Gut microbiome composition and metabolomic profiles of wild western lowland gorillas (Gorilla gorilla gorilla) reflect host ecology.

The metabolic activities of gut microbes significantly influence host physiology; thus, characterizing the forces that modulate this micro-ecosystem is key to understanding mammalian biology and fitness. To investigate the gut microbiome of wild primates and determine how these microbial communities respond to the host's external environment, we characterized faecal bacterial communities and, for the first time, gut metabolomes of four wild lowland gorilla groups in the Dzanga-Sangha Protected Areas, Central African Republic. Results show that geographical range may be an important modulator of the gut microbiomes and metabolomes of these gorilla groups. Distinctions seemed to relate to feeding behaviour, implying energy harvest through increased fruit consumption or fermentation of highly fibrous foods. These observations were supported by differential abundance of metabolites and bacterial taxa associated with the metabolism of cellulose, phenolics, organic acids, simple sugars, lipids and sterols between gorillas occupying different geographical ranges. Additionally, the gut microbiomes of a gorilla group under increased anthropogenic pressure could always be distinguished from that of all other groups. By characterizing the interplay between environment, behaviour, diet and symbiotic gut microbes, we present an alternative perspective on primate ecology and on the forces that shape the gut microbiomes of wild primates from an evolutionary context.

Intestinal bacterial population of healthy rats during the administration of chitosan and chitooligosaccharides.

The effect of the administration of chitosan (CS) and chitooligosaccharides (COS) on rat fecal microbiota was analyzed in this study. The profile of total bacterial population was monitored during 3 weeks of CS or COS application using denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene amplicons. Quantitative PCR was used for monitoring possible changes in the levels of total bacteria and the levels of individual bacterial groups: Bifidobacteria, Clostridium leptum, Enterobacteriaceae, Lactobacillus-Streptococcus-Enterobacter, and Bacteroides-Prevotella. The DGGE profiles revealed a high complexity and individuality of each tested subject, and variations in the composition of band pattern were observed. CS or COS per os administration changed the profile and structure of the microbial ecosystem of the gastrointestinal tract of healthy rats. COS have, in most cases, an opposite effect compared with CS; only the Bacteroides-Prevotella bacterial group and Enterobacteriaceae were influenced in the same way. The Bifidobacteria group was not influenced by the administration CS and COS.

Excretome of the chitinolytic bacterium Clostridium paraputrificum J4.

A strictly anaerobic mesophilic chitinolytic bacterial strain identified as Clostridium paraputrificum J4 was isolated from human feces. In response to various types of growth substrates, the bacterium produced an array of chitinolytic enzymes representing significant components of the J4 strain secretome. The excreted active proteins were characterized by estimating the enzymatic activities of endochitinase, exochitinase, and N-acetylglucosaminidase induced by cultivation in medium M-10 with colloidal chitin. The enzyme activities produced by J4 strain cultivated in medium M-10 with glucose were significantly lower. The spectrum of extracellularly excreted proteins was separated by SDS-PAGE. The chitinase variability was confirmed on zymograms of renatured SDS-PAGE. The enzymes were visualized under ultraviolet light by using 4-methylumbelliferyl derivatives of N-acetyl-ß-D: -glucosaminide, N,N´-diacetyl-ß-D: -chitobiose, or N,N´,N˝-triacetyl-ß-D: -chitotriose for ß-N-acetylglucosaminidase, chitobiosidase, or endochitinase activities, respectively. Protein components of the secretome were separated by 2D-PAGE analysis. The distinct protein bands were excised, isolated, and subsequently characterized by using MALDI-TOF/TOF tandem mass spectrometry. The final identification was performed according to sequence homology by database searching.

The antimicrobial action of chitosan, low molar mass chitosan, and chitooligosaccharides on human colonic bacteria.

Antibacterial effect of chitooligosaccharides (COS) and low molar mass chitosans (LMWC) is considered as one of the most important characteristics of chitosan (CS) hydrolysates. Here, we show the in vitro effect of different COS, LMWC, and CS on representative anaerobic bacteria isolated from human colon as a possibility of targeting modification of colonic microflora composition by supplementation of dietary CS products by humans. Specific growth rate of seven selected nonpathogenic anaerobic bacterial strains (Clostridium paraputrificum, Clostridium beijerinckii, Roseburia intestinalis, Bacteroides vulgatus, Bacteriodes thetaiotaomicron, Faecalibacterium prausnitzii and Blautia coccoides) was determined in the presence of 0.25 and 0.5% COS (2, 3, and 6 kDa), 0.025 and 0.05% of LMWC (10 and 16 kDa), and 0.025 and 0.1% of CS in vitro. The growth rate decreased in all strains in the presence of COS and LMWC in higher concentrations in comparison to control incubations. A relatively higher resistance to CS hydrolyzates was detected in R. intestinalis and F. prausnitzii, and more susceptible were bacteria belonging to Bacteoides sp. and Clostridium sp. The antimicrobial activity, minimum inhibitory concentrations (MIC), and minimal bactericidal concentrations (MBC) were determined. The antimicrobial activity increased with the degree of polymerization (DP). MIC ranged from 0.25 to 4.5% in dependence on bacterial strain and DP of CS/LMWC. MBC also decreased with DP. The most effective antimicrobial action was detected in LMWC with 16 kDa and CS. Weak antimicrobial activity was found in COS with small molecules (2 and 3 kDa).

Chitinolytic enzymes from bacterium inhabiting human gastrointestinal tract -- critical parameters of protein isolation from anaerobic culture.

The object of this study are chitinolytic enzymes produced by bacterium Clostridium paraputrificum J4 isolated from the gastrointestinal tract of a healthy human. In particular, we focus on the development of purification protocols, determination of properties of the enzymes and their activity profiles. The process of bacteria cultivation and isolation of chitinolytic complex of enzymes showing specific activities of endo-, exo-chitinase and N-acetyl-ß-glucosaminidase was optimized. A range of various purification procedures were used such as ultrafiltration, precipitation, chromatographic separations (ion-exchange, size exclusion, chromatofocusing) in altered combinations. The optimal purification protocol comprises two or three steps. Individual samples were analyzed by SDS/PAGE electrophoresis and after renaturation their activity could be detected using zymograms. Mass spectroscopy peptide fragment analysis and MALDI analysis of the purest samples indicate presence of endochitinase B (molecular mass about 85 kDa) and of 60-kDa endo- and exochitinases.

Relation between phylogenetic position, lipid metabolism and butyrate production by different Butyrivibrio-like bacteria from the rumen.

The Butyrivibrio group comprises Butyrivibrio fibrisolvens and related Gram-positive bacteria isolated mainly from the rumen of cattle and sheep. The aim of this study was to investigate phenotypic characteristics that discriminate between different phylotypes. The phylogenetic position, derived from 16S rDNA sequence data, of 45 isolates from different species and different countries was compared with their fermentation products, mechanism of butyrate formation, lipid metabolism and sensitivity to growth inhibition by linoleic acid (LA). Three clear sub-groups were evident, both phylogenetically and metabolically. Group VA1 typified most Butyrivibrio and Pseudobutyrivibrio isolates, while Groups VA2 and SA comprised Butyrivibrio hungatei and Clostridium proteoclasticum, respectively. All produced butyrate but strains of group VA1 had a butyrate kinase activity <40 U (mg protein)(-1), while strains in groups VA2 and SA all exhibited activities >600 U (mg protein)(-1). The butyrate kinase gene was present in all VA2 and SA bacteria tested but not in strains of group VA1, all of which were positive for the butyryl-CoA CoA-transferase gene. None of the bacteria tested possessed both genes. Lipase activity, measured by tributyrin hydrolysis, was high in group VA2 and SA strains and low in Group VA1 strains. Only the SA group formed stearic acid from LA. Linoleate isomerase activity, on the other hand, did not correspond with phylogenetic position. Group VA1 bacteria all grew in the presence of 200 microg LA ml(-1), while members of Groups VA2 and SA were inhibited by lower concentrations, some as low as 5 microg ml(-1). This information provides strong links between phenotypic and phylogenetic properties of this group of clostridial cluster XIVa Gram-positive bacteria.