of SARS-CoV-2 RNA and a local IgG and IgA immune response in tear fluid. These data confirm the possibility of SARS-CoV-2 transmission through tear fluid and the importance of the eye as a first defence against SARS-CoV-2, indicating the potential of tears as a non-invasive surrogate for serum in monitoring the host immune response., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Recent EACVI recommendations described the importance of limiting cardiovascular imaging during the COVID-19 pandemic in order to reduce virus transmission, protect healthcare professionals from contamination, and reduce consumption of personal protective equipment. However, an elevated troponin remains a frequent request for cardiac imaging in COVID-19 patients, partly because it signifies cardiac injury due to a variety of causes and partly because it is known to convey a worse prognosis. The present paper aims to provide guidance to clinicians regarding the appropriateness of cardiac imaging in the context of troponin elevation and myocardial injury, how best to decipher the mechanism of myocardial injury, and how to guide patient management., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.permissions@oup.com.)

Introduction: Viral remission after analytical treatment interruption (ATI), termed post-treatment control, has been described in a small proportion of HIV-positive patients. This phenomenon has been separately associated to both low levels of HIV-1 proviral DNA as well as cell-associated RNA. We investigated whether the combination of both parameters could help predict delayed viral rebound after treatment interruption (TI)., Methods: We conducted an open single-arm ATI study in four Belgian HIV reference centres from January 2016 to July 2018. Eligible participants were adults who had fewer than 50 HIV-1 RNA copies/mL for more than two years, more than 500 CD4 cells/µL for more than three months, and were in general good health. Consenting participants who had fewer than 66 copies total HIV-1 DNA (t-DNA) and fewer than 10 copies cell-associated HIV-1 unspliced RNA (US-RNA) per million peripheral blood mononuclear cells (PBMCs), interrupted therapy and were monitored closely. Antiretroviral therapy (ART) was resumed after two consecutive viral loads exceeding 1000 copies or one exceeding 10,000 copies/mL. The primary outcome was the proportion of participants with fewer than 50 HIV-1 RNA copies/mL 48 weeks after TI. Secondary outcomes were time to viral rebound, the frequency of serious adverse events (AEs) and evolution of t-DNA and US-RNA after TI., Results: All 16 consenting participants who interrupted therapy experienced rapid viral rebound two to eight weeks after TI. No serious AEs were observed. Levels of t-DNA and US-RNA increased after TI but returned to pre-ATI levels after treatment restart. None of the studied demographic, clinical and biological parameters were predictive of time of viral rebound., Conclusions: The combination of low levels of t-DNA and US-RNA in PBMCs, corresponding respectively to a small and transcriptionally silent viral reservoir, is not predictive of viral remission after TI in patients on ART., (© 2020 The Authors. Journal of the International AIDS Society published by John Wiley & Sons Ltd on behalf of the International AIDS Society.)

Background Since 1 June 2017, oral pre-exposure prophylaxis (PrEP) could be prescribed and reimbursed in Belgium as prophylactic medication for people who are at increased risk of HIV acquisition. The aim of this study was to determine the uptake of daily and event-driven PrEP in Belgium during the first 9 months of roll-out., Methods: Routine aggregated data on the number of reimbursement requests and the number of boxes of Truvada (Gilead Sciences, Cambridge, UK) delivered for PrEP through the Belgian pharmacies were obtained from the National Institute for Health and Disability Insurance. We also collected aggregated data from seven Aids Reference Centres (ARCs) currently providing most of the PrEP care in Belgium., Results: From 1 June 2017 to 28 February 2018, 1352 requests for reimbursement were approved by the National Institute for Health and Disability Insurance. Almost 98% of those who bought at least one box of 30 tablets of emtricitabine 200mg/tenofovir disoproxil fumarate 300mg (FTC/TDF) in a Belgian pharmacy were male, and most (67%) were between 30 and 50 years of age. According to data obtained from ARCs, the proportion of those choosing event-driven PrEP initially ranged between 29% and 73%., Conclusions: The uptake of PrEP in Belgium since the start of the roll-out in June 2017 has been high, and almost entirely limited to men who have sex with men, of whom 43% initially prefer a non-daily regimen. A better understanding is needed as to why other populations, such as sub-Saharan African migrants, are not accessing PrEP, as well as the development of a more sustainable PrEP delivery model.

In this review of the current challenges in the fight against HIV, we describe the state of the HIV epidemic and the framework put in place using the 90-90-90 objectives to try and curb the epidemic worldwide. There are numerous effective and evidence-based prevention measures against the spread of HIV, but the biggest challenges lie in the lack of political commitment, reluctance to address issues of sexuality and reproduction, and criminalization of key populations that are at the highest risk of HIV. Access to HIV treatment and continued care without stigmatization should be as easy and cheap as possible for those who are tested and diagnosed with HIV to achieve the best results worldwide. Regarding the treatment of HIV, the last decades have been very successful in dramatically improving the quality of life of people living with HIV, reducing the transmission rate and decreasing HIV-associated morbidity and mortality. It could even be argued that the next milestone will be a strategy that allows individuals to stop combination antiretroviral therapy safely before a cure is discovered. Despite great progress, people with HIV have shorter life expectancy than those without the virus, and the underlying causes are probably multifactorial, including premature aging, drug toxicities, and comorbidities. Even if challenges remain, hope should too, with the ultimate goal to end the HIV epidemic.

We present the updated Belgian guidelines for the use of non-occupational HIV post-exposure prophylaxis (NONOPEP). This document is inspired by UK guidelines 2015, adapted to the Belgian situation and approved by all AIDS reference centers in Belgium. When recommended, NONOPEP should be initiated as soon as possible, preferably within 24 h of exposure but can be offered up to 72 h. The duration of NONOPEP should be 28 days. These current guidelines include epidemiologic estimations, which can be used to calculate the risk of infection after a potential exposure and help to decide whether or not to start prophylaxis. We review which medications to use in the context of the last Belgian NONOPEP convention, provide a checklist for initial assessment, and make recommendations for monitoring individuals receiving NONOPEP.

Objectives: This study aimed to evaluate HIV sequence evolution in whole genes and in CD8 T-cell epitope regions following immunotherapy and subsequent analytical treatment interruption (ATI). A second objective of this study was to analyze associations between vaccine-specific immune responses and epitope mutation rates., Design: HIV-1-infected patients on combined antiretroviral therapy (cART) were subjected to immunotherapy by the administration of an autologous dendritic cell-based therapeutic vaccine expressing Tat, Rev, and Nef and subsequent ATI., Methods: HIV-1 genes were amplified and sequenced from plasma RNA obtained before initiation of cART as well as during ATI. Control sequences for virus evolution in untreated HIV-1-infected individuals were obtained from the HIV Sequence Database (Los Alamos). CD8 T-cell epitope regions were defined based on literature data and prediction models. HIV-1-specific immune responses were evaluated to analyze their impact on sequence evolution., Results: Viral sequence evolution in the tat, rev, and nef genes of vaccinated patients was similar to that of controls. The number of mutations observed inside and outside CD8 T-cell epitopes was comparable for vaccine-targeted and nontargeted proteins. We found no evidence for an impact of vaccine-induced or enhanced immune responses on the number of mutations inside or outside epitopes., Conclusion: Therapeutic vaccination of HIV-1-infected patients with a dendritic cell-based vaccine targeting Tat, Rev, and Nef did not affect virus evolution at the whole gene level nor at the CD8 T-cell epitope level.

Recently, it has been demonstrated that disease progression during HIV infection is not determined merely by the number of HIV-specific T cells but also by their quality (J. R. Almeida, et al., J. Exp. Med. 204:2473-2485, 2007; C. T. Berger, et al., J. Virol. 85:9334-9345, 2011; M. R. Betts, et al., Blood 107:4781-4789, 2006; V. V. Ganusov, et al., J. Virol. 85:10518-10528, 2011; P. Kiepiela, et al., Nat. Med. 13:46-53, 2007; and F. Pereyra, et al., J. Infect. Dis. 197:563-571, 2008). Therefore, strategies to specifically enhance or induce high-quality, HIV-specific T-cell responses are necessary to develop effective immune therapies. Thalidomide, lenalidomide, and pomalidomide have a strong capacity to boost immune responses and are therefore referred to as immunomodulatory drugs (IMiDs). We evaluated the effects of lenalidomide and pomalidomide on HIV-specific T cells. We found that the presence of IMiDs during in vitro T-cell stimulation with dendritic cells electroporated with Gag- or Nef-encoding mRNA resulted in higher numbers of cytokine-secreting HIV-specific CD8(+) T cells, particularly inducing polyfunctional HIV-specific CD8(+) T cells with an enhanced lytic capacity. Furthermore, CD8(+) T-cell responses were detected upon stimulation with lower antigenic peptide concentrations, and a higher number of Gag epitopes was recognized upon addition of IMiDs. Finally, IMiDs reduced the proliferation of the HIV-specific CD4(+) T cells while increasing the number of polyfunctional CD4(+) T cells. These results provide new information about the effects of IMiDs on antigen-specific T cells and suggest that these drugs increase the efficacy of immune therapies for infectious diseases and cancer.

Antigen-specific immunity is crucially important for containing viral replication in human immunodeficiency virus (HIV)-1-infected hosts. Several epitopes have been predicted for the early expressed HIV-1 proteins Tat and Rev, but few have been studied in detail. We characterized the human leukocyte antigen (HLA)-B44-restricted Rev epitope EELLKTVRL (EL9) in an HIV-1-infected subject treated with antiretroviral therapy. Interestingly, a high sequence similarity was found between the EL9 epitope and the human nucleolar protein 6 (NOL6). However, this similarity does not seem to impede immunogenicity as CD8(+) T-cells, previously stimulated with EL9-pulsed dendritic cells, were able to specifically recognize the HIV-1 Rev epitope without cross-recognizing the human self-antigen NOL6. After the subject interrupted antiretroviral therapy and virus rebounded, mutations within the EL9 epitope were identified. Although the emerging mutations resulted in decreased or abolished T-cell recognition, they did not impair Rev protein function. Mutations leading to escape from T-cell recognition persisted for up to 124 weeks following treatment interruption. This study shows that the HLA-B44-restricted Rev CD8(+) T-cell epitope EL9 is immunogenic notwithstanding its close resemblance to a human peptide. The epitope mutates as a consequence of dynamic interaction between T-cells and HIV-1. Clinical status, CD4(+) T-cell count and viral load remained stable despite escape from T-cell recognition., (© 2012 John Wiley & Sons A/S.)

In a phase I/IIa clinical trial, 17 HIV-1 infected patients, stable on cART, received 4 vaccinations with autologous dendritic cells electroporated with mRNA encoding Tat, Rev and Nef, after which cART was interrupted. Vaccination was safe and feasible. During the analytical treatment interruption (ATI), no serious adverse events were observed. Ninety-six weeks following ATI, 6/17 patients remained off therapy. Although induced and/or enhanced CD4(+) and CD8(+) T-cell responses specific for the immunogens were observed in most of the patients, we found no correlation with the number of weeks off cART. Moreover, CD4(+) T-cell counts, plasma viral load and the time remaining off cART following ATI did not differ from historical control data. To conclude, the vaccine was safe, well tolerated and resulted in vaccine-specific immune responses. Since no correlation with clinical parameters could be found, these results warrant further research in order to optimize the efficacy of vaccine-induced T-cell responses., (Copyright © 2011 Elsevier Inc. All rights reserved.)

One of the consequences of HIV infection is a progressive loss of T cell functions, resulting in decreased cytokine secretion and proliferation and an increased sensitivity to apoptosis. Therefore, successful therapeutic vaccination approaches should aim at restoring the functionality of existing HIV-specific T cells, as well as to efficiently induce potent, HIV-specific T cells from naïve T cells. In this study, we wanted to determine the stimulatory capacity of DCs coelectroporated with mRNA encoding for different costimulatory molecules of the TNFSF, together with HIV antigen-encoding mRNA. We show that DCs electroporated with 4-1BBL can enhance the proliferation, functionality, cytokine production, and survival of HIV-specific CD8(+) T cells. Furthermore, we are the first to show that a combination of 4-1BBL and CD40L overexpression on DCs dramatically enhances CD4(+) and CD8(+) T cell responses. Finally, we demonstrate that signaling through 4-1BB, but not through CD40, can alleviate the suppressive effect of Tregs on CD8(+) T cell proliferation. Thus, the combination of 4-1BBL and CD40L enhances HIV-specific CD8(+) T cell responses in a synergistic way, resulting in enhanced proliferation of CD4(+) and CD8(+) T cell subsets, an increased cytokine secretion, and a reduced sensitivity to Treg-mediated immune suppression.

The limitations of highly active anti-retroviral therapy (HAART) have necessitated the development of alternative therapeutic strategies. One of the approaches that has gained prominence in recent years is therapeutic vaccination. We decided to assess the capacity of mature dendritic cells, derived from blood monocytes of HIV-1 infected patients, to generate functional T-cell responses. For this purpose, we constructed a chimeric mRNA encoding the proteins Tat, Rev and Nef. The TaReNef encoding information was linked to the HLA class II-targeting sequence of DC-LAMP. Broadly directed HIV-specific CD4(+) and CD8(+) cytotoxic T cells exhibiting a poly-functional cytokine secretion pattern were generated by co-culturing with autologous chimeric mRNA electroporated dendritic cells. Thus, administration of ex vivo generated dendritic cells expressing the early proteins Tat, Rev and Nef might offer a promising approach for therapeutic vaccination in HIV-1 infection.