Gut microbiota-derived short-chain fatty acids promote prostate cancer progression via inducing cancer cell autophagy and M2 macrophage polarization.

We have previously demonstrated abnormal gut microbial composition in castration-resistant prostate cancer (CRPC) patients, here we revealed the mechanism of gut microbiota-derived short-chain fatty acids (SCFAs) as a mediator linking CRPC microbiota dysbiosis and prostate cancer (PCa) progression. By using transgenic TRAMP mouse model, PCa patient samples, in vitro PCa cell transwell and macrophage recruitment assays, we examined the effects of CRPC fecal microbiota transplantation (FMT) and SCFAs on PCa progression. Our results showed that FMT with CRPC patients' fecal suspension increased SCFAs-producing gut microbiotas such as Ruminococcus, Alistipes, Phascolarctobaterium in TRAMP mice, and correspondingly raised their gut SCFAs (acetate and butyrate) levels. CRPC FMT or SCFAs supplementation significantly accelerated mice's PCa progression. In vitro, SCFAs enhanced PCa cells migration and invasion by inducing TLR3-triggered autophagy that further activated NF-κB and MAPK signalings. Meanwhile, autophagy of PCa cells released higher level of chemokine CCL20 that could reprogramme the tumor microenvironment by recruiting more macrophage infiltration and simultaneously polarizing them into M2 type, which in turn further strengthened PCa cells invasiveness. Finally in a cohort of 362 PCa patients, we demonstrated that CCL20 expression in prostate tissue was positively correlated with Gleason grade, pre-operative PSA, neural/seminal vesical invasion, and was negatively correlated with post-operative biochemical recurrence-free survival. Collectively, CRPC gut microbiota-derived SCFAs promoted PCa progression via inducing cancer cell autophagy and M2 macrophage polarization. CCL20 could become a biomarker for prediction of prognosis in PCa patients. Intervention of SCFAs-producing microbiotas may be a useful strategy in manipulation of CRPC.

Dietary Eicosapentaenoic Acid and Docosahexaenoic Acid Ethyl Esters Influence the Gut Microbiota and Bacterial Metabolites in Rats.

Dietary fish oil containing eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) has been reported to affect the diversity and composition of gut microbiota and bacterial metabolites. However, few reports have focused on the effects of EPA and DHA on gut microbiota diversity and bacterial metabolites. This study evaluated the effects of dietary EPA-ethyl ester (EE) and DHA-EE on steroid metabolism, gut microbiota, and bacterial metabolites in Wistar rats. Male rats were fed the experimental diets containing 5% (w/w) soybean oil-EE (SOY diet), EPA-EE (EPA diet), and DHA-EE (DHA diet) for four weeks. The lipid contents in the serum and liver, mRNA expression levels in the liver, and the diversity, composition, and metabolites of the gut microbiota were evaluated. The EPA and DHA diets decreased serum and liver cholesterol contents compared to the SOY diet. In addition, there were no significant changes in gene expression levels related to steroid metabolism in the liver between the EPA and DHA groups. Rats fed the DHA diet had lower microbiota diversity indices, such as Simpson and Shannon indices, than rats fed the SOY and EPA diets. In addition, rats fed EPA and DHA had significant differences in the relative abundance of microbiota at the genus level, such as Phascolarctobacterium, Turicibacter, and [Eubacterium]. Therefore, it was concluded that EPA and DHA have different effects on the diversity and composition of gut microbiota under the experimental conditions employed herein.

Exploring the signature gut and oral microbiome in individuals of specific Ayurveda prakriti.

Diagnosis and treatment of various diseases in Ayurveda, the Indian system of medicine, relies on 'prakriti' phenotyping of individuals into predominantly three constitutions, kapha, pitta and vata. Recent studies propose that microbiome play an integral role in precision medicine. A study of the relationship between prakriti - the basis of personalized medicine in Ayurveda and that of gut microbiome, and possible biomarker of an individual's health, would vastly improve precision therapy. Towards this, we analyzed bacterial metagenomes from buccal (oral microbiome) and fecal (gut microbiome) samples of 272 healthy individuals of various predominant prakritis. Major bacterial genera from gut microbiome included Prevotella, Bacteroides and Dialister while oral microbiome included Streptococcus, Neisseria, Veilonella, Haemophilus, Porphyromonas and Prevotella. Though the core microbiome was shared across all individuals, we found prakriti specific signatures such as preferential presence of Paraprevotella and Christensenellaceae in vata individuals. A comparison of core gut microbiome of each prakriti with a database of 'healthy' microbes identified microbes unique to each prakriti with functional roles similar to the physiological characteristics of various prakritis as described in Ayurveda. Our findings provide evidence to Ayurvedic interventions based on prakriti phenotyping and possible microbial biomarkers that can stratify the heterogenous population and aid in precision therapy.

Microbial interaction between the succinate-utilizing bacterium Phascolarctobacterium faecium and the gut commensal Bacteroides thetaiotaomicron.

A large variety of microbes are present in the human gut, some of which are considered to interact with each other. Most of these interactions involve bacterial metabolites. Phascolarctobacterium faecium hardly uses carbohydrates for growth and instead uses succinate as a substrate. This study investigated the growth behavior of the co-culture of the succinate-specific utilizer P. faecium and the succinogenic gut commensal Bacteroides thetaiotaomicron. Succinate production by B. thetaiotaomicron supported the growth of P. faecium and concomitant propionate production via the succinate pathway. The succinate produced was completely converted to propionate. This result was comparable with the monoculture of P. faecium in the medium supplemented with 1% (w/v) succinate. We analyzed the transcriptional response (RNA-Seq) between the mono- and co-culture of P. faecium and B. thetaiotaomicron. Comparison of the expression levels of genes of P. faecium between the mono- and co-cultured conditions highlighted that the genes putatively involved in the transportation of succinate were notably expressed under the co-cultured conditions. Differential expression analysis showed that the presence of P. faecium induced changes in the B. thetaiotaomicron transcriptional pattern, for example, expression changes in the genes for vitamin B12 transporters and reduced expression of glutamate-dependent acid resistance system-related genes. Also, transcriptome analysis of P. faecium suggested that glutamate and succinate might be used as sources of succinyl-CoA, an intermediate in the succinate pathway. This study revealed some survival strategies of asaccharolytic bacteria, such as Phascolarctobacterium spp., in the human gut.

Phascolarctobacterium wakonense sp. nov., isolated from common marmoset (Callithrix jacchus) faeces.

Two strictly anaerobic strains (MB11T and MB56) were isolated from common marmoset (Callithrixjacchus) faeces. Cells of the two strains were Gram-stain-negative, pleomorphic short (strain MB11T) or long (strain MB56) rods. Phylogenetic analysis based on 16S rRNA gene sequences revealed that both isolates were related to the genus Phascolarctobacterium. They had 16S rRNA gene sequences similarities lower than 93 % to previously described species, Phascolarctobacterium faecium ACM 3679T and Phascolarctobacterium succinatutens YIT 12067T, and 98.7 % between themselves. DNA-DNA hybridization values showed that strains MB11T and MB56 were the same species. The genomic DNA G+C content of strains MB11T and MB56 were 47.3-47.4 mol% and 47.7-48.0 mol%. The isolates had different enzymatic activities compared with P. succinatutens JCM 16074T and different major cellular fatty acids compared with P. faecium ACM 3679T. Substrate availability revealed that they utilized not only succinate, but also pyruvate. With pyruvate supplementation, they produced both propionate and acetate, while only propionate production occured with succinate. As suggested by the phylogenic and physiological properties of strains MB11T and MB56, we propose the name Phascolarctobacteriumwakonense sp. nov. with the type strain MB11T (=JCM 32899T=DSM 107697T).

Effects of partial replacement of maize in the diet with crude glycerin and/or soyabean oil on ruminal fermentation and microbial population in Nellore steers.

The objective of this study was to determine whether a combination of crude glycerin (CG) and soyabean oil (SO) could be used to partially replace maize in the diet of Nellore steers while maintaining optimum feed utilisation. Eight castrated Nellore steers fitted with ruminal and duodenal cannulas were used in a double 4×4 Latin square design balanced for residual effects, in a factorial arrangement (A×B), when factor A corresponded to the provision of SO, and factor B to the provision of CG. Steers feed SO and CG showed similar DM intake, DM, organic matter and neutral-detergent fibre digestibility to that of steers fed diets without oil and without glycerine (P>0·05). Both diets with CG additions reduced the acetate:propionate ratio and increased the proportion of iso-butyrate, butyrate, iso-valerate and valerate (P<0·05). Steers fed diets containing SO had less total N excretion (P<0·001) and showed greater retained N expressed as % N intake (P=0·022). SO and CG diet generated a greater ruminal abundance of Prevotella, Succinivibrio, Ruminococcus, Syntrophococcus and Succiniclasticum. Archaea abundance (P=0·002) and total ciliate protozoa were less in steers fed diets containing SO (P=0·011). CG associated with lipids could be an energy source, which is a useful strategy for the partial replacement of maize in cattle diets, could result in reduced total N excretion and ruminal methanogens without affecting intake and digestibility.

Host-microbiome interactions in human type 2 diabetes following prebiotic fibre (galacto-oligosaccharide) intake.

Aberrant microbiota composition and function have been linked to several pathologies, including type 2 diabetes. In animal models, prebiotics induce favourable changes in the intestinal microbiota, intestinal permeability (IP) and endotoxaemia, which are linked to concurrent improvement in glucose tolerance. This is the first study to investigate the link between IP, glucose tolerance and intestinal bacteria in human type 2 diabetes. In all, twenty-nine men with well-controlled type 2 diabetes were randomised to a prebiotic (galacto-oligosaccharide mixture) or placebo (maltodextrin) supplement (5·5 g/d for 12 weeks). Intestinal microbial community structure, IP, endotoxaemia, inflammatory markers and glucose tolerance were assessed at baseline and post intervention. IP was estimated by the urinary recovery of oral 51Cr-EDTA and glucose tolerance by insulin-modified intravenous glucose tolerance test. Intestinal microbial community analysis was performed by high-throughput next-generation sequencing of 16S rRNA amplicons and quantitative PCR. Prebiotic fibre supplementation had no significant effects on clinical outcomes or bacterial abundances compared with placebo; however, changes in the bacterial family Veillonellaceae correlated inversely with changes in glucose response and IL-6 levels (r -0·90, P=0·042 for both) following prebiotic intake. The absence of significant changes to the microbial community structure at a prebiotic dosage/length of supplementation shown to be effective in healthy individuals is an important finding. We propose that concurrent metformin treatment and the high heterogeneity of human type 2 diabetes may have played a significant role. The current study does not provide evidence for the role of prebiotics in the treatment of type 2 diabetes.

Interactions between oil substrates and glucose on pure cultures of ruminal lipase-producing bacteria.

The hydrolysis of free fatty acids from lipids is a prerequisite for biohydrogenation, a process that effectively saturates free fatty acids. Anaerovibrio lipolyticus 5s and Butyrivibrio fibrisolvens have long been thought to be the major contributors to ruminal lipolysis; however, Propionibacterium avidum and acnes recently have been identified as contributing lipase activity in the rumen. In order to further characterize the lipase activity of these bacterial populations, each was grown with three different lipid substrates, olive oil, corn oil, and flaxseed oil (3 %). Because different finishing rations contain varying levels of glycogen (a source of free glucose) this study also documented the effects of glucose on lipolysis. P. avidum and A. lipolyticus 5s demonstrated the most rapid rates (P < 0.05) of lipolysis for cultures grown with olive oil and flaxseed oil, respectively. A. lipolyticus, B. fibrisolvens, and P. avidum more effectively hydrolyzed flaxseed oil than olive oil or corn oil, especially in the presence of 0.02 % glucose. Conversely, P. acnes hydrolyzed corn oil more readily than olive oil or flaxseed oil and glucose had no effect on lipolytic rate. Thus, these bacterial species demonstrated different specificities for oil substrates and different sensitivities to glucose.

Draft genome sequences for two metal-reducing Pelosinus fermentans strains isolated from a Cr(VI)-contaminated site and for type strain R7.

Pelosinus fermentans 16S rRNA gene sequences have been reported from diverse geographical sites since the recent isolation of the type strain. We present the genome sequence of the P. fermentans type strain R7 (DSM 17108) and genome sequences for two new strains with different abilities to reduce iron, chromate, and uranium.

Microbial community succession during lactate amendment and electron acceptor limitation reveals a predominance of metal-reducing Pelosinus spp.

The determination of the success of in situ bioremediation strategies is complex. By using controlled laboratory conditions, the influence of individual variables, such as U(VI), Cr(VI), and electron donors and acceptors on community structure, dynamics, and the metal-reducing potential can be studied. Triplicate anaerobic, continuous-flow reactors were inoculated with Cr(VI)-contaminated groundwater from the Hanford, WA, 100-H area, amended with lactate, and incubated for 95 days to obtain stable, enriched communities. The reactors were kept anaerobic with N(2) gas (9 ml/min) flushing the headspace and were fed a defined medium amended with 30 mM lactate and 0.05 mM sulfate with a 48-h generation time. The resultant diversity decreased from 63 genera within 12 phyla to 11 bacterial genera (from 3 phyla) and 2 archaeal genera (from 1 phylum). Final communities were dominated by Pelosinus spp. and to a lesser degree, Acetobacterium spp., with low levels of other organisms, including methanogens. Four new strains of Pelosinus were isolated, with 3 strains being capable of Cr(VI) reduction while one also reduced U(VI). Under limited sulfate, it appeared that the sulfate reducers, including Desulfovibrio spp., were outcompeted. These results suggest that during times of electron acceptor limitation in situ, organisms such as Pelosinus spp. may outcompete the more-well-studied organisms while maintaining overall metal reduction rates and extents. Finally, lab-scale simulations can test new strategies on a smaller scale while facilitating community member isolation, so that a deeper understanding of community metabolism can be revealed.

Effects of freeze-dried Mitsuokella jalaludinii culture and Natuphos(®) phytase supplementation on the performance and nutrient utilisation of broiler chickens.

BACKGROUND: Phytate-bound phosphorus (P) in poultry diets is poorly available to chickens. Hence exogenous phytase is often added to their diets. Mitsuokella jalaludinii is a rumen bacterial species that produces high phytase activity. In this study the effects of freeze-dried active M. jalaludinii culture (FD-AMJC) and Natuphos(®) phytase (phytase N) supplementations on the growth performance and nutrient utilisation of broiler chickens fed a low-available P (aP) diet were evaluated. RESULTS: Supplementation of FD-AMJC or phytase N to the low-aP diet improved the feed intake, feed conversion rate, body weight gain, dry matter (DM) digestibility and P, Ca and Mn retention, increased the tibia bone ash content, Ca and P concentrations in tibia DM and P and Zn concentrations in plasma and reduced the P excretion of broiler chickens. However, the feed conversion rate, P and Ca retention, DM digestibility and reduction of P excretion were better with FD-AMJC than phytase N supplementation. Supplementation of FD-AMJC to the low-aP diet also improved the apparent metabolisable energy value of the diet, Cu and Zn retention and crude protein digestibility, but phytase N supplementation did not. CONCLUSION: FD-AMJC supplementation was more efficient in improving nutrient utilisation and reducing P excretion in chickens than phytase N supplementation.

Characterization of Phascolarctobacterium succinatutens sp. nov., an asaccharolytic, succinate-utilizing bacterium isolated from human feces.

Isolation, cultivation, and characterization of the intestinal microorganisms are important for understanding the comprehensive physiology of the human gastrointestinal (GI) tract microbiota. Here, we isolated two novel bacterial strains, YIT 12067(T) and YIT 12068, from the feces of healthy human adults. Phylogenetic analysis indicated that they belonged to the same species and were most closely related to Phascolarctobacterium faecium ACM 3679(T), with 91.4% to 91.5% 16S rRNA gene sequence similarities, respectively. Substrate availability tests revealed that the isolates used only succinate; they did not ferment any other short-chain fatty acids or carbohydrates tested. When these strains were cocultured with the xylan-utilizing and succinate-producing bacterium Paraprevotella xylaniphila YIT 11841(T), in medium supplemented with xylan but not succinate, their cell numbers became 2 to 3 orders of magnitude higher than those of the monoculture; succinate became undetectable, and propionate was formed. Database analysis revealed that over 200 uncultured bacterial clones from the feces of humans and other mammals showed high sequence identity (>98.7%) to YIT 12067(T). Real-time PCR analysis also revealed that YIT 12067(T)-like bacteria were present in 21% of human fecal samples, at an average level of 3.34 × 10(8) cells/g feces. These results indicate that YIT 12067(T)-like bacteria are distributed broadly in the GI tract as subdominant members that may adapt to the intestinal environment by specializing to utilize the succinate generated by other bacterial species. The phylogenetic and physiological properties of YIT 12067(T) and YIT 12068 suggest that these strains represent a novel species, which we have designated Phascolarctobacterium succinatutens sp. nov.

Desulfosporomusa polytropa gen. nov., sp. nov., a novel sulfate-reducing bacterium from sediments of an oligotrophic lake.

Five strains of sulfate-reducing bacteria were isolated from the highest positive dilutions of a most probable number (MPN) series supplemented with lactate and inoculated with sediments from the oligotrophic Lake Stechlin. The isolates were endospore-forming and were motile by means of laterally inserted flagella. They stained Gram-negative and contained b-type cytochromes. CO difference spectra indicated the presence of P582 as a sulfite reductase. Phylogenetic analyses of the 16S rDNA sequences revealed that the isolates were very closely affiliated with the genus Sporomusa. However, sulfate and amorphous Fe(OH)(3), but not sulfite, elemental sulfur, MnO(2), or nitrate were used as terminal electron acceptors. Homoacetogenic growth was found with H(2)/CO(2) gas mixture, formate, methanol, ethanol, and methoxylated aromatic compounds. The strains grew autotrophically with H(2) plus CO(2) in the presence or absence of sulfate. Formate, butyrate, several alcohols, organic acids, carbohydrates, some amino acids, choline, and betaine were also utilized as substrates. The growth yield with lactate and sulfate as substrate was 7.0 g dry mass/mol lactate and thus two times higher than in sulfate-free fermenting cultures. All isolates were able to grow in a temperature range of 4-37 degrees C. Physiologically and by the presence of a Gram-negative cell wall, the new isolates resemble known Desulfosporosinus species. However, phylogenetically they are affiliated with the Gram-negative genus Sporomusa belonging to the Selenomonas subgroup of the Firmicutes. Therefore, the new isolates reveal a new phylogenetic lineage of sulfate-reducing bacteria. A new genus and species, Desulfosporomusa polytropa gen. nov., sp. nov. is proposed.

Production of 2-aminobutyrate by Megasphaera elsdenii.

Production of the amino acid 2-aminobutyrate was studied in four strains of Megasphaera elsdenii grown on a lactate-based growth medium containing Bacto-casamino acids and yeast extract. Supplementation with threonine increased the production of 2-aminobutyrate in three of the four strains, but no substantial increase in production was noted with serine, methionine, or aspartate, all of which are potential sources for the precursor of 2-aminobutyrate, 2-oxobutyrate. L-Cycloserine, an inhibitor of alanine transaminases, decreased both alanine and 2-aminobutyrate production, suggesting that 2-aminobutyrate synthesis may share the same metabolic pathway as alanine synthesis or that 2-oxobutyrate can act as a substrate for alanine transaminases. Decreases in the production of 2-aminobutyrate were associated with a reduction in the catabolism of branched-chain amino acids in two of the four strains.

Inhibition of the autoxidation of ascorbate and norepinephrine by extracts of Clostridium butyricum, Megasphaera elsdenii and Escherichia coli.

The autoxidation of ascorbate and of norepinephrine in Krebs Ringer phosphate medium, pH 7.4, was studied. The autoxidation of the two substances was determined spectrophotometrically at 265 and 480 nm respectively. The effect of dialyzed extracts (m.w. greater than 12,000) from Escherichia coli (aerobe), Megasphaera elsdenii, and Clostridium butyricum (obligate anaerobes) was examined and compared to similarly prepared extracts from rat serum and cerebral cortex. The assay medium contained cellular components diluted 10(3)-10(6)-fold. Up to 10(4)-fold dilution there was a substantial reduction in the rate of both autoxidation reactions, but the preparations from M. elsdenii and C. butyricum were conspicuously less effective. After 5 min heat treatment at 100 degrees C the anaerobic preparations produced less than 20% inhibition, while the activity of the other preparations remained unchanged at 75-95% inhibition. These and earlier experiments involving additional mammalian species (Mishra and Kovachich, Neurosci. Lett., 43: 103-108, 1983) and plants (Mishra and Kovachich, Life Sci., 34: 2207-2212, 1984) suggest that a high level of heat-stable antioxidant activity in one or both of these autoxidation tests (denatured plant extracts only inhibit ascorbate autoxidation) is a general characteristic of organisms that thrive in oxygen-rich atmosphere.