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Background Rectal douching is the practice of rinsing the anus and rectum prior to, or after, anal sex. Among gay, bisexual, and other men who have sex with men (sexual minority men), rectal douching is likely to enhance HIV acquisition by amplifying inflammation of the rectal mucosa. This study evaluated the association of rectal douching with rectal inflammation among sexual minority men at risk for HIV infection. Methods HIV uninfected sexual minority men who reported receptive anal intercourse in the past 3 months (N = 197) were recruited from four STI clinics in South Florida, the leading region for new HIV infections among sexual minority men in the United States. A brief survey assessed rectal douching practices and sexual behaviors in the previous 3 months. Rectal inflammation was assessed by measuring 13 human rectal inflammatory cytokines/chemokines in rectal swabs using the LEGENDplex Human Inflammation Panel in a sample of 38 participants who reported douching and condomless receptive anal intercourse (CRAI). Results Average age was 35.8 years (SD=15.4). The sample was multi-ethnic: 42% Caucasian, 38% Hispanic/Latino, 13% Black/African American, and 7% other ethnic minority. Approximately two-thirds of participants (65%) reported any rectal douching in the past 3 months and the median number times participants douched was five. Participants who douched had more CRAI partners (Cohen's d = 0.4587; P < 0.01), and more CRAI partners to ejaculation (Cohen's d = 0.4813; P < 0.01) compared with participants who did not douche. Participants who reported douching five or more times in the past 3 months displayed significantly higher levels of IL-8 (Cohen's d = 0.79; P = 0.02) than those who douched less than five times. Conclusion Among sexual minority men who engage in CRAI, more frequent douching is associated with higher levels of rectal inflammation. Assessment of rectal douching should be included when evaluating HIV prevention interventions among sexual minority men, as those who douche are more likely to engage in CRAI and are at higher risk for acquisition of HIV and other sexually transmitted infection. Further mechanistic studies are needed to assess the role of rectal douching in promoting rectal inflammation. Disclosures All authors: No reported disclosures. [ABSTRACT FROM AUTHOR]

Antibiotic therapies are known to disrupt gastrointestinal (GI) bacterial communities. HIV and pathogenic simian immunodeficiency virus (SIV) infections have also been associated with disrupted GI bacterial communities. We administered a combination antibiotic therapy to six SIV-infected rhesus macaques and collected colon biopsies, stool samples and rectal swabs before and after antibiotics, and evaluated the bacterial communities at each sample site using high-throughput 16S rRNA gene sequencing. The colon mucosa and stool samples displayed different bacterial communities, while the rectal swabs showed a mixture of the mucosal and stool-associated bacteria. Antibiotics disrupted the native bacterial communities at each sample site. The colon mucosa showed depleted abundances of the dominant Helicobacteraceae, while we found depleted abundances of the dominant Ruminococcaceae sp. in the stool. The rectal swabs showed similar trends as the colon mucosa, but were more variable. After the antibiotic treatment, there were increased abundances of similar taxa of facultative anaerobic bacteria, including Lactobacillaceae and Enterobacteriaceae at each sample site. [ABSTRACT FROM AUTHOR]

Despite the efficacy of antiretroviral-based pre-exposure prophylactics (PrEP) in men who have sex with men, studies in women have produced widely varying outcomes. Recent evidence demonstrates that vaginal microbial communities are associated with increased HIV acquisition risk and may impact PrEP efficacy. Here, we investigate the mechanisms underlying how vaginal bacteria alter PrEP drug levels and impact HIV infection rates ex vivo. Using cervicovaginal lavages (CVLs) from women with or without bacterial vaginosis (BV), we identified microbial metabolism of PrEP drugs in BV samples through LC-MS/MS analysis of soluble drug levels and metabolite formation in dual T-cell cultures. CVL samples were assessed for microbiome analysis using sequencing of bacterial 16S rRNA genes. We also observed non-Lactobacillus bacteria that are associated with BV may potentially impact PrEP efficacy through increased HIV infection rates in co-cultures containing Lactobacillus or BV bacteria, PrEP drugs, CEM-GFP cells, and HIV-1LAI virus. Finally, we used these data to develop a novel predictive mathematical simulation modeling system to predict these drug interactions for future trials. These studies demonstrate how dysbiotic vaginal microbiota may impact PrEP drugs and provides evidence linking vaginal bacteria to PrEP efficacy in women. Author summary: HIV prevention strategies with antiretroviral drugs as pre-exposure prophylactics (PrEP) are not efficacious in women, in part due to biological factors such as the vaginal microbiome. Lactobacillus spp. prevent the colonization of dysbiotic bacteria (i.e. bacterial vaginosis), which is associated with increased HIV transmission. However, the role these microbes play in altering the pharmacokinetics of PrEP drugs is currently unknown. Here we show that primary bacteria from women with dysbiosis impact PrEP drug levels and kinetics. We found that dysbiotic microbes metabolize PrEP drugs faster than target cell uptake, and significantly alter HIV infection rates in vitro, and this can be predicted with novel computational modeling. Our results demonstrate that the vaginal microbiome may play a key role in HIV prevention through altering therapeutic drug levels via metabolism. Thus, better measurements and interventions for vaginal dysbiosis will be critical in improving the efficacy of HIV prevention efforts in women. [ABSTRACT FROM AUTHOR]

Simian-human immunodeficiency viruses (SHIVs) have been utilized to test vaccine efficacy and characterize mechanisms of viral transmission and pathogenesis. However, the majority of SHIVs currently available have significant limitations in that they were developed using sequences from chronically HIV-infected individuals or uncommon HIV subtypes or were optimized for the macaque model by serially passaging the engineered virus in vitro or in vivo. Recently, a newly developed SHIV, SHIV.C.CH505.375H.dCT (SHIV.CH505), which incorporates vpu-env (gp140) sequences from a transmitted/founder HIV-1 subtype C strain, was shown to retain attributes of primary HIV-1 strains. However, a comprehensive analysis of the immunopathology that results from infection with this virus, especially in critical tissue compartments like the intestinal mucosa, has not been completed. In this study, we evaluated the viral dynamics and immunopathology of SHIV.CH505 in rhesus macaques. In line with previous findings, we found that SHIV.CH505 is capable of infecting and replicating efficiently in rhesus macaques, resulting in peripheral viral kinetics similar to that seen in pathogenic SIV and HIV infection. Furthermore, we observed significant and persistent depletions of CCR5+ and CCR6+ CD4+ T cells in mucosal tissues, decreases in CD4+ T cells producing Th17 cell-associated cytokines, CD8+ T cell dysfunction, and alterations of B cell and innate immune cell function, indicating that SHIV.CH505 elicits intestinal immunopathology typical of SIV/HIV infection. These findings suggest that SHIV.CH505 recapitulates the early viral replication dynamics and immunopathogenesis of HIV-1 infection of humans and thus can serve as a new model for HIV-1 pathogenesis, treatment, and prevention research. [ABSTRACT FROM AUTHOR]

Gastrointestinal (GI) mucosal dysfunction predicts and likely contributes to non-infectious comorbidities and mortality in HIV infection and persists despite antiretroviral therapy. However, the mechanisms underlying this dysfunction remain incompletely understood. Neutrophils are important for containment of pathogens but can also contribute to tissue damage due to their release of reactive oxygen species and other potentially harmful effector molecules. Here we used a flow cytometry approach to investigate increased neutrophil lifespan as a mechanism for GI neutrophil accumulation in chronic, treated HIV infection and a potential role for gastrointestinal dysbiosis. We report that increased neutrophil survival contributes to neutrophil accumulation in colorectal biopsy tissue, thus implicating neutrophil lifespan as a new therapeutic target for mucosal inflammation in HIV infection. Additionally, we characterized the intestinal microbiome of colorectal biopsies using 16S rRNA sequencing. We found that a reduced Lactobacillus: Prevotella ratio associated with neutrophil survival, suggesting that intestinal bacteria may contribute to GI neutrophil accumulation in treated HIV infection. Finally, we provide evidence that Lactobacillus species uniquely decrease neutrophil survival and neutrophil frequency in vitro, which could have important therapeutic implications for reducing neutrophil-driven inflammation in HIV and other chronic inflammatory conditions. [ABSTRACT FROM AUTHOR]

Background. Cannabis is a widely used drug in the United States, and the frequency of cannabis use in the human immunodeficiency virus (HIV)-infected population is disproportionately high. Previous human and macaque studies suggest that cannabis may have an impact on plasma viral load; however, the relationship between cannabis use and HIV-associated systemic inflammation and immune activation has not been well defined. Methods. The impact of cannabis use on peripheral immune cell frequency, activation, and function was assessed in 198 HIV- infected, antiretroviral-treated individuals by flow cytometry. Individuals were categorized into heavy, medium, or occasional cannabis users or noncannabis users based on the amount of the cannabis metabolite 11-nor-carboxy-tetrahydrocannabinol (THC- COOH) detected in plasma by mass spectrometry. Results. Heavy cannabis users had decreased frequencies of human leukocyte antigen (HLA)-DR+CD38+CD4+ and CD8+ T-cell frequencies, compared to frequencies of these cells in non-cannabis-using individuals. Heavy cannabis users had decreased frequencies of intermediate and nonclassical monocyte subsets, as well as decreased frequencies of interleukin 23- and tumor necrosis factor-α-producing antigen-presenting cells. Conclusions. While the clinical implications are unclear, our findings suggest that cannabis use is associated with a potentially beneficial reduction in systemic inflammation and immune activation in the context of antiretroviral-treated HIV infection. [ABSTRACT FROM AUTHOR]

Liver disease is a leading contributor to morbidity and mortality during HIV infection, despite the use of combination antiretroviral therapy (cART). The precise mechanisms of liver disease during HIV infection are poorly understood partially due to the difficulty in obtaining human liver samples as well as the presence of confounding factors (e.g. hepatitis co-infection, alcohol use). Utilizing the simian immunodeficiency virus (SIV) macaque model, a controlled study was conducted to evaluate the factors associated with liver inflammation and the impact of cART. We observed an increase in hepatic macrophages during untreated SIV infection that was associated with a number of inflammatory and fibrosis mediators (TNFα, CCL3, TGFβ). Moreover, an upregulation in the macrophage chemoattractant factor CCL2 was detected in the livers of SIV-infected macaques that coincided with an increase in the number of activated CD16+ monocyte/macrophages and T cells expressing the cognate receptor CCR2. Expression of Mac387 on monocyte/macrophages further indicated that these cells recently migrated to the liver. The hepatic macrophage and T cell levels strongly correlated with liver SIV DNA levels, and were not associated with the levels of 16S bacterial DNA. Utilizing in situ hybridization, SIV-infected cells were found primarily within portal triads, and were identified as T cells. Microarray analysis identified a strong antiviral transcriptomic signature in the liver during SIV infection. In contrast, macaques treated with cART exhibited lower levels of liver macrophages and had a substantial, but not complete, reduction in their inflammatory profile. In addition, residual SIV DNA and bacteria 16S DNA were detected in the livers during cART, implicating the liver as a site on-going immune activation during antiretroviral therapy. These findings provide mechanistic insights regarding how SIV infection promotes liver inflammation through macrophage recruitment, with implications for in HIV-infected individuals. [ABSTRACT FROM AUTHOR]

An altered intestinal microbiome during chronic human immunodeficiency virus (HIV) infection is associated with mucosal dysfunction, inflammation, and disease progression. We performed a preclinical evaluation of the safety and efficacy of fecal microbiota transplantation (FMT) as a potential therapeutic in HIV-infected individuals. Antiretroviral-treated, chronically simian immunodeficiency virus (SIV)-infected rhesus macaques received antibiotics followed by FMT. The greatest microbiota shift was observed after antibiotic treatment. The bacterial community composition at 2 weeks post-FMT resembled the pre-FMT community structure, although differences in the abundances of minor bacterial populations remained. Immunologically, we observed significant increases in the number of peripheral Th17 and Th22 cells and reduced CD4+ T cell activation in gastrointestinal tissues post-FMT. Importantly, the transplant was well tolerated with no negative clinical side effects. Although this pilot study did not control for the differential contributions of antibiotic treatment and FMT to the observed results, the data suggest that FMT may have beneficial effects that should be further evaluated in larger studies. [ABSTRACT FROM AUTHOR]

Given the critical role of mucosal surfaces in susceptibility to infection, it is imperative that effective mucosal responses are induced when developing efficacious vaccines and prevention strategies for infection. Modulating the microbiota in the gastrointestinal (GI) tract through the use of probiotics (PBio) is a safe and well-tolerated approach to enhance mucosal and overall health. We assessed the longitudinal impact of daily treatment with the VSL#3 probiotic on cellular and humoral immunity and inflammation in healthy macaques. PBio therapy resulted in significantly increased frequencies of B cells expressing IgA in the colon and lymph node (LN), likely because of significantly increased LN T follicular helper cell frequencies and LN follicles. Increased frequencies of IL-23+ APCs in the colon were found post-PBio treatment, which correlated with LN T follicular helper cells. Finally, VSL#3 significantly downmodulated the response of TLR2-, TLR3-, TLR4-, and TLR9-expressing HEK293 cells to stimulation with Pam3CSK4, polyinosinic-polycytidylic acid, LPS, and ODN2006, respectively. These data provide a mechanism for the beneficial impact of PBio on mucosal health and implicates the use of PBio therapy in the context of vaccination or preventative approaches to enhance protection from mucosal infection by improving immune defenses at the mucosal portal of entry. [ABSTRACT FROM AUTHOR]

Here we describe the dynamic effects of cumulus clouds on microclimate and shoot-level photosynthetic gas exchange in saplings of two treeline conifer species— Picea engelmannii and Abies lasiocarpa. Measurements were made during both clear-sky and partly cloudy conditions (∼10–70% cumulus cloud cover) throughout the 2012 growing season within an alpine-treeline ecotone. Cumulus clouds generated dynamic fluctuations in photosynthetically active radiation (PAR), higher maximum PAR (>2500 μmol m −2 s −1 ), 2–4 fold increases in diffuse PAR, reduced daily mean and cumulative PAR, lower needle temperatures, and reduced leaf-to-air vapor pressure differences relative to clear-sky conditions. Onset of cloud-shade corresponded with declines in photosynthesis, needle temperatures, and evapotranspiration, which were proportional to cloud duration and opacity. Despite increased diffuse light and greater sunlight intensity during cloud-gaps, photosynthesis was never higher on partly cloudy days compared to clear days in either species, during cloud-gaps or cloud-shade. However, reduced transpiration paired with photosynthesis comparable with clear-sky levels during cloud-gaps resulted in greater instantaneous water use efficiency relative to clear-sky measurements. There was no apparent photoinhibition of photosynthesis reflected in gas exchange measurements in response to abrupt and dramatic changes in PAR levels caused by cumulus clouds. We conclude that cumulus clouds reduce instantaneous and daily carbon gain, although lower needle temperatures and associated reductions in transpirational water loss may alleviate daily and seasonal water stress, thereby enhancing carbon gain over the growing season. [ABSTRACT FROM AUTHOR]