Changes in fecal microbiota of healthy dogs administered amoxicillin

The effect of oral amoxicillin treatment on fecal microbiota of seven healthy adult dogs was determined with a focus on the prevalence of bacterial antibiotic resistance and changes in predominant bacterial populations. After 4–7 days of exposure to amoxicillin, fecal Escherichia coli expressed resistance to multiple antibiotics when compared with the pre-exposure situation. Two weeks postexposure, the susceptibility pattern had returned to pre-exposure levels in most dogs. A shift in bacterial populations was confirmed by molecular fingerprinting of fecal bacterial populations using denaturing gradient gel electrophoresis (PCR-DGGE) of the 16S V3 rRNA gene region. Much of the variation in DGGE profiles could be attributed to dog-specific factors. However, permutation tests indicated that amoxicillin exposure significantly affected the DGGE profiles after controlling for the dog effect ( P =0.02), and pre-exposure samples were clearly separated from postexposure samples. Sequence analysis of DGGE bands and real-time PCR quantification indicated that amoxicillin exposure caused a shift in the intestinal ecological balance toward a Gram-negative microbiota including resistant species in the family Enterobacteriaceae .     

健康与腹泻林麝肠道菌群的多态性

目的 分析圈养状态下正常林麝和腹泻林麝粪便菌群的组成差异,找到能够影响肠道总菌群稳态的标志菌,为减少林麝腹泻提供防治新策略。 方法 测序分析54只健康林麝(正常组)和18只有腹泻症状的林麝(腹泻组)肠道菌群总DNA中16S rRNA基因V3-V4区,注释微生物种类和数量。计算Chao1、Ace、Shannon和Simpson等多样性指数,进行聚类分析。 结果 正常组和腹泻组之间共有的可操作分类单元(OTUs)2 366个,正常组特有的OTUs 409个,腹泻组特有的OTUs 150个。按组计算α多样性指数,结果表明腹泻组Chao1指数(1 352.766±75.497,t=10.798 7,P结论 腹泻发生时,林麝的肠道菌群丰富度和多样性有所下降,菌群总量显著增加。拟杆菌门物种丰富度极显著性减少的同时,作为潜在致病菌标志的变形菌门(Proteobacteria)比例上升,Chryseobacterium数量极显著升高,研究结果丰富了对林麝腹泻基于肠道菌群变化的认识。     

Differences of subcutaneous adipose tissue topography between type-2 diabetic men and healthy controls

Men with noninsulin-dependent diabetes mellitus (type 2 DM) provide a different subcutaneous body fat distribution and a concentration of fatness on the upper trunk compared with healthy subjects. However, subcutaneous fat distribution is always measured in an inaccurate and/or very simplified way (e.g., by caliper), and to date, there exists no study reporting on the exact and complete subcutaneous adipose tissue distribution of type 2 DM men. A new optical device, the LIPOMETER, enables the nonivasive, quick, and safe determination of the thickness of subcutaneous adipose tissue layers at any given site of the human body. The specification of 15 evenly distributed body sites allows the precise measurement of subcutaneous body fat distribution, so-called subcutaneous adipose tissue topography (SAT-Top). SAT-Tops of 21 men with clinically proven type 2 DM (mean age of 57.5 +/- 6.7 years) and 111 healthy controls of similar age (mean age 59.0 +/- 5.4 years) were measured. In this paper, we describe the precise SAT-Top differences of these two groups and we present the multidimensional SAT-Top information condensed in a two-dimensional factor value plot. In type 2 DM men, especially in the upper trunk, SAT-Top is significantly increased (up to +50.7% at the neck) compared with their healthy controls. One hundred eleven of the 132 individuals (84.1%) are correctly classified (healthy or type 2 DM) by their subcutaneous fat pattern by stepwise discriminant analysis.     

Molecular typing of fecal eukaryotic microbiota of human infants and their respective mothers

The micro-eukaryotic diversity from the human gut was investigated using universal primers directed towards 18S rRNA gene, fecal samples being the source of DNA. The subjects in this study included two breast-fed and two formula-milk-fed infants and their mothers. The study revealed that the infants did not seem to harbour any microeukaryotes in their gut. In contrast, there were distinct eukaryotic microbiota present in the mothers. The investigation is the first of its kind in the comparative study of the human feces to reveal the presence of micro-eukaryotic diversity variance in infants and adults from the Indian subcontinent. The micro-eukaryotes encountered during the investigation include known gut colonizers like Blastocystis and some fungi species. Some of these micro-eukaryotes have been speculated to be involved in clinical manifestations of various diseases. The study is an attempt to highlight the importance of micro-eukaryotes in the human gut.     

中国深圳和芬兰艾斯堡地区 健康婴幼儿肠道菌群的比较

目的探索中国与欧洲沿海发达地区婴幼儿肠道菌群的结构差异及随年龄增长的变化趋势。方法招募深圳地区健康婴幼儿35名并收集其粪便样本,采用Illumina的MiSeq平台对肠道菌群的16S rRNA V4序列进行测序。同时,结合40名芬兰艾斯堡地区健康婴幼儿肠道菌群数据,进行两地儿童菌群结构比较。结果中国与芬兰地区幼儿肠道菌群结构较为一致,在物种多样性及均匀度方面差异无统计学意义。菌属水平分析中,两组幼儿肠道内均呈现以拟杆菌属、双歧杆菌属为主导的菌群结构。回归分析表明,幼儿肠道内菌群多样性随年龄的增长明显增加(R~2=0.174,P0.05),0～1岁与1～3岁幼儿的菌群结构差异有统计学意义(R~2=0.079,P0.05)。韦荣球菌、肠球菌和布劳特菌等菌属随年龄增长丰度明显降低,而粪杆菌、瘤胃球菌、罗斯菌等菌属显著增加。结论生命早期肠道菌群的发育主要与年龄因素密切相关,地区因素为次要因素。     

Differences in developing intestinal microbiota between allergic and non-allergic infants: a pilot study in Japan

The bacterial compositions of feces were monitored in the first 2 months for 15 infants born in Japan, including eight subjects who developed allergy by the age of 2 years. Primer sets targeting six predominant bacterial groups in the infant intestine, Bacteroidaceae, Enterobacteriaceae, bifidobacteria, enterococci, lactobacilli, and the Clostridium perfringens group, were used for real-time PCR to quantitate each population in the feces. The population of Bacteroidaceae was significantly higher in the allergic group at the ages of 1 month (P=0.03) and 2 months (P=0.05) than in the non-allergic group, while no statistically significant difference was observed for the other bacterial populations.     

Molecular monitoring of the fecal microbiota of healthy human subjects during administration of lactulose and Saccharomyces boulardii

Diet is a major factor in maintaining a healthy human gastrointestinal tract, and this has triggered the development of functional foods containing a probiotic and/or prebiotic component intended to improve the host's health via modulation of the intestinal microbiota. In this study, a long-term placebo-controlled crossover feeding study in which each subject received several treatments was performed to monitor the effect of a prebiotic substrate (i.e., lactulose), a probiotic organism (i.e., Saccharomyces boulardii), and their synbiotic combination on the fecal microbiota of three groups of 10 healthy human subjects differing in prebiotic dose and/or intake of placebo versus synbiotic. For this purpose, denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene amplicons was used to detect possible changes in the overall bacterial composition using the universal V(3) primer and to detect possible changes at the subpopulation level using group-specific primers targeting the Bacteroides fragilis subgroup, the genus Bifidobacterium, the Clostridium lituseburense group (cluster XI), and the Clostridium coccoides-Eubacterium rectale group (cluster XIVa). Although these populations remained fairly stable based on DGGE profiling, one pronounced change was observed in the universal fingerprint profiles after lactulose ingestion. Band position analysis and band sequencing revealed that a band appearing or intensifying following lactulose administration could be assigned to the species Bifidobacterium adolescentis. Subsequent analysis with real-time PCR (RT-PCR) indicated a statistically significant increase (P < 0.05) in total bifidobacteria in one of the three subject groups after lactulose administration, whereas a similar but nonsignificant trend was observed in the other two groups. Combined RT-PCR results from two subject groups indicated a borderline significant increase (P = 0.074) of B. adolescentis following lactulose intake. The probiotic yeast S. boulardii did not display any detectable universal changes in the DGGE profiles, nor did it influence the bifidobacterial levels. This study highlighted the capacity of an integrated approach consisting of DGGE analysis and RT-PCR to monitor and quantify pronounced changes in the fecal microbiota of healthy subjects upon functional food administration.     

Culture-independent analysis of fecal microbiota in infants, with special reference to Bifidobacterium species

Fecal microbiota of 31 breast-fed, 26 mix-fed, and 11 bottle-fed infants were analyzed by using terminal restriction fragment length polymorphism (T-RFLP), and culture method. We first determined the total and cultivated bacterial counts in infant fecal microbiota. Only approximately 30% of bacteria present in fecal microbiota were cultivable while the remainder was yet-to-be cultured bacteria. Sixty-eight fecal samples were divided into two clusters (I and II) by T-RFLP analysis, and then subdivided into five subclusters (Ia, Ib, IIa, IIb and IIc). There was no clear relationship between clusters and feeding method. A proportion of bifidobacteria was detected in the fecal material by PCR method using species-specific primers. The predominant Bifidobacterium spp. was Bifidobacterium longum longum type (43 samples (63.2%)), followed by B. longum infantis type (23 samples (33.8%)) and B. breve (16 samples (23.5%)). The distribution of Bifidobacterium spp. was similar in the three feeding groups. In contrast, the high incidence of B. breve in cluster I, especially subcluster Ia and B. longum longum type in cluster II, especially subcluster IIa and IIc were characterized by T-RFLP method. Our results showed that the colonization of Bifidobacterium spp. in infant feces correlated with the T-RFLP clusters.     

Differences in the gut microbiota between Cercopithecinae and Colobinae in captivity

The gut microbiome of captive primates can provide a window into their health and disease status. The diversity and composition of gut microbiota are influenced by not only host phylogeny, but also host diet. Old World monkeys (Cercopithecidae) are divided into two subfamilies: Cercopithecinae and Colobinae. The diet and physiological digestive features differ between these two subfamilies. Accordingly, highthroughput sequencing was used to examine gut microbiota differences between these two subfamilies, using data from 29 Cercopithecinae individuals and 19 Colobinae individuals raised in captivity. Through a comparative analysis of operational taxonomic units (OTUs), significant differences in the diversity and composition of gut microbiota were observed between Cercopithecinae and Colobinae. In particular, the gut microbiota of captive Old World monkeys clustered strongly by the two subfamilies. The Colobinae microbial diversity was higher than that of Cercopithecinae. Additionally, Firmicutes, Lactobacillaceae, Veillonellaceae, and Prevotella abundance were higher in Cercopithecinae, while Bacteroidetes, Ruminococcaceae, Christensenellaceae, Bacteroidaceae, and Acidaminococcaceae abundance were higher in Colobinae. PICRUSt analysis revealed that the predicted metagenomes of metabolic pathways associated with proteins, carbohydrates, and amino acids were significantly higher in Colobinae. In the context of host phylogeny, these differences between Cercopithecinae and Colobinae could reflect adaptations associated with their respective diets. This well-organized dataset is a valuable resource for future related research on primates and gut microbiota. Moreover, this study may provide useful insight into animal management practices and primate conservation.

     

粪便菌群移植的研究与应用

粪便菌群移植早在半个世纪前就有报道应用于临床治疗,但一直没有受到人们的重视。过去的几年间,粪便菌群移植又在针对Clostridium difficile(艰难梭状芽胞杆菌)感染的治疗中逐渐兴起,并且有非常良好的治疗效果,对Clostridium difficile感染性肠炎的病症完全消失。因此,越来越多的患者开始尝试粪便菌群移植的治疗方法,以代替传统的药物治疗方法。肠道菌群的研究逐渐深入我们意识到,肠道菌群已经不仅仅是人体肠道共生微生物而已,而是可以视为人体一个独立且密不可分的人体器官,承担着免疫系统与代谢系统中重要的生理活动。这些关于肠道菌群的研究对粪便菌群移植有着强有力的推动。许多疾病的发病根源,尤其是与微生物紊乱有直接联系的代谢疾病,都可以从微生物与人体的互作模式中寻找答案。本课题综述了国内外近年来粪便菌群移植在人体疾病研究中的应用研究。     

粪微生态制品保存方法

目的 探索粪微生态制品体外保存方法,为粪微生态制品的保存及运输提供依据。方法 将新鲜粪微生态制品按照加入甘油、蔗糖保护剂的不同以及制备冻干粉的方法分为6组,用10倍稀释法稀释粪菌液,微量点样法将各浓度的粪菌液涂于Wilkins-Chalgren培养基,30℃恒温箱中培养并计数。在保存2周、1个月、2个月、3个月时复苏部分微生态制品,用同样的方法培养并计数。结果 粪微生态制品活菌数随时间递减,在2周内下降速度最快,递减105~107数量级。结论 甘油、蔗糖这两种保护剂以及冻干粉的制备对粪微生态制品的保存差异不大,新鲜粪微生态制品仍为最佳选择。     

Comparison of fecal microbiota and polyamine concentration in adult patients with intractable atopic dermatitis and healthy adults

Fecal microbiota and polyamine concentration obtained from eleven intractable adult-type atopic dermatitis (AD) patients and thirteen healthy adults were compared. Fecal microbiota were analyzed using terminal-restriction fragment length polymorphism. The fecal microbiota of volunteers were divided into two clusters, A (n=16) and B (n=8), and the number of AD patients was found to be higher in Cluster B than Cluster A, suggesting that there are relationships between the obstinacy of intractable adult-type AD and intestinal microbiota in Cluster B. Fecal spermidine concentration in Cluster B were lower than that in Cluster A significantly (P<0.05). Fecal putrescine concentration in Cluster B also tended to be lower than that in Cluster A. Terminal-restriction fragment (T-RF) of 122 bp generated by digestion with Hha I, which were predicted as unknown bacteria, were detected characteristically in Cluster A. In contrast, T-RFs of 368/9 bp generated by digestion with Hha I, which were predicted as Enterobacteriaceae, were detected characteristically in Cluster B. These bacteria are closely associated with intestinal polyamine concentration. These findings raise the possibility that a low concentration of intestinal polyamines produced by intestinal microbiota is one of the important factors in the onset of intractable adult-type AD.     

Influence of milk-feeding type and genetic risk of developing coeliac disease on intestinal microbiota of infants: the PROFICEL study

Interactions between environmental factors and predisposing genes could be involved in the development of coeliac disease (CD). This study has assessed whether milk-feeding type and HLA-genotype influence the intestinal microbiota composition of infants with a family history of CD. The study included 164 healthy newborns, with at least one first-degree relative with CD, classified according to their HLA-DQ genotype by PCR-SSP DQB1 and DQA1 typing. Faecal microbiota was analysed by quantitative PCR at 7 days, and at 1 and 4 months of age. Significant interactions between milk-feeding type and HLA-DQ genotype on bacterial numbers were not detected by applying a linear mixed-model analysis for repeated measures. In the whole population, breast-feeding promoted colonization of C. leptum group, B. longum and B. breve, while formula-feeding promoted that of Bacteroides fragilis group, C. coccoides-E. rectale group, E. coli and B. lactis. Moreover, increased numbers of B. fragilis group and Staphylococcus spp., and reduced numbers of Bifidobacterium spp. and B. longum were detected in infants with increased genetic risk of developing CD. Analyses within subgroups of either breast-fed or formula-fed infants indicated that in both cases increased risk of CD was associated with lower numbers of B. longum and/or Bifidobacterium spp. In addition, in breast-fed infants the increased genetic risk of developing CD was associated with increased C. leptum group numbers, while in formula-fed infants it was associated with increased Staphylococcus and B. fragilis group numbers. Overall, milk-feeding type in conjunction with HLA-DQ genotype play a role in establishing infants' gut microbiota; moreover, breast-feeding reduced the genotype-related differences in microbiota composition, which could partly explain the protective role attributed to breast milk in this disorder.     

粪菌移植的现状及未来

粪菌移植是一种直接改变受体肠道微生物群以使其正常化,从而获得治疗效益的方法。自2013年美国食品和药物管理局批准粪菌移植用于治疗复发性和难治性艰难梭菌感染以来,粪菌移植成为研究热点。从此,粪菌移植应用的范围迅速扩大,不仅用于胃肠道疾病,而且用于胃肠外疾病。虽然目前的证据认为粪菌移植是一种普遍安全、副作用少的治疗方法,但其扩大适应症及安全性问题尚未形成共识,需要更深入的研究。本文就目前粪菌移植与胃肠内外疾病的关系及临床安全性问题进行综述,为临床更好地开展粪菌移植提供参考。     

Culture-independent analysis of fecal microbiota in cattle

The phylogenetic diversity of the fecal bacterial community in Holstein cattle was determined by 16S ribosomal RNA gene sequence analysis. The sequences were affiliated with the following phyla: Firmicutes (81.3%), Bacteroidetes (14.4%), Actinobacteria (2.5%), and Proteobacteria (1.4%). The Clostridium leptum subgroup was the most phylogenetically diverse group in cattle feces. In addition, a number of previously uncharacterized and unidentified bacteria were recognized in clone libraries.     

Differences in intestinal microbiota between avian brood parasites and their hosts

The intestinal microbiota determines the effectiveness of digestion in vertebrates, and is influenced by the external environment (mainly the diet), gut characteristics, and phylogeny. Avian brood-parasitic nestlings of the sub-family Cuculinae develop in nests of phylogenetically distant passerines and can be fed with the host diet. If the shaping of bacterial communities is dominated by phylogenetic constraints, and therefore the microbiota of parasitic nestlings differs from that of host nestlings, the energy and micronutrients that parasites and hosts obtain from a similar amount of food would be different. In this case, the bacterial communities of parasitic and host nestlings would have important consequences with respect to brood parasite development. By experimentally creating mixed broods of magpies ( Pica pica ) and great spotted cuckoos ( Clamator glandarius ), we investigated their cloacal microbiota using ribosomal intergenic spacer analysis. We found significant differences in bacterial assemblages of the parasitic and host nestlings, although none of the phylotypes were specific in either great spotted cuckoos or magpies. Cuckoos presented more complex communities, which could help the brood parasitic life style and allow the digestion of food provided by different potential hosts. Moreover, the intestinal morphology is different between the two species due to phylogenetic differences in the two taxa, which would influence the dissimilar bacterial assemblages. The detected differences in microbiota of great spotted cuckoo and magpie nestlings, which might occur in other brood parasite–host systems, may imply a lower digestion efficiency in parasites. Thus, the higher level requirements of cuckoo nestlings may be explained, at least in part, by cuckoos having a suboptimal bacterial community for processing the host diet.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 406–414.     

山西省健康人群的肠道菌群组成特征

目的探索山西省健康人群的肠道菌群组成特征及性别和年龄对肠道菌群组成的影响。方法应用16S rRNA基因测序技术,对山西省99名健康个体的粪便细菌DNA进行测序分析。结果山西省健康人群的肠道菌群在属水平分为两个集群,相对丰度最高的分别是拟杆菌属和普氏菌属;在区分这两个集群中,拟杆菌属与普氏菌属的曲线下面积（AUC）分别是0.97、1.00。男性组与女性组的物种丰富度（richness）和多样性（diversity）差异都无统计学意义（均P>0.05）,基于Bray-Curtis距离的主坐标分析（PCoA）图显示两组人群的样本分布没有明显分离（相似性分析：r=-0.0296,P>0.05）,LEfSe分析显示与性别分组有关的细菌很少。30-39岁、40-49岁和50-59岁三组人群的物种丰富度和多样性差异都无统计学意义（均P>0.05）,基于Bray-Curtis距离的PCoA图显示三组人群的样本分布没有明显分离（相似性分析：r=0.0109,P>0.05）,LEfSe分析显示几乎没有与年龄分组有关的细菌。结论山西省健康人群的肠道菌群更倾向分为两种肠型（拟杆菌型和普氏菌型）。性别对肠道菌群组成可能没有显著影响,30-59岁人群的肠道菌群组成比较稳定     

Effect of a multi-species synbiotic formulation on fecal bacterial microbiota of healthy cats and dogs as evaluated by pyrosequencing

The effect of a multi-species synbiotic on the fecal microbiota of healthy cats (n?=?12) and dogs (n?=?12) was evaluated. The synbiotic (containing 5?×?10(9) CFU of a mixture of seven probiotic strains, and a blend of fructooligosaccharides and arabinogalactans) was administered daily for 21?days. Fecal and serum samples were collected before, during, and up to 3?weeks after administration. Changes in the fecal microbiota were analyzed using denaturing gradient gel electrophoresis, 16S rRNA gene libraries, quantitative real-time PCR, and 16S rRNA gene 454-pyrosequencing. Probiotic species were detectable in 10/12 dogs and 11/12 cats during product administration. Abundances of Enterococcus and Streptococcus spp. were significantly increased in at least one time point during administration, and returned to baseline abundance after treatment was discontinued. No changes in the major bacterial phyla were identified on 454-pyrosequencing. No adverse gastrointestinal effects were recorded and no significant changes in gastrointestinal function or immune markers were observed during the study period. This study shows that while the ingestion of probiotics and prebiotics does not appear to alter the predominant bacterial phyla present in feces, supplementation with the investigated synbiotic leads to an increased abundance of probiotic bacteria in the feces of healthy cats and dogs.     

Structural segregation of gut microbiota between colorectal cancer patients and healthy volunteers

Despite a long-suspected role in the development of human colorectal cancer (CRC), the composition of gut microbiota in CRC patients has not been adequately described. In this study, fecal bacterial diversity in CRC patients (n=46) and healthy volunteers (n=56) were profiled by 454 pyrosequencing of the V3 region of the 16S ribosomal RNA gene. Both principal component analysis and UniFrac analysis showed structural segregation between the two populations. Forty-eight operational taxonomic units (OTUs) were identified by redundancy analysis as key variables significantly associated with the structural difference. One OTU closely related to Bacteroides fragilis was enriched in the gut microbiota of CRC patients, whereas three OTUs related to Bacteroides vulgatus and Bacteroides uniformis were enriched in that of healthy volunteers. A total of 11 OTUs belonging to the genera Enterococcus, Escherichia/Shigella, Klebsiella, Streptococcus and Peptostreptococcus were significantly more abundant in the gut microbiota of CRC patients, and 5 OTUs belonging to the genus Roseburia and other butyrate-producing bacteria of the family Lachnospiraceae were less abundant. Real-time quantitative PCR further validated the significant reduction of butyrate-producing bacteria in the gut microbiota of CRC patients by measuring the copy numbers of butyryl-coenzyme A CoA transferase genes (Mann–Whitney test, P<0.01). Reduction of butyrate producers and increase of opportunistic pathogens may constitute a major structural imbalance of gut microbiota in CRC patients.