Genetic mutation in Escherichia coli genome during adaptation to the murine intestine is optimized for the host diet.

Mammalian gut microbes colonize the intestinal tract of their host and adapt to establish a microbial ecosystem. The host diet changes the nutrient profile of the intestine and has a high impact on microbiota composition. Genetic mutations in Escherichia coli, a prevalent species in the human gut, allow for adaptation to the mammalian intestine, as reported in previous studies. However, the extent of colonization fitness in the intestine elevated by genetic mutation and the effects of diet change on these mutations in E. coli are still poorly known. Here, we show that notable mutations in sugar metabolism-related genes (gatC, araC, and malI) were detected in the E. coli K-12 genome just 2 weeks after colonization in the germ-free mouse intestine. In addition to elevated fitness by deletion of gatC, as previously reported, deletion of araC and malI also elevated E. coli fitness in the murine intestine in a host diet-dependent manner. In vitro cultures of medium containing nutrients abundant in the intestine (e.g., galactose, N-acetylglucosamine, and asparagine) also showed increased E. coli fitness after deletion of the genes-of-interest associated with their metabolism. Furthermore, the host diet was found to influence the developmental trajectory of gene mutations in E. coli. Taken together, we suggest that genetic mutations in E. coli are selected in response to the intestinal environment, which facilitates efficient utilization of nutrients abundant in the intestine under laboratory conditions. Our study offers some insight into the possible adaptation mechanisms of gut microbes.IMPORTANCEThe gut microbiota is closely associated with human health and is greatly impacted by the host diet. Bacteria such as Escherichia coli live in the gut all throughout the life of a human host and adapt to the intestinal environment. Adaptive mutations in E. coli are reported to enhance fitness in the mammalian intestine, but to what extent is still poorly known. It is also unknown whether the host diet affects what genes are mutated and to what extent fitness is affected. This study suggests that genetic mutations in the E. coli K-12 strain are selected in response to the intestinal environment and facilitate efficient utilization of abundant nutrients in the germ-free mouse intestine. Our study provides a better understanding of these intestinal adaptation mechanisms of gut microbes.

Gut environment changes due to androgen deprivation therapy in patients with prostate cancer.

BACKGROUND: It is estimated that by 2040 there will be 1,017,712 new cases of prostate cancer worldwide. Androgen deprivation therapy (ADT) is widely used as a treatment option for all disease stages. ADT, and the resulting decline in androgen levels, may indirectly affect gut microbiota. Factors affecting gut microbiota are wide-ranging; however, literature is scarce on the effects of ADT on gut microbiota and metabolome profiles in patients with prostate cancer. METHODS: To study the changes of gut microbiome by ADT, this 24-week observational study investigated the relationship between testosterone levels and changes in gut microbiota in Japanese patients with prostate cancer undergoing ADT. Sequential faecal samples were collected 1 and 2 weeks before ADT, and 1, 4, 12, and 24 weeks after ADT. Blood samples were collected at almost the same times. Bacterial 16 S rRNA gene-based microbiome analyses and capillary electrophoresis-time-of-flight mass spectrometry-based metabolome analyses were performed. RESULTS: In total, 23 patients completed the study. The α- and ß-diversity of gut microbiota decreased significantly at 24 weeks after ADT (p = 0.017, p < 0.001, respectively). Relative abundances of Proteobacteria, Gammaproteobacteria, Pseudomonadales, Pseudomonas, and concentrations of urea, lactate, butyrate, 2-hydroxyisobutyrate and S-adenosylmethionine changed significantly after ADT (p < 0.05). There was a significant positive correlation between the abundance of Proteobacteria, a known indicator of dysbiosis, and the concentration of lactate (R = 0.49, p < 0.01). CONCLUSIONS: The decline in testosterone levels resulted in detrimental changes in gut microbiota. This dysbiosis may contribute to an increase in frailty and an increased risk of adverse outcomes in patients with prostate cancer.

Draft Genome Sequences of Bifidobacterium animalis Consecutively Isolated from Healthy Japanese Individuals.

Bifidobacterium species are well recognized as probiotics and colonized in various parts of the human body. Here, we report the draft genome sequences of Bifidobacterium animalis isolated from two healthy Japanese volunteers, one of which was sampled twice before and after a 10-year interval. A core genome phylogeny analysis indicated that the strains isolated from the same volunteer were closely related. This paper is the first report of multiple draft genome sequences of B. animalis independently isolated from the same individual and provides insight into the probiotic potential of a member of this species.

The guanylate cyclase C agonist linaclotide ameliorates the gut-cardio-renal axis in an adenine-induced mouse model of chronic kidney disease.

BACKGROUND: Cardiorenal syndrome is a major cause of mortality in patients with chronic kidney disease (CKD). However, the involvement of detrimental humoral mediators in the pathogenesis of cardiorenal syndrome is still controversial. Trimethylamine-N-oxide (TMAO), a hepatic metabolic product of trimethylamine generated from dietary phosphatidylcholine or carnitine derived by the gut microbiota, has been linked directly with progression of cardiovascular disease and renal dysfunction. Thus, targeting TMAO may be a novel strategy for the prevention of cardiovascular disease and chronic kidney disease. METHODS: Linaclotide, a guanylate cyclase C agonist, was administered to adenine-induced renal failure (RF) mice and changes in renal function and levels of gut-derived uremic toxins, as well as the gut microbiota community, were analyzed using metabolomic and metagenomic methods to reveal its cardiorenal effect. RESULTS: Linaclotide decreased the plasma levels of TMAO at a clinically used low dose of 10 µg/kg in the adenine-induced RF mouse model. At a high concentration of 100 µg/kg, linaclotide clearly improved renal function and reduced the levels of various uremic toxins. A reduction in TMAO levels following linaclotide treatment was also observed in a choline-fed pro-atherosclerotic model. Linaclotide ameliorated renal inflammation and fibrosis and cardiac fibrosis, as well as decreased the expression of collagen I, transforming growth factor-ß, galectin-3 (Gal-3) and ST2 genes. Plasma levels of Gal-3 and ST2 were also reduced. Because exposure of cardiomyocytes to TMAO increased fibronectin expression, these data suggest that linaclotide reduced the levels of TMAO and various uremic toxins and may result in not only renal, but also cardiac, fibrosis. F4/80-positive macrophages were abundant in small intestinal crypts in RF mice, and this increased expression was decreased by linaclotide. Reduced colonic claudin-1 levels were also restored by linaclotide, suggesting that linaclotide ameliorated the 'leaky gut' in RF mice. Metagenomic analysis revealed that the microbial order Clostridiales could be responsible for the change in TMAO levels. CONCLUSION: Linaclotide reduced TMAO and uremic toxin levels and could be a powerful tool for the prevention and control of the cardiorenal syndrome by modification of the gut-cardio-renal axis.

Cutibacterium acnes (Propionibacterium acnes) 16S rRNA Genotyping of Microbial Samples from Possessions Contributes to Owner Identification.

The human skin surface harbors huge numbers of microbes. The skin microbiota interacts with its host and forms a skin microbiome profile that is specific for each individual. It has been reported that the skin microbiota that is left on an individual's possessions can act as a sort of "fingerprint" and be used for owner identification. However, this approach needs to be improved to take into account any long-term instability of skin microbiota and contamination from nonspecific bacteria. Here, we took advantage of single-nucleotide polymorphisms (SNPs) in the 16S-encoding rRNA gene of Cutibacterium acnes, the most common and abundant bacterium on human skin, to perform owner identification. We first developed a high-throughput genotyping method based on next-generation sequencing to characterize the SNPs of the C. acnes 16S rRNA gene and found that the genotype composition of C. acnes 16S rRNA is individual specific. Owner identification accuracy of around 90% based on random forest machine learning was achieved by using a combination of C. acnes 16S rRNA genotype and skin microbiome profile data. Furthermore, our study showed that the C. acnes 16S rRNA genotype remained more stable over time than the skin microbiome profile. This characteristic of C. acnes was further confirmed by the analysis of publicly available human skin metagenome data. Our approach, with its high precision, good reproducibility, and low costs, thus provides new possibilities in the field of microbiome-based owner identification and forensics in general.IMPORTANCE Cutibacterium acnes is the most common and abundant bacterial species on human skin, and the gene that encodes its 16S rRNA has multiple single-nucleotide polymorphisms. In this study, we developed a method to efficiently determine the C. acnes 16S rRNA genotype composition from microbial samples taken from the hands of participants and from their possessions. Using the C. acnes 16S rRNA genotype composition, we could predict the owner of a possession with around 90% accuracy when the 16S rRNA gene-based microbiome profile was included. We also showed that the C. acnes 16S rRNA genotype composition was more stable over time than the skin microbiome profile and thus is more suitable for owner identification.

Draft Genome Sequences of Enterococcus faecalis Strains Isolated from Healthy Japanese Individuals.

Enterococcus faecalis is a common commensal of the intestines of humans and other mammals but is also a frequent cause of serious ailments. Here, we report 14 draft genome sequences of strains of Enterococcus faecalis, a normal inhabitant and Gram-positive bacterium that was isolated from 7 healthy Japanese volunteers.

Complete Genome Sequence of Halomonas olivaria, a Moderately Halophilic Bacterium Isolated from Olive Processing Effluents, Obtained by Nanopore Sequencing.

Here, we report the complete genome sequence of Halomonas olivaria strain TYRC17, a moderately halophilic, Gram-negative bacterium that was isolated from olive processing effluents. The 5-Mbp genome consists of 7,375 protein-coding sequences, including a variety of genes involved in tyrosol metabolism, nitrate respiration, and the production of polysaccharides.

Gut microbiome-derived phenyl sulfate contributes to albuminuria in diabetic kidney disease.

Diabetic kidney disease is a major cause of renal failure that urgently necessitates a breakthrough in disease management. Here we show using untargeted metabolomics that levels of phenyl sulfate, a gut microbiota-derived metabolite, increase with the progression of diabetes in rats overexpressing human uremic toxin transporter SLCO4C1 in the kidney, and are decreased in rats with limited proteinuria. In experimental models of diabetes, phenyl sulfate administration induces albuminuria and podocyte damage. In a diabetic patient cohort, phenyl sulfate levels significantly correlate with basal and predicted 2-year progression of albuminuria in patients with microalbuminuria. Inhibition of tyrosine phenol-lyase, a bacterial enzyme responsible for the synthesis of phenol from dietary tyrosine before it is metabolized into phenyl sulfate in the liver, reduces albuminuria in diabetic mice. Together, our results suggest that phenyl sulfate contributes to albuminuria and could be used as a disease marker and future therapeutic target in diabetic kidney disease.

A Metabolomic-Based Evaluation of the Role of Commensal Microbiota throughout the Gastrointestinal Tract in Mice.

Commensal microbiota colonize the surface of our bodies. The inside of the gastrointestinal tract is one such surface that provides a habitat for them. The gastrointestinal tract is a long organ system comprising of various parts, and each part possesses various functions. It has been reported that the composition of intestinal luminal metabolites between the small and large intestine are different; however, comprehensive metabolomic and commensal microbiota profiles specific to each part of the gastrointestinal lumen remain obscure. In this study, by using capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS)-based metabolome and 16S rRNA gene-based microbiome analyses of specific pathogen-free (SPF) and germ-free (GF) murine gastrointestinal luminal profiles, we observed the different roles of commensal microbiota in each part of the gastrointestinal tract involved in carbohydrate metabolism and nutrient production. We found that the concentrations of most amino acids in the SPF small intestine were higher than those in the GF small intestine. Furthermore, sugar alcohols such as mannitol and sorbitol accumulated only in the GF large intestine, but not in the SPF large intestine. On the other hand, pentoses, such as arabinose and xylose, gradually accumulated from the cecum to the colon only in SPF mice, but were undetected in GF mice. Correlation network analysis between the gastrointestinal microbes and metabolites showed that niacin metabolism might be correlated to Methylobacteriaceae. Collectively, commensal microbiota partially affects the gastrointestinal luminal metabolite composition based on their metabolic dynamics, in cooperation with host digestion and absorption.

Intestinal barrier regulates immune responses in the liver via IL-10-producing macrophages.

The gut-liver axis is of clinical importance as a potential therapeutic target in a wide range of liver diseases; however, the mechanisms underlying interactions between microbial products and immune responses in the liver remain unknown. In this study, we demonstrated that IL-10-producing macrophages contribute to immune tolerance in the inflamed liver under intestinal barrier disruption in a murine tandem model of dextran sulfate sodium (DSS) colitis and concanavalin A (Con A) hepatitis. Intestinal barrier disruption protected mice from subsequent liver injury, and the severity of colitis directly affected susceptibility to such injury. The protective effect of DSS-Con A was canceled in gut-sterilized mice, suggesting that gut microbiota play a substantial role in this process. Altered gut microbiota and their metabolites, along with a disrupted intestinal barrier, directly gave rise to immunological permissiveness in the inflamed liver. We identified 1-methylnicotinamide (1-MNA) as a candidate metabolite capable of suppressing liver injury with the potential to induce IL-10-producing macrophages. Consistently, expression of nicotinamide N-methyltransferase, which converts nicotinamide to 1-MNA, was upregulated in the liver of DSS-Con A mice, and this effect was abrogated by gut sterilization. Collectively, our results provide a mechanistic insight into the regulation of immunological balance in the liver via the gut-liver axis.

Proton Pump Inhibitors Increase the Susceptibility of Mice to Oral Infection with Enteropathogenic Bacteria.

BACKGROUND AND AIMS: Proton pump inhibitors (PPIs) are among the most frequently prescribed medications. Side effects including an increased risk of intestinal infections have been reported. It is assumed that PPIs can increase susceptibility to enteropathogens; however, the underlying mechanisms are unknown. Here in this study, we explored whether Lansoprazole (Laz), one of the PPIs, increases the susceptibility to enteropathogens, and further investigated the mechanism of it. METHODS: Mice were administered Laz intraperitoneally once daily and orally infected with Citrobacter rodentium (C. rodentium). The establishment of intestinal infection was assessed by histology and inflammatory cytokine expression levels measured by quantitative PCR. To test whether Laz changes the intestinal environment to influence the susceptibility, intestinal pH, microbiota, metabolites and immune cell distributions were evaluated via pH measurement, 16S rRNA gene sequencing, metabolome, and flow cytometry analyses after Laz administration. RESULTS: Colitis was induced with less C. rodentium in Laz-treated mice as compared with the controls. We found that increased numbers of C. rodentium could reach the cecum following Laz administration. Laz increased pH in the stomach but not in the intestines. It induced dysbiosis and changed the metabolite content of the small intestine. However, these changes did not lead to alterations of immune cell distribution. CONCLUSIONS: Laz raised susceptibility to C. rodentium as increased numbers of the pathogen reach the site of infection. Our results suggest that it was due to increased stomach pH which allowed more peroral enteropathogens to pass the stomach, but not because of changes of intestinal environment.

Canagliflozin reduces plasma uremic toxins and alters the intestinal microbiota composition in a chronic kidney disease mouse model.

Accumulation of uremic toxins, which exert deleterious effects in chronic kidney disease, is influenced by the intestinal environment; the microbiota contributes to the production of representative uremic toxins, including p-cresyl sulfate and indoxyl sulfate. Canagliflozin is a sodium-glucose cotransporter (SGLT) 2 inhibitor, and it also exerts a modest inhibitory effect on SGLT1. The inhibition of intestinal SGLT1 can influence the gastrointestinal environment. We examined the effect of canagliflozin on the accumulation of uremic toxins in chronic kidney disease using adenine-induced renal failure mice. Two-week canagliflozin (10 mg/kg po) treatment did not influence the impaired renal function; however, it significantly reduced the plasma levels of p-cresyl sulfate and indoxyl sulfate in renal failure mice (a 75% and 26% reduction, respectively, compared with the vehicle group). Additionally, canagliflozin significantly increased cecal short-chain fatty acids in the mice, suggesting the promotion of bacterial carbohydrate fermentation in the intestine. Analysis of the cecal microbiota showed that canagliflozin significantly altered microbiota composition in the renal failure mice. These results indicate that canagliflozin exerts intestinal effects that reduce the accumulation of uremic toxins including p-cresyl sulfate. Reduction of accumulated uremic toxins by canagliflozin could provide a potential therapeutic option in chronic kidney disease.