Molecular analysis of the bacterial community in digestive tract of rabbit

This work aimed to study the stability over time of the bacterial community in cæcum and fæces of the rabbit (diversity index and structure) without experimental disturbance and to evaluate its relationships with environmental parameters. Soft and hard fæces of 14 rabbits were sampled for 5 weeks while cæcal content was sampled on the 3rd week (by surgery) and the 5th week (at slaughter). Bacterial communities were assessed by studying CE-SSCP profiles of 16S rRNA genes fragments. Redox potential, pH, NH3-N concentration and volatile fatty acid concentrations were measured in the cæcum. Data showed that bacterial communities of soft and hard fæces barely differed from that of the cæcum (ANOSIM-R < 0.25; p < 0.05). Without disturbance, the bacterial communities of fæces were stable over time (ANOSIM-R < 0.25; p < 0.001). However, the bacterial communities of cæcum and fæces were affected by the surgery (ANOSIM-R = 0.22–0.33; p < 0.001). The cæcal content was an acidic (pH = 6.03 ± 0.33) and an anaerobic environment (redox potential = ?160 ± 43 mV). Only the redox potential was correlated with the diversity index of the bacterial community of the cæcum (R² = 0.35; p < 0.05) and no environmental parameters were correlated to its structure.     

Probiotics shown to change bacterial community structure in the avian gastrointestinal tract.

Culturing and molecular techniques were used to monitor changes in the bacterial flora of the avian gastrointestinal (GI) tract following introduction of genetically modified (GM) and unmodified probiotics. Community hybridization of amplified 16S ribosomal DNA demonstrated that the bacterial flora of the GI tract changed significantly in response to the probiotic treatments. The changes were not detected by culturing. Although both GM and non-GM strains of Enterococcus faecium NCIMB 11508 changed the bacterial flora of the chicken GI tract, they did so differently. Probing the community DNA with an Enterococcus faecalis-specific probe showed that the relative amount of E. faecalis in the total eubacterial population increased in the presence of the non-GM strain and decreased in the presence of the GM probiotic compared with the results obtained with an untreated control group.     

Metagenomic bacterial community profiles of chicken embryo gastrointestinal tract by using T-RFLP analysis

Thirty microbial phylotypes of microorganisms were found in the gastrointestinal tract of chicken belonging to the Hajseks White breed, and 38 phylotypes were found in the gastrointestinal tract of chicken belonging to the Hajseks Brown breed. The microbiome of the gastrointestinal tract of the chicken embryos of the Hajseks White breed was dominated by the typical representatives of avian intestinal microflora—bacteria of the family Enterobacteriaceae (47.3%), orders Actinomycetales (13.6%) and Bifidobacteriales (20.6%), and the family Lachnospiraceae (1.1%). The microbiome of the gastrointestinal tract of the chicken embryos of the Hajseks Brown breed was dominated by the pathogenic bacteria of the order Rickettsiales (94.8%). The metagenome of gastrointestinal tract of both breeds also contained a small number of genes of unidentified bacteria.     

Postnatal development of bacterial population in the gastrointestinal tract of calves

Bacterial 16S rDNA from fecal samples of two calves were amplified by PCR and analyzed by denaturing gradient gel electrophoresis; selected bands were sequenced. Escherichia coli and Bifidobacterium animalis were the initial colonizers, followed by species closely related to the genera Bacteroides, Clostridium and Faecalibacterium. Change of diet was connected with shifts of bacterial population and with the occurrence of many bacterial species that have not been cultured up to now. The diet change corresponded with an alteration in a volatile-fatty-acid concentration in fecal samples.     

Enumeration of anaerobic bacterial microflora of the equine gastrointestinal tract

Samples from the duodenum, jejunum, and ileum, as well as from the cecum and colon, were obtained from 11 mature grass-fed horses. Viable counts of total culturable and proteolytic bacteria were made on habitat-simulating media containing 40% clarified ruminal fluid. The mean pHs in the duodenum, jejunum, and ileum were 6.32, 7.10, and 7.47, respectively; the mean pH decreased to 6.7 in the hindgut. The acetate concentration increased along the length of the small intestine and was the only volatile fatty acid present in this gut segment. Molar proportions of acetate, propionate, and butyrate in the hindgut were 85:10:3. Differences in bacterial counts on habitat-simulating media containing equine cecal fluid or clarified ruminal fluid were negligible. Bacterial counts showed a substantial population in the duodenum (ca. 2.9 x 10(6) per g [wet weight] of sample), and this increased to 29.0 x 10(6) in the jejunum and 38.4 x 10(6) in the ileum. Proteolytic bacteria formed a high proportion of the total culturable bacteria, especially in duodenal samples. Counts of proteolytic bacteria per gram (wet weight) of sample were 3.0 x 10(6), 15.6 x 10(6), and 22.0 x 10(6) in the duodenum, jejunum, and ileum, respectively. There was a close relationship between lumenal and mucosal bacterial counts, although actual values were lower in mucosal samples. The mucosal bacterial population in the duodenum was high relative to the lumenal population. Although the comparison of bacterial populations in the hindgut of the horse and white rhino was limited to a single animal, the results were of interest. Counts were higher in the cecum than in the colon for both the horse and the white rhino.(ABSTRACT TRUNCATED AT 250 WORDS)     

胃肠道微生物及其分子生态学技术研究进展

由于与人类健康和疾病密切相关,胃肠道微生物研究已成为当今的热点研究领域。目前研究的分子手段已经从单纯的分析微生物群落结构组成,发展到通过宏转录组学、宏蛋白组学和代谢组学等"组学"技术揭示胃肠道微生物群落的相应功能。本文结合我们的研究工作,综述了国内外胃肠道微生物及其分子生态学技术的研究进展。     

Enumeration of anaerobic bacterial microflora of the equine gastrointestinal tract.

Samples from the duodenum, jejunum, and ileum, as well as from the cecum and colon, were obtained from 11 mature grass-fed horses. Viable counts of total culturable and proteolytic bacteria were made on habitat-simulating media containing 40% clarified ruminal fluid. The mean pHs in the duodenum, jejunum, and ileum were 6.32, 7.10, and 7.47, respectively; the mean pH decreased to 6.7 in the hindgut. The acetate concentration increased along the length of the small intestine and was the only volatile fatty acid present in this gut segment. Molar proportions of acetate, propionate, and butyrate in the hindgut were 85:10:3. Differences in bacterial counts on habitat-simulating media containing equine cecal fluid or clarified ruminal fluid were negligible. Bacterial counts showed a substantial population in the duodenum (ca. 2.9 x 10(6) per g [wet weight] of sample), and this increased to 29.0 x 10(6) in the jejunum and 38.4 x 10(6) in the ileum. Proteolytic bacteria formed a high proportion of the total culturable bacteria, especially in duodenal samples. Counts of proteolytic bacteria per gram (wet weight) of sample were 3.0 x 10(6), 15.6 x 10(6), and 22.0 x 10(6) in the duodenum, jejunum, and ileum, respectively. There was a close relationship between lumenal and mucosal bacterial counts, although actual values were lower in mucosal samples. The mucosal bacterial population in the duodenum was high relative to the lumenal population. Although the comparison of bacterial populations in the hindgut of the horse and white rhino was limited to a single animal, the results were of interest. Counts were higher in the cecum than in the colon for both the horse and the white rhino.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of RNA from bacterial samples of the human gastrointestinal tract

The human gastrointestinal (GI) tract contains a complex microbial community that consists of numerous uncultured microbes. Therefore, nucleic-acid-based approaches have been introduced to study microbial diversity and activity, and these depend on the proper isolation of DNA, rRNA and mRNA. Here, we present an RNA isolation protocol that is suitable for a wide variety of GI tract samples. The procedure for isolating DNA from GI tract samples is described in another Nature Protocols article. One of the benefits of our RNA isolation protocol is that sampling can be performed outside the laboratory, which offers possibilities for implementation in large intervention studies. The RNA isolation is based on mechanical disruption, followed by isolation of nucleic acids using phenol:chloroform:isoamylalcohol extraction and removal of DNA. In our laboratory, this protocol has resulted in the isolation of rRNA and mRNA of sufficient quality and quantity for microbial diversity and activity studies. Depending on the number of samples, the sample type and the quenching procedure chosen, the whole procedure can be performed within 2.5-4 h.     

Isolation of DNA from bacterial samples of the human gastrointestinal tract

The human gastrointestinal (GI) tract contains a complex microbial community that develops in time and space. The most widely used approaches to study microbial diversity and activity are all based on the analysis of nucleic acids, DNA, rRNA and mRNA. Here, we present a DNA isolation protocol that is suitable for a wide variety of GI tract samples, including biopsies with minute amounts of material. The protocol is set up in such a way that sampling can be performed outside the laboratory, which offers possibilities for implementation in large intervention studies. The DNA isolation is based on mechanical disruption, followed by isolation of nucleic acids using phenol:chloroform:isoamylalcohol extraction. In addition, it includes an alternative DNA isolation protocol that is based on a commercial kit. These protocols have all been successfully used in our laboratory, resulting in isolation of DNA of sufficient quality for microbial diversity studies. Depending on the number of samples and sample type, the whole procedure will take approximately 2.5-4 hours.     

Assessment of mouse strain on bacterial translocation from the gastrointestinal tract

The translocation of indigenous bacteria from the gastrointestinal tract to the mesenteric lymphnodes was compared in ten strains of mice. Indigenous Escherichia coli were cultured from the mesenteric lymphnodes of only two of the six mouse strains examined. Thus, spontaneous translocation of indigenous enteric bacteria across the intestinal barrier did not occur to any significant extent in any of the mouse strains examined. Since bacterial overgrowth in the gastrointestinal tract promotes bacterial translocation, bacterial translocation was tested in ten mouse strains including B10 series after antibiotic-decontaminated and subsequent colonization with streptomycin-resistant E. coli C25. E. coli C25 populated the ceca of the mice at levels of 10(8) to 10(9) per gram and translocated to 90-100% of the mesenteric lymphnodes with mean of 10(1.13) to 10(1.86) per mesenteric lymphnode. However, there were no significant differences between mouse strains as to the translocation incidence or the numbers of viable E. coli C25 per mesenteric lymphnode. Thus, genetic differences between mouse strains did not influence bacterial translocation from the gastrointestinal tract to the mesenteric lymphnodes.     

Indigenous bacteria and bacterial metabolic products in the gastrointestinal tract of broiler chickens

The gastrointestinal tract is a dynamic ecosystem containing a complex microbial community. In this paper, the indigenous intestinal bacteria and the microbial fermentation profile particularly short chain fatty acids (SCFA), lactate, and ammonia concentrations are reviewed. The intestinal bacterial composition changes with age. The bacterial density of the small intestine increases with age and comprises of lactobacilli, streptococci, enterobacteria, fusobacteria and eubacteria. Strict anaerobes (anaerobic gram-positive cocci, Eubacterium spp., Clostridium spp., Lactobacillus spp., Fusobacterium spp. and Bacteroides) are predominating caecal bacteria in young broilers. Data from culture-based studies showed that bifidobacteria could not be isolated from young birds, but were recovered from four-week-old broilers. Caecal lactobacilli accounted for 1.5-24% of the caecal bacteria. Gene sequencing of caecal DNA extracts showed that the majority of bacteria belonged to Clostridiaceae. Intestinal bacterial community is influenced by the dietary ingredients, nutrient levels and physical structure of feed. SCFA and other metabolic products are affected by diet formulation and age. Additional studies are required to know the bacterial metabolic activities together with the community analysis of the intestinal bacteria. Feed composition and processing have great potential to influence the activities of intestinal bacteria towards a desired direction in order to support animal health, well-being and microbial safety of broiler meat.     

Effect of age on bacterial translocation from the gastrointestinal tract in mice.

To clarify the effects of age on bacterial translocation from the gastrointestinal tract, mice at the age of 1, 2, 4, 6, 12, and 15 months were antibiotic-decontaminated for 4 days and then inoculated orally with streptomycin-resistant Escherichia coli C25. Mice treated with cyclophosphamide and untreated controls were tested for bacterial translocation to the mesenteric lymph nodes (MLN) 2 days later. The population levels of E. coli C25 in cyclophosphamide-treated and untreated mice were approximately 10(9.3) and 10(9.5) per gram of cecum, respectively, at each tested age. There were no significant differences in the incidence of translocation of E. coli C25 to MLN at any of the tested ages, whereas the number of E. coli C25 detected in MLN was higher in young mice than in aged mice in both the cyclophosphamide-treated and untreated groups. These findings suggest that bacterial translocation from the GI tract may be a more important problem in young animals than in aged animals.     

Spatial and temporal variations of the bacterial community in the bovine digestive tract

Aims:  Improved knowledge of the bacterial community of the digestive tract is required to enhance the efficiency of digestion in herbivores. This work aimed to study spatial and temporal variations of the bacterial communities in the bovine digestive tract and their correlation with gut environmental parameters.
Methods and Results:  Rumen content and faeces of five cows were sampled for 3 weeks. In addition, reticulum content was sampled during the third week. Bacterial communities were assessed by studying capillary electrophoresis single-stranded conformation polymorphism (CE-SSCP) profiles of 16S rRNA genes. The bacterial community structure differed between the forestomach and faecal contents. The abundance of several operational taxonomic units changed from week to week. Bacterial community structure of the rumen was correlated to propionic acid and NH₃–N concentrations.
Conclusions:  The bacterial community of the bovine digestive tract varied in space and time.
Significance and Impact of the Study:  The study of the bacterial communities of the digestive tract in herbivores should be widened from the rumen to the large intestine. The amplitude and origin of the temporal variation of the ruminal bacterial community need to be better understood to improve the control of the fermentative activity in herbivores.     

DNA-based monitoring of total bacterial community structure in environmental samples

Determining the structure of bacterial communities and their response to stimuli is key to understanding community function and the interactions that occur between microorganisms and the environment. However, bacterial communities often comprise complex assemblages of large numbers of different bacterial populations. An approach is presented which allows bacterial community structure to be determined by fractionation of the complex mixture of total bacterial community DNA using the DNA-binding dye bisbenzimidazole which imposes G + C-dependent changes in the buoyant density of DNA. Bacterial community structure presented as percentage of total DNA vs. percentage G + C content of DNA is an indication of the relative abundance of phylogenetic groups of bacteria. Changes in the composition of a soil bacterial community in response to perturbations in the form of carbon amendment and altered water status were monitored.     

Influence of human gastrointestinal tract bacterial pathogens on host cell apoptosis

Several pathogenic bacteria are able to trigger apoptosis in the host cell, but the mechanisms by which it occurs differ, and the resulting pathology can take different courses. Induction and/or blockage of programmed cell death upon infection is a result of complex interaction of bacterial proteins with cellular proteins involved in signal transduction and apoptosis. In this review we focus on pro/anti-apoptotic activities exhibited by two enteric pathogens Salmonella enterica, Yersinia spp. and gastric pathogen Helicobacter pylori. We present current knowledge on how interaction between mammalian and bacterial cell relates to the molecular pathways of apoptosis, and what is the role of apoptosis in pathogenesis.     

Molecular microbial ecology of the gastrointestinal tract: from phylogeny to function

During the past decade it became evident that anaerobic cultivation-based approaches provides an incomplete picture of the microbial diversity in the GI tract, since at present only a minority of microbes can be obtained in culture. The application of molecular, mainly 16S ribosomal RNA (rRNA)-based approaches enables researchers to bypass the cultivation step and has proven its usefulness in studying the microbial composition in a variety of ecosystems, including the gastrointestinal (GI) tract. This critical review summarizes the impact of these culture-independent approaches on our knowledge of the ecology of the GI tract and provides directions for future studies which should emphasize function of specific strains, species and groups of microbes.     

The effect of cocoa and polydextrose on bacterial fermentation in gastrointestinal tract simulations

Effects of cocoa mass and supplemented dietary fiber (polydextrose) on microbial fermentation were studied by combining digestion simulations of stomach and small intestine with multi-staged colon simulations. During the four phases of digestion, concentrations of available soluble proteins and reducing sugars reflected in vivo absorption of nutrients in small intestine. In colon simulation vessels, addition of polydextrose to digested cocoa mass significantly increased concentrations of total short-chain fatty acids and butyric acid, from 103 to 468 mM (P<0.01) and from 12 to 22 mM (P<0.01), respectively. Long-chain fatty acid concentrations (decreasing from 1,222 to 240 mM) were mainly affected by the presence of digested cocoa mass. Cocoa mass with or without polydextrose addition significantly decreased production of cadaverine (P<0.02) and branched-chain fatty acids compared to control during colon simulations. Results indicate beneficial effects on metabolism of colonic microbiota after digestion of cocoa mass, and even more so with polydextrose addition.