Longitudinal transkingdom gut microbial approach towards decompensation in outpatients with cirrhosis.

OBJECTIVE: First decompensation development is a critical milestone that needs to be predicted. Transkingdom gut microbial interactions, including archaeal methanogens, may be important targets and predictors but a longitudinal approach is needed. DESIGN: Cirrhosis outpatients who provided stool twice were included. Group 1: compensated, group 2: 1 decompensation (decomp), group 3: >1 decompensationwere followed and divided into those who remained stable or decompensated. Bacteria, viral and archaeal presence, α/ß diversity and taxa changes over time adjusted for clinical variables were analysed. Correlation networks between kingdoms were analysed. RESULTS: 157 outpatients (72 group 1, 33 group 2 and 52 group 3) were followed and 28%-47% developed outcomes. Baseline between those who remained stable/developed outcome: While no α/ß diversity differences were seen, commensals were lower and pathobionts were higher in those who decompensated. After decompensation: those experiencing their first decompensation showed greater decrease in α/ß-diversity, bacterial change (↑Lactobacillus spp, Streptococcus parasanguinis and ↓ beneficial Lachnospiraceae and Eubacterium hallii) and viral change (↑Siphoviridae, ↓ Myoviridae) versus those with further decompensation. Archaea: 19% had Methanobacter brevii, which was similar between/within groups. Correlation networks: Baseline archaeal-viral-bacterial networks were denser and more homogeneous in those who decompensated versus the rest. Archaea-bacterial correlations collapsed post first decompensation. Lactobacillus phage Lc Nu and C2-like viruses were negatively linked with beneficial bacteria. CONCLUSION: In this longitudinal study of cirrhosis outpatients, the greatest transkingdom gut microbial changes were seen in those reaching the first decompensation, compared with subsequent decompensating events. A transkingdom approach may refine prediction and provide therapeutic targets to prevent cirrhosis progression.

Impact of bacterial infections and spontaneous bacterial peritonitis prophylaxis on phage-bacterial dynamics in cirrhosis.

BACKGROUND AND AIMS: Gut microbiota, including bacteria and phages, are altered in cirrhosis, but their role during infections and spontaneous bacterial peritonitis (SBP) prophylaxis is unclear. Our aim was determine metagenomic changes in gut bacteria; phages and their linkages centered around Gram-negative and Gram-positive pathobionts in patients with cirrhosis with/without infections or SBP prophylaxis. APPROACH AND RESULTS: We included uninfected (n = 231) and infected (n = 30, SBP n = 19 and urinary tract infection n = 11 before antibiotics) patients who gave stool for bacterial and phage metagenomics. We matched uninfected to infected patients 1:1 on a model for end-stage liver disease (MELD). We also analyzed subgroups of patients with ascites matched on an MELD (n = 73) to patients on SBP prophylaxis (n = 7) and then to SBP infection. Phage and bacterial taxa differences (DESeq2) and correlation networks centered around Escherichia coli and Enterococcus faecium were analyzed. Infections were mostly due to Enterobacteriaceae and Enterococcus spp. On metagenomics, higher fold changes of Enterobacteriaceae members, Enterococcus and Streptococcus spp., and Escherichia phages were seen in infected patients. Correlation networks showed more complex bacteria-phage linkages in infected patients compared with uninfected ones overall and centered around E. coli and E. faecium. SBP prophylaxis induced higher Gram-positive bacteria. In SBP, Enterococcus and Escherichia were higher versus ascites. Correlation networks around E. coli were complex in ascites but sparse with SBP prophylaxis, whereas the reverse was seen with E. faecium. Lytic phages and those associated with antibiotic resistance were correlated with E. faecium. CONCLUSION: In cirrhosis, there are significant changes in phage-bacterial linkages in infected patients and those on SBP prophylaxis compared to the remaining patients. SBP prophylaxis enriches complexity of E. faecium-centered but induces a collapse in E. coli-centered phage-bacterial correlations.

Detection of SARS-CoV-2 RNA in wastewater from dormitory buildings in a university campus: comparison with individual testing results.

Wastewater-based epidemiology (WBE) for monitoring COVID-19 has been largely used to detect the spread of the disease at the community level. From February to December 2022, we collected 24-h composite sewage samples from dormitory buildings in George Mason University (Fairfax, Virginia, USA) housing approximately 5,200 resident students. SARS-CoV-2 RNA extraction was achieved using an automated system based on magnetic nanoparticles. Analysis of SARS-CoV-2 RNA was performed using reverse transcription quantitative PCR based on the Centers for Disease Control and Prevention (CDC) N1 and N2 assays. From the 362 samples collected, 86% showed positive detection of SARS-CoV-2 RNA. Wastewater monitoring was able to detect SARS-CoV-2 RNA in 96% of the samples from buildings housing students with COVID-19. Over the period of study, we observed significant correlations between the SARS-CoV-2 concentration (copy number mL-1) in wastewater and the number of positive cases on campus based on individual saliva testing. Although several reports have been published on the wastewater monitoring of COVID-19 in university campuses, our study is one of the very few that provides results that were obtained during the last phase of the pandemic (roughly the year 2022), when the large majority of students were vaccinated and back on campus.

Stool microbiota are superior to saliva in distinguishing cirrhosis and hepatic encephalopathy using machine learning.

BACKGROUND & AIMS: Saliva and stool microbiota are altered in cirrhosis. Since stool is logistically difficult to collect compared to saliva, it is important to determine their relative diagnostic and prognostic capabilities. We aimed to determine the ability of stool vs. saliva microbiota to differentiate between groups based on disease severity using machine learning (ML). METHODS: Controls and outpatients with cirrhosis underwent saliva and stool microbiome analysis. Controls vs. cirrhosis and within cirrhosis (based on hepatic encephalopathy [HE], proton pump inhibitor [PPI] and rifaximin use) were classified using 4 ML techniques (random forest [RF], support vector machine, logistic regression, and gradient boosting) with AUC comparisons for stool, saliva or both sample types. Individual microbial contributions were computed using feature importance of RF and Shapley additive explanations. Finally, thresholds for including microbiota were varied between 2.5% and 10%, and core microbiome (DESeq2) analysis was performed. RESULTS: Two hundred and sixty-nine participants, including 87 controls and 182 patients with cirrhosis, of whom 57 had HE, 78 were on PPIs and 29 on rifaximin were included. Regardless of the ML model, stool microbiota had a significantly higher AUC in differentiating groups vs. saliva. Regarding individual microbiota: autochthonous taxa drove the difference between controls vs. patients with cirrhosis, oral-origin microbiota the difference between PPI users/non-users, and pathobionts and autochthonous taxa the difference between rifaximin users/non-users and patients with/without HE. These were consistent with the core microbiome analysis results. CONCLUSIONS: On ML analysis, stool microbiota composition is significantly more informative in differentiating between controls and patients with cirrhosis, and those with varying cirrhosis severity, compared to saliva. Despite logistic challenges, stool should be preferred over saliva for microbiome analysis. LAY SUMMARY: Since it is harder to collect stool than saliva, we wanted to test whether microbes from saliva were better than stool in differentiating between healthy people and those with cirrhosis and, among those with cirrhosis, those with more severe disease. Using machine learning, we found that microbes in stool were more accurate than saliva alone or in combination, therefore, stool should be preferred for analysis and collection wherever possible.

Impact of Antibiotic Resistance Genes in Gut Microbiome of Patients With Cirrhosis.

BACKGROUND AND AIMS: Cirrhosis is associated with changes in intestinal microbiota that can lead to hepatic encephalopathy (HE) and infections, especially with antibiotic-resistant organisms. However, the impact of gut microbial antibiotic resistance gene (ARG) burden on clinical outcomes is unclear. The aims of the study were to determine the impact of ARGs in cirrhosis-related gut metagenome on outcomes and disease progression, study the effect of rifaximin on ARG burden, and compare ARGs in cirrhosis with chronic kidney disease (CKD) and diabetes. METHODS: In outpatients with cirrhosis who underwent metagenomics, we evaluated change in ARG abundances with progression and their multivariable impact on 90-day hospitalizations and deaths over 1 year. We also studied ARGs pre- and 8 weeks post-rifaximin in patients with compensated cirrhosis in an open-label trial. Finally, ARGs from CKD and diabetes studies were compared with cirrhosis on machine learning. RESULTS: A total of 163 patients with cirrhosis (43 compensated, 20 ascites-only, 30 HE-only, 70 both) and 40 controls were included. ARG abundances were higher in cirrhosis versus controls and worsened with advancing cirrhosis severity; 44 patients were hospitalized and 14 died. ARG abundances were associated with hospitalizations and mortality while controlling for cirrhosis complications, medications, and demographics. Rifaximin trial: ARG abundance patterns were minimally affected in 19 patients post-rifaximin. CKD/diabetes comparison: ARG abundance patterns in cirrhosis are distinguishable on machine learning and include more gram-positive ARGs. CONCLUSIONS: Cirrhosis is associated with high gut microbial ARG gene burden compared with controls, which worsens with disease progression and may be different from CKD and diabetes. ARGs are not affected by rifaximin and are associated with hospitalizations and death.

Interaction of Microbiome, Diet, and Hospitalizations Between Brazilian and American Patients With Cirrhosis.

BACKGROUND & AIMS: Gut microbiota are affected by diet, country, and affect outcomes in cirrhosis. Western diets are associated with dysbiosis. Comparisons with other diets is needed. We aimed to compare cirrhosis patients from the United States with cirrhosis patients from Brazil with respect to diet, microbiota, and impact on hospitalizations. METHODS: Healthy controls and compensated/decompensated outpatients with cirrhosis from the United States and Brazil underwent dietary recall and stool for 16S ribosomal RNA sequencing. Demographics and medications/cirrhosis details were compared within and between countries. Patients with cirrhosis were followed up for 90-day hospitalizations. Regression for Shannon diversity was performed within cirrhosis. Regression for hospitalizations adjusting for clinical and microbial variables was performed. RESULTS: Model for end-stage liver disease (MELD), diabetes, ascites, and albumin were similar, but more Americans were men, had higher hepatic encephalopathy and alcohol/hepatitis C etiology, with lower nonalcoholic fatty liver disease than Brazilians. Brazilians had higher cereal, rice, and yogurt intake vs the United States. As disease progressed, cereals, rice/beans, coffee, and chocolate consumption was reduced. Microbial diversity was higher in Brazilians. Within cirrhosis, high diversity was related to Brazilian origin (P < .0001), age, and cereal intake (P = .05), while high MELD scores (P = .009) and ascites (P = .05) did the reverse. Regardless of stage, beneficial taxa and taxa associated with grant and yogurt intake were higher (Ruminococcaceae, Christensenellacae, and Prevotellaceae), while pathobionts (Porphyromonadaceae, Sutterellaceae, and Enterobacteriaceae) were lower in Brazilians. More Americans were hospitalized vs Brazilians (P = .002). On regression, MELD (P = .001) and ascites (P = .001) were associated with higher hospitalizations, while chocolate (P = .03) and Brazilian origin (P = .001) were associated with lower hospitalizations with/without microbiota inclusion. CONCLUSIONS: Brazilian cirrhotic patients follow a diet richer in cereals and yogurt, which is associated with higher microbial diversity and beneficial microbiota and could contribute toward lower hospitalizations compared with a Western-diet-consuming American cohort.

Association of serum metabolites and gut microbiota at hospital admission with nosocomial infection development in patients with cirrhosis.

Cirrhosis is complicated by a high rate of nosocomial infections (NIs), which result in poor outcomes and are challenging to predict using clinical variables alone. Our aim was to determine predictors of NI using admission serum metabolomics and gut microbiota in inpatients with cirrhosis. In this multicenter inpatient cirrhosis study, serum was collected on admission for liquid chromatography-mass spectrometry metabolomics, and a subset provided stool for 16SrRNA analysis. Hospital course, including NI development and death, were analyzed. Metabolomic analysis using analysis of covariance (ANCOVA) (demographics, Model for End-Stage Liver Disease [MELD] admission score, white blood count [WBC], rifaximin, and infection status adjusted) and random forest analyses for NI development were performed. Additional values of serum metabolites over clinical variables toward NI were evaluated using logistic regression. Stool microbiota and metabolomic correlations were compared in patients with and without NI development. A total of 602 patients (231 infection admissions) were included; 101 (17%) developed NIs, which resulted in worse inpatient outcomes, including intensive care unit transfer, organ failure, and death. A total of 127 patients also gave stool samples, and 20 of these patients developed NIs. The most common NIs were spontaneous bacterial peritonitis followed by urinary tract infection, Clostridioides difficile, and pneumonia. A total of 247 metabolites were significantly altered on ANCOVA. Higher MELD scores (odds ratio, 1.05; p < 0.0001), admission infection (odds ratio, 3.54; p < 0.0001), and admission WBC (odds ratio, 1.05; p = 0.04) predicted NI (area under the curve, 0.74), which increased to 0.77 (p = 0.05) with lower 1-linolenoyl-glycerolphosphocholine (GPC) and 1-stearoyl-GPC and higher N-acetyltryptophan and N-acetyl isoputreanine. Commensal microbiota were lower and pathobionts were higher in those who developed NIs. Microbial-metabolite correlation networks were complex and dense in patients with NIs, especially sub-networks centered on Ruminococcaceae and Pseudomonadaceae. NIs are common and associated with poor outcomes in cirrhosis. Admission gut microbiota in patients with NIs showed higher pathobionts and lower commensal microbiota. Microbial-metabolomic correlations were more complex, dense, and homogeneous among those who developed NIs, indicating greater linkage strength. Serum metabolites and gut microbiota on admission are associated with NI development in cirrhosis.

A Randomized Clinical Trial of Fecal Microbiota Transplant for Alcohol Use Disorder.

BACKGROUND AND AIMS: Alcohol use disorder (AUD) is associated with microbial alterations that worsen with cirrhosis. Fecal microbiota transplant (FMT) could be a promising approach. APPROACH AND RESULTS: In this phase 1, double-blind, randomized clinical trial, patients with AUD-related cirrhosis with problem drinking (AUDIT-10 > 8) were randomized 1:1 into receiving one placebo or FMT enema from a donor enriched in Lachnospiraceae and Ruminococcaceae. Six-month safety was the primary outcome. Alcohol craving questionnaire, alcohol consumption (urinary ethylglucuronide/creatinine), quality of life, cognition, serum IL-6 and lipopolysaccharide-binding protein, plasma/stool short-chain fatty acids (SCFAs), and stool microbiota were tested at baseline and day 15. A 6-month follow-up with serious adverse event (SAE) analysis was performed. Twenty patients with AUD-related cirrhosis (65 ± 6.4 years, all men, Model for End-Stage Liver Disease 8.9 ± 2.7) with similar demographics, cirrhosis, and AUD severity were included. Craving reduced significantly in 90% of FMT versus 30% in placebo at day 15 (P = 0.02) with lower urinary ethylglucuronide/creatinine (P = 0.03) and improved cognition and psychosocial quality of life. There was reduction in serum IL-6 and lipopolysaccharide-binding protein and increased butyrate/isobutyrate compared with baseline in FMT but not placebo. Microbial diversity increased with higher Ruminococcaceae and other SCFAs, producing taxa following FMT but not placebo, which were linked with SCFA levels. At 6 months, patients with any SAEs (8 vs. 2, P = 0.02), AUD-related SAEs (7 vs. 1, P = 0.02), and SAEs/patient (median [interquartile range], 1.5 [1.25] vs. 0 [0.25] in FMT, P = 0.02) were higher in placebo versus FMT. CONCLUSIONS: This phase 1 trial shows that FMT is safe and associated with short-term reduction in alcohol craving and consumption with favorable microbial changes versus placebo in patients with alcohol-associated cirrhosis with alcohol misuse. There was also a reduction in AUD-related events over 6 months in patients assigned to FMT.

The Presence of Denitrifiers In Bacterial Communities of Urban Stormwater Best Management Practices (BMPs).

Stormwater best management practices (BMPs) are engineered structures that attempt to mitigate the impacts of stormwater, which can include nitrogen inputs from the surrounding drainage area. The goal of this study was to assess bacterial community composition in different types of stormwater BMP soils to establish whether a particular BMP type harbors more denitrification potential. Soil sampling took place over the summer of 2015 following precipitation events. Soils were sampled from four bioretention facilities, four dry ponds, four surface sand filters, and one dry swale. 16S rRNA gene analysis of extracted DNA and RNA amplicons indicated high bacterial diversity in the soils of all BMP types sampled. An abundance of denitrifiers was also indicated in the extracted DNA using presence/absence of nirS, nirK, and nosZ denitrification genes. BMP soil bacterial communities were impacted by the surrounding soil physiochemistry. Based on the identification of a metabolically-active community of denitrifiers, this study has indicated that denitrification could potentially occur under appropriate conditions in all types of BMP sampled, including surface sand filters that are often viewed as providing low potential for denitrification. The carbon content of incoming stormwater could be providing bacterial communities with denitrification conditions. The findings of this study are especially relevant for land managers in watersheds with legacy nitrogen from former agricultural land use.

Interaction of bacterial metagenome and virome in patients with cirrhosis and hepatic encephalopathy.

OBJECTIVE: Altered bacterial composition is associated with disease progression in cirrhosis but the role of virome, especially phages, is unclear. DESIGN: Cross-sectional and pre/post rifaximin cohorts were enrolled. Cross-sectional: controls and cirrhotic outpatients (compensated, on lactulose (Cirr-L), on rifaximin (Cirr-LR)) were included and followed for 90-day hospitalisations. Pre/post: compensated cirrhotics underwent stool collection pre/post 8 weeks of rifaximin. Stool metagenomics for bacteria and phages and their correlation networks were analysed in controls versus cirrhosis, within cirrhotics, hospitalised/not and pre/post rifaximin. RESULTS: Cross-sectional: 40 controls and 163 cirrhotics (63 compensated, 43 Cirr-L, 57 Cirr-LR) were enrolled. Cirr-L/LR groups were similar on model for end-stage liver disease (MELD) score but Cirr-L developed greater hospitalisations versus Cirr-LR (56% vs 30%, p=0.008). Bacterial alpha/beta diversity worsened from controls through Cirr-LR. While phage alpha diversity was similar, beta diversity was different between groups. Autochthonous bacteria linked negatively, pathobionts linked positively with MELD but only modest phage-MELD correlations were seen. Phage-bacterial correlation network complexity was highest in controls, lowest in Cirr-L and increased in Cirr-LR. Microviridae and Faecalibacterium phages were linked with autochthonous bacteria in Cirr-LR, but not Cirr-L hospitalised patients had greater pathobionts, lower commensal bacteria and phages focused on Streptococcus, Lactococcus and Myoviridae. Pre/post: No changes in alpha/beta diversity of phages or bacteria were seen postrifaximin. Phage-bacterial linkages centred around urease-producing Streptococcus species collapsed postrifaximin. CONCLUSION: Unlike bacteria, faecal phages are sparsely linked with cirrhosis characteristics and 90-day outcomes. Phage and bacterial linkages centred on urease-producing, ammonia-generating Streptococcus species were affected by disease progression and rifaximin therapy and were altered in patients who experienced 90-day hospitalisations.

Sex is associated with differences in gut microbial composition and function in hepatic encephalopathy.

BACKGROUND & AIMS: Altered microbiota can affect the gut-liver-brain axis in cirrhosis and hepatic encephalopathy (HE), but the impact of sex on these changes is unclear. We aimed to determine differences in fecal microbiota composition/functionality between men and women with cirrhosis and HE on differing treatments. METHODS: Cross-sectional stool microbiome composition (16s rRNA sequencing) and microbial functional analyses were performed in men and women with cirrhosis, and controls. Patients with HE on rifaximin+lactulose (HE-Rif), patients with HE on lactulose only (HE-Lac) and those with cirrhosis without HE (No-HE) were compared to controls using random forest classifier. Men and women were also compared. RESULTS: A total of 761 individuals were included, 619 with cirrhosis (466 men, 153 women) and 142 controls (92 men, 50 women). Men were older and more frequently used proton pump inhibitors (PPIs), but model for end-stage liver disease score, No-HE (n = 319), HE-lac (n = 130) and HE-Rif (n = 170) proportions were similar. PPI/age-adjusted AUC of differentiation between controls vs. all cirrhosis, and controls vs. No-HE were higher within women than men, but the adjusted AUC for No-HE vs. HE-Rif was higher in men. Control vs. HE-Rif differentiation was similar across sexes. Men vs. women were different in all cirrhosis, No-HE and HE-Lac but not HE-Rif on PERMANOVA and AUC analyses. Autochthonous taxa decreased and pathobionts increased with disease progression regardless of sex. In men, Lactobacillaceae were higher in HE-Lac but decreased in HE-Rif, along with Veillonellaceae. Pathways related to glutamate and aromatic compound degradation were higher in men at all stages. Degradation of androstenedione, an estrogenic precursor, was lower in men vs. women in HE-Rif, likely enhancing feminization. CONCLUSIONS: There are differences in gut microbial function and composition between men and women with cirrhosis, which could be implicated in differential responses to HE therapies. Further studies linking these differences to sex-specific outcomes are needed. LAY SUMMARY: Patients with cirrhosis develop changes in their brain function, and men often develop feminization with disease progression. However, the interaction between sex, microbiota and disease severity is unclear. We found that as disease progressed in men, their microbial composition began to approach that observed in women, with changes in specific microbes that are associated with male hormone metabolism.

Serum Levels of Metabolites Produced by Intestinal Microbes and Lipid Moieties Independently Associated With Acute-on-Chronic Liver Failure and Death in Patients With Cirrhosis.

BACKGROUND & AIMS: Inpatients with cirrhosis have high rates of acute-on-chronic failure (ACLF) development and high mortality within 30 days of admission to the hospital. Better biomarkers are needed to predict these outcomes. We performed metabolomic analyses of serum samples from patients with cirrhosis at multiple centers to determine whether metabolite profiles might identify patients at high risk for ACLF and death. METHODS: We performed metabolomic analyses, using liquid chromatography, of serum samples collected at time of admission to 12 North American tertiary hepatology centers from 602 patients in the North American Consortium for the Study of End-Stage Liver Disease sites from 2015 through 2017 (mean age, 56 years; 61% men; mean model for end-stage liver disease score, 19.5). We performed analysis of covariance, adjusted for model for end-stage liver disease at time of hospital admission, serum levels of albumin and sodium, and white blood cell count, to identify metabolites that differed between patients who did vs did not develop ACLF and patients who did vs did not die during hospitalization and within 30 days. We performed random forest analysis to identify specific metabolite(s) that were associated with outcomes and area under the curve (AUC) analyses to analyze them in context of clinical parameters. We analyzed microbiomes of stool samples collected from 133 patients collected at the same time and examined associations with serum metabolites. RESULTS: Of the 602 patients analyzed, 88 developed ACLF (15%), 43 died in the hospital (7%), and 72 died within 30 days (12%). Increased levels of compounds of microbial origin (aromatic compounds, secondary or sulfated bile acids, and benzoate) and estrogen metabolites, as well as decreased levels of phospholipids, were associated with development of ACLF, inpatient, and 30-day mortality and were also associated with fecal microbiomes. Random forest analysis and logistic regression showed that levels of specific microbially produced metabolites identified patients who developed ACLF with an AUC of 0.84 (95% confidence interval [CI] 0.78-0.88; P = .001), patients who died while in the hospital with an AUC of 0.81 (95% CI 0.74-0.85; P = .002), and patients who died within 30 days with an AUC of 0.77 (95% CI 0.73-0.81; P = .02). The metabolites were significantly additive to clinical parameters for predicting these outcomes. Metabolites associated with outcomes were also correlated with microbiomes of stool samples. CONCLUSIONS: In an analysis of serum metabolites and fecal microbiomes of patients hospitalized with cirrhosis at multiple centers, we associated metabolites of microbial origin and lipid moieties with development of ACLF and death as an inpatient or within 30 days, after controlling for clinical features.

Gut Microbial Signature of Hepatocellular Cancer in Men With Cirrhosis.

The gut microbiome is altered in cirrhosis. Recent evidence has suggested a key role for the gut microbiota in the progression of cirrhosis and the development of hepatocellular carcinoma (HCC). We studied the differences in the microbial composition in patients with cirrhosis with prior and future HCC in the context of other complications (eg, infections, hepatic encephalopathy). The following 2 cohorts were recruited prospectively: the prior HCC cohort, in which outpatients with HCC within 2 years were age-matched, sex-matched, and Model for End-Stage Liver Disease (MELD) score-matched with those without HCC; and the future HCC cohort, in which patients were followed for 2 years and divided into future HCC versus no HCC after age, sex, and MELD-score matching and other complications were also recorded. Microbiota composition and predicted function were analyzed with ribosomal RNA sequencing and Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PiCRUST)and compared between (1) prior HCC versus none and (2) future HCC versus none, and in the future cohort, comparisons were also made between those patients who developed (1) HCC only versus without complications, (2) HCC only versus non-HCC complications only, and (3) HCC + other complications versus non-HCC complications only. A total of 142 men (76 total in the prior cohort [38 with/38 without HCC] and 66 total in the future cohort [33 with/33 without future HCC]) were included. The groups had similar etiology, lactulose/rifaximin/proton pump inhibitor use, diabetes mellitus, and non-HCC complications. Microbial diversity was similar between prior HCC/not or future HCC/not. On DESeq2 higher Clostridium sensu stricto and Anaerotruncus were significantly associated with protection from HCC, whereas the reverse was seen with Raoultella and Haemophilus regardless of prior/future HCC comparisons. Functions focused on urea cycle, bioenergetics, tryptophan, and toluene metabolism were different between the groups. Rothia was specific for other complications. Despite age, sex, and MELD-score matching and accounting for other complications, gut microbiota composition and the predicted function are different in men with cirrhosis with and without prior HCC and can be extended toward future HCC development.

Berberine alters gut microbial function through modulation of bile acids.

BACKGROUND: Berberine (BBR) is a plant-based nutraceutical that has been used for millennia to treat diarrheal infections and in contemporary medicine to improve patient lipid profiles. Reduction in lipids, particularly cholesterol, is achieved partly through up-regulation of bile acid synthesis and excretion into the gastrointestinal tract (GI). The efficacy of BBR is also thought to be dependent on structural and functional alterations of the gut microbiome. However, knowledge of the effects of BBR on gut microbiome communities is currently lacking. Distinguishing indirect effects of BBR on bacteria through altered bile acid profiles is particularly important in understanding how dietary nutraceuticals alter the microbiome. RESULTS: Germfree mice were colonized with a defined minimal gut bacterial consortium capable of functional bile acid metabolism (Bacteroides vulgatus, Bacteroides uniformis, Parabacteroides distasonis, Bilophila wadsworthia, Clostridium hylemonae, Clostridium hiranonis, Blautia producta; B4PC2). Multi-omics (bile acid metabolomics, 16S rDNA sequencing, cecal metatranscriptomics) were performed in order to provide a simple in vivo model from which to identify network-based correlations between bile acids and bacterial transcripts in the presence and absence of dietary BBR. Significant alterations in network topology and connectivity in function were observed, despite similarity in gut microbial alpha diversity (P = 0.30) and beta-diversity (P = 0.123) between control and BBR treatment. BBR increased cecal bile acid concentrations, (P < 0.05), most notably deoxycholic acid (DCA) (P < 0.001). Overall, analysis of transcriptomes and correlation networks indicates both bacterial species-specific responses to BBR, as well as functional commonalities among species, such as up-regulation of Na+/H+ antiporter, cell wall synthesis/repair, carbohydrate metabolism and amino acid metabolism. Bile acid concentrations in the GI tract increased significantly during BBR treatment and developed extensive correlation networks with expressed genes in the B4PC2 community. CONCLUSIONS: This work has important implications for interpreting the effects of BBR on structure and function of the complex gut microbiome, which may lead to targeted pharmaceutical interventions aimed to achieve the positive physiological effects previously observed with BBR supplementation.

Neuroinflammation in Murine Cirrhosis Is Dependent on the Gut Microbiome and Is Attenuated by Fecal Transplant.

Cirrhosis and hepatic encephalopathy (HE) is associated with an altered gut-liver-brain axis. Fecal microbial transplant (FMT) after antibiotics improves outcomes in HE, but the impact on brain function is unclear. The aim of this study is to determine the effect of colonization using human donors in germ-free (GF) mice on the gut-liver-brain axis. GF and conventional mice were made cirrhotic using carbon tetrachloride and compared with controls in GF and conventional state. Additional GF mice were colonized with stool from controls (Ctrl-Hum) and patients with cirrhosis (Cirr-Hum). Stools from patients with HE cirrhosis after antibiotics were pooled (pre-FMT). Stools from the same patients 15 days after FMT from a healthy donor were also pooled (post-FMT). Sterile supernatants were created from pre-FMT and post-FMT samples. GF mice were colonized using stools/sterile supernatants. For all mice, frontal cortex, liver, and small/large intestines were collected. Cortical inflammation, synaptic plasticity and gamma-aminobutyric acid (GABA) signaling, and liver inflammation and intestinal 16s ribosomal RNA microbiota sequencing were performed. Conventional cirrhotic mice had higher degrees of neuroinflammation, microglial/glial activation, GABA signaling, and intestinal dysbiosis compared with other groups. Cirr-Hum mice had greater neuroinflammation, microglial/glial activation, and GABA signaling and lower synaptic plasticity compared with Ctrl-Hum mice. This was associated with greater dysbiosis but no change in liver histology. Pre-FMT material colonization was associated with neuroinflammation and microglial activation and dysbiosis, which was reduced significantly with post-FMT samples. Sterile pre-FMT and post-FMT supernatants did not affect brain parameters. Liver inflammation was unaffected. Conclusion: Fecal microbial colonization from patients with cirrhosis results in higher degrees of neuroinflammation and activation of GABAergic and neuronal activation in mice regardless of cirrhosis compared with those from healthy humans. Reduction in neuroinflammation by using samples from post-FMT patients to colonize GF mice shows a direct effect of fecal microbiota independent of active liver inflammation or injury.

Extensive photobiont sharing in a rapidly radiating cyanolichen clade.

Recent studies have uncovered remarkable diversity in Dictyonema s.lat. basidiolichens, here recognized as subtribe Dictyonemateae. This group includes five genera and 148 species, but hundreds more await description. The photobionts of these lichens belong to Rhizonema, a recently resurrected cyanobacterial genus known by a single species. To further investigate photobiont diversity within Dictyonemateae, we generated 765 new cyanobacterial sequences from 635 specimens collected from 18 countries. The ITS barcoding locus supported the recognition of 200 mycobiont (fungal) species among these samples, but the photobiont diversity was comparatively low. Our analyses revealed three main divisions of Rhizonema, with two repeatedly recovered as monophyletic (proposed as new species), and the third mostly paraphyletic. The paraphyletic lineage corresponds to R. interruptum and partnered with mycobionts from all five genera in Dictyonemateae. There was no evidence of photobiont-mycobiont co-speciation, but one of the monophyletic lineages of Rhizonema appears to partner predominantly with one of the two major clades of Cora (mycobiont) with samples collected largely from the northern Andes. Molecular clock estimations indicate the Rhizonema species are much older than the fungal species in the Dictyonemateae, suggesting that these basidiolichens obtained their photobionts from older ascolichen lineages and the photobiont variation in extant lineages of Dictyonemateae is the result of multiple photobiont switches. These results support the hypothesis of lichens representing "fungal farmers," in which diverse mycobiont lineages associate with a substantially lower diversity of photobionts by sharing those photobionts best suited for the lichen symbiosis among multiple and often unrelated mycobiont lineages.

Vaginal microbiome: normalcy vs dysbiosis.

It has been long understood that the vaginal microflora is crucial in maintaining a normal physiological environment for the host and its involvement is deemed indispensable for reproductive success. A global concept of normalcy vs. dysbiosis of vaginal microbiome is debatable as women of different races have a unique vaginal microflora with regional variations. Vaginal microflora is a dynamic microenvironment affected by gestational status, menstrual cycle, sexual activity, age, and contraceptive use. Normal vaginal flora is dominated by lactobacilli especially in women of European descent vs. African American women. These microbes confer the host vagina protection from potentially pathogenic microbes that may lead to urinary tract infections and sexually transmitted diseases. Changes in the vaginal microbiota including reduced lactobacilli abundance and increased facultative and anaerobic organism populations result in bacterial vaginosis, that predisposes the host to several conditions like low birth weight and increased risk of contracting bacterial infections. On the other hand, the vaginal microbiome is also reshaped during pregnancy, with less microbial diversity with a dominance of Lactobacillus species. However, an altered vaginal microbiota with low lactobacilli abundance especially during pregnancy may result in induction of excessive inflammation and pre-term labor. Since the vaginal microbiome plays an important role during embryo implantation, it is not surprising that bacterial vaginosis is more common in infertile women and associated with reduced rates of conception. Probiotic has great success in treating bacterial vaginosis and restoring the normal microbiome in recent. This report, reviewed the relationships between the vaginal microbiome and women's reproductive health.

Cathelicidin-related antimicrobial peptide alleviates alcoholic liver disease through inhibiting inflammasome activation.

Alcoholic liver disease (ALD) is associated with gut dysbiosis and hepatic inflammasome activation. While it is known that antimicrobial peptides (AMPs) play a critical role in the regulation of bacterial homeostasis in ALD, the functional role of AMPs in the alcohol-induced inflammasome activation is unclear. The aim of this study was to determine the effects of cathelicidin-related antimicrobial peptide (CRAMP) on inflammasome activation in ALD. CRAMP knockout (Camp-/-) and wild-type (WT) mice were subjected to binge-on-chronic alcohol feeding and synthetic CRAMP peptide was administered. Serum/plasma and hepatic tissue samples from human subjects with alcohol use disorder and/or alcoholic hepatitis were analyzed. CRAMP deficiency exacerbated ALD with enhanced inflammasome activation as shown by elevated serum interleukin (IL)-1ß levels. Although Camp-/- mice had comparable serum endotoxin levels compared to WT mice after alcohol feeding, hepatic lipopolysaccharide (LPS) binding protein (LBP) and cluster of differentiation (CD) 14 were increased. Serum levels of uric acid (UA), a Signal 2 molecule in inflammasome activation, were positively correlated with serum levels of IL-1ß in alcohol use disorder patients with ALD and were increased in Camp-/- mice fed alcohol. In vitro studies showed that CRAMP peptide inhibited LPS binding to macrophages and inflammasome activation stimulated by a combination of LPS and UA. Synthetic CRAMP peptide administration decreased serum UA and IL-1ß concentrations and rescued the liver from alcohol-induced damage in both WT and Camp-/- mice. In summary, CRAMP exhibited a protective role against binge-on-chronic alcohol-induced liver damage via regulation of inflammasome activation by decreasing LPS binding and UA production. CRAMP administration may represent a novel strategy for treating ALD. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Comparative study of the hemolymph microbiome between live and recently dead American lobsters Homarus americanus.

Lobsters and other crustaceans do not have sterile hemolymph. Despite this, little is known about the microbiome in the hemolymph of the lobster Homarus americanus. The purpose of this study was to characterize the hemolymph microbiome in lobsters. The lobsters were part of a larger study on the effect of temperature on epizootic shell disease, and several died during the course of the study, providing an opportunity to examine differences in the microbiomes between live and recently dead (1-24 h) animals. The hemolymph microbiomes of live lobsters was different from those in dead animals and both were different from the tank microbiome in which the animals had been held. The microbiomes of live lobsters were more diverse and had a different suite of bacteria than those from dead animals. The dominant taxa in live lobsters belonged to Flavobacteriaceae and Rhodobacteraceae, whereas Vibrionaceae and Enterobacteriaceae were predominant in the dead lobsters. Although aquarium microbiomes overlapped with the hemolymph microbiomes, there was less overlap and lower abundance of taxa in comparison with hemolymph from live lobsters. Previous studies reporting bacteria in the digestive tract of lobsters suggested that Vibrionaceae and Enterobacteriaceae had invaded the hemolymph via the gut. Our study suggests that hemolymph bacteria abundant in live lobsters do not originate from the tank milieu and comprise a rich, natural, or native background of bacterial constituents.

Fecal Microbial Transplant Capsules Are Safe in Hepatic Encephalopathy: A Phase 1, Randomized, Placebo-Controlled Trial.

Hepatic encephalopathy (HE) can cause major morbidity despite standard of care (SOC; rifaximin/lactulose). Fecal microbial transplant (FMT) enemas postantibiotics are safe, but the effect of FMT without antibiotics using the capsular route requires investigation. The aim of this work was to determine the safety, tolerability, and impact on mucosal/stool microbiota and brain function in HE after capsular FMT in a randomized, single-blind, placebo-controlled clinical trial in Virginia. Patients with cirrhosis with recurrent HE with MELD (Model for End-Stage Liver Disease) <17 on SOC were randomized 1:1 into receiving 15 FMT capsules versus placebo from a single donor enriched in Lachnospiraceae and Ruminococcaceae. Endoscopies with duodenal and sigmoid biopsies, stool analysis, cognition, serum lipopolysaccharide-binding protein (LBP), and duodenal antimicrobial peptide (AMP) expression at baseline were used. Clinical follow-up with SOC maintenance was performed until 5 months. FMT-assigned patients underwent repeat endoscopies 4 weeks postenrollment. Twenty subjects on lactulose/rifaximin were randomized 1:1. MELD score was similar at baseline (9.6 vs. 10.2) and study end (10.2 vs. 10.5). Six patients in the placebo group required hospitalizations compared to 1 in FMT, which was deemed unrelated to FMT. Infection/HE episodes were similar between groups. Baseline microbial diversity was similar in all tissues between groups. Post-FMT, duodenal mucosal diversity (P = 0.01) increased with higher Ruminococcaceae and Bifidobacteriaceae and lower Streptococcaceae and Veillonellaceae. Reduction in Veillonellaceae were noted post-FMT in sigmoid (P = 0.04) and stool (P = 0.05). Duodenal E-cadherin (P = 0.03) and defensin alpha 5 (P = 0.03) increased whereas interleukin-6 (P = 0.02) and serum LBP (P = 0.009) reduced post-FMT. EncephalApp performance improved post-FMT only (P = 0.02). Conclusion: In this phase 1 study, oral FMT capsules are safe and well tolerated in patients with cirrhosis and recurrent HE. FMT was associated with improved duodenal mucosal diversity, dysbiosis, and AMP expression, reduced LBP, and improved EncephalApp performance. Further studies are needed to prove efficacy.