Inhibitory effect of a copper-dipeptide complex on the establishment of a Clostridium perenne strain in the intestinal tract of gnotobiotic mice.

A semisynthetic diet fed to axenic mice was found to prevent the establishment of a Clostridium perenne strain in their intestinal tract. This inhibitory effect did not occur when axenic mice were preinoculated with a strain of Clostridium difficile. The inhibitory effect was related to the presence in the intestinal contents of axenic mice of both dietary copper and a dipeptide, aspartic-epsilon-lysine. When C. difficile was inoculated into axenic mice, the dipeptide disappeared from the digesta, and C. perenne became established even in the presence of high concentrations of copper.     

凝结芽胞杆菌TBC 169株对肠道致病菌的抑菌作用

目的 研究凝结芽胞杆菌TBC 169株对大肠埃希菌、痢疾志贺菌、伤寒沙门菌、普通变形杆菌、铜绿假单胞菌和金黄色葡萄球菌的抑制作用。方法 先将大肠埃希菌、痢疾杆菌等6种菌分别进行单独培养,测定不同培养时间内的pH和活菌数,然后将凝结芽胞杆菌TBC 169株分别和致病菌进行混合培养,再测pH和活菌数,并与单独培养时的测定情况进行比较。结果 凝结芽胞杆菌TBC 169株对大肠埃希菌、痢疾志贺菌、伤寒沙门菌等6种菌均有明显的抑制作用,尤其是对伤寒沙门菌和铜绿假单胞菌的抑制作用更强。结论 凝结芽胞杆菌TBC 169株对肠道致病菌有显著的抑制作用。     

Modulation of cytotoxin production by Clostridium difficile in the intestinal tracts of gnotobiotic mice inoculated with various human intestinal bacteria

Gnotobiotic mice died 2 days after inoculation of a cytotoxigenic Clostridium difficile strain. Protection occurred when mice were previously inoculated with a strain of Escherichia coli or Bifidobacterium bifidum. Intestinal cytotoxin production was highly reduced in the surviving mice, whereas the C. difficile population level did not decrease to a great extent.     

Modulation of cytotoxin production by Clostridium difficile in the intestinal tracts of gnotobiotic mice inoculated with various human intestinal bacteria.

Gnotobiotic mice died 2 days after inoculation of a cytotoxigenic Clostridium difficile strain. Protection occurred when mice were previously inoculated with a strain of Escherichia coli or Bifidobacterium bifidum. Intestinal cytotoxin production was highly reduced in the surviving mice, whereas the C. difficile population level did not decrease to a great extent.     

Clostridium innocuum: a glucoseureide-splitting inhabitant of the human intestinal tract

Glycosylureides were recently described as non-invasive markers of intestinal transit time. The underlying principle is an enzymatic splitting of (13)C-labelled ureides by intestinal bacteria. The (13)CO(2) released from the urea moiety of the glycosylureides can be measured in breath samples when the ingested tracer substrate reaches the caecum that is colonised with microbes. To date, the microbes that degrade glycosylureides are unknown. In order to identify the glucoseureide (GU)-splitting bacteria, 174 different strains of intestinal microbes obtained from five healthy adults were checked for their ability to degrade GU. The results of the microbial cultures and thin layer chromatography revealed that GU was exclusively degraded by Clostridium innocuum, belonging to the normal human intestinal microflora. C. innocuum probably synthesises a yet unknown enzyme that splits the glucose-urea bond. We suggest that the term glucoseureidehydrolase is the appropriate designation for this enzyme.     

Prevention of Clostridium difficile induced mortality in gnotobiotic mice by Saccharomyces boulardii

Oral preventive treatment of gnotobiotic mice by Saccharomyces boulardii significantly decreased mortality following Clostridium difficile infection. A single S. boulardii ingestion protected 16% of mice, whereas 56% were protected when S. boulardii was given continuously in the drinking water. No direct antagonistic effect of the yeast on C. difficile numbers was detected, whereas a modulation of fecal cytotoxin production was demonstrated.     

Inhibitory effect of taurolipids on Clostridium perfringens sialidase

Taurolipids A and B, which are detergent-type compounds isolated from protozoan Tetrahymena cells, were demonstrated to inhibit strongly the activity of Clostridium perfringens sialidase. On addition of 280 pmol of taurolipid B to 20 mU of the enzyme, the sialidase activity was decreased to 7% of the original activity at pH 5.1 as the optimum pH. The inhibition was non-competitive. Effective inhibition was observed at the acidic region from the isoelectric point of the sialidase, and at a low ionic strength. Both the long chain acyl and sulfonic acid groups of taurolipids were required for the inhibition of the sialidase activity. A mechanism is postulated for the inhibition.     

Inhibitory impact of bifidobacteria on the transfer of beta-lactam resistance among Enterobacteriaceae in the gnotobiotic mouse digestive tract

While looking for new means to limit the dissemination of antibiotic resistance, we evaluated the role of potentially probiotic bifidobacteria on the transfer of resistance genes between enterobacteria. Transfers of bla genes encoding extended-spectrum beta-lactamases (SHV-5 and CTX-M-15) were studied in the absence or presence of bifidobacteria. In vitro, transfer frequencies of these bla genes decreased significantly in the presence of three of five tested strains, i.e., Bifidobacterium longum CUETM-89-215, Bifidobacterium bifidum CIP-56.7T, and Bifidobacterium pseudocatenulatum CIP-104168T. Four transfer experiments were conducted in the digestive tract of gnotobiotic mice, the first three observing the effect of B. longum CUETM-89-215, B. bifidum CIP-56.7T, and B. pseudocatenulatum CIP-104168T on blaSHV-5 transfer and the fourth experiment studying the effect of B. bifidum CIP-56.7T on blaCTX-M-15 transfer. These experiments revealed significant decreases in the transconjugant levels (up to 3 logs) in mice having received B. bifidum CIP-56.7T or B. pseudocatenulatum CIP-104168T compared to control mice. Bifidobacteria appear to have an inhibitory impact on the transfer of antibiotic resistance genes. The inhibitory effect is associated to specific bifidobacterial strains and may be related to the production of thermostable metabolites by these strains.     

消毒剂对环境菌株抑制作用的研究

目的 研究企业常用消毒剂对于洁净室环境监测分离的菌株样本的抑制作用。方法 通过VITEK2-COMPACT全自动细菌鉴定及药敏分析系统鉴定收集到的环境菌株。对3种消毒剂(碘伏、无水乙醇和苯扎溴铵)进行梯度稀释,利用打孔法研究3种消毒剂在不同含量下对环境菌株的抑制作用。结果 共检出革兰阳性菌8种、革兰阴性菌2种、酵母菌1种、芽孢杆菌2种;苯扎溴铵对革兰阳性菌的抑菌能力都较强,随着含量的降低,抑菌作用逐渐减弱;碘伏对革兰阴性菌及酵母菌的抑制作用都非常强,随着含量降低,抑菌作用逐渐降低。3种消毒剂对2种芽孢杆菌的抑制作用均有限。结论 用1.00%苯扎溴铵和0.50%的碘伏抑菌作用都非常强。另外,应配合使用杀孢子剂,避免芽孢杆菌孢子在空气中传播。     

Adaptation of protein expression by Escherichia coli in the gastrointestinal tract of gnotobiotic mice

The gastrointestinal tract of mammals is inhabited by several hundred bacterial species. While the effects of the gut microbiota upon the host have been widely studied, the microbial response to host factors has only recently attracted attention. In order to investigate the influence of the host on the physiology of gastrointestinal bacteria, a simplified model of host–bacteria interaction was created by associating germfree mice with commensal Escherichia coli . Here we demonstrate the feasibility of analysing the bacterial response to the conditions in the digestive system by a proteomics-based approach. Two-dimensional gel electrophoresis (2D-GE) followed by electrospray ionization-tandem mass spectrometry (ESI-MS/MS) was used to identify bacterial proteins from caecal and faecal samples. In a set of 60 arbitrarily chosen spots of stably and differentially expressed proteins, 50 different bacterial proteins were identified. Their ascribed functions suggest that the host-associated bacteria adapt their metabolism to the conditions in the intestine by utilizing arginine, asparagine and aspartate as well as glucose/galactose, ribose, maltose, glucuronate, galacturonate and gluconate as substrates. Thirteen proteins not previously detected on 2D-gels and 10 proteins with unknown or poorly characterized physiological function were identified, while the existence of three proteins had so far only been inferred from predictions or by homology.     

Effect of toxins produced by various Clostridium difficile strains on cecum size reduction in gnotobiotic mice

Inoculation of axenic mice with Clostridium difficile strains induced a significant reduction in ceca weight (dry or wet), whereas a nontoxinogenic strain led to a partial reduction. A strain, which produces cytotoxin and no enterotoxin in vivo, caused a reduction similar to that observed with a nontoxinogenic strain. Simultaneous cytotoxin and enterotoxin production by various C. difficile strains caused the cecum size to diminish to that observed for conventional control mice.     

Inhibitory effect of TNF-alpha on the intestinal absorption of galactose

Sepsis is a systemic response to infection in which toxins, such as bacterial lipopolysaccharide (LPS), stimulate the production of inflammatory mediators like the cytokine tumor necrosis factor alpha (TNF-alpha). Previous studies from our laboratory have revealed that LPS inhibits the intestinal absorption of L-leucine and D-fructose in rabbit when it was intravenously administered, and that TNF-alpha seems to mediate this effect on amino acid absorption. To extend this work, the present study was designed to evaluate the possible effect of TNF-alpha on D-galactose intestinal absorption, identify the intracellular mechanisms involved and establish whether this cytokine mediates possible LPS effects. Our findings indicate that TNF-alpha decreases D-galactose absorption both in rabbit intestinal tissue preparations and brush-border membrane vesicles. Western blot analysis revealed reduced amounts of the Na+/glucose cotransporter (SGLT1) protein in the plasma membrane attributable to the cytokine. On the contrary, TNF-alpha increased SGLT1 mRNA levels. Specific inhibitors of the secondary messengers PKC, PKA, the MAP kinases p38 MAP, JNK, MEK1/2 as well as the proteasome, diminished the TNF-alpha-evoked inhibitory effect. LPS inhibition of the uptake of the sugar was blocked by a TNF-alpha antagonist. In conclusion, TNF-alpha inhibits D-galactose intestinal absorption by decreasing the number of SGLT1 molecules at the enterocyte plasma membrane through a mechanism in which several protein-like kinases are involved.     

The effect of new antimicrobial preparations used in combination with complete gnotobiotic isolation on the survivability and intestinal microflora of total-body irradiated mice

The aim of these investigations was the study of the effect of different antimicrobial preparations on the survival rate and intestinal microflora of mice with experimental acute radiation sickness. These investigations revealed that the survival rate of the animals increased 3.1 times with the use of Supramycin, 2.4 times with the use of Tacef and 1.6 times with the use of Spizef and Pen-bristol. The study of the influence of these preparations on the intestinal microflora revealed that Spizef not only decreased the number of opportunistic microorganisms, but also led to a sharp drop in the level of lactobacteria. The use of Pen-bristol and Tacef led to practically complete elimination of enterobacteria enterococci, staphylococci, as well as lactobacteria. Supramycin essentially suppressed the number of opportunistic microorganisms and did not affect the level of lactobacteria. This was probably the cause of the highest effectiveness of Supramycin in comparison with Tacef, Spizef and Pen-bristol.     

Isolation of a bile salt sulfatase-producing Clostridium strain from rat intestinal microflora.

Bile acid sulfates, formed in human and rat livers, are desulfated by the intestinal microflora. In our study we first isolated from conventional rat feces an unnamed bacterium, termed strain S1, which desulfated the 5 beta-bile salt 3 alpha-sulfates in vitro and in vivo after association with gnotobiotic rats. Strain S1 also possessed 12 alpha-hydroxysteroid dehydrogenase and bile salt-deconjugating activities. The strain was a strict anaerobic, CO2-requiring, gram-negative, sporeforming rod and was designated as belonging to the genus Clostridium. Growth was scarce in culture media, unless in the presence of 0.1% taurine, a sulfur-containing amino acid. Addition of this substance raised the number of bacteria in thioglycolate and peptone yeast media from 10(4) per ml to 10(6) to 10(7) per ml and increased the colony diameter on agar medium from 0.2 mm to 0.5 to 0.9 mm. Sulfatase activity was specific for the 5 beta-bile salt sulfates, leaving the 5 alpha-bile salt sulfates unchanged. In addition, the sulfatase activity was cell bound, and its production was dependent on the composition of the culture medium, although no minimal sulfur medium was required for sulfatase activity.     

香菇多糖对微生态失调小鼠肠道菌群及免疫功能的调节作用

目的 探讨香菇多糖对微生态失调小鼠肠道菌群及免疫功能的调节作用.方法 经盐酸林可霉素灌胃建立肠道微生态失调小鼠模型,香菇多糖灌胃治疗,同时设正常对照组、自然恢复组和丽珠肠乐组.7d后处死各组小鼠,进行肠道菌群定量、免疫器官体重及其淋巴细胞转化率检测.结果 用香菇多糖对肠道微生态失调小鼠进行治疗后,小鼠肠道双歧杆菌、乳酸杆菌数量显著增加,而肠杆菌和肠球菌的数量显著降低;脾脏指数明显增加,对胸腺指数无影响;显著增强了淋巴细胞转化率.结论 盐酸林克霉素灌胃能诱导微生态失调小鼠模型的有效建立.香菇多糖能调整小鼠肠道菌群及免疫功能.     

马齿苋多糖对肠道微生态失调小鼠的调整作用研究

目的研究应用马齿苋多糖对肠道微生态失调小鼠进行调整治疗,达到从微生态学角度防治感染的目的。方法应用林可霉素灌胃建立肠道微生态失调小鼠模型,然后用马齿苋多糖进行治疗,同时设正常对照组、阳性对照组和阴性对照组,于给药7 d后处死小鼠,进行肠道菌群定量、肠内容物挥发性脂肪酸检测及肠黏膜电镜观察。结果林可霉素灌胃3 d后,小鼠肠道菌群失调,肠内容物挥发性脂肪酸含量明显下降,肠黏膜损伤严重。持续7 d治疗后,治疗组小鼠肠道双歧杆菌和乳酸杆菌数量明显上升,肠内容物挥发性脂肪酸含量明显上升,损伤的肠黏膜基本修复。结论应用林可霉素可以成功建立肠道微生态失调动物模型;马齿苋多糖具有扶植肠道正常菌群生长,调整菌群失调,防治感染的作用,是理想的中药微生态调节剂。     

Evolution of commensal bacteria in the intestinal tract of mice

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Characteristics of a new cellulolytic Clostridium sp. isolated from pig intestinal tract.

Gram-positive, spore-forming, motile, cellulolytic rods were isolated from 10(7) dilutions of pig fecal samples. The pigs had previously been fed pure cultures of the ruminal cellulolytic organism Clostridium longisporum. Isolates formed terminal to subterminal spores, and a fermentable carbohydrate was required for growth. Besides cellulose, the isolates utilized cellobiose, glycogen, maltose, and starch. However, glucose, fructose, sucrose, pectin, and xylose were not used as energy sources. Major fermentation products included formate and butyrate. The isolates did not digest proteins from gelatin or milk. Unlike C. longisporum, which has limited ability to degrade cell wall components from grasses (switchgrass, bromegrass, and ryegrass), the swine isolates were equally effective in degrading these components from both alfalfa and grasses. The extent of degradation was equal to or better than that observed with the predominant ruminal cellulolytic organisms. On the basis of morphology, motility, spore formation, fermentation products, and the ability to hydrolyze cellulose, the isolates are considered to be a new species of the genus Clostridium. It is unclear whether C. longisporum played a role in the establishment or occurrence of this newly described cellulolytic species. This is the first report of a cellulolytic Clostridium sp. isolated from the pig intestinal tract.