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BACKGROUNDMechanisms underlying persistent cardiopulmonary symptoms following SARS-CoV-2 infection (post-acute sequelae of COVID-19 “PASC” or “Long COVID”) remain unclear. This study sought to elucidate mechanisms of cardiopulmonary symptoms and reduced exercise capacity using advanced cardiac testing.METHODSWe performed cardiopulmonary exercise testing (CPET), cardiac magnetic resonance imaging (CMR) and ambulatory rhythm monitoring among adults > 1 year after confirmed SARS-CoV-2 infection in Long-Term Impact of Infection with Novel Coronavirus cohort (LIINC; substudy ofNCT04362150). Adults who completed a research echocardiogram (at a median 6 months after SARS-CoV-2 infection) without evidence of heart failure or pulmonary hypertension were asked to complete additional cardiopulmonary testing approximately 1 year later. Although participants were recruited as a prospective cohort, to account for selection bias, the primary analyses were as a case-control study comparing those with and without persistent cardiopulmonary symptoms. We also correlated findings with previously measured biomarkers. We used logistic regression and linear regression models to adjust for potential confounders including age, sex, body mass index, time since SARS-CoV-2 infection, and hospitalization for acute SARS-CoV-2 infection, with sensitivity analyses adjusting for medical history.RESULTSSixty participants (unselected for symptoms, median age 53, 42% female, 87% non- hospitalized) were studied at median 17.6 months following SARS-CoV-2 infection. On maximal CPET, 18/37 (49%) with symptoms had reduced exercise capacity (peak VO22was 5.2 ml/kg/min (95%CI 2.1-8.3; p=0.001) or 16.9% lower actual compared to predicted (95%CI 4.3- 29.6; p=0.02) among those with symptoms compared to those without symptoms. Chronotropic incompetence was present among 12/21 (57%) with reduced VO2including 11/37 (30%) with symptoms and 1/19 (5%) without (p=0.04). Inflammatory markers (hsCRP, IL-6, TNF-α) and SARS-CoV-2 antibody levels measured early in PASC were negatively correlated with peak VO2more than 1 year later. Late-gadolinium enhancement on CMR and arrhythmias on ambulatory monitoring were not present.CONCLUSIONSWe found evidence of objectively reduced exercise capacity among those with cardiopulmonary symptoms more than 1 year following COVID-19, which was associated with elevated inflammatory markers early in PASC. Chronotropic incompetence may explain exercise intolerance among some with cardiopulmonary phenotype Long COVID.Graphical AbstractKey PointsLong COVID symptoms were associated with reduced exercise capacity on cardiopulmonary exercise testing more than 1 year after SARS-CoV-2 infection. The most common abnormal finding was chronotropic incompetence. Reduced exercise capacity was associated with early elevations in inflammatory markers.

Limited data are available on the long-term clinical and immunologic consequences of SARS-CoV-2 infection in people with HIV (PWH).We measured SARS-CoV-2-specific humoral and cellular responses in people with and without HIV recovering from COVID-19 ( n = 39 and n = 43, respectively) using binding antibody, surrogate virus neutralization, intracellular cytokine staining, and inflammatory marker assays. We identified individuals experiencing postacute sequelae of SARS-CoV-2 infection (PASC) and evaluated immunologic parameters. We used linear regression and generalized linear models to examine differences by HIV status in the magnitude of inflammatory and virus-specific antibody and T-cell responses, as well as differences in the prevalence of PASC.Among PWH, we found broadly similar SARS-CoV-2-specific antibody and T-cell responses as compared with a well matched group of HIV-negative individuals. PWH had 70% lower relative levels of SARS-CoV-2-specific memory CD8 + T cells ( P = 0.007) and 53% higher relative levels of PD-1+ SARS-CoV-2-specific CD4 + T cells ( P = 0.007). Higher CD4 + /CD8 + ratio was associated with lower PD-1 expression on SARS-CoV-2-specific CD8 + T cells (0.34-fold effect, P = 0.02). HIV status was strongly associated with PASC (odds ratio 4.01, P = 0.008), and levels of certain inflammatory markers (IL-6, TNF-alpha, and IP-10) were associated with persistent symptoms.We identified potentially important differences in SARS-CoV-2-specific CD4 + and CD8 + T cells in PWH and HIV-negative participants that might have implications for long-term immunity conferred by natural infection. HIV status strongly predicted the presence of PASC. Larger and more detailed studies of PASC in PWH are urgently needed.

BACKGROUNDThe presence and reactivation of chronic viral infections, such as EBV, CMV, and HIV, have been proposed as potential contributors to long COVID (LC), but studies in well-characterized postacute cohorts of individuals with COVID-19 over a longer time course consistent with current case definitions of LC are limited.METHODSIn a cohort of 280 adults with prior SARS-CoV-2 infection, we assessed the presence and types of LC symptoms and prior medical history (including COVID-19 history and HIV status) and performed serological testing for EBV and CMV using a commercial laboratory. We used covariate-adjusted binary logistic regression models to identify independent associations between variables and LC symptoms.RESULTSWe observed that LC symptoms, such as fatigue and neurocognitive dysfunction, at a median of 4 months following initial diagnosis were independently associated with serological evidence suggesting recent EBV reactivation (early antigen-diffuse IgG positivity) or high nuclear antigen (EBNA) IgG levels but not with ongoing EBV viremia. Serological evidence suggesting recent EBV reactivation (early antigen-diffuse IgG positivity) was most strongly associated with fatigue (OR = 2.12). Underlying HIV infection was also independently associated with neurocognitive LC (OR = 2.5). Interestingly, participants who had serologic evidence of prior CMV infection were less likely to develop neurocognitive LC (OR = 0.52).CONCLUSIONOverall, these findings suggest differential effects of chronic viral coinfections on the likelihood of developing LC and association with distinct syndromic patterns. Further assessment during the acute phase of COVID-19 is warranted.TRIAL REGISTRATIONLong-term Impact of Infection with Novel Coronavirus; ClinicalTrials.gov NCT04362150.FUNDINGThis work was supported by NIH/National Institute of Allergy and Infectious Diseases grants (3R01AI141003-03S1, R01AI158013, and K24AI145806); the Zuckerberg San Francisco General Hospital Department of Medicine and Division of HIV, Infectious Diseases, and Global Medicine; and the UCSF-Bay Area Center for AIDS Research (P30-AI027763).

From 2 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) household transmission studies (enrolling April 2020 to January 2022) with rapid enrollment and specimen collection for 14 days, 61% (43/70) of primary cases had culturable virus detected ≥6 days post-onset. Risk of secondary infection among household contacts tended to be greater when primary cases had culturable virus detected after onset. Regardless of duration of culturable virus, most secondary infections (70%, 28/40) had serial intervals

Interferon (IFN)–specific autoantibodies have been implicated in severe coronavirus disease 2019 (COVID-19) and have been proposed as a potential driver of the persistent symptoms characterizing “long COVID,” a type of postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We report that only 2 of 215 participants with convalescent SARS-CoV-2 infection tested over 394 time points, including 121 people experiencing long COVID symptoms, had detectable IFN-α2 antibodies. Both had been hospitalized during the acute phase of the infection. These data suggest that persistent anti-IFN antibodies, although a potential driver of severe COVID-19, are unlikely to contribute to long COVID symptoms in the postacute phase of the infection.

Background: Antiretroviral therapy (ART) non-adherence and methamphetamine use are associated with higher HIV drug resistance prevalence. How they affect drug resistance mutations accumulation is less studied. Objective: We assessed factors associated with drug resistance mutations accumulation. Methods: We evaluated HIV chronically-infected patients from a clinic-based research cohort on first-line ART regimens with genotype results within 30 days of baseline. Methamphetamine use and ART adherence were self-reported at each study visit. High ART adherence was defined as 0-5% missed doses in the last 30 days. Results: One-hundred twenty-five patients contributed 496 study visits. At baseline, 81% of patients reported high ART adherence; 90% reported no methamphetamine use in the prior 4 months, 8% used monthly or less and 2% used daily or weekly. Methamphetamine users and non-users had similarly high ART adherence (p=0.93). Adjusted incidence rate ratio (aIRR) of drug resistance mutations accumulation was 2.04 (95% CI 0.64, 6.46) for daily/weekly users and 1.71 (95% CI 0.66, 4.42) for patients with monthly or less users, compared to non-users. aIRR was 0.71 (95% CI 0.44, 1.15) with >5-10% missed ART doses and 1.21 (95% CI 0.80, 1.83) with >10% missed doses compared to 0-5% missed doses. Conclusions: We found no strong evidence for the effect of methamphetamine use and ART adherence on drug resistance mutations accumulation. Research cohort patients may have been more engaged in care and treatment adherent than non-cohort patients. Our findings suggest methamphetamine use might not lead to treatment failure among HIV patients who are otherwise engaged in care.

Background SARS-CoV-2 nucleocapsid antigen can be detected in plasma, but little is known about its performance as a diagnostic test for acute SARS-CoV-2 infection or infectious viral shedding among nonhospitalized individuals. Methods We used data generated from anterior nasal and blood samples collected in a longitudinal household cohort of SARS-CoV-2 cases and contacts. Participants were classified as true positives if polymerase chain reaction (PCR) positive for SARS-CoV-2 and as true negatives if PCR negative and seronegative. Infectious viral shedding was determined by the cytopathic effect from viral culture. Stratified by 7 days after symptom onset, we constructed receiver operating characteristic (ROC) curves to describe optimized accuracy (Youden index), optimized sensitivity, and specificity. Results Of 80 participants, 58 (73%) were true positives while 22 (27%) were true negatives. Using the manufacturer's cutoff of 1.25 pg/mL for evaluating infection, sensitivity was higher from 0 to 7 days (77.6% [95% confidence interval {CI}, 64%–88.2%]) than from 8 to 14 days (43.2% [95% CI, 31.1%–54.5%]) after symptom onset; specificity was unchanged at 100% (95% CI, 88.1%–100%). This test had higher sensitivity (100% [95% CI, 88.4%–100%]) and lower specificity (65% [95% CI, 40.8%–84.6%]) for infectious viral shedding as compared with infection, particularly within the first week of symptom onset. Although the presence of N-antigen correlated with infectious viral shedding (r = 0.63; P < .01), sensitivity still declined over time. Additional cutoffs from ROC curves were identified to optimize sensitivity and specificity. Conclusions We found that this SARS-CoV-2 N-antigen test was highly sensitive for detecting early but not late infectious viral shedding, making it a viable screening test for community-dwelling individuals to inform isolation practices.

BackgroundAlthough many patients with Kaposi sarcoma (KS) in sub-Saharan Africa are diagnosed with AIDS Clinical Trials Group (ACTG) T1 disease, T1 staging insufficiently captures clinical heterogeneity of advanced KS. Using a representative community-based sample, we detailed disease severity at diagnosis to inform KS staging and treatment in sub-Saharan Africa.MethodsWe performed rapid case ascertainment on people living with HIV, aged 18 years or older, newly diagnosed with KS from 2016 to 2019 at 3 clinic sites in Kenya and Uganda to ascertain disease stage as close as possible to diagnosis. We reported KS severity using ACTG and WHO staging criteria and detailed measurements that are not captured in the current staging systems.ResultsWe performed rapid case ascertainment within 1 month for 241 adults newly diagnosed with KS out of 389 adult patients with suspected KS. The study was 68% men with median age 35 years and median CD4 count 239. Most of the patients had advanced disease, with 82% qualifying as ACTG T1 and 64% as WHO severe/symptomatic KS. The most common ACTG T1 qualifiers were edema (79%), tumor-associated ulceration (24%), extensive oral KS (9%), pulmonary KS (7%), and gastrointestinal KS (4%). There was marked heterogeneity within T1 KS, with 25% of patients having 2 T1 qualifying symptoms and 3% having 3 or more.ConclusionMost of the patients newly diagnosed with KS had advanced stage disease, even in the current antiretroviral therapy "treat-all" era. We observed great clinical heterogeneity among advanced stage patients, leading to questions about whether all patients with advanced KS require the same treatment strategy.

We aimed to characterize the variability in the illness experience and recovery process from COVID-19. We conducted in-depth individual interviews with participants enrolled in the Long-term Immunological Impact of Novel Coronavirus (LIINC) cohort study in San Francisco, California from June through October of 2020. Participants were adults who had a previously confirmed positive SARV-CoV-2 nucleic acid amplification test result, had recovered or were recovering from acute infection, and underwent serial evaluations at our clinical research center. We purposefully sampled 24 English- and Spanish-speaking adults with asymptomatic, mild and severe symptomatic infection, including those who were hospitalized, and those with HIV co-infection. Half of our sample (50.0%) identified as Latinx/Hispanic and most of the participants were men (62.5%). We used thematic analysis to characterize the illness experience, recovery process, and mental health impact of experiencing COVID-19 and present clinical data for each participant. Emergent themes were: (1) across symptom profiles and severity, experiencing COVID-19 was associated with psychological distress, (2) among participants with symptomatic infection, the illness experience was characterized by uncertainty in terms of managing symptoms and recovery, and (3) despite wide-ranging illness experiences, participants shared many common characteristics, including health information-seeking behavior facilitated by access to medical care, and uncertainty regarding the course of their illness and recovery. COVID-19 was associated with elevated levels of psychological distress, regardless of symptoms.

Background There is limited evidence to evaluate screening algorithms with rapid antigen testing and exposure assessments as identification strategies for paucisymptomatic or asymptomatic Ebola virus (EBOV) infection and unrecognized EBOV disease (EVD). Methods We used serostatus and self-reported postexposure symptoms from a cohort study to classify contact-participants as having no infection, paucisymptomatic or asymptomatic infection, or unrecognized EVD. Exposure risk was categorized as low, intermediate, or high. We created hypothetical scenarios to evaluate the World Health Organization (WHO) case definition with or without rapid diagnostic testing (RDT) or exposure assessments. Results This analysis included 990 EVD survivors and 1909 contacts, of whom 115 (6%) had paucisymptomatic or asymptomatic EBOV infection, 107 (6%) had unrecognized EVD, and 1687 (88%) were uninfected. High-risk exposures were drivers of unrecognized EVD (adjusted odds ratio, 3.5 [95% confidence interval, 2.4–4.9]). To identify contacts with unrecognized EVD who test negative by the WHO case definition, the sensitivity was 96% with RDT (95% confidence interval, 91%–99%), 87% with high-risk exposure (82%–92%), and 97% with intermediate- to high-risk exposures (93%–99%). The proportion of false-positives was 2% with RDT and 53%–93% with intermediate- and/or high-risk exposures. Conclusion We demonstrated the utility and trade-offs of sequential screening algorithms with RDT or exposure risk assessments as identification strategies for contacts with unrecognized EVD.

Long COVID, a type of post-acute sequelae of SARS-CoV-2 (PASC), has been associated with sustained elevated levels of immune activation and inflammation. However, the mechanisms that drive this inflammation remain unknown. Inflammation during acute coronavirus disease 2019 could be exacerbated by microbial translocation (from the gut and/or lung) to blood. Whether microbial translocation contributes to inflammation during PASC is unknown. We did not observe a significant elevation in plasma markers of bacterial translocation during PASC. However, we observed higher levels of fungal translocation - measured as β-glucan, a fungal cell wall polysaccharide - in the plasma of individuals experiencing PASC compared with those without PASC or SARS-CoV-2-negative controls. The higher β-glucan correlated with higher inflammation and elevated levels of host metabolites involved in activating N-methyl-d-aspartate receptors (such as metabolites within the tryptophan catabolism pathway) with established neurotoxic properties. Mechanistically, β-glucan can directly induce inflammation by binding to myeloid cells (via Dectin-1) and activating Syk/NF-κB signaling. Using a Dectin-1/NF-κB reporter model, we found that plasma from individuals experiencing PASC induced higher NF-κB signaling compared with plasma from negative controls. This higher NF-κB signaling was abrogated by piceatannol (Syk inhibitor). These data suggest a potential targetable mechanism linking fungal translocation and inflammation during PASC.

A multiplexed enzyme-linked immunosorbent assay (ELISA) that simultaneously measures antibody binding to multiple antigens can extend the impact of serosurveillance studies, particularly if the assay approaches the simplicity, robustness, and accuracy of a conventional single-antigen ELISA. Here, we report on the development of multiSero, an open-source multiplex ELISA platform for measuring antibody responses to viral infection. Our assay consists of three parts: (1) an ELISA against an array of proteins in a 96-well format; (2) automated imaging of each well of the ELISA array using an open-source plate reader; and (3) automated measurement of optical densities for each protein within the array using an open-source analysis pipeline. We validated the platform by comparing antibody binding to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) antigens in 217 human sera samples, showing high sensitivity (0.978), specificity (0.977), positive predictive value (0.978), and negative predictive value (0.977) for classifying seropositivity, a high correlation of multiSero determined antibody titers with commercially available SARS-CoV-2 antibody tests, and antigen-specific changes in antibody titer dynamics upon vaccination. The open-source format and accessibility of our multiSero platform can contribute to the adoption of multiplexed ELISA arrays for serosurveillance studies, for SARS-CoV-2 and other pathogens of significance.

OBJECTIVE Dolutegravir (DTG) is now a preferred component of first-line antiretroviral therapy (ART). However, prevalence data on natural resistance to integrase inhibitors [integrase strand transfer inhibitors (INSTIs)] in circulating non-subtype B HIV-1 in sub-Saharan Africa is scarce. Our objective is to report prevalence of pre-treatment integrase polymorphisms associated with resistance to INSTIs in an ART-naive cohort with diverse HIV-1 subtypes. DESIGN We retrospectively examined HIV-1 integrase sequences from Uganda. METHODS Plasma samples were derived from the Uganda AIDS Rural Treatment Outcomes (UARTO) cohort, reflecting enrollment from 2002 to 2010, prior to initiation of ART. HIV-1 integrase was amplified using nested-PCR and Sanger-sequenced (HXB2 4230-5093). Stanford HIVdb v8.8 was used to infer clinically significant INSTI-associated mutations. Human leukocyte antigen (HLA) typing was performed for all study participants. RESULTS Plasma samples from 511 ART-naive individuals (subtype: 48% A1, 39% D) yielded HIV-1 integrase genotyping results. Six out of 511 participants (1.2%) had any major INSTI-associated mutations. Of these, two had E138T (subtype A1), three had E138E/K (subtype D), and one had T66T/I (subtype D). No participants had mutations traditionally associated with high levels of INSTI resistance. HLA genotypes A∗02:01/05/14, B∗44:15, and C∗04:07 predicted the presence of L74I, a mutation recently observed in association with long-acting INSTI cabotegravir virologic failure. CONCLUSION We detected no HIV-1 polymorphisms associated with high levels of DTG resistance in Uganda in the pre-DTG era. Our results support widespread implementation of DTG but careful monitoring of patients on INSTI with virologic failure is warranted to determine if unique mutations predict failure for non-B subtypes of HIV-1.

Cognitive post-acute sequelae of SARS-CoV-2 (PASC) can occur after mild COVID-19. Detailed clinical characterizations may inform pathogenesis. We evaluated 22 adults reporting cognitive PASC and 10 not reporting cognitive symptoms after mild SARS-CoV-2 infection through structured interviews, neuropsychological testing, and optional cerebrospinal fluid (CSF) evaluations (53%). Delayed onset of cognitive PASC occurred in 43% and associated with younger age. Cognitive PASC participants had a higher number of pre-existing cognitive risk factors (2.5 vs. 0; p = 0.03) and higher proportion with abnormal CSF findings (77% vs. 0%; p = 0.01) versus controls. Cognitive risk factors and immunologic mechanisms may contribute to cognitive PASC pathogenesis.

It has been demonstrated that activated mast cells (MCs) are enriched in Kaposi sarcoma (KS) tumors and contribute to the inflammatory microenvironment. Mechanisms driving MC activation, however, are incompletely understood. We sought to understand whether immunoglobulin E (IgE), a potent activator of MCs, was associated with KS incidence and severity. In a cross-sectional study of untreated human immunodeficiency virus (HIV)–infected adults with or without KS in Uganda, we found that patients with KS had higher plasma IgE levels than those without KS. After adjustment for age, sex, CD4+ T-cell count, and HIV RNA levels, there was a dose-response relationship between plasma IgE levels and the presence and severity of KS. Higher eosinophil counts were also associated with IgE levels, and plasma interleukin 33 concentrations were higher in individuals with KS. These findings suggest that IgE-driven atopic inflammation may contribute the pathogenesis of KS. Therapies targeting IgE-mediated MC activation thus might represent a novel approach for treatment or prevention of KS.

Background: Efforts to understand the impact of SARS-CoV-2 variants, vaccine status, and treatment on the development and persistence of Long COVID have intensified. Methods: We report 4 sequential cases from a post-COVID cohort study demonstrating variability in outcomes following differentially timed nirmatrelvir therapy, received as part of clinical care. Results: In the first case, the participant experienced symptomatic rebound and developed Long COVID despite early initiation of antiviral therapy. In the next 2 cases, participants reported improvement in persistent COVID symptoms when nirmatrelvir was taken 25 and 60 days following initial symptom onset. In the final case, an individual with presumed Long COVID for 2 years reported substantial improvement in chronic symptoms when taking nirmatrelvir following SARS-CoV-2 re-infection. Conclusions: These anecdotes suggest that systematic study of antiviral therapy for Long COVID is warranted.

BACKGROUND: Residual systemic inflammation, which is associated with non-AIDS clinical outcomes, may persist despite viral suppression. We assessed the effect of antiretroviral (ART) adherence interruptions on systemic inflammation among Ugandans living with HIV who were virally suppressed. SETTING: We evaluated adults initiating first-line ART at a regional referral hospital clinic in Mbarara, Uganda. METHODS: Plasma concentrations of interleukin-6 (IL-6), D-dimer, soluble sCD14, sCD163, the kynurenine/tryptophan (K/T) ratio, and CD8(+) T-cell activation (HLA-DR(+)/CD38(+) co-expression) were measured at baseline and 6 months following ART initiation among participants who achieved viral suppression (VL

Objective Individuals with HIV suffer a higher burden of cardiovascular diseases. Traditional cardiovascular risk scores consistently underestimate cardiovascular risk in this population. Subsets of microRNAs (miRNAs) are differentially expressed among individuals with cardiovascular disease and individuals infected with HIV. However, no study has clarified whether specific miRNAs may be biomarkers for cardiovascular disease in individuals with HIV. Design/methods We compared the miRNA expression profiles of 34 HIV-positive individuals who had experienced clinically adjudicated type I myocardial infarctions (MI) with the profiles of 76 HIV-positive controls matched by traditional cardiovascular risk factors and HIV-specific measures. Using the elastic net algorithm, we selected miRNAs most strongly associated with incident MI and then used conditional Cox proportional hazards regression and cross-validation to evaluate miRNAs and their association with incident MI. We evaluated whether miRNA markers would improve risk classification relative to the Framingham Risk Score. Results Higher miR-125a-5p and miR-139-5p expression levels were each associated with increased risk of developing MI after adjustment for Framingham Risk Score and HIV-related factors (hazard ratio 2.43, P = 0.018; hazard ratio 2.13, P = 0.048, respectively). Compared with the Framingham Risk Score alone, adding expression levels of miR-125a-5p or miR-139-5p resulted in an integrated discrimination improvement of 10.1 or 5.8%, respectively. Conclusion MiR-125a-5p and miR-139-5p, transcripts known to be differentially expressed in HIV-positive individuals, may serve as unique biomarkers predictive of cardiovascular disease in these patients and may help clarify processes because of HIV infection that contribute to cardiovascular disorders in this population.

Background There is mounting evidence for the presence of postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (PASC), but there is limited information on the spectrum, magnitude, duration, and patterns of these sequelae as well as their influence on quality of life. Methods We assembled a cohort of adults with a documented history of SARS-CoV-2 RNA positivity at ≥2 weeks past onset of coronavirus disease 2019 (COVID-19) symptoms or, if asymptomatic, first positive test. At 4-month intervals, we queried physical and mental health symptoms and quality of life. Results Of the first 179 participants enrolled, 10 were asymptomatic during the acute phase of SARS-CoV-2 infection, 125 were symptomatic but not hospitalized, and 44 were symptomatic and hospitalized. During the postacute phase, fatigue, shortness of breath, concentration problems, headaches, trouble sleeping, and anosmia/dysgeusia were most common through 8 months of observation. Symptoms were typically at least somewhat bothersome and sometimes exhibited a waxing-and-waning course. Some participants experienced symptoms of depression, anxiety, and post-traumatic stress, as well as difficulties with performance of usual activities. The median visual analogue scale rating of general health was lower at 4 and 8 months compared with pre-COVID-19. Two clusters of symptom domains were identified. Conclusions Many participants report bothersome symptoms following onset of COVID-19 with variable patterns of persistence and impact on quality of life. The substantial variability suggests the existence of multiple subphenotypes of PASC. A rigorous approach to the prospective measurement of symptoms and functional manifestations sets the stage for the next phase of research focusing on the pathophysiologic causes of the various subgroups of PASC.

BackgroundIn HIV infection, even under long-term antiretroviral therapy (ART), up to 20% of HIV-infected individuals fail to restore CD4+ T cell counts to the levels similar to those of healthy controls. The mechanisms of poor CD4+ T cell reconstitution on suppressive ART are not fully understood.MethodsHere, we tested the hypothesis that lipopolysaccharide (LPS) from bacteria enriched in the plasma from immune non-responders (INRs) contributes to blunted CD4+ T cell recovery on suppressive ART in HIV. We characterized plasma microbiome in HIV INRs (aviremic, CD4+ T cell counts < 350 cells/μl), immune responders (IRs, CD4+ T cell counts > 500 cells/μl), and healthy controls. Next, we analyzed the structure of the lipid A domain of three bacterial species identified by mass spectrometry (MS) and evaluated the LPS function through LPS induced proinflammatory responses and CD4+ T cell apoptosis in PBMCs. In comparison, we also evaluated plasma levels of proinflammatory cytokine and chemokine patterns in these three groups. At last, to study the causality of microbiome-blunted CD4+ T cell recovery in HIV, B6 mice were intraperitoneally (i.p.) injected with heat-killed Burkholderia fungorum, Serratia marcescens, or Phyllobacterium myrsinacearum, twice per week for total of eight weeks.FindingsINRs exhibited elevated plasma levels of total microbial translocation compared to the IRs and healthy controls. The most enriched bacteria were Burkholderia and Serratia in INRs and were Phyllobacterium in IRs. Further, unlike P. myrsinacearum LPS, B. fungorum and S. marcescens LPS induced proinflammatory responses and CD4+ T cell apoptosis in PBMCs, and gene profiles of bacteria-mediated cell activation pathways in THP-1 cells in vitro. Notably, LPS structural analysis by mass spectrometry revealed that lipid A from P. myrsinacearum exhibited a divergent structure consistent with weak toll-like receptor (TLR) 4 agonism, similar to the biological profile of probiotic bacteria. In contrast, lipid A from B. fungorum and S. marcescens showed structures more consistent with canonical TLR4 agonists stemming from proinflammatory bacterial strains. Finally, intraperitoneal (i.p.) injection of inactivated B. fungorum and S. marcescens but not P. myrsinacearum resulted in cell apoptosis in mesenteric lymph nodes of C57BL/6 mice in vivo.InterpretationThese results suggest that the microbial products are causally associated with INR phenotype. In summary, variation in blood microbial LPS immunogenicity may contribute to immune reconstitution in response to suppressive ART. Collectively, this work is consistent with immunologically silencing microbiome being causal and targetable with therapy in HIV.FundingThis work was supported by the National Institute of Allergy and Infectious Diseases (NIAID; R01 AI128864, Jiang) (NIAID; P30 AI027767, Saag/Health), the Medical Research Service at the Ralph H. Johnson VA Medical Center (merit grant VA CSRD MERIT I01 CX-002422, Jiang), and the National Institute of Aging (R21 AG074331, Scott). The SCOPE cohort was supported by the UCSF/Gladstone Institute of Virology & Immunology CFAR (P30 AI027763, Gandhi) and the CFAR Network of Integrated Clinical Systems (R24 AI067039, Saag). The National Center for Advancing Translational Sciences of the National Institutes of Health under Award Number UL1TR001450 (the pilot grant, Jiang). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Background The biological processes associated with postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (PASC) are unknown. Methods We measured soluble markers of inflammation in a SARS-CoV-2 recovery cohort at early (90 days) timepoints. We defined PASC as the presence of 1 or more coronavirus disease 2019 (COVID-19)–attributed symptoms beyond 90 days. We compared fold-changes in marker values between those with and without PASC using mixed-effects models with terms for PASC and early and late recovery time periods. Results During early recovery, those who went on to develop PASC generally had higher levels of cytokine biomarkers including tumor necrosis factor–α (1.14-fold higher mean ratio [95% confidence interval {CI}, 1.01–1.28]; P = .028) and interferon-γ–induced protein 10 (1.28-fold higher mean ratio [95% CI, 1.01–1.62]; P = .038). Among those with PASC, there was a trend toward higher interleukin 6 levels during early recovery (1.29-fold higher mean ratio [95% CI, .98–1.70]; P = .07), which became more pronounced in late recovery (1.44-fold higher mean ratio [95% CI, 1.11–1.86]; P Conclusions Persistent immune activation may be associated with ongoing symptoms following COVID-19. Further characterization of these processes might identify therapeutic targets for those experiencing PASC.

Shortness of breath, chest pain, and palpitations occur as postacute sequelae of COVID-19, but whether symptoms are associated with echocardiographic abnormalities, cardiac biomarkers, or markers of systemic inflammation remains unknown. In a cross-sectional analysis, we assessed symptoms, performed echocardiograms, and measured biomarkers among adults more than 8 weeks after confirmed SARS-CoV-2 infection. We modeled associations between symptoms and baseline characteristics, echocardiographic findings, and biomarkers using logistic regression. We enrolled 102 participants at a median of 7.2 months following COVID-19 onset; 47 individuals reported dyspnea, chest pain, or palpitations. Median age was 52 years, and 41% of participants were women. Female sex, hospitalization, IgG antibody against SARS-CoV-2 receptor binding domain, and C-reactive protein were associated with symptoms. Regarding echocardiographic findings, 4 of 47 participants (9%) with symptoms had pericardial effusions compared with 0 of 55 participants without symptoms; those with effusions had a median of 4 symptoms compared with a median of 1 symptom in those without effusions. There was no strong evidence for a relationship between symptoms and echocardiographic functional parameters or other biomarkers. Among adults more than 8 weeks after SARS-CoV-2 infection, SARS-CoV-2 RBD antibodies, markers of inflammation, and, possibly, pericardial effusions are associated with cardiopulmonary symptoms. Investigation into inflammation as a mechanism underlying postacute sequelae of COVID-19 is warranted.

Background We assessed whether key biomarkers of endothelial activation and hemostasis/thrombosis were elevated in individuals receiving effective antiretroviral therapy (ART) in the year before ischemic stroke. Methods We conducted a case-control study nested in the CNICS cohort, comparing 42 adjudicated cases with ischemic stroke to 83 controls matched for ART regimen. Angiopoietin-1, angiopoietin-2, C-reactive protein, interleukin-6, plasminogen activation inhibitor-1, P-selectin, serum amyloid-A, soluble CD14, ICAM-1, VCAM-1, apolipoprotein A1, ADAMTS13, and von Willebrand factor (VWF) were measured in stored plasma collected before the stroke event. We used conditional logistic regression to identify associations with ischemic stroke, with and without adjustment for Atherosclerotic Cardiovascular Disease (ASCVD) and Veterans Aging Cohort Study (VACS) scores. Results After adjustment for age and sex, higher plasma viral load and higher angiopoeitin-2, soluble CD14, and VWF were associated with increased odds of ischemic stroke; higher nadir CD4 count was associated with decreased odds of ischemic stroke. VWF remained associated with subsequent ischemic stroke after adjustment for ASCVD score (adjusted odds 1.74, 95%CI 1.01–2.98 per log2 increment). In a separate model adjusting for VACS score, only VWF (adjusted odds 1.80, 95% CI 1.04–3.12 per log2 increment) was associated with subsequent ischemic stroke. In a sensitivity analysis excluding participants with viral load ≥400 copies/mL, associations between VWF and ischemic stroke were attenuated, with risk estimates ranging from 1.59–1.64 per log2 increment. Conclusions Endothelial activation and related release and attachment of VWF may play an important role in ischemic stroke among persons living with treated HIV infection.

Future HIV-1 curative therapies require a thorough understanding of the distribution of genetically-intact HIV-1 within T-cell subsets during antiretroviral therapy (ART) and the cellular mechanisms that maintain this reservoir. Therefore, we sequenced near-full-length HIV-1 genomes and identified genetically-intact and genetically-defective genomes from resting naive, stem-cell memory, central memory, transitional memory, effector memory, and terminally-differentiated CD4+ T-cells with known cellular half-lives from 11 participants on ART. We find that a higher infection frequency with any HIV-1 genome was significantly associated with a shorter cellular half-life, such as transitional and effector memory cells. A similar enrichment of genetically-intact provirus was observed in these cells with relatively shorter half-lives. We found that effector memory and terminally-differentiated cells also had significantly higher levels of expansions of genetically-identical sequences, while only transitional and effector memory cells contained genetically-intact proviruses that were part of a cluster of identical sequences. Expansions of identical sequences were used to infer cellular proliferation from clonal expansion. Altogether, this indicates that specific cellular mechanisms such as short half-life and proliferative potential contribute to the persistence of genetically-intact HIV-1. IMPORTANCE The design of future HIV-1 curative therapies requires a more thorough understanding of the distribution of genetically-intact HIV-1 within T-cell subsets as well as the cellular mechanisms that maintain this reservoir. These genetically-intact and presumably replication-competent proviruses make up the latent HIV-1 reservoir. Our investigations into the possible cellular mechanisms maintaining the HIV-1 reservoir in different T-cell subsets have revealed a link between the half-lives of T-cells and the level of proviruses they contain. Taken together, we believe our study shows that more differentiated and proliferative cells, such as transitional and effector memory T-cells, contain the highest levels of genetically-intact proviruses, and the rapid turnover rate of these cells contributes to the expansion of genetically-intact proviruses within them. Therefore, our study delivers an in-depth assessment of the cellular mechanisms, such as cellular proliferation and half-life, that contribute to and maintain the latent HIV-1 reservoir.

Background Although ≈70% of the world's population of people living with HIV reside in sub‐Saharan Africa, there are minimal prospective data on the contributions of HIV infection to atherosclerosis in the region. Methods and Results We conducted a prospective observational cohort study of people living with HIV on antiretroviral therapy >40 years of age in rural Uganda, along with population‐based comparators not infected with HIV. We collected data on cardiovascular disease risk factors and carotid ultrasound measurements annually. We fitted linear mixed effects models, adjusted for cardiovascular disease risk factors, to estimate the association between HIV serostatus and progression of carotid intima media thickness (cIMT). We enrolled 155 people living with HIV and 154 individuals not infected with HIV and collected cIMT images at 1045 visits during a median of 4 annual visits per participant (interquartile range 3–4, range 1–5). Age (median 50.9 years) and sex (49% female) were similar by HIV serostatus. At enrollment, there was no difference in mean cIMT by HIV serostatus (0.665 versus 0.680 mm, P =0.15). In multivariable models, increasing age, blood pressure, and non–high‐density lipoprotein cholesterol were associated with greater cIMT ( P P =0.25). Conclusions In rural Uganda, treated HIV infection was not associated with faster cIMT progression. These results do not support classification of treated HIV infection as a risk factor for subclinical atherosclerosis progression in rural sub‐Saharan Africa. Registration URL: https://www.ClinicalTrials.gov ; Unique identifier: NCT02445079.

Background The updated Veterans Aging Cohort Study (VACS) Index 2.0 combines general and human immunodeficiency virus (HIV)–specific biomarkers to generate a continuous score that accurately discriminates risk of mortality in diverse cohorts of persons with HIV (PWH), but a score alone is difficult to interpret. Using data from the North American AIDS Cohort Collaboration (NA-ACCORD), we translate VACS Index 2.0 scores into validated probability estimates of mortality. Methods Because complete mortality ascertainment is essential for accurate calibration, we restricted analyses to cohorts with mortality from the National Death Index or equivalent sources. VACS Index 2.0 components were ascertained from October 1999 to April 2018. Mortality was observed up to March 2019. Calibration curves compared predicted (estimated by fitting a gamma model to the score) to observed mortality overall and within subgroups: cohort (VACS/NA-ACCORD subset), sex, age 500 copies/mL, CD4 count Results Among 37230 PWH in VACS and 8061 PWH in the NA-ACCORD subset, median age was 53 and 44 years; 3% and 19% were women; and 48% and 39% were black. Discrimination in NA-ACCORD (C-statistic = 0.842 [95% confidence interval {CI}, .830–.854]) was better than in VACS (C-statistic = 0.813 [95% CI, .809–.817]). Predicted and observed mortality largely overlapped in VACS and the NA-ACCORD subset, overall and within subgroups. Conclusions Based on this validation, VACS Index 2.0 can reliably estimate probability of all-cause mortality, at various follow-up times, among PWH in North America.

Background Pregnant women with HIV (PWWH) have high postpartum morbidity and mortality from infections like tuberculosis. Immunologic changes during pregnancy and postpartum periods may contribute to these risks, particularly the immunoregulatory kynurenine pathway of tryptophan catabolism, which contributes to both HIV and tuberculosis pathogenesis and increases in the early postpartum period. Methods Women with HIV initiating antiretroviral therapy (ART) in the Uganda AIDS Rural Treatment Outcomes (UARTO) cohort who were pregnant at enrollment or became pregnant during observation were studied (n = 54). Plasma kynurenine/tryptophan (KT) ratio, soluble CD14 (sCD14), sCD163, sCD27, interferon-inducible protein 10 (IP-10), D-dimer, interleukin-6, and intestinal fatty-acid binding protein levels were assessed through the first year of ART and at 3-month intervals throughout pregnancy and 1 year postpartum. Biomarker changes were assessed with linear mixed models adjusted for ART duration. Hemoglobin concentration changes were used to estimate pregnancy-related changes in plasma volume. Results The median pre-ART CD4 count was 134. D-dimer increased through the third trimester before returning to baseline postpartum, while most other biomarkers declined significantly during pregnancy, beyond what would be expected from pregnancy-associated plasma volume expansion. IP-10 and sCD14 remained suppressed for at least 12 months postpartum. KT ratio was the only biomarker that increased above prepregnancy baseline postpartum (mean + 30%; P Conclusions Several immune activation markers decline during pregnancy and remain suppressed postpartum, but the kynurenine pathway of tryptophan catabolism increases above baseline for ≥9 months postpartum. The mechanisms underlying postpartum kynurenine pathway activity are incompletely understood but may contribute to increased tuberculosis risk in this setting.

Key Points Question How often do persons with new febrile illness access coronavirus testing and receive a test result within 7 days of illness onset? Findings In this cohort study, generally low rates of coronavirus testing were observed in 2679 participants reporting new onset of febrile illness. Although testing rates improved somewhat during the study period, timely coronavirus test results were sought and received by only 25.9% of newly febrile persons at the end of the study analysis period in late October 2020. Meaning Our results suggest systematic underuse of coronavirus testing in patients with febrile illness that may contribute to community transmission., This cohort study examines rates of testing for SARS-CoV-2 and receipt of test results within 7 days of febrile illness onset using data from smartphone-based weekly surveys administered as part of the COVID-19 Citizen Science Study., Importance Active SARS-CoV-2 (coronavirus) transmission continues in the US. It is unclear whether better access to coronavirus testing and more consistent use of testing could substantially reduce transmission. Objective To describe coronavirus testing in persons with new onset of febrile illness and analyze whether there are changes over time and differences by race and ethnicity. Design, Setting, and Participants This cohort study used data from the COVID-19 Citizen Science Study, launched in March 2020, which recruited participants via press release, word-of-mouth, and partner organizations. Participants completed daily surveys about COVID-19 symptoms and weekly surveys about coronavirus testing. All adults (aged at least 18 years) with a smartphone were eligible to join. For this analysis, US participants with new onset of febrile illness from April 2020 to October 2020 were included. Data analysis was performed from November 2020 to March 2021. Main Outcomes and Measures Receipt of a coronavirus test result within 7 days of febrile illness onset. Results Of the 2679 participants included in this analysis, the mean (SD) age was 46.3 (13.4) years, 1983 were female (74%), 2017 were college educated (75%), and a total of 3865 distinct new febrile illness episodes were reported (300 episodes [7.8%] from Hispanic participants, 71 episodes [1.8%] from Black participants, and 3494 episodes [90.4%] from not Black, not Hispanic participants) between April 2 and October 23, 2020. In weekly surveys delivered during the 14 days after fever onset, 12% overall (753 participants) indicated receipt of a test result. Using serial survey responses and parametric time-to-event modeling, it was estimated that by 7 days after onset of febrile illness, a total of 20.5% (95% CI, 19.1%-22.0%) had received a test result. This proportion increased from 9.8% (95% CI, 7.5%-12.0%) early in the epidemic to 24.1% (95% CI, 21.5%-26.7%) at the end of July, but testing rates did not substantially improve since then, increasing to 25.9% (95% CI; 21.6%-30.3%) in late October at the start of the winter surge. Black participants reported receiving a test result about half as often as others (7% [7 of 103] of survey responses vs 12% [53 of 461] for Hispanic vs 13% [693 of 5516] for not Black, not Hispanic; P = .03). This association was not statistically significant in adjusted time-to-event models (hazard ratio = 0.59 vs not Black, not Hispanic participants; 95% CI, 0.26-1.34). Conclusions and Relevance Systematic underuse of coronavirus testing was observed in this cohort study through late October 2020, at the beginning of the winter COVID-19 surge, which may have contributed to preventable coronavirus transmission.

Background and ObjectivesThe biologic mechanisms underlying neurologic postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (PASC) are incompletely understood.MethodsWe measured markers of neurologic injury (glial fibrillary acidic protein [GFAP], neurofilament light chain [NfL]) and soluble markers of inflammation among a cohort of people with prior confirmed SARS-CoV-2 infection at early and late recovery after the initial illness (defined as less than and greater than 90 days, respectively). The primary clinical outcome was the presence of self-reported CNS PASC symptoms during the late recovery time point. We compared fold changes in marker values between those with and without CNS PASC symptoms using linear mixed-effects models and examined relationships between neurologic and immunologic markers using rank linear correlations.ResultsOf 121 individuals, 52 reported CNS PASC symptoms. During early recovery, those who went on to report CNS PASC symptoms had elevations in GFAP (1.3-fold higher mean ratio, 95% CI 1.04–1.63, p = 0.02), but not NfL (1.06-fold higher mean ratio, 95% CI 0.89–1.26, p = 0.54). During late recovery, neither GFAP nor NfL levels were elevated among those with CNS PASC symptoms. Although absolute levels of NfL did not differ, those who reported CNS PASC symptoms demonstrated a stronger downward trend over time in comparison with those who did not report CNS PASC symptoms (p = 0.041). Those who went on to report CNS PASC also exhibited elevations in interleukin 6 (48% higher during early recovery and 38% higher during late recovery), monocyte chemoattractant protein 1 (19% higher during early recovery), and tumor necrosis factor α (19% higher during early recovery and 13% higher during late recovery). GFAP and NfL correlated with levels of several immune activation markers during early recovery; these correlations were attenuated during late recovery.DiscussionSelf-reported neurologic symptoms present approximately 4 months after SARS-CoV-2 infection are associated with elevations in markers of neurologic injury and inflammation at earlier time points. Some inflammatory pathways seem to be involved months after acute infection. Additional work will be needed to better characterize these processes and to identify interventions to prevent or treat this condition.

Single-nucleotide polymorphisms (SNPs) in CYP2B6 have been shown to predict variation in plasma efavirenz concentrations, but associations between these SNPs and efavirenz-mediated depression and viral suppression are less well described. We evaluated three SNPs in CYP2B6 (rs3745274, rs28399499, and rs4803419) in Ugandan persons living with HIV. To define exposure, we used previously published pharmacokinetic modeling data to categorize participants as normal, intermediate, and poor efavirenz metabolizers. Our outcomes were probable depression in the first 2 years after antiretroviral therapy (ART) initiation (mean score of >1.75 on the Hopkins Symptom Depression Checklist) and viral suppression 6 months after ART initiation. We fit generalized estimating equation and modified Poisson regression models adjusted for demographic, clinical, and psychosocial characteristics with or without individuals with depression at the time of ART initiation. Among 242 participants, there were no differences in the pre-ART depression or viral load by efavirenz metabolism strata (p > .05). Participants were classified as normal (32%), intermediate (50%), and poor (18%) metabolizers. Seven percent (56/242) of follow-up visits met criteria for depression. Eighty-five percent (167/202) of participants who completed a 6-month visit achieved viral suppression. CYP2B6 metabolizer strata did not have a statistically significant association with either depression [adjusted risk ratio (aRR) comparing intermediate or poor vs. normal, 1.46; 95% confidence interval (CI), 0.72-2.95] or 6-month viral suppression (aRR, 1.01; 95% CI, 0.88-1.15). However, in analyses restricted to participants without pre-ART depression, poorer CYP2B6 metabolism was associated with increased odds of depression (adjusted odds ratio, 4.11; 95% CI, 1.04-16.20). Efavirenz-metabolizing allele patterns are strongly associated with risk of incident depression. Future work should elucidate further region-specific gene-environment interactions and whether alternate polymorphisms may be associated with efavirenz metabolism.

BACKGROUNDRapid case ascertainment (RCA) refers to the expeditious and detailed examination of patients with a potentially rapidly fatal disease shortly after diagnosis. RCA is frequently performed in resource-rich settings to facilitate cancer research. Despite its utility, RCA is rarely implemented in resource-limited settings and has not been performed for malignancies. One cancer and context that would benefit from RCA in a resource-limited setting is HIV-related Kaposi sarcoma (KS) in sub-Saharan Africa.METHODSTo determine the feasibility of RCA of KS, we searched for all potential newly diagnosed KS among HIV-infected adults attending three community-based facilities in Uganda and Kenya. Searching involved querying of electronic medical records, pathology record review, and notification by clinicians. Upon identification, a team verified eligibility and attempted to locate patients to perform RCA, which included epidemiologic, clinical and laboratory measurements.RESULTSWe identified 593 patients with suspected new KS. Of the 593, 171 were ineligible, mainly because biopsy failed to confirm KS (65%) or KS was not new (30%). Among the 422 remaining, RCA was performed within 1 month for 56% of patients and within 3 months for 65% (95% confidence interval: 59 to 70%). Reasons for not performing RCA included intervening death (47%), inability to contact (44%), refusal/unsuitable to consent (8.3%), and patient re-location (0.7%).CONCLUSIONSWe found that RCA — an important tool for cancer research in resource-rich settings — is feasible for the investigation of community-representative KS in East Africa. Feasibility of RCA for KS suggests feasibility for other cancers in Africa.

Immune activation complicates HIV despite antiretroviral therapy (ART). Indoleamine 2,3 dioxygenase (IDO) catabolizes tryptophan (T) to kynurenine (K), regulating immune activity, and IDO activity increases with age. This study examines the relationship of IDO activity, bacterial translocation, and aging in people living with HIV (PLWH) on ART. Samples and data from PLWH on ART from the Centers for AIDS Research Network of Integrated Clinical Systems and from matched HIV-uninfected patients (controls) from the Multicenter AIDS Cohort Study and the Women's Interagency HIV Study were analyzed. The ratio of K to T (K:T) and neopterin were indicators of inflammation; 16S ribosomal DNA (16S rDNA) and lipopolysaccharide (LPS) were markers of bacterial translocation. Samples and data from 205 PLWH and 99 controls were analyzed. PLWH had higher K:T values across all ages, with a significant relationship between age and K:T for both groups. CD4 count or CD4 nadir had no association with K:T. There was no positive association between level of 16S rDNA or LPS detection and K:T. K:T and neopterin were associated. PLWH had elevated IDO activity, at younger ages, despite ART. This study suggests K:T ratio increases with age in both groups and is elevated in PLWH at all ages compared with age-matched controls.

Antibody detection of prior SARS-CoV-2 infection depends on severity, assay, and timing with implications for serosurveillance., Interpretation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serosurveillance studies is limited by poorly defined performance of antibody assays over time in individuals with different clinical presentations. We measured antibody responses in plasma samples from 128 individuals over 160 days using 14 assays. We found a consistent and strong effect of disease severity on antibody magnitude, driven by fever, cough, hospitalization, and oxygen requirement. Responses to spike protein versus nucleocapsid had consistently higher correlation with neutralization. Assays varied substantially in sensitivity during early convalescence and time to seroreversion. Variability was dramatic for individuals with mild infection, who had consistently lower antibody titers, with sensitivities at 6 months ranging from 33 to 98% for commercial assays. Thus, the ability to detect previous infection by SARS-CoV-2 is highly dependent on infection severity, timing, and the assay used. These findings have important implications for the design and interpretation of SARS-CoV-2 serosurveillance studies.

As cases of coronavirus disease 2019 (COVID-19) mount worldwide, attention is needed on potential long-term neurologic impacts for the majority of patients who experience mild to moderate illness managed as outpatients. To date, there has not been discussion of persistent neurocognitive deficits in patients with milder COVID-19. We present two cases of non-hospitalized patients recovering from COVID-19 with persistent neurocognitive symptoms. Commonly used cognitive screens were normal, while more detailed testing revealed working memory and executive functioning deficits. An observational cohort study of individuals recovering from COVID-19 (14 or more days following symptom onset) identified that among the first 100 individuals enrolled, 14 were non-hospitalized patients reporting persistent cognitive issues. These 14 participants had a median age of 39 years (interquartile range: 35–56), and cognitive symptoms were present for at least a median of 98 days (interquartile range: 71–120 following acute COVID-19 symptoms); no participants with follow-up evaluation reported symptom resolution. We discuss potential mechanisms to be explored in future studies, including direct viral effects, indirect consequences of immune activation, and immune dysregulation causing auto-antibody production.

The precise role of CD4 T cell turnover in maintaining HIV persistence during antiretroviral therapy (ART) has not yet been well characterized. In resting CD4 T cell subpopulations from 24 HIV-infected ART-suppressed and 6 HIV-uninfected individuals, we directly measured cellular turnover by heavy water labeling, HIV reservoir size by integrated HIV-DNA (intDNA) and cell-associated HIV-RNA (caRNA), and HIV reservoir clonality by proviral integration site sequencing. Compared to HIV-negatives, ART-suppressed individuals had similar fractional replacement rates in all subpopulations, but lower absolute proliferation rates of all subpopulations other than effector memory (TEM) cells, and lower plasma IL-7 levels (p = 0.0004). Median CD4 T cell half-lives decreased with cell differentiation from naïve to TEM cells (3 years to 3 months, p, Author summary HIV infection is a life-long disease for which we do not currently have a cure. One reason for this is that the virus goes into a dormant state and hides in CD4 T cell lymphocytes. Many of the less mature infected cells slowly self-renew (i.e., replace themselves) over time, whereas other types of infected cells rapidly proliferate and expand, and are replaced more rapidly. In our study, we directly measured how rapidly different types of CD4 T cells divide in treated people with HIV. In addition to confirming that less mature CD4 T cell populations self-renew at a very slow rate, and that more mature cells proliferate and expand at a more rapid rate, we also found that cell types that divide more rapidly are more likely to contain HIV. By carefully defining how rapidly these different cell populations are replaced, we help inform studies of HIV cure interventions that specifically target these discrete populations of HIV-infected cells.

As the SARS-CoV-2 pandemic continues, reports have demonstrated neurologic sequelae following COVID-19 recovery. Mechanisms to explain long-term neurological sequelae are unknown and need to be identified. Plasma from 24 individuals recovering from COVID-19 at 1 to 3 months after initial infection were collected for cytokine and antibody levels and neuronal-enriched extracellular vesicle (nEV) protein cargo analyses. Plasma cytokine IL-4 was increased in all COVID-19 participants. Volunteers with self-reported neurological problems (nCoV, n = 8) had a positive correlation of IL6 with age or severity of the sequalae, at least one co-morbidity and increased SARS-CoV-2 antibody compared to those COVID-19 individuals without neurological issues (CoV, n = 16). Protein markers of neuronal dysfunction including amyloid beta, neurofilament light, neurogranin, total tau, and p-T181-tau were all significantly increased in the nEVs of all participants recovering from COVID-19 compared to historic controls. This study suggests ongoing peripheral and neuroinflammation after COVID-19 infection that may influence neurological sequelae by altering nEV proteins. Individuals recovering from COVID-19 may have occult neural damage while those with demonstrative neurological symptoms additionally had more severe infection. Longitudinal studies to monitor plasma biomarkers and nEV cargo are warranted to assess persistent neurodegeneration and systemic effects.

The current severe acute respiratory syndrome coronavirus 2 testing policy and practice limits testing as a prevention tool. Radical shifts are required to increase the scale of rapid testing strategies and improve dissemination and implementation of venue-based and self-testing approaches. Attention to the full translation pipeline is required to reach high-risk segments of the population., We propose a broad translation perspective be applied to COVID-19 testing development with the goals of improving not only scalability, but also test dissemination, implementation, and reach.

HIV Nef counteracts cellular host restriction factors SERINC3 and SERINC5, but our understanding of how naturally occurring global Nef sequence diversity impacts these activities is limited. Here, we quantify SERINC3 and SERINC5 internalization function for 339 Nef clones, representing the major pandemic HIV-1 group M subtypes A, B, C and D. We describe distinct subtype-associated hierarchies for Nef-mediated internalization of SERINC5, for which subtype B clones display the highest activities on average, and of SERINC3, for which subtype B clones display the lowest activities on average. We further identify Nef polymorphisms that modulate its ability to counteract SERINC proteins, including substitutions in the N-terminal domain that selectively impair SERINC3 internalization. Our findings demonstrate that the SERINC antagonism activities of HIV Nef differ markedly among major viral subtypes and between individual isolates within a subtype, suggesting that variation in these functions may contribute to global differences in viral pathogenesis., Author summary Cellular restriction factors dampen viral replication and reduce pathogenesis. In response, HIV has evolved a variety of mechanisms to counteract restriction factors, including the ability of its Nef protein to internalize Serine incorporator (SERINC) proteins 3 and 5 from the infected cell surface, thereby enhancing infectivity of viral particles. Nef displays substantial sequence diversity, but how this impacts SERINC antagonism remains unclear. To examine this, we used cell culture models to measure SERINC internalization function for 339 participant-derived Nef clones, representing four globally relevant HIV-1 group M subtypes. We observed significant variability in Nef function among circulating viral strains and subtypes. We also identified naturally occurring Nef mutations that modulate its ability to counteract SERINC proteins, including some that selectively impair SERINC3 internalization. These findings uncover viral features that may contribute to global differences in HIV pathogenesis and provide new insight into the interactions between Nef and SERINC restriction factors that can inform future mechanistic studies.

The HIV-1 accessory protein Vpu enhances viral spread by downregulating CD4 and BST-2/tetherin on the surface of infected cells. Natural variability in these Vpu functions may contribute to HIV-1 pathogenesis, but this has not been investigated among the diverse viral subtypes that contribute to the HIV-1 pandemic. In this study, we found that Vpu function differs significantly among HIV-1 subtypes A, B, C, and D. On average, subtype C clones displayed the lowest ability to downregulate both CD4 and tetherin, while subtype B and D clones were more functional. We also identified Vpu polymorphisms that associate with functional differences among HIV-1 isolates and subtypes. Our study suggests that genetic diversity in Vpu may play an important role in the differential pathogenesis and/or spread of HIV-1., Downregulation of BST-2/tetherin and CD4 by HIV-1 viral protein U (Vpu) promotes viral egress and allows infected cells to evade host immunity. Little is known however about the natural variability in these Vpu functions among the genetically diverse viral subtypes that contribute to the HIV-1 pandemic. We collected Vpu isolates from 332 treatment-naive individuals living with chronic HIV-1 infection in Uganda, Rwanda, South Africa, and Canada. Together, these Vpu isolates represent four major HIV-1 group M subtypes (A [n = 63], B [n = 84], C [n = 94], and D [n = 59]) plus intersubtype recombinants and uncommon strains (n = 32). The ability of each Vpu clone to downregulate endogenous CD4 and tetherin was quantified using flow cytometry following transfection into an immortalized T-cell line and compared to that of a reference Vpu clone derived from HIV-1 subtype B NL4.3. Overall, the median CD4 downregulation function of natural Vpu isolates was similar to that of NL4.3 (1.01 [interquartile range {IQR}, 0.86 to 1.18]), while the median tetherin downregulation function was moderately lower than that of NL4.3 (0.90 [0.79 to 0.97]). Both Vpu functions varied significantly among HIV-1 subtypes (Kruskal-Wallis P

BACKGROUND: The relative stabilities of the intact and defective HIV genomes over time during effective antiretroviral therapy (ART) have not been fully characterized. METHODS: We used the intact proviral DNA assay (IPDA) to estimate the rate of change of intact and defective proviruses in HIV-infected adults on ART. We used linear spline models with a knot at seven years and a random intercept and slope up to the knot. We estimated the influence of covariates on rates of change. RESULTS: We studied 81 individuals for a median of 7.3 (IQR 5.9-9.6) years. Intact genomes declined more rapidly from initial suppression through seven years (15.7% per year decline; 95% CI -22.8%, -8.0%) and more slowly after seven years (3.6% per year; 95% CI -8.1%, +1.1%). The estimated half-life of the reservoir was 4.0 years (95% CI 2.7-8.3) until year seven and 18.7 years (95% CI 8.2-infinite) thereafter. There was substantial variability between individuals in the rate of decline until year seven. Intact provirus declined more rapidly than defective provirus (P < 0.001) and showed a faster decline in individuals with higher CD4(+) T cell nadirs. CONCLUSION: The biology of the replication-competent (intact) reservoir differs from that of the replication-incompetent (non-intact) pool of proviruses. The IPDA will likely be informative when investigating the impact of interventions targeting the reservoir. FUNDING: Delaney AIDS Research Enterprise, UCSF/Gladstone Institute of Virology & Immunology CFAR, CFAR Network of Integrated Systems, amfAR Institute for HIV Cure Research, I4C and Beat-HIV Collaboratories, Howard Hughes Medical Institute, Gilead Sciences, Bill and Melinda Gates Foundation.

Although many HIV cure strategies seek to expand HIV-specific CD8+ T cells to control the virus, all are likely to fail if cellular exhaustion is not prevented. A loss in stem-like memory properties (i.e., the ability to proliferate and generate secondary effector cells) is a key feature of exhaustion; little is known, however, about how these properties are regulated in human virus-specific CD8+ T cells. We found that virus-specific CD8+ T cells from humans and nonhuman primates naturally controlling HIV/SIV infection express more of the transcription factor TCF-1 than noncontrollers. HIV-specific CD8+ T cell TCF-1 expression correlated with memory marker expression and expansion capacity and declined with antigenic stimulation. CRISPR-Cas9 editing of TCF-1 in human primary T cells demonstrated a direct role in regulating expansion capacity. Collectively, these data suggest that TCF-1 contributes to the regulation of the stem-like memory property of secondary expansion capacity of HIV-specific CD8+ T cells, and they provide a rationale for exploring the enhancement of this pathway in T cell-based therapeutic strategies for HIV.

Background Kaposi’s sarcoma (KS) is one of the most common HIV-associated malignancies in sub-Saharan Africa. Worldwide, the availability of antiretroviral therapy (ART) has improved KS survival. In resource-rich settings, survival has also benefited from chemotherapy, which is widely available. Little is known, however, about the epidemiology of chemotherapy use for HIV-associated KS in resource-limited regions such as sub-Saharan Africa. Methods We identified all patients newly diagnosed with HIV-related KS from 2009 to 2012 in the 26-clinic AMPATH network, a large community-based care network in Kenya. We ascertained disease severity at diagnosis, frequency of initiation of chemotherapy, and distribution of chemotherapeutic regimens used. Indications for chemotherapy included AIDS Clinical Trial Group T1 stage and/or “severe” disease defined by WHO KS treatment guidelines. Results Of 674 patients diagnosed with KS, charts were available for 588; 61% were men, median age was 35 years, and median CD4 at KS diagnosis was 185 cells/μl. At time of diagnosis, 58% had at least one chemotherapy indication, and 22% had more than one indication. For patients with a chemotherapy indication, cumulative incidence of chemotherapy initiation (with death as a competing event) was 37% by 1 month and 56% by 1 year. Median time from diagnosis to chemotherapy initiation was 25 days (IQR 1–50 days). In multivariable regression, patients with > 3 chemotherapy indications at time of diagnosis had a 2.30 (95% CI 1.46–3.60) increased risk of rapid chemotherapy initiation (within 30 days of diagnosis) compared to those with only one chemotherapy indication (p Conclusions A substantial fraction of patients with KS in East Africa are diagnosed at advanced disease stage. For patients with chemotherapy indications, nearly half did not receive chemotherapy by one year. Liposomal anthracyclines, often used in resource-rich settings, were not first line. These findings emphasize challenges in East Africa cancer care, and highlight the need for further advocacy for improved access to higher quality chemotherapy in this setting.

Background People living with human immunodeficiency virus (HIV; PLWH) have a markedly elevated anal cancer risk, largely due to loss of immunoregulatory control of oncogenic human papillomavirus infection. To better understand anal cancer development and prevention, we determined whether recent, past, cumulative, or nadir/peak CD4+ T-cell count (CD4) and/or HIV-1 RNA level (HIV RNA) best predict anal cancer risk. Methods We studied 102 777 PLWH during 1996–2014 from 21 cohorts participating in the North American AIDS Cohort Collaboration on Research and Design. Using demographics-adjusted, cohort-stratified Cox models, we assessed associations between anal cancer risk and various time-updated CD4 and HIV RNA measures, including cumulative and nadir/peak measures during prespecified moving time windows. We compared models using the Akaike information criterion. Results Cumulative and nadir/peak CD4 or HIV RNA measures from approximately 8.5 to 4.5 years in the past were generally better predictors for anal cancer risk than their corresponding more recent measures. However, the best model included CD4 nadir (ie, the lowest CD4) from approximately 8.5 years to 6 months in the past (hazard ratio [HR] for Conclusions Our results are consistent with anal cancer promotion by severe, prolonged HIV-induced immunosuppression. Nadir and cumulative CD4 may represent useful markers for identifying PLWH at higher anal cancer risk.

BackgroundThe utility and validity of cardiovascular diseases (CVD) risk scores are not well studied in sub-Saharan Africa. We compared and correlated CVD risk scores with carotid intima media thickness (c-IMT) among HIV-infected and uninfected people in Uganda.MethodsWe first calculated CVD risk using the (1) Framingham laboratory-based score; (2) Framingham nonlaboratory score (FRS-BMI); (3) Reynolds risk score; (4) American College of Cardiology and American Heart Association score; and (5) the Data collection on Adverse Effects of Anti-HIV Drugs score. We then compared absolute risk scores and risk categories across each score using Pearson correlation and kappa statistics, respectively. Finally, we fit linear regression models to estimate the strength of association between each risk score and c-IMT.ResultsOf 205 participants, half were females and median age was 49 years [interquartile range (IQR) 46-53]. Median CD4 count was 430 cells/mm (IQR 334-546), with median 7 years of antiretroviral therapy exposure (IQR 6.4-7.5). HIV-uninfected participants had a higher median systolic blood pressure (121 vs. 110 mm Hg), prevalent current smokers (18% vs. 4%, P = 0.001), higher median CVD risk scores (P < 0.003), and greater c-IMT (0.68 vs. 0.63, P = 0.003). Overall, FRS-BMI was highly correlated with other risk scores (all rho >0.80). In linear regression models, we found significant correlations between increasing CVD risk and higher c-IMT (P < 0.01 in all models).ConclusionsIn this cross-sectional study from Uganda, the FRS-BMI correlated well with standard risk scores and c-IMT. HIV-uninfected individuals had higher risk scores than HIV-infected individuals, and the difference seemed to be driven by modifiable factors.

BACKGROUND: Age-associated conditions are increasingly common among persons living with human immunodeficiency virus (HIV) (PLWH). A longitudinal investigation of their accrual is needed given their implications on clinical care complexity. We examined trends in the co-occurrence of age-associated conditions among PLWH receiving clinical care, and differences in their prevalence by demographic subgroup. METHODS: This cohort study was nested within the North American AIDS Cohort Collaboration on Research and Design. Participants from HIV outpatient clinics were antiretroviral therapy–exposed PLWH receiving clinical care (ie, ≥1 CD4 count) in the United States during 2000–2009. Multimorbidity was irreversible, defined as having ≥2: hypertension, diabetes mellitus, chronic kidney disease, hypercholesterolemia, end-stage liver disease, or non–AIDS-related cancer. Adjusted prevalence ratios (aPR) and 95% confidence intervals (CIs) comparing demographic subgroups were obtained by Poisson regression with robust error variance, using generalized estimating equations for repeated measures. RESULTS: Among 22969 adults, 79% were male, 36% were black, and the median baseline age was 40 years (interquartile range, 34–46 years). Between 2000 and 2009, multimorbidity prevalence increased from 8.2% to 22.4% (P(trend) < .001). Adjusting for age, this trend was still significant (P < .001). There was no difference by sex, but blacks were less likely than whites to have multimorbidity (aPR, 0.87; 95% CI, .77–.99). Multimorbidity was the highest among heterosexuals, relative to men who have sex with men (aPR, 1.16; 95% CI, 1.01–1.34). Hypertension and hypercholesterolemia most commonly co-occurred. CONCLUSIONS: Multimorbidity prevalence has increased among PLWH. Comorbidity prevention and multisubspecialty management of increasingly complex healthcare needs will be vital to ensuring that they receive needed care.

We describe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T cell responses, soluble markers of inflammation, and antibody levels and neutralization capacity longitudinally in 70 individuals with PCR-confirmed SARS-CoV-2 infection. Participants represent a spectrum of illness and recovery, including some with persistent viral shedding in saliva and many experiencing post-acute sequelae of SARS-CoV-2 infection (PASC). T cell responses remain stable for up to 9 months. Whereas the magnitude of early CD4+ T cell immune responses correlates with severity of initial infection, pre-existing lung disease is independently associated with higher long-term SARS-CoV-2-specific CD8+ T cell responses. Among participants with PASC 4 months following coronavirus disease 2019 (COVID-19) symptom onset, we observe a lower frequency of CD8+ T cells expressing CD107a, a marker of degranulation, in response to Nucleocapsid (N) peptide pool stimulation, and a more rapid decline in the frequency of N-specific interferon-γ-producing CD8+ T cells. Neutralizing antibody levels strongly correlate with SARS-CoV-2-specific CD4+ T cell responses., Graphical abstract, CD4+ and CD8+ T cell responses following natural infection with COVID-19 are stable over 8 months. Individuals with PASC demonstrate a lower frequency of CD8+ T cells expressing CD107a, a marker of degranulation, and a more rapid decline in the frequency of N-specific interferon-γ-producing CD8+ T cells.

Despite a poor toxicity profile, zidovudine supersedes abacavir (ABC) as an alternative first-line agent in most international treatment guidelines because of concerns about HLA-B*57:01-related ABC-hypersensitivity. We detected one case of HLA-B*57:01 carriage among 513 HIV-infected individuals in Uganda, which, in combination with previous reports, supports the safety of ABC in the region.

Objective: Long-acting rilpivirine is a candidate for preexposure prophylaxis (PrEP) for prevention of HIV-1 infection. However, rilpivirine resistance mutations at reverse transcriptase codon 138 (E138X) occur naturally in a minority of HIV-1-infected persons; in particular those expressing human leukocyte antigen (HLA)-B*18 where reverse transcriptase-E138X arises as an immune escape mutation. We investigate the global prevalence, B*18-linkage and replicative cost of reverse transcriptase-E138X and its regional implications for rilpivirine PrEP. Methods: We analyzed linked reverse transcriptase-E138X/HLA data from 7772 antiretroviral-naive patients from 16 cohorts spanning five continents and five HIV-1 subtypes, alongside unlinked global reverse transcriptase-E138X and HLA frequencies from public databases. E138X-containing HIV-1 variants were assessed for in-vitro replication as a surrogate of mutation stability following transmission. Results: Reverse transcriptase-E138X variants, where the most common were rilpivirine resistance-associated mutations E138A/G/K, were significantly enriched in HLA-B*18-positive individuals globally (P = 3.5 × 10−20) and in all HIV-1 subtypes except A. Reverse transcriptase-E138X and B*18 frequencies correlated positively in 16 cohorts with linked HIV/HLA genotypes (Spearman's R = 0.75; P = 7.6 × 10−4) and in unlinked HIV/HLA data from 43 countries (Spearman's R = 0.34, P = 0.02). Notably, reverse transcriptase-E138X frequencies approached (or exceeded) 10% in key epidemic regions (e.g. sub-Saharan Africa, Southeastern Europe) where B*18 is more common. This, along with the observation that reverse transcriptase-E138X variants do not confer in-vitro replicative costs, supports their persistence, and ongoing accumulation in circulation over time. Conclusions: Results illustrate the potential for a natural immune-driven HIV-1 polymorphism to compromise antiretroviral-based prevention, particularly in key epidemic regions. Regional reverse transcriptase-E138X surveillance should be undertaken before use of rilpivirine PrEP.

Preventable cardiovascular disease (CVD) risk factors are responsible for the majority of CVD-related deaths, and are increasingly recognized as a cause of morbidity and mortality for HIV-infected persons taking antiretroviral therapy (ART). Simplified tools such as the American Heart Association's ideal cardiovascular health (iCVH) construct may identify and prognosticate CVD risk in resource-limited settings. No studies have evaluated iCVH metrics in sub-Saharan Africa or among HIV-infected adults. Thus, the central aim of this study was to compare levels of iCVH metrics and their correlations with carotid atherosclerosis for HIV-infected adults versus uninfected controls in a well-phenotyped Ugandan cohort. We analyzed the prevalence of iCVH metrics in a mixed cohort of HIV-infected persons on stable ART and uninfected, population-based comparators in Mbarara, Uganda. We also assessed the validity of iCVH by correlating iCVH values with common carotid intima media thickness (CCIMT). HIV-infected persons had a mean of 4.9 (SD 1.1) iCVH metrics at ideal levels versus 4.3 (SD 1.2) for uninfected controls (p = .002). This difference was largely driven by differences in blood pressure, blood glucose, and diet. In multivariable-adjusted linear regression models, each additional iCVH metric at an ideal level was associated with a significant 0.024 mm decrease in CCIMT (p