The precise effects of HIV-1 on the gut microbiome are unclear. Initial cross-sectional studies provided contradictory associations between microbial richness and HIV serostatus and suggested shifts from Bacteroides to Prevotella predominance following HIV-1 infection, which have not been found in animal models or in studies matched for HIV-1 transmission groups. In two independent cohorts of HIV-1-infected subjects and HIV-1-negative controls in Barcelona (n = 156) and Stockholm (n = 84), men who have sex with men (MSM) predominantly belonged to the Prevotella-rich enterotype whereas most non-MSM subjects were enriched in Bacteroides, independently of HIV-1 status, and with only a limited contribution of diet effects. Moreover, MSM had a significantly richer and more diverse fecal microbiota than non-MSM individuals. After stratifying for sexual orientation, there was no solid evidence of an HIV-specific dysbiosis. However, HIV-1 infection remained consistently associated with reduced bacterial richness, the lowest bacterial richness being observed in subjects with a virological-immune discordant response to antiretroviral therapy. Our findings indicate that HIV gut microbiome studies must control for HIV risk factors and suggest interventions on gut bacterial richness as possible novel avenues to improve HIV-1-associated immune dysfunction.

HIV-1 infection is characterized by altered intestinal barrier, gut microbiota dysbiosis, and systemic inflammation. We hypothesized that changes of the gut microbiota predict immune dysfunction and HIV-1 progression, and that antiretroviral therapy (ART) partially restores the microbiota composition. An observational study including 28 viremic patients, three elite controllers, and nine uninfected controls. Blood and stool samples were collected at baseline and for 19 individuals at follow-up (median 10 months) during ART. Microbiota composition was determined by 16S rRNA sequencing (Illumina MiSeq). Soluble markers of microbial translocation and monocyte activation were analyzed by Limulus Amebocyte Lysate assay or ELISA. Several alpha-diversity measures, including number of observed bacterial species and Shannon index, were significantly lower in viremic patients compared to controls. The alpha diversity correlated with CD4 T-cell counts and inversely with markers of microbial translocation and monocyte activation. In multivariate linear regression, for every age and sex-adjusted increase in the number of bacterial species, the CD4 T-cell count increased with 0.88 (95% confidence interval 0.35-1.41) cells/μl (P = 0.002). After introduction of ART, microbiota alterations persisted with further reduction in alpha diversity. The microbiota composition at the genus level was profoundly altered in viremic patients, both at baseline and after ART, with Prevotella reduced during ART (P < 0.007). Gut microbiota alterations are closely associated with immune dysfunction in HIV-1 patients, and these changes persist during short-term ART. Our data implicate that re-shaping the microbiota may be an adjuvant therapy in patients commencing successful ART.

CD8⁺ T cell exhaustion is a hallmark of many cancers and chronic infections. In mice, T cell factor 1 (TCF-1) maintains exhausted CD8⁺ T cell responses, whereas thymocyte selection-associated HMG box (TOX) is required for the epigenetic remodeling and survival of exhausted CD8⁺ T cells. However, it has remained unclear to what extent these transcription factors play analogous roles in humans. In this study, we mapped the expression of TOX and TCF-1 as a function of differentiation and specificity in the human CD8⁺ T cell landscape. Here, we demonstrate that circulating TOX⁺ CD8⁺ T cells exist in most humans, but that TOX is not exclusively associated with exhaustion. Effector memory CD8⁺ T cells generally expressed TOX, whereas naive and early-differentiated memory CD8⁺ T cells generally expressed TCF-1. Cytolytic gene and protein expression signatures were also defined by the expression of TOX. In the context of a relentless immune challenge, exhausted HIV-specific CD8⁺ T cells commonly expressed TOX, often in clusters with various activation markers and inhibitory receptors, and expressed less TCF-1. However, polyfunctional memory CD8⁺ T cells specific for cytomegalovirus (CMV) or Epstein-Barr virus (EBV) also expressed TOX, either with or without TCF-1. A similar phenotype was observed among HIV-specific CD8⁺ T cells from individuals who maintained exceptional immune control of viral replication. Collectively, these data demonstrate that TOX is expressed by most circulating effector memory CD8⁺ T cell subsets and not exclusively linked to exhaustion.

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Mucosa-associated invariant T (MAIT) cells represent a large innate-like evolutionarily conserved antimicrobial T-cell subset in humans. MAIT cells recognize microbial riboflavin metabolites from a range of microbes presented by MR1 molecules. MAIT cells are impaired in several chronic diseases including HIV-1 infection, where they show signs of exhaustion and decline numerically. Here, we examined the broader effector functions of MAIT cells in this context and strategies to rescue their functions. Residual MAIT cells from HIV-infected patients displayed aberrant baseline levels of cytolytic proteins, and failed to mobilize cytolytic molecules in response to bacterial antigen. In particular, the induction of granzyme B (GrzB) expression was profoundly defective. The functionally impaired MAIT cell population exhibited abnormal T-bet and Eomes expression patterns that correlated with the deficiency in cytotoxic capacity and cytokine production. Effective antiretroviral therapy (ART) did not fully restore these aberrations. Interestingly, IL-7 was capable of arming resting MAIT cells from healthy donors into cytotoxic GrzB+ effector T cells capable of killing bacteria-infected cells and producing high levels of pro-inflammatory cytokines in an MR1-dependent fashion. Furthermore, IL-7 treatment enhanced the sensitivity of MAIT cells to detect low levels of bacteria. In HIV-infected patients, plasma IL-7 levels were positively correlated with MAIT cell numbers and function, and IL-7 treatment in vitro significantly restored MAIT cell effector functions even in the absence of ART. These results indicate that the cytolytic capacity in MAIT cells is severely defective in HIV-1 infected patients, and that the broad-based functional defect in these cells is associated with deficiency in critical transcription factors. Furthermore, IL-7 induces the arming of effector functions and enhances the sensitivity of MAIT cells, and may be considered in immunotherapeutic approaches to restore MAIT cells. The mucosa-associated invariant T (MAIT) cells recognize antigens that are byproducts of the riboflavin biosynthesis pathway shared by many microbes. These antigens are presented by the MHC class I-like MR1 molecules and trigger rapid activation of MAIT cells in an innate-like fashion with deployment of effector mechanisms including cytokine production and cytolysis. Here, we investigated the MAIT cell response to bacteria in humans infected with HIV-1, and possible means to restore functionality to these cells. MAIT cell dysfunction in HIV-infected patients included an inability to express components of the cytolytic effector machinery. Impairment of the MAIT cell population involved the loss of expression of the transcription factors T-bet and Eomes. Interestingly, IL-7 had strong effects on MAIT cells, including the antigen-independent arming of cytolytic function and enhanced sensitivity for low levels of bacteria. In HIV-infected patients, plasma IL-7 levels were positively associated with the size of the MAIT cell population, and IL-7 could rescue their function. These findings indicate that MAIT cell impairment in HIV-1 infection is broad-based, includes loss of critical transcription factors, and loss of cytolytic function. Furthermore, the data support the notion that IL-7 is a strong candidate for immunotherapy in diseases associated with MAIT cell loss.

Gut microbiota dysbiosis features progressive HIV infection and is a potential target for intervention. Herein, we explored the microbiome of 16 elite controllers (EC), 32 antiretroviral therapy naive progressors and 16 HIV negative controls. We found that the number of observed genera and richness indices in fecal microbiota were significantly higher in EC versus naive. Genera Succinivibrio, Sutterella, Rhizobium, Delftia, Anaerofilum and Oscillospira were more abundant in EC, whereas Blautia and Anaerostipes were depleted. Additionally, carbohydrate metabolism and secondary bile acid synthesis pathway related genes were less represented in EC. Conversely, fatty acid metabolism, PPAR-signalling and lipid biosynthesis proteins pathways were enriched in EC vs naive. The kynurenine pathway of tryptophan metabolism was altered during progressive HIV infection, and inversely associated with microbiota richness. In conclusion, EC have richer gut microbiota than untreated HIV patients, with unique bacterial signatures and a distinct metabolic profile which may contribute to control of HIV.