Some aspects of the gastrointestinal microflora of germfree mice associated with cultured microfloras

Attempts are described to 'normalize' germfree mice by association with 3, 21 and 71 different intestinal bacterial cultures isolated from mice with an SPF flora. Germfree mice associated naturally with an SPF flora served as controls. Vital bacterial counts were determined by aerobic and anaerobic culture. Stomach and small intestine contained fewer bacteria per gram than caecum and large intestine. Aerobic vital counts from caecum and large intestine were higher in the experimental groups than in control mice. The aerobic and anaerobic flora in stomach and small intestine comprised mainly Gram-positive non-fusiform shaped rods. In the caecum and colon Gram-positive cocci predominated in the aerobic culture while in the anaerobic culture fusiform-shaped rods were prominent. Scanning electron microscopy of oesophagus, ileum, caecum and faeces demonstrated colonization of the oesophageal epithelium only after association with 71 bacterial strains; the filamentous bacteria present in the ileum of SPF mice were not found in the experimental groups and caecum and faeces contained mainly fusiform-shaped bacteria. Non-bacterial matter decreased in the caecum and faeces with increase in the complexity of the flora.     

几种常用中药对地鼠肠道正常菌群的影响

我们选择了几种药性不同的中药,使用各种选择性培养基,对给药组和正常动物组的地鼠肠菌群进行了研究,结果表明,黄芪组与正常动物组比较需氧菌的量有所增加,统计学差异显著(P<0.001)。而其他几味苦寒药(大黄、黄芩、白芍等)对需氧菌的作用不明显。在各类厌氧菌的分离结果中,各给中药组与正常组比较均有明显差异。给药组之间比较,药性相同的组之间没有显著变化,药性不同的组之间变化显著。通过实验我们发现中药对正常地鼠的肠道菌群是有明显影响的,与正常动物组相比及不同药性组间相比各类菌的增长或减少都具有统计学意义。     

虎斑颈槽蛇肠道微生物组成和分布特征

蛇类是一种研究能量平衡的模型动物,而肠道微生物则帮助宿主获取能量和营养。大多数肠道微生物的研究集中在圈养动物上,而对于野生蛇类的研究很少。本研究从3只野生虎斑颈槽蛇（Rhabdophis tigrinus）的大肠、小肠和泄殖腔内容物提取微生物总DNA,进行16S rRNA基因V4区域扩增子测序,分析虎斑颈槽蛇肠道核心微生物组成和分布特征。结果表明,阿尔法指数在不同肠道区域的多样性不存在显著性差异。大肠、小肠和泄殖腔3个肠道部位共享534个操作分类单元（OTUs）,大肠包含最多特有OTUs（388个）。门水平,梭杆菌门（Fusobacteria）在小肠（52.87% ± 14.49%）、大肠（41.12% ± 22.60%）和泄殖腔（65.70% ± 10.44%）均为优势菌。属水平,鲸杆菌属（Cetobacterium）在小肠（46.36% ± 13.86%）、大肠（21.95% ± 9.82%）和泄殖腔（58.18% ± 14.29%）为优势菌,柠檬酸杆菌属（Citrobacter）在小肠和泄殖腔中有显著性差异。此外,在其肠道检测到很多潜在的致病菌,例如,柠檬酸杆菌属、明串珠菌属（Trichococcus）和丹毒丝菌属（Erysipelothrix）,了解了潜在致病菌在虎斑颈槽蛇肠道中的分布情况。在聚类分析中发现泄殖腔的数据重复性较好,将本研究泄殖腔数据与前人研究过的食鱼蝮（Agkistrodon piscivorus）泄殖腔高通量测序数据进行联合分析,为更广泛开展不同蛇类肠道微生物组成的差异性分析提供理论借鉴依据。     

Succinate accumulation in pig large intestine during antibiotic-associated diarrhea and the constitution of succinate-producing flora

Succinate was the major organic acid detected in the hindgut content of pigs suffering from antibiotic-associated diarrhea. Antibiotic-associated diarrhea was induced by an oral dose of polymyxin B sulfate (3,000,000 units/day) or an intramuscular injection of enrofloxacin (0.6 g enrofloxacin/day). In the large intestine of enrofloxacin-treated pigs, Gram-negative facultative anaerobic rods phylogenetically related to Escherichia coli and Gram-positive facultative anaerobic non-spore-forming rods phylogenetically related to Lactobacilli were isolated as succinate producers. Succinate-producing Lactobacilli were only isolated as the succinate producer in polymyxin B sulfate-treated pigs. In contrast to antibiotic-associated diarrhea pigs, bacteria belonging to Bacteroidaceae, Fusobacteria, and Enterobacteriaceae were detected as succinate producers in a non-treated pig. In antibiotic-associated diarrhea conditions, antibiotic-resistant Enterobacteria, E. coli in particular, and Lactobacilli may contribute to an abnormal succinate accumulation and may affect water absorption in the hindgut that relates to an expression of antibiotic-associated diarrhea.     

海水混养池塘虾蛤肠道与养殖环境的微生物多样性

【背景】海水混养池塘环境微生物以及动物肠道微生物的群落结构已有研究,但对混养环境中多品种动物肠道与环境微生物群落的关系尚未见报道。【目的】研究海水虾蛤混养环境中微生物多样性以及与养殖动物健康之间的关系。【方法】采用Illumina高通量测序技术测定冬季莆田市北江养殖区2个混养池塘中水体、底泥以及虾蛤肠道的菌群结构。【结果】同一池塘水体与底泥之间、不同池塘水体或底泥之间的微生物结构存在一定的差异;同一养殖区2个混养池塘虾与蛤肠道微生物结构之间具有极高的相似性,与养殖环境存在显著的差异。微生物多样性和丰富度差异很大,表现出底泥水体肠道;虾蛤肠道微生物以厚壁细菌和γ-变形细菌为主;池塘水体以放线菌、α-变形细菌以及拟杆菌为主,底泥以γ-变形细菌和δ-变形细菌为主。养殖动物肠道微生物主要优势种为乳球菌属和假单胞菌属,池塘环境内存在较高丰度的黄杆菌类潜在致病菌,而在虾和蛤的肠道中基本未检出。2个池塘底泥硫还原细菌含量较高,增加了底质产生硫化氢等有害物质的风险。【结论】比较混养池塘中水体、底泥以及虾蛤肠道三者之间微生物群落结构的差异,揭示虾、贝混养模式微生物与养殖环境的关系,为池塘养殖虾、贝疾病防治和混养结构的优化提供参考。     

急性胰腺炎患者舌苔微生物研究

目的观察急性胰腺炎（ＡＰ）患者舌苔微生物的变化,总结不同舌象、不同病情的ＡＰ患者舌苔微生物变化规律。方法将纳入的５２例ＡＰ患者按分级标准分为轻症（ＭＡＰ）组（ｎ＝３３）,重症（ＳＡＰ）组（ｎ＝１９）,分别于治疗前、第３天及第９天清晨观察并采集舌苔,作细菌培养、鉴定、定量,测溶菌酶（ＬＺＭ）含量。２５例健康薄白苔作为对照组。结果ＡＰ患者舌苔菌落总数减少。厚苔菌落总数及Ｇ－厌氧杆菌多于薄苔,口腔优势菌在厚苔变化表现为厌氧菌检出率增高而需氧菌下降。ＳＡＰ组肠道杆菌的检出率显著增加;病程早期除口腔链球菌、Ｇ＋厌氧杆菌减少外,ＳＡＰ组Ｇ－厌氧杆菌也减少;病程第９天时,ＭＡＰ组基本恢复正常,而ＳＡＰ组各种异常无改变。ＡＰ患者舌苔ＬＺＭ均显著升高。结论ＡＰ病程中口腔舌面出现微生态失调现象,严重程度与病情及舌苔的变化有关。ＡＰ厚苔菌落总数多于薄苔,主要为Ｇ－厌氧杆菌增加。厚苔厌氧菌增加,而需氧菌减少。     

咖啡摄入量对小鼠肠道微生物及酶活性的影响

目的研究摄入不同咖啡浓度对小鼠酶活性、肠道微生物、体重的影响,从分析肠道四种微生物变化和四种酶的活性变化来讨论摄入咖啡对健康的影响。方法 SPF级昆明小鼠随机分为4组,即低、中、高咖啡浓度组和正常对照组,所有实验组小鼠日常给予蒸馏水饮用,实验低、中、高浓度组每天2次分别灌胃给予0.0045g/mL、0.0090g/mL、0.0180g/mL的咖啡溶液0.4mL/(只·次),对照组给予蒸馏水灌胃,实验时间为1个月,测小鼠体重、肠道微生物和酶活性等指标并进行分析。结果与对照组相比,低浓度咖啡摄入组乳杆菌、双歧杆菌、大肠埃希菌数显著增加(P0.01或P0.05),需氧和厌氧细菌数显著降低(P0.01);中浓度咖啡摄入组乳杆菌、双歧杆菌数显著低于对照组和低浓度组(P0.01),大肠埃希菌显著高于对照组(P0.05);高浓度咖啡摄入组乳杆菌和大肠埃希菌总数显著低于对照组(P0.01或P0.05);低浓度摄入组的木聚糖酶、纤维素酶、淀粉酶、蛋白酶的活性均显著高于(Ps0.01)对照组和中浓度咖啡摄入组;各组小鼠体重差异无统计学意义(P0.05)。结论适当摄入咖啡小鼠肠道双歧杆菌和乳杆菌等益生菌数量增加显著,小鼠肠道中具有代表性的淀粉酶、纤维素酶、蛋白酶、木聚糖酶的活性均显著增加,但咖啡摄入对小鼠体重影响不显著。     

大肠菌群与氮营养素互作及其对机体健康的影响

蛋白质不仅是构建机体组织的主要原料,而且对动物新陈代谢活动至关重要。数目庞大的肠道细菌在机体营养素,尤其是氮营养素的代谢过程中发挥重要作用。小肠细菌能代谢部分氨基酸,进而影响宿主整体氨基酸的代谢。与小肠相比,大肠拥有更为丰富的菌群和更长的蠕动时间。一方面,进入大肠的氮营养素会影响大肠菌群的代谢和群落结构;另一方面,大肠菌群也能广泛参与氮营养素的代谢与利用,生成许多代谢产物,进而影响机体健康。本文主要综述了日粮蛋白质对大肠菌群的影响、大肠菌群代谢氨基酸的产物及其对肠道生理和机体健康的影响。     

Microbial succession and intestinal enzyme activities in the developing rat

J. CHANG, R.W. CHADWICK, J.C. ALLISON, Y.O. HAYES, D.L. TALLEY AND C.E. AUTRY. 1994. The succession of gut bacteria and selected intestinal enzyme activities in developing 7–35-d-old rats was studied. Aerobes and anaerobes were identified as members of four broad major bacterial groups, i.e. Gram-positive rods, Gram-positive cocci, Gram-negative rods and obligate anaerobes. The enzyme activities of nitro and azo reductases, β-glucuronidase, dechlorinase and dehydrochlorinase were determined by anaerobic incubation of intestinal homogenates with 3,4-dichloronitrobenzene, methyl orange, ***p-nitrophenyl-β-D-glucuronide, and ***p, p-DDT respectively. Nitroreductase and azo reductase activities increased significantly with the appearance of anaerobes in the large intestine. No increase in either nitroreductase or azo reductase activities in the small intestine was found. The early and high level of β-glucuronidase activity in the small and large intestines coincided with high numbers of coliforms recovered in 7 and 14 d animals. Dehydrochlorinase activity appeared early but was undetectable at both 21 and 28 d. Its activity increased at 35 d. Dechlorinase activity was variable in development. The rapid changes in the microbial flora and intestinal enzyme activities may influence the susceptibility of pre-pubescent rats to a variety of toxicants. Therefore, age-dependent toxicity may be important in the risk assessment of some environmental chemicals.     

肠道需氧革兰氏阴性杆菌与门静脉血内毒素水平关系的初步探讨

本实验以普通 Webster 大鼠为动物模型,探讨肠道需氧革兰氏阴性杆菌(Gram ne-gative,G~-杆菌)与门静脉血内毒素的关系。以抗生素联合灌胃使大鼠肠道脱污染,降低肠道定植抗力,再以4种需氧 G~-杆菌混合液灌胃,行肠道再污染,然后再分别测定正常鼠(组),脱污染鼠(组)及再污染鼠(组)不同肠段和粪便需氧 G~-杆菌及相应鼠门静脉血内毒素水平。结果表明,肠道需氧 G~-杆菌的变化与相应门静脉血内毒素水平的变化基本一致.由此可见,上消化道需氧 G~-杆菌过生长,可能会成为门静脉血内毒素水平增加的原因之一。     

Extremely high circulating levels of 1α,25-dihydroxyvitamin D3 in the marmoset,a new world monkey

Compared to most mammals, the marmoset, a new world monkey, requires particularly large amounts of vitamin D to maintain normal growth. We compared serum concentrations of vitamin D metabolites in marmosets with rhesus monkeys and humans. The circulating levels of 1α,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃] in marmosets were 4 to 10 times higher than those in rhesus monkeys and humans. But none of the marmosets exhibited hypercalcemia. In two marmosets which had suffered bone fractures, the 1α,25-(OH)₂D₃ levels were particularly elevated. These results suggest that the marmoset has an end-organ resistance to 1α,25(OH)₂D₃.     

Intestinal microflora of monkeys, normal and in pathology, and its correction by preparations of live bacteria from human biocenosis (coli bacteria, bifikol and bifidum bacterin)

The study of the qualitative and quantitative composition of intestinal microflora in 15 anthropoid apes, 60 lower primates and 72 monkeys with clinically pronounced dysbacteriosis was made, which revealed the prevalence of microorganisms belonging to the genus Proteus, a decrease in the biological activity of normal Escherichia coli and in the content of bifido- and lactobacteria. The treatment of 39 rhesus and pig-tailed macaques with the preparations of live bacteria occurring in normal human microflora led to clinical convalescence and the normalization of the bacteriocenosis, while in 33 control animals no positive shifts in clinical and bacteriological data were observed. The essential similarity of the composition of intestinal microflora in higher and lower primates and in man makes it possible to use these animals as models for testing the effectiveness of new biological preparations and determines the expediency of using bacterial therapy for the treatment of monkeys at the period of acclimatization.     

Microbial colonization and succession in the large intestine of newborn infants during their stay with mothers at the maternity hospital]

Formation of microflora in the large intestine of 5-day old infants was studied in one of the Moscow maternity homes. The up-to-date procedures for isolation and identification of aerobic and anaerobic organisms were used in the study and the findings were processed on a computer. In the newborns of the maternity home of the "mother-infant" type there was observed colonization of the large intestine with aerobic and anaerobic organisms. A wave-like dynamics in the formation of the symbiotic microflora was revealed. It reflected the phenomenon of the microbial succession in the infants. The attempts to detect microbial interference between the species colonizing the large intestine showed that it was extremely rare in the 5-day old infants. This was likely the reason of the low intestine resistance to the colonization in the newborns which in its turn defined the frequent colonization of the intestine mucosa with S. aureus and the organisms of the Klebsiella, Enterobacter and Citrobacter group.     

Gut microflora of vervet and samango monkeys in relation to diet

The microflora in the gastrointestinal tracts of wild vervet and samango monkeys (Cercopithecus aethiops and C. mitis, respectively) were studied, using fermentation acid analysis, electron microscopy, and culturing methods. The diets of the two species of monkey differ considerably, with that of the samango including a greater proportion of cellulose-rich leaf material, and this is reflected in the microflora. Volatile fatty acid measurements along the gut of both species showed that these end products of bacterial metabolism were concentrated in the cecum and colon. Electron microscopy indicated that morphologically similar bacteria were present in the cecum and colon of both species, but the samango possessed a distinct stomach microflora. Bacteria in the lumina of the four main regions of the gut of the monkeys (stomach, small intestine, cecum, and colon) were plated on a number of anaerobic media (Mann, Rogosa, and Sharp; clostridial basal; and complex media). The cecum and colon were found to contain higher numbers of microbes per gram (wet weight) of gut content than the stomach and small intestine. Microbial isolates were able to catabolize carboxymethyl cellulose and other polymers. This may aid the monkeys, particularly samangos, in the digestion of fibrous dietary components such as leaves.     

Microflora of the large intestine in patients with connective tissue dysplasia

The specific features of intestinal aerobic and anaerobic intestinal microflora in children with nondifferentiated connective tissue dysplasia were under study. A high rate of dysbiotic disturbances was noted in patients with connective tissue dysplasia in comparison with healthy persons. Profound quantitative and qualitative changes in the biocenosis of the intestine were detected in patients with the pathology of the gastrointestinal tract as well as that of the locomotor system. Changes in the species composition and the persistence properties of the intestinal microflora may serve as pathogenetic factors in the development of connective tissue dysplasia.     

Dysbacteriosis in patients with colon polyposis

The study revealed the most profound changes in the composition of intestinal microflora in patients with polyposis of the large intestine. In these patients anaerobic microflora (bifidobacteria, lactic acid bacteria) was more often suppressed than in other examined groups, in particular, patients with cholelithic disease. The associations of hemolytic Escherichia with Klebsiella pneumoniae, Pseudomonas aeruginosa, Citrobacter freundii were often observed as well as the increased content of enterococci and fungi of the genus Candida. The determination of frequency and degree of manifestations showed that dysbacteriosis was registered in the absolute majority of patients (97.4%) with polyposis of the large intestine. Among patients with cholelithic disease disturbances in microbiocenosis were detected in 60.0% of cases, the profundity and character of the microflora composition changes being less pronounced.     

Comparative study of the oxidation of propranolol enantiomers in hepatic and small intestinal microsomes from cynomolgus and marmoset monkeys

Oxidative metabolism of propranolol (PL) enantiomers (R-PL and S-PL) to 4-hydroxypropranolol (4-OH-PL), 5-OH-PL and N-deisopropylpropranolol (NDP) was examined in hepatic microsomes from cynomolgus and marmoset monkeys and in small intestinal microsomes from monkeys and humans. In hepatic microsomes, levels of oxidation activities were similar between the two monkey species, and substrate enantioselectivity (R-PL < S-PL) was observed in the formation of 5-OH-PL and/or NDP. Kinetic experiments revealed that the formation of all metabolites was biphasic in cynomolgus monkeys, whereas only the formation of NDP was biphasic in marmosets. Inhibition experiments employing human CYP antibodies and chemical inhibitors suggested that mainly CYP2D enzymes and partially CYP1A and 2C enzymes are involved in the oxidation of PL in both monkey liver microsomes. In small intestinal microsomes, activity levels were much higher in cynomolgus monkeys than in marmosets and humans and reversed substrate enantioselectivity (R-PL > S-PL) was seen in the formation of NDP in cynomolgus monkeys and humans and in the formation of 5-OH-PL in marmosets. The formation of the three metabolites in cynomolgus monkeys and the formation of NDP in marmosets were biphasic, while the formation of 4-OH-PL in humans was monophasic. From the inhibition experiments using CYP antibodies, CYP2C9 and 2C19 were thought to be involved as N-deisopropylases and CYP2D6 and 3A4 as 4-hydroxylases in human small intestine. Furthermore, CYP1A, 2C and 3A enzymes could be involved in cynomolgus monkeys and CYP2C and 3A enzymes in marmosets. These results indicate that the oxidative profile of PL in hepatic and small intestinal microsomes differ considerably among cynomolgus monkeys, marmosets and humans.     

The prevention of the translocation of intestinal microflora after the rescue of an organism from a terminal state]

In experiments on guinea pigs, the animals were resuscitated from clinical death caused by the acute loss of blood and subsequently treated intragastrically with enterosorbents: activated carbon fibrous material, alone and in combination with polymyxin B, polyphepan (a lignin derivative), polymethyloxan hydrogel and the sorbent Enterocat. In the animals, not treated during the postresuscitation period, a high population level of enterobacteria and Gram-positive aerobic cocci was registered in the contents of the small and large intestines and their translocation to mesenteric lymph nodes, liver, spleen and blood was observed. The amount of lactobacilli in the small intestine was decreased. Enterosorbents were found to decrease a high population level of intestinal microflora, to prevent the translocation of Gram-positive aerobic cocci and to inhibit the penetration of enterobacteria through the enteric barrier in the postresuscitation period. Combined use of activated carbon fibrous material with polymyxin B proved to be most effective for the elimination of enterobacteria.     

Gut microflora of vervet and samango monkeys in relation to diet.

The microflora in the gastrointestinal tracts of wild vervet and samango monkeys (Cercopithecus aethiops and C. mitis, respectively) were studied, using fermentation acid analysis, electron microscopy, and culturing methods. The diets of the two species of monkey differ considerably, with that of the samango including a greater proportion of cellulose-rich leaf material, and this is reflected in the microflora. Volatile fatty acid measurements along the gut of both species showed that these end products of bacterial metabolism were concentrated in the cecum and colon. Electron microscopy indicated that morphologically similar bacteria were present in the cecum and colon of both species, but the samango possessed a distinct stomach microflora. Bacteria in the lumina of the four main regions of the gut of the monkeys (stomach, small intestine, cecum, and colon) were plated on a number of anaerobic media (Mann, Rogosa, and Sharp; clostridial basal; and complex media). The cecum and colon were found to contain higher numbers of microbes per gram (wet weight) of gut content than the stomach and small intestine. Microbial isolates were able to catabolize carboxymethyl cellulose and other polymers. This may aid the monkeys, particularly samangos, in the digestion of fibrous dietary components such as leaves.     

水溶性壳聚糖对小鼠肠道菌群的影响

目的应用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术研究水溶性壳聚糖对正常小鼠肠道菌群的影响。方法使用SPF级昆明小鼠18只,随机分为3组,每组6只,依次为正常对照组(NC)、水溶性壳聚糖高(WSC-H)和低浓度组(WSC-L),灌服对应药物30d后收取鼠便,细菌基因组DNA提取,PCR-DGGE电泳得到肠道菌群图谱,计算菌群结构多样性指数,聚类分析相似性并鉴定优势条带序列。结果水溶性壳聚糖处理后小鼠肠道菌群结构发生改变,多样性指数、丰富度和均匀度较低,乳杆菌成为肠道中的优势菌型。结论水溶性壳聚糖能够增加肠道中乳杆菌的种类和数量,起到益生元的作用来调节肠道微生态平衡。