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IntroductionLatent tuberculosis infection (LTBI) is a common coinfection in people living with HIV (PWH). How LTBI and HIV exposure in utero influence the development of infant humoral immunity is not well characterized. To address this question, we assessed the relationship between maternal humoral responses in pregnant women with HIV or with HIV/LTBI on humoral responses in infants to BCG vaccination and TB acquisition.MethodsPlasma samples were obtained from mother infant pairs during pregnancy (14-34 wks gestation) and in infants at 12 and 44 wks of age from the IMPAACT P1078 clinical trial. LTBI was established by Interferon gamma release assay (IGRA). Progression to active TB (ATB) disease was observed in 5 women at various times after giving birth. All infants were BCG vaccinated at birth and tested for IGRA at 44 weeks. Mtb (PPD, ESAT6/CFP10, Ag85A, LAM), HIV (GP120), and Influenza (HA) specific IgG, IgM, and IgA were measured in plasma samples using a bead based Luminex assay with Flexmap 3D.ResultsIn maternal plasma there were no differences in Mtb-specific antibodies or viral antibodies in relation to maternal IGRA status. ATB progressors showed increases in Mtb-specific antibodies at diagnosis compared to study entry. However, when compared to the non-progressors at entry, progressors had higher levels of Ag85A IgG and reduced ESAT6/CFP10 IgG and LAM IgG, IgM, and IgA1. All infants showed a decrease in IgG to viral antigens (HIV GP120 and HA) from 12 to 44 weeks attributed to waning of maternally transferred antibody titers. However, Mtb-specific (PPD, ESAT6/CFP10, Ag85A, and LAM) IgG and IgM increased from 12 to 44 weeks. HIV and HA IgG levels in maternal and 12-week infant plasma were highly correlated, and ESAT6/CFP10 IgG and LAM IgG showed a relationship between maternal and infant Abs. Finally, in the subset of infants that tested IGRA positive at 44 weeks, we observed a trend for lower LAM IgM compared to IGRA- infants at 44 weeks.DiscussionThe results from our study raise the possibility that antibodies to LAM are associated with protection from progression to ATB and support further research into the development of humoral immunity against TB through infection or vaccination.

Summary: In perinatal HIV infection, early antiretroviral therapy (ART) initiation is recommended but questions remain regarding infant immune responses to HIV and its impact on immune development. Using single cell transcriptional and phenotypic analysis we evaluated the T cell compartment at pre-ART initiation of infants with perinatally acquired HIV from Maputo, Mozambique (Towards AIDS Remission Approaches cohort). CD8+ T cell maturation subsets exhibited altered distribution in HIV exposed infected (HEI) infants relative to HIV exposed uninfected infants with reduced naive, increased effectors, higher frequencies of activated T cells, and lower frequencies of cells with markers of self-renewal. Additionally, a cluster of CD8+ T cells identified in HEI displayed gene profiles consistent with cytotoxic T lymphocytes and showed evidence for hyper expansion. Longitudinal phenotypic analysis revealed accelerated maturation of CD8+ T cells was maintained in HEI despite viral control. The results point to an HIV-directed immune response that is likely to influence reservoir establishment.

BackgroundBoth alcohol consumption and HIV infection are associated with worse brain, cognitive, and clinical outcomes in older adults. However, the extent to which brain and cognitive dysfunction is reversible with reduction or cessation of drinking is unknown. ObjectiveThe 30-Day Challenge study was designed to determine whether reduction or cessation of drinking would be associated with improvements in cognition, reduction of systemic and brain inflammation, and improvement in HIV-related outcomes in adults with heavy drinking. MethodsThe study design was a mechanistic experimental trial, in which all participants received an alcohol reduction intervention followed by repeated assessments of behavioral and clinical outcomes. Persons were eligible if they were 45 years of age or older, had weekly alcohol consumption of 21 or more drinks (men) or 14 or more drinks (women), and were not at high risk of alcohol withdrawal. After a baseline assessment, participants received an intervention consisting of contingency management (money for nondrinking days) for at least 30 days followed by a brief motivational interview. After this, participants could either resume drinking or not. Study questionnaires, neurocognitive assessments, neuroimaging, and blood, urine, and stool samples were collected at baseline, 30 days, 90 days, and 1 year after enrollment. ResultsWe enrolled 57 persons with heavy drinking who initiated the contingency management protocol (mean age 56 years, SD 4.6 years; 63%, n=36 male, 77%, n=44 Black, and 58%, n=33 people with HIV) of whom 50 completed 30-day follow-up and 43 the 90-day follow-up. The planned study procedures were interrupted and modified due to the COVID-19 pandemic of 2020-2021. ConclusionsThis was the first study seeking to assess changes in brain (neuroimaging) and cognition after alcohol intervention in nontreatment-seeking people with HIV together with people without HIV as controls. Study design strengths, limitations, and lessons for future study design considerations are discussed. Planned analyses are in progress, after which deidentified study data will be available for sharing. Trial RegistrationClinicalTrials.gov NCT03353701; https://clinicaltrials.gov/study/NCT03353701 International Registered Report Identifier (IRRID)DERR1-10.2196/53684

The objectives of this study were to (1) investigate the association between human blood type and COVID-19 in both inpatient and longitudinal populations and (2) identify the association between blood type and severity of COVID-19 via presence of cellular biomarkers of severe infection in hospitalized individuals at our institution in South Florida. This study consisted of (1) a single-center retrospective analysis of 669 out of 2741 COVID-19-positive, screened patients seen from 1 January 2020 until 31 March 2021 at the University of Miami Emergency Department (ED) who tested positive for COVID-19 and had a documented ABO blood type and (2) a longitudinal SARS-CoV-2 immunity study (“CITY”) at the University of Miami Miller School of Medicine, consisting of 185 survey participants. In an inpatient setting, blood type appeared to be associated with COVID-19 severity and mortality. Blood type O sustained less risk of COVID-19 mortality, and blood type O- demonstrated less risk of developing COVID-19 pneumonia. Inpatients with O- blood type exhibited less biomarkers of severe infection than did other blood types. In a longitudinal setting, there was no association found between blood type and SARS-CoV-2 infection.

False positive reactive plasmin reagin (RPR) reactivity following a COVID-19 vaccine has been reported, and it is therefore conceivable that individuals who receive frequent coronavirus disease 2019 (COVID-19) vaccinations may exhibit durable RPR responses. Here, we sought to investigate the extent to which repeated mRNA COVID-19 vaccines can elicit chronic false RPR reactivity in a longitudinal cohort. Participants (n = 119) in an IRB-approved (#20201026), longitudinal SARS-CoV-2 cohort study were screened for RPR reactivity via manual RPR card assays. Samples with reactive results underwent additional testing, including follow-on RPR screening at additional timepoints, confirmatory fluorescent treponemal antibody (FTA-ABS) testing and anti-nuclear antibody (ANA) testing. Medical histories were collected. We observed (n = 2) screen-positive RPR results (1.7% [2/119]) following booster vaccination, for which two individuals exhibited chronic, vaccine-induced RPR reactivity for up to 9 months following booster vaccination. Both participants were ANA-negative. It is imperative for clinicians to be mindful of the potential immunologic interference of COVID-19 vaccines with standard infectious disease assays, including RPR testing. Detailed medical histories and clinical contexts, including recent vaccination, should be reviewed prior to proceeding with distressing and invasive workups.

Post vaccine immunity following COVID-19 mRNA vaccination may be driven by extrinsic, or controllable and intrinsic, or inherent health factors. Thus, we investigated the effects of extrinsic and intrinsic on the peak antibody response following COVID-19 primary vaccination and on the trajectory of peak antibody magnitude and durability over time. Participants in a longitudinal cohort attended visits every 3 months for up to 2 years following enrollment. At baseline, participants provided information on their demographics, recreational behaviors, and comorbid health conditions which guided our model selection process. Blood samples were collected for serum processing and spike antibody testing at each visit. Cross-sectional and longitudinal models (linear-mixed effects models) were generated to assess the relationship between selected intrinsic and extrinsic health factors on peak antibody following vaccination and to determine the influence of these predictors on antibody over time. Following cross-sectional analysis, we observed higher peak antibody titers after primary vaccination in females, those who reported recreational drug use, younger age, and prior COVID-19 history. Following booster vaccination, females and Hispanics had higher peak titers after the 3rd and 4th doses, respectively. Longitudinal models demonstrated that Moderna mRNA-1273 recipients, females, and those previously vaccinated had increased peak titers over time. Moreover, drug users and half-dose Moderna mRNA-1273 recipients had higher peak antibody titers over time following the first booster, while no predictive factors significantly affected post-second booster antibody responses. Overall, both intrinsic and extrinsic health factors play a significant role in shaping humoral immunogenicity after initial vaccination and the first booster. The absence of predictive factors for second booster immunogenicity suggests a more robust and consistent immune response after the second booster vaccine administration.

People with HIV (PWH) frequently suffer from Opioid (OP) Use Disorder (OUD). In an investigation of the impact of OUD on underlying immune dysfunction in PWH, we previously reported that OP use exacerbates inflammation in virally controlled PWH followed in the Infectious Diseases Elimination Act (IDEA) Syringe Services Program (SSP). Unexpectedly, Flu vaccination-induced antibody responses in groups with OUD were superior to PWH without OUD. Here, we investigated the profile of 48 plasma biomarkers comprised of TNF and Ig superfamily (SF) molecules known to impact interactions between T and B cells in 209 participants divided into four groups: (1) HIV+OP+, (2) HIV−OP+, (3) HIV+OP−, and (4) HIV−OP−. The differential expression of the top eight molecules ranked by median values in individual Groups 1–3 in comparison to Group 4 was highly significant. Both OP+ groups 1 and 2 had higher co-stimulatory TNF SF molecules, including 4-1BB, OX-40, CD40, CD30, and 4-1BBL, which were found to positively correlate with Flu Ab titers. In contrast, HIV+OP− exhibited a profile dominant in Ig SF molecules, including PDL-2, CTLA-4, and Perforin, with PDL-2 showing a negative correlation with Flu vaccine titers. These findings are relevant to vaccine development in the fields of HIV and OUD.

IntroductionPeople with HIV (PWH) are known to have underlying inflammation and immune activation despite virologic control. Substance use including opioid dependence is common in this population and is associated with increased morbidity and reduced lifespan. The primary objective of the present study termed opioid immunity study (OPIS), was to investigate the impact of chronic opioids in PWH.MethodsThe study recruited people with and without HIV who had opioid use disorder (OUD). Study participants (n=221) were categorized into four groups: HIV+OP+, n=34; HIV-OP+, n=66; HIV+OP-, n=55 and HIV-OP-, n=62 as controls. PWH were virally suppressed on ART and those with OUD were followed in a syringe exchange program with confirmation of OP use by urine drug screening. A composite cytokine score was developed for 20 plasma cytokines that are linked to inflammation. Cellular markers of immune activation (IA), exhaustion, and senescence were determined in CD4 and CD8 T cells. Regression models were constructed to examine the relationships of HIV status and opioid use, controlling for other confounding factors.ResultsHIV+OP+ participants exhibited highest inflammatory cytokines and cellular IA, followed by HIV-OP+ for inflammation and HIV+OP- for IA. Inflammation was found to be driven more by opioid use than HIV positivity while IA was driven more by HIV than opioid use. In people with OUD, expression of CD38 on CD28-CD57+ senescent-like T cells was elevated and correlated positively with inflammation.DiscussionGiven the association of inflammation with a multitude of adverse health outcomes, our findings merit further investigations to understand the mechanistic pathways involved.

Background: The persistence of HIV-1-infected cells during antiretroviral therapy is well documented but may be modulated by early initiation of antiretroviral therapy in infants. Methods: Here, we longitudinally analyzed the proviral landscape in nine infants with vertical HIV-1 infection from Mozambique over a median period of 24 months, using single-genome, near full-length, next-generation proviral sequencing. Results: We observed a rapid decline in the frequency of intact proviruses, leading to a disproportional under-representation of intact HIV-1 sequences within the total number of HIV-1 DNA sequences after 12-24 months of therapy. In addition, proviral integration site profiling in one infant demonstrated clonal expansion of infected cells harboring intact proviruses and indicated that viral rebound was associated with an integration site profile dominated by intact proviruses integrated into genic and accessible chromatin locations. Conclusion: Together, these results permit rare insight into the evolution of the HIV-1 reservoir in infants infected with HIV-1 and suggest that the rapid decline of intact proviruses, relative to defective proviruses, may be attributed to a higher vulnerability of genome-intact proviruses to antiviral immunity. Technologies to analyze combinations of intact proviral sequences and corresponding integration sites permit a high-resolution analysis of HIV-1 reservoir cells after early antiretroviral treatment initiation in infants.

Summary: Background: Despite antiretroviral treatment (ART), immune dysfunction persists in children with perinatal HIV infection (HEI). Here we investigated the impact of HIV status on maternal antibody (Ab) passage, long-term vaccine induced immunity and B-cell maturation. Methods: 46 HIV Exposed Uninfected (HEU), 43 HEI, and 15 HIV unexposed uninfected (HUU) infants were vaccinated with 3 doses of DTaP-HepB-Hib-PCV10-OP at 2, 3, and 4 months at Matola Provincial Hospital, Maputo, Mozambique. Tetanus toxoid specific (TT) IgG, HIV Ab and B-cell phenotype characteristics were evaluated at entry, pre-ART, 5, 10, and 18 months in this longitudinal cohort study. Findings: Baseline (maternal) plasma TT Ab levels were significantly lower in HEI compared to both HEU and HUU and a faster decay of TT Ab was observed in HEI compared to HEU with significantly lower TT Ab levels at 10 and 18 months of age. TT unprotected (UP) (≤0.1 IU/mL) HEI showed higher HIV-RNA at entry and higher longitudinal HIV viremia (Area Under the Curve) compared to TT protected (P) HEI. A distinct HIV-Ab profile was found at entry in HEI compared to HEU. B-cell phenotype showed a B-cell perturbation in HEI vs HEU infants at entry (mean age 40.8 days) with lower transitional CD10+CD19+ B-cells and IgD+CD27− naive B-cells and an overall higher frequency of IgD−CD27− double negative B-cell subsets in HEI. Interpretation: B-cell perturbation, presenting with higher double negative IgD−CD27− B-cells was observed in neonatal age and may play a major role in the B-cell exhaustion in HEI. The ability to maintain TT protective Ab titers over time is impaired in HEI with uncontrolled viral replication and the current vaccination schedule is insufficient to provide long-term protection against tetanus. Funding: This work was supported by: NIH grant to SP (5R01AI127347-05); Children’s Hospital Bambino Gesú (Ricerca corrente 2019) to NC, and Associazione Volontari Bambino Gesù to PP.

Abstract Background Understanding the immune correlates of cardiovascular disease (CVD) risk in HIV infection is an important area of investigation in the current era of aging with HIV infection. Less is known about CVD risk and HIV infection in developing nations where additional risk factors may be playing a role in the CVD development. In this study, we assessed the effects of systemic inflammation, microbial translocation (MT), T cell immune activation (IA), and nadir CD4 counts on cardiac function and arterial stiffness as markers of subclinical atherosclerosis in HIV-infected individuals. Methods People with HIV (PWH) who were ART naïve (n = 102) or virally suppressed on ART (n = 172) were stratified on nadir CD4 counts and compared to HIV-uninfected controls (n = 64). Determination was made of cardiac function via radial pulse wave and carotid intima thickness (C-IMT) measurements. Plasma biomarkers of inflammation and MT by ELISA or multiplex assays, and immune activation (IA) of T cells based HLA-DR and CD38 expression were investigated by flow cytometry. T-test, Mann–Whitney U test, and Spearman correlation were used to analyze study parameters. Results Reduction in cardiac function with lower cardiac ejection time (p

Mpox is an infectious disease caused by the monkeypox virus (MPXV) belonging to the Orthopoxvirus (OPXV) genus, which includes smallpox and vaccinia virus (VACV). A global mpox outbreak which began in May 2022 has infected more than 88,000 people. VACV-based vaccines provide protection against mpox disease but complicate the use of serological assays for disease surveillance. We tested the reactivity of serum IgG from Modified Vaccinia Ankara-Bavarian Nordic (MVA-BN)-vaccinated (n = 12) and convalescent mpox-infected (n = 5) individuals and uninfected, non-vaccinated controls (n = 32) to MPXV/VACV proteins A27, A29, A30, A35, B16, B21, C19, D6, E8, H3, I1, and L1. Using a subset of MPXV antigen-based assays (A35, B16, E8, H3, and I1), we conducted a mpox antibody survey of serum from 214 individuals, including 117 (54.7%) people with HIV (PWH) collected between June 2022 and January 2023, excluding individuals who reported recent mpox vaccination or infection, and 32 young, pre-pandemic controls. The convalescent sera reacted strongly to most tested antigens. Vaccine sera responses were limited to A35, E8, H3, and I1. IgG antibody to E8 was markedly elevated in all vaccinated individuals. B16 IgG showed high sensitivity (100% [95% CI: 56.55–100.0%]) and specificity (91.67% [64.61–99.57%]) for distinguishing infection from MVA-BN vaccination, while E8 IgG showed 100% [75.75–100] sensitivity and 100% [79.61–100] specificity for detecting and distinguishing vaccinated individuals from controls. We identified 11/214 (5.1%) recent serum samples and 1/32 (3.1%) young, pre-pandemic controls that were seropositive for ≥2 MPXV antibodies, including 6.8% of PWH. Seropositivity was 10/129 (7.8%) among males compared to 1/85 (1.2%) among females. Our findings provide insight into the humoral immune response to mpox and demonstrate the usefulness of inexpensive, antigen-based serosurveillance in identifying asymptomatic or unreported infections.

IntroductionThe influence of pre-existing humoral immunity, inter-individual demographic factors, and vaccine-associated reactogenicity on immunogenicity following COVID vaccination remains poorly understood.MethodsTen-fold cross-validated least absolute shrinkage and selection operator (LASSO) and linear mixed effects models were used to evaluate symptoms experienced by COVID+ participants during natural infection and following SARS-CoV-2 mRNA vaccination along with demographics as predictors for antibody (AB) responses to recombinant spike protein in a longitudinal cohort study.ResultsIn previously infected individuals (n=33), AB were more durable and robust following primary vaccination when compared to natural infection alone. Higher AB were associated with experiencing dyspnea during natural infection, as was the total number of symptoms reported during the COVID-19 disease course. Both local and systemic symptoms following 1st and 2nd dose (n=49 and 48, respectively) of SARS-CoV-2 mRNA vaccines were predictive of higher AB after vaccination. Lastly, there was a significant temporal relationship between AB and days since infection or vaccination, suggesting that vaccination in COVID+ individuals is associated with a more robust immune response.DiscussionExperiencing systemic and local symptoms post-vaccine was suggestive of higher AB, which may confer greater protection.

Abstract Background HIV infection induces inflammaging and chronic immune activation (IA), which are negatively associated with protective humoral immunity. Similar to HIV, aging is also associated with increased inflammaging and IA. The metabolic requirements of B cell responses in HIV infected (HIV+) individuals are not known, although metabolic abnormalities have been reported in these individuals. How these metabolic abnormalities are exacerbated by aging is also not known. Methods B cells were isolated by magnetic sorting from the blood of young and elderly HIV + individuals, as well as from the blood of age-matched healthy controls. We evaluated the composition of the B cell pool by flow cytometry, the expression of RNA for pro-inflammatory and metabolic markers by qPCR and their metabolic status using a Seahorse XFp extracellular flux analyzer. Results In this study we have evaluated for the first time the metabolic phenotype of B cells from young and elderly HIV + individuals as compared to those obtained from age-matched healthy controls. Results show that the B cell pool of HIV + individuals is enriched in pro-inflammatory B cell subsets, expresses higher levels of RNA for pro-inflammatory markers and is hyper-metabolic, as compared to healthy controls, and more in elderly versus young HIV + individuals, suggesting that this higher metabolic phenotype of B cells is needed to support B cell IA. We have identified the subset of Double Negative (DN) B cells as the subset mainly responsible for this hyper-inflammatory and hyper-metabolic profile. Conclusions Our results identify a relationship between intrinsic B cell inflammation and metabolism in HIV + individuals and suggest that metabolic pathways in B cells from HIV + individuals may be targeted to reduce inflammaging and IA and improve B cell function and antibody responses.

ABSTRACT To understand reinfection rates and correlates of protection for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), we established eight different longitudinal cohorts in 2020 under the umbrella of the PARIS (Protection Associated with Rapid Immunity to SARS-CoV-2)/SPARTA (SARS SeroPrevalence And Respiratory Tract Assessment) studies. Here, we describe the PARIS/SPARTA cohorts, the harmonized assays and analysis that are performed across the cohorts, as well as case definitions for SARS-CoV-2 infection and reinfection that have been established by the team of PARIS/SPARTA investigators. IMPORTANCE Determining reinfection rates and correlates of protection against SARS-CoV-2 infection induced by both natural infection and vaccination is of high significance for the prevention and control of coronavirus disease 2019 (COVID-19). Furthermore, understanding reinfections or infection after vaccination and the role immune escape plays in these scenarios will inform the need for updates of the current SARS-CoV-2 vaccines and help update guidelines suitable for the postpandemic world.

BackgroundDespite a successful antiretroviral therapy (ART), adolescents living with perinatally acquired HIV (PHIV) experience signs of B-cell hyperactivation with expansion of ‘namely’ atypical B-cell phenotypes, including double negative (CD27-IgD-) and termed age associated (ABCs) B-cells (T-bet+CD11c+), which may result in reduced cell functionality, including loss of vaccine-induced immunological memory and higher risk of developing B-cells associated tumors. In this context, perinatally HIV infected children (PHIV) deserve particular attention, given their life-long exposure to chronic immune activation.MethodsWe studied 40 PHIV who started treatment by the 2nd year of life and maintained virological suppression for 13.5 years, with 5/40 patients experiencing transient elevation of the HIV-1 load in the plasma (Spike). We applied a multi-disciplinary approach including immunological B and T cell phenotype, plasma proteomics analysis, and serum level of anti-measles antibodies as functional correlates of vaccine-induced immunity.ResultsPhenotypic signs of B cell hyperactivation were elevated in subjects starting ART later (%DN T-bet+CD11c+ p=0.03; %AM T-bet+CD11c+ p=0.02) and were associated with detectable cell-associated HIV-1 RNA (%AM T-bet+CD11c+ p=0.0003) and transient elevation of the plasma viral load (spike). Furthermore, B-cell hyperactivation appeared to be present in individuals with higher frequency of exhausted T-cells, in particular: %CD4 TIGIT+ were associated with %DN (p=0.008), %DN T-bet+CD11c+ (p=0.0002) and %AM T-bet+CD11c+ (p=0.002) and %CD4 PD-1 were associated with %DN (p=0.048), %DN T-bet+CD11c+ (p=0.039) and %AM T-bet+CD11c+ (p=0.006). The proteomic analysis revealed that subjects with expansion of these atypical B-cells and exhausted T-cells had enrichment of proteins involved in immune inflammation and complement activation pathways. Furthermore, we observed that higher levels of ABCs were associated a reduced capacity to maintain vaccine-induced antibody immunity against measles (%B-cells CD19+CD10- T-bet+, p=0.035).ConclusionWe identified that the levels of hyperactivated B cell subsets were strongly affected by time of ART start and associated with clinical, viral, cellular and plasma soluble markers. Furthermore, the expansion of ABCs also had a direct impact on the capacity to develop antibodies response following routine vaccination.

Introduction: HIV-positive (HIV+) kidney transplant recipients exhibit a 2- to 3-fold increased risk of allograft rejection. Dysregulated immune activation in HIV infection persists despite successful antiretroviral therapy and is associated with non-AIDS morbidity, including renal disease. We hypothesized that the pathological levels of inflammation and immune activation associated with chronic HIV infection could have clinical utility in the prediction of rejection in HIV+ kidney recipients. Methods: Prospective cohort study of 22 HIV-negative (HIV−; donor) to HIV+ (recipient) kidney transplant recipients who underwent biomarker assessment pretransplant and were subsequently followed for development of acute rejection. Plasma levels of markers of inflammation (soluble tumor necrosis factor receptor 1 [sTNF-R1] and C-reactive protein [CRP]) and microbial translocation (soluble CD14 and lipopolysaccharide) were measured by enzyme-linked immunosorbent assay or chromogenic endpoint assay. Levels of activated (CD38+HLADR+) CD4+ and CD8+ T cells, and T regulatory cells (CD4+CD25highFoxP3+) were measured by flow cytometry. Results: Among the biomarkers evaluated, only the pretransplant levels of sTNF-R1, CRP, and frequencies of CD38+HLADR+ CD8 T cells, were found to be at significantly higher levels among patients who experienced biopsy-proven acute rejection. Confirming our hypothesis, patients with high pretransplant levels of sTNF-R1 or activated CD8+ T cells had a significantly increased 200-day cumulative incidence of biopsy-proven acute rejection (0 vs. 38% for both; P = 0.01). Similarly, pretransplant CRP levels higher than 5 μg/ml were associated with increased risk of acute rejection within the first 6 months post-transplant (0 vs. 43%; P = 0.01). Conclusion: Biomarker-based identification of HIV+ recipients at increased risk for rejection might facilitate individualized induction immunosuppression regimens in this vulnerable patient population. Keywords: C-reactive protein, HIV, kidney transplant, rejection, sTNF-R1, T-cell activation

Natural aging and HIV infection are associated with chronic low-grade systemic inflammation, immune senescence, and impaired antibody responses to vaccines such as the influenza (flu) vaccine. We investigated the role of IL-21, a CD4+ T follicular helper cell (Tfh) regulator, on flu vaccine antibody response in nonhuman primates (NHPs) in the context of age and controlled SIV mac239 infection. Three doses of the flu vaccine with or without IL-21–IgFc were administered at 3-month intervals in aged SIV+ NHPs following virus suppression with antiretroviral therapy. IL-21–treated animals demonstrated higher day 14–postboost antibody responses, which associated with expanded CD4+ T central memory cells and peripheral Tfh–expressing (pTfh–expressing) T cell immunoreceptor with Ig and ITIM domains (TIGIT), expanded activated memory B cells, and contracted CD11b+ monocytes. Draining lymph node (LN) tissue from IL-21–treated animals revealed direct association between LN follicle Tfh density and frequency of circulating TIGIT+ pTfh cells. We conclude that IL-21 enhances flu vaccine–induced antibody responses in SIV+ aged rhesus macaques (RMs), acting as an adjuvant modulating LN germinal center activity. A strategy to supplement IL-21 in aging could be a valuable addition in the toolbox for improving vaccine responses in an aging HIV+ population.

Aging is associated with declining immunity and inflammation as well as alterations in the gut microbiome with a decrease of beneficial microbes and increase in pathogenic ones. The aim of this study was to investigate the age associated gut microbiome in relation to immunologic and metabolic profile in a non-human primate (NHP) model. 12 geriatric (age 19-24 years) and 4 young adult (age 3-4 years) Rhesus macaques were included in this study. Immune cell subsets were characterized in peripheral blood mononuclear cells (PBMC) by flow cytometry and plasma cytokines levels were determined by bead based multiplex cytokine analysis. Stool samples were collected by ileal loop and investigated for microbiome analysis by shotgun metagenomics. Serum, gut microbial lysate, and microbe-free fecal extract were subjected to metabolomic analysis by mass-spectrometry. Our results showed that the gut microbiome in geriatric animals had higher abundance of Archaeal and Proteobacterial species and lower Firmicutes than the young adults. Highly abundant microbes in the geriatric animals showed a direct association with plasma biomarkers of inflammation and immune activation such as neopterin, CRP, TNF, IL-2, IL-6, IL-8 and IFN-γ. Significant enrichment of metabolites that contribute to inflammatory and cytotoxic pathways was observed in serum and feces of geriatric animals compared to the young adults. We conclude that aging NHP undergo immunosenescence and age associated alterations in the gut microbiome that has a distinct metabolic profile. Aging NHP can serve as a model for investigating the relationship of the gut microbiome to particular age-associated comorbidities and for strategies aimed at modulating the microbiome.

Early initiation of antiretroviral therapy and adherence to achieve viral load suppression (VLS) are crucial for reducing morbidity and mortality of perinatally HIV-infected infants. In this descriptive cohort study of 39 HIV perinatally infected infants, who started treatment at one month of life in Mozambique, we aimed to describe the viral response over 2 years of follow up. VLS ≤ 400 copies/mL, sustained VLS and viral rebound were described using a Kaplan–Meier estimator. Antiretroviral drug transmitted resistance was assessed for a sub-group of non-VLS infants. In total, 61% of infants reached VLS, and 50% had a rebound. Cumulative probability of VLS was 36%, 51%, and 69% at 6, 12 and 24 months of treatment, respectively. The median duration of VLS was 7.4 months (IQR 12.6) and the cumulative probability of rebound at 6 months was 30%. Two infants had resistance biomarkers to drugs included in their treatment regimen. Our findings point to a low rate of VLS and high rate of viral rebound. More frequent viral response monitoring is advisable to identify infants with rebound and offer timely adherence support. It is urgent to tailor the psychosocial support model of care to this specific age group and offer differentiated service delivery to mother–baby pairs.

HIV eradication is hindered by the existence of latent HIV reservoirs in CD4+ T cells. Therapeutic strategies targeting latent cells are required to achieve a functional cure, however the study of latently infected cells from HIV infected persons is extremely challenging due to the lack of biomarkers that uniquely characterize them. In this study, the dual reporter virus HIVGKO was used to investigate latency establishment and maintenance in lymphoid-derived CD4+ T cells. Single cell technologies to evaluate protein expression, host gene expression, and HIV transcript expression were integrated to identify and analyze latently infected cells. FDA-approved, JAK1/2 inhibitors were tested in this system as a potential therapeutic strategy to target the latent reservoir. Latent and productively infected tonsillar CD4+ T cells displayed similar activation profiles as measured by expression of CD69, CD25, and HLADR, however latent cells showed higher CXCR5 expression 3 days post-infection. Single cell analysis revealed a small set of genes, including HIST1-related genes and the inflammatory cytokine, IL32, that were upregulated in latent compared to uninfected and productively infected cells suggesting a role for these molecular pathways in persistent HIV infection. In vitro treatment of HIV-infected CD4+ T cells with physiological concentrations of JAK1/2 inhibitors, ruxolitinib and baricitinib, used in clinical settings to target inflammation, reduced latent and productive infection events when added 24 hr after infection and blocked HIV reactivation from latent cells. Our methods using an established model of HIV latency and lymphoid-derived cells shed light on the biology of latency in a crucial anatomical site for HIV persistence and provides key insights about repurposing baricitinib or ruxolitinib to target the HIV reservoir.

RTS,S/AS01 (GSK) is the world’s first malaria vaccine. However, despite initial efficacy of almost 70% over the first 6 months of follow-up, efficacy waned over time. A deeper understanding of the immune features that contribute to RTS,S/AS01-mediated protection could be beneficial for further vaccine development. In two recent controlled human malaria infection (CHMI) trials of the RTS,S/AS01 vaccine in malaria-naïve adults, MAL068 and MAL071, vaccine efficacy against patent parasitemia ranged from 44% to 87% across studies and arms (each study included a standard RTS,S/AS01 arm with three vaccine doses delivered in four-week-intervals, as well as an alternative arm with a modified version of this regimen). In each trial, RTS,S/AS01 immunogenicity was interrogated using a broad range of immunological assays, assessing cellular and humoral immune parameters as well as gene expression. Here, we used a predictive modeling framework to identify immune biomarkers measured at day-of-challenge that could predict sterile protection against malaria infection. Using cross-validation on MAL068 data (either the standard RTS,S/AS01 arm alone, or across both the standard RTS,S/AS01 arm and the alternative arm), top-performing univariate models identified variables related to Fc effector functions and titer of antibodies that bind to the central repeat region (NANP6) of CSP as the most predictive variables; all NANP6-related variables consistently associated with protection. In cross-study prediction analyses of MAL071 outcomes (the standard RTS,S/AS01 arm), top-performing univariate models again identified variables related to Fc effector functions of NANP6-targeting antibodies as highly predictive. We found little benefit–with this dataset–in terms of improved prediction accuracy in bivariate models vs. univariate models. These findings await validation in children living in malaria-endemic regions, and in vaccinees administered a fourth RTS,S/AS01 dose. Our findings support a “quality as well as quantity” hypothesis for RTS,S/AS01-elicited antibodies against NANP6, implying that malaria vaccine clinical trials should assess both titer and Fc effector functions of anti-NANP6 antibodies.

The size of the latent HIV reservoir is associated with the timing of therapeutic interventions and overall health of the immune system. Here, we demonstrate that T cell phenotypic signatures associate with viral reservoir size in a cohort of HIV vertically infected children and young adults under durable viral control, and who initiated anti-retroviral therapy (ART)

The number of patients affected by chronic diseases with special vaccination needs is burgeoning. In this scenario, predictive markers of immunogenicity, as well as signatures of immune responses are typically missing even though it would especially improve the identification of personalized immunization practices in these populations. We aimed to develop a predictive score of immunogenicity to Influenza Trivalent Inactivated Vaccination (TIV) by applying deep machine learning algorithms using transcriptional data from sort-purified lymphocyte subsets after in vitro stimulation. Peripheral blood mononuclear cells (PBMCs) collected before TIV from 23 vertically HIV infected children under ART and virally controlled were stimulated in vitro with p09/H1N1 peptides (stim) or left unstimulated (med). A multiplexed-qPCR for 96 genes was made on fixed numbers of 3 B cell subsets, 3 T cell subsets and total PBMCs. The ability to respond to TIV was assessed through hemagglutination Inhibition Assay (HIV) and ELIspot and patients were classified as Responders (R) and Non Responders (NR). A predictive modeling framework was applied to the data set in order to define genes and conditions with the higher predicted probability able to inform the final score. Twelve NR and 11 R were analyzed for gene expression differences in all subsets and 3 conditions [med, stim or Δ (stim-med)]. Differentially expressed genes between R and NR were selected and tested with the Adaptive Boosting Model to build a prediction score. The score obtained from subsets revealed the best prediction score from 46 genes from 5 different subsets and conditions. Calculating a combined score based on these 5 categories, we achieved a model accuracy of 95.6% and only one misclassified patient. These data show how a predictive bioinformatic model applied to transcriptional analysis deriving from in-vitro stimulated lymphocytes subsets may predict poor or protective vaccination immune response in vulnerable populations, such as HIV-infected individuals. Future studies on larger cohorts are needed to validate such strategy in the context of vaccination trials.

With advancement, prompt use, and increasing accessibility of antiretroviral therapy, people with HIV are living longer and have comparable lifespans to those negative for HIV. However, people living with HIV experience tradeoffs with quality of life often developing age-associated co-morbid conditions such as cancers, cardiovascular diseases, or neurodegeneration due to chronic immune activation and inflammation. This creates a discrepancy in chronological and physiological age, with HIV-infected individuals appearing older than they are, and in some contexts ART-associated toxicity exacerbates this gap. The complexity of the accelerated aging process in the context of HIV-infection highlights the need for greater understanding of biomarkers involved. In this review, we discuss markers identified in different anatomical sites of the body including periphery, brain, and gut, as well as markers related to DNA that may serve as reliable predictors of accelerated aging in HIV infected individuals as it relates to inflammatory state and immune activation.

Malaria-071, a controlled human malaria infection trial, demonstrated that administration of three doses of RTS,S/AS01 malaria vaccine given at one-month intervals was inferior to a delayed fractional dose (DFD) schedule (62.5% vs 86.7% protection, respectively). To investigate the underlying immunologic mechanism, we analyzed the B and T peripheral follicular helper cell (pTfh) responses. Here, we show that protection in both study arms was associated with early induction of functional IL-21-secreting circumsporozoite (CSP)-specific pTfh cells, together with induction of CSP-specific memory B cell responses after the second dose that persisted after the third dose. Data integration of key immunologic measures identified a subset of non-protected individuals in the standard (STD) vaccine arm who lost prior protective B cell responses after receiving the third vaccine dose. We conclude that the DFD regimen favors persistence of functional B cells after the third dose.

Chronic infection with human immunodeficiency virus (HIV) and hepatitis C virus (HCV) affects an estimated 35 million and 75 million individuals worldwide, respectively. These viruses induce persistent inflammation which often drives the development or progression of organ-specific diseases and even cancer including Hepatocellular Carcinoma (HCC). In this study, we sought to examine inflammatory responses following HIV or HCV stimulation of macrophages or Kupffer cells (KCs), that may contribute to virus mediated inflammation and subsequent liver disease. KCs are liver-resident macrophages and reports have provided evidence that HIV can stimulate and infect them. In order to characterize HIV-intrinsic innate immune responses that may occur in the liver, we performed microarray analyses on KCs following HIV stimulation. Our data demonstrate that KCs upregulate several innate immune signaling pathways involved in inflammation, myeloid cell maturation, stellate cell activation, and Triggering Receptor Expressed on Myeloid cells 1 (TREM1) signaling. TREM1 is a member of the immunoglobulin superfamily of receptors and it is reported to be involved in systemic inflammatory responses due to its ability to amplify activation of host defense signaling pathways. Our data demonstrate that stimulation of KCs with HIV or HCV induces the upregulation of TREM1. Additionally, HIV viral proteins can upregulate expression of TREM1 mRNA through NF-кB signaling. Furthermore, activation of the TREM1 signaling pathway, with a targeted agonist, increased HIV or HCV-mediated inflammatory responses in macrophages due to enhanced activation of the ERK1/2 signaling cascade. Silencing TREM1 dampened inflammatory immune responses elicited by HIV or HCV stimulation. Finally, HIV and HCV infected patients exhibit higher expression and frequency of TREM1 and CD68 positive cells. Taken together, TREM1 induction by HIV contributes to chronic inflammation in the liver and targeting TREM1 signaling may be a therapeutic option to minimize HIV induced chronic inflammation.

Antigen-primed cluster of differentiation (CD) 4+ T follicular helper (Tfh) cells interact with B cells in the germinal centers (GCs) of lymph nodes to generate vaccine-induced antibody (Ab) responses. In the circulation, peripheral Tfh (pTfh) cells, a subset of memory CD4 T cells, serve as surrogates for GC Tfh because of several functional and phenotypic similarities between them. We investigated features of H1N1 influenza antigen-specific pTfh (Ag.pTfh) in virologically controlled HIV+ volunteers on antiretroviral therapy (ART) and healthy control (HC) participants selected from a seasonal influenza vaccine responsiveness study. Selection of the participants was made based on age, defined as young (18-40 y) and old (>60 y) and on their classification as a vaccine responder (VR) or vaccine nonresponder (VNR). VRs demonstrated expansion of CD40L+ and CD69+ Ag.pTfh, with induction of intracellular interleukin 21 (IL-21) and inducible costimulator (ICOS) post vaccination; these responses were strongest in young HC VRs and were less prominent in HIV+ individuals of all ages. Ag.pTfh in VNRs exhibited dramatically different characteristics from VRs, displaying an altered phenotype and a cytokine profile dominated by cytokines IL-2, tumor necrosis factor alpha (TNF-α), or IL-17 but lacking in IL-21. In coculture experiments, sorted pTfh did not support the B cell IgG production in VNRs and were predominantly an inflammatory T helper 1 (Th1)/T helper 17 (Th17) phenotype with lower ICOS and higher programmed cell death protein 1 (PD1) expression. Induction of IL-21 and ICOS on Ag.pTfh cells are negatively affected by both aging and HIV infection. Our findings demonstrate that dysfunctional Ag.pTfh cells with an altered IL-21/IL-2 axis contribute to inadequate vaccine responses. Approaches for targeting inflammation or expanding functional Tfh may improve vaccine responses in healthy aging and those aging with HIV infection.

Recent findings point to a role of Checkpoint Inhibitor (CPI) receptors at the tissue level in immune homeostasis. Here we investigated the role of CPI molecules on immune cells in relation to cardiac function. Participants recruited in Chennai, India consisted of HIV+ ART naive viremic (Gp1 n = 102), HIV+ on ART, virologically suppressed (Gp2, n = 172) and HIV negative healthy controls (Gp3, n = 64). A cross-sectional analysis of cardiac function, arterial resistance and immunologic assessment of CPI expressing T cells was performed. Data indicate that ART naive exhibited cardiac function impairment and greater arterial stiffness than the other groups. Frequencies of CD4+ T cells expressing LAG-3 and PD1 were higher in ART naïve while TIGIT and TIM3 were similar among the patient groups. LAG-3+, PD1+ and dual LAG-3+PD1+ CD4 T cells were inversely correlated with cardiac function and arterial elasticity and directly with arterial stiffness in ART naïve participants and with arterial elasticity in virally suppressed group on ART. We conclude that HIV induced upregulation of LAG-3 singly or in combination with PD1 in immune cells may regulate cardiac health and warrant mechanistic investigations. The implications of these findings have bearing for the potential utility of anti-LAG-3 immunotherapy for cardiac dysfunction in chronic HIV infection.

Modern technologies and their increased accessibility have shifted ‘benchtop’ medical research to the larger dimension of ‘omics. The huge amount of data derived from gene expression and sequencing experiments has propelled physicians, basic scientists and bioinformaticians towards a common goal to transform ‘big data’ into predictive constructs that are readily available and will offer clinical utility. Although most of the studies available in the literature have been performed on healthy subjects and in peripheral blood mononuclear cells (PBMC), which are a heterogenous and extremely variable pool of cells, scientists are now trying to address mechanistic questions in purified cell subsets in pathological conditions. In the field of HIV, few attempts have been made to comprehensively evaluate gene-expression profiles of infected patients with different disease status. With the view of discovering a path towards remission or viral eradication, perinatally HIV-infected children represent a unique model. In fact the well-defined time of infection and the resulting opportunity to start early treatment, thereby generating a smaller size of viral reservoir and a more intact immune system, allow for investigation of therapeutic strategies to defeat the virus. In this scenario, ‘transcriptomic’ or gene expression technologies and supporting bioinformatics applications need to be strategically integrated to provide novel information about immune correlates of virus control following treatment interruption. Here we review modern techniques for gene expression analysis and discuss the best transcriptomic strategies applicable to the field of functional cure in paediatric HIV infection.

T follicular helper (Tfh) cells are a subset of CD4 T cells that provide critical signals to antigen-primed B cells in germinal centers to undergo proliferation, isotype switching, and somatic hypermutation to generate long-lived plasma cells and memory B cells during an immune response. The quantity and quality of Tfh cells therefore must be tightly controlled to prevent immune dysfunction in the form of autoimmunity and, on the other hand, immune deficiency. Both Tfh and B cell perturbations appear during HIV infection resulting in impaired antibody responses to vaccines such as seasonal trivalent influenza vaccine, also seen in biologic aging. Although many of the HIV-associated defects improve with antiretroviral therapy (ART), excess immune activation and antigen-specific B and T cell responses including Tfh function are still impaired in virologically controlled HIV-infected persons on ART. Interestingly, HIV infected individuals experience increased risk of age-associated pathologies. This review will discuss Tfh and B cell dysfunction in HIV infection and highlight the impact of chronic HIV infection and aging on Tfh–B cell interactions.

Despite effective antiretroviral therapy (ART), HIV-infected individuals with apparently similar clinical and immunological characteristics can vary in responsiveness to vaccinations. However, molecular mechanisms responsible for such impairment, as well as biomarkers able to predict vaccine responsiveness in HIV-infected children, remain unknown. Following the hypothesis that a B cell qualitative impairment persists in HIV-infected children (HIV) despite effective ART and phenotypic B cell immune reconstitution, the aim of the current study was to investigate B cell gene expression of HIV compared to age-matched healthy controls (HCs) and to determine whether distinct gene expression patterns could predict the ability to respond to influenza vaccine. To do so, we analyzed prevaccination transcriptional levels of a 96-gene panel in equal numbers of sort-purified B cell subsets (SPBS) isolated from peripheral blood mononuclear cells using multiplexed RT-PCR. Immune responses to H1N1 antigen were determined by hemaglutination inhibition and memory B cell ELISpot assays following trivalent-inactivated influenza vaccination (TIV) for all study participants. Although there were no differences in terms of cell frequencies of SPBS between HIV and HC, the groups were distinguishable based upon gene expression analyses. Indeed, a 28-gene signature, characterized by higher expression of genes involved in the inflammatory response and immune activation was observed in activated memory B cells (CD27+CD21−) from HIV when compared to HC despite long-term viral control (>24 months). Further analysis, taking into account H1N1 responses after TIV in HIV participants, revealed that a 25-gene signature in resting memory (RM) B cells (CD27+CD21+) was able to distinguish vaccine responders from non-responders (NR). In fact, prevaccination RM B cells of responders showed a higher expression of gene sets involved in B cell adaptive immune responses (APRIL, BTK, BLIMP1) and BCR signaling (MTOR, FYN, CD86) when compared to NR. Overall, these data suggest that a perturbation at a transcriptional level in the B cell compartment persists despite stable virus control achieved through ART in HIV-infected children. Additionally, the present study demonstrates the potential utility of transcriptional evaluation of RM B cells before vaccination for identifying predictive correlates of vaccine responses in this population.

In pathogenic HIV and SIV infections of humans and rhesus macaques (RMs), preferential depletion of CD4⁺ Th17 cells correlates with mucosal immune dysfunction and disease progression. Interleukin (IL)-21 promotes differentiation of Th17 cells, long-term maintenance of functional CD8⁺ T cells, and differentiation of memory B cells and antibody-secreting plasma cells. We hypothesized that administration of IL-21 will improve mucosal function in the context of pathogenic HIV/SIV infections. To test this hypothesis, we infected 12 RMs with SIV(mac239) and at day 14 post-infection treated six of them with rhesus rIL-21-IgFc. IL-21-treatment was safe and did not increase plasma viral load or systemic immune activation. Compared to untreated animals, IL-21-treated RMs showed (i) higher expression of perforin and granzyme B in total and SIV-specific CD8⁺ T cells and (ii) higher levels of intestinal Th17 cells. Remarkably, increased levels of Th17 cells were associated with reduced levels of intestinal T cell proliferation, microbial translocation and systemic activation/inflammation in the chronic infection. In conclusion, IL-21-treatment in SIV-infected RMs improved mucosal immune function through enhanced preservation of Th17 cells. Further preclinical studies of IL-21 may be warranted to test its potential use during chronic infection in conjunction with antiretroviral therapy.

Persistent immune activation and microbial translocation associated with HIV infection likely place HIV-infected aging women at high risk of developing chronic age-related diseases. We investigated immune activation and microbial translocation in HIV-infected aging women in the post-menopausal ages.Twenty-seven post-menopausal women with HIV infection receiving antiretroviral treatment with documented viral suppression and 15 HIV-negative age-matched controls were enrolled. Levels of immune activation markers (T cell immune phenotype, sCD25, sCD14, sCD163), microbial translocation (LPS) and biomarkers of cardiovascular disease and impaired cognitive function (sVCAM-1, sICAM-1 and CXCL10) were evaluated.T cell activation and exhaustion, monocyte/macrophage activation, and microbial translocation were significantly higher in HIV-infected women when compared to uninfected controls. Microbial translocation correlated with T cell and monocyte/macrophage activation. Biomarkers of cardiovascular disease and impaired cognition were elevated in women with HIV infection and correlated with immune activation.HIV-infected antiretroviral-treated aging women who achieved viral suppression are in a generalized status of immune activation and therefore are at an increased risk of age-associated end-organ diseases compared to uninfected age-matched controls.

Aging and HIV infection are independently associated with excessive immune activation and impaired immune responses to vaccines, but their relationships have not been examined.For selecting an aging population we enrolled 28 post-menopausal women including 12 healthy volunteers and 16 HIV-infected women on antiretroviral treatment with