Abundance and diversity of mucosa-associated hydrogenotrophic microbes in the healthy human colon

Hydrogenotrophic microbiota have a significant impact on colonic health; however, little is known about their diversity and ecology in situ. Here, molecular-based methods and multivariate analyses were used to examine the abundance and diversity of mucosa-associated hydrogenotrophic microbes in 90 biopsies collected from right colon, left colon and rectum of 25 healthy subjects. Functional genes of all three hydrogenotrophic groups were detected in at least one colonic region of all subjects. Methanogenic archaea (MA) constituted approximately one half of the hydrogenotrophic microbiota in each colonic region. Sulfate-reducing bacteria (SRB) were more abundant than acetogens in right colon, while acetogens were more abundant than SRB in left colon and rectum. MA genotypes exhibited low diversity, whereas SRB genotypes were diverse and generally similar across the three regions within subject but significantly variable among subjects. Multivariate cluster analysis defined subject-specific patterns for the diversity of SRB genotypes; however, neither subject- nor region-specific clusters were observed for the abundance of hydrogenotrophic functional genes. Sequence analyses of functional gene clones revealed that mucosa-associated SRB were phylogenetically related to Desulfovibrio piger, Desulfovibrio desulfuricans and Bilophila wadsworthia; whereas MA were related to Methanobrevibacter spp., Mb. smithii and the order Methanomicrobiales. Together these data demonstrate for the first time that the human colonic mucosa is persistently colonized by all three groups of hydrogenotrophic microbes, which exhibit segmental and interindividual variation in abundance and diversity.     

新疆特殊生境岩石内生细菌末端限制性片段长度多态性技术分析

【目的】了解新疆特殊生境不同类型岩石内生细菌的组成及多样性。【方法】采用末端限制性片段长度多态性技术(Terminal Restriction Fragment Length Polymorphism,T-RFLP),分析新疆乌苏花岗岩(1号样)、一号冰川和木垒变质岩(2,3号样)、裕民和托克逊岩石漆(4,5号样)内生细菌群落。【结果】样品间多样性指数变化不大;聚类分析表明岩石类型相同,其相似性较高,2号样和3号样聚为一支并与1号样再聚为一支,4号样与5号样聚为一支;各样品共有种群为厚壁菌门(Firmicutes)、放线菌门(Actinobacteria)、变形菌门(Proteobacteria)和拟杆菌门(Bacteroidetes),1号样存在酸杆菌门(Acidobacteria),2号样存在浮霉菌门(Planctomycetes);除5号样优势类群为放线菌门(29.3%),其它4个样品均为变形菌门,只是所占比例略有不同。【结论】生境不同的同类型岩石的内生细菌群落组成存在差异,各类岩石中可能存在大量未知细菌新种。     

Diversity of the particulate methane monooxygenase gene in methanotrophic samples from different rice field soils in China and the Philippines

Methanotrophic bacteria play a crucial role in regulating the emission of CH4 from rice fields into the atmosphere. We investigated the CH4 oxidation activity together with the diversity of methanotrophic bacteria in ten rice field soils from different geographic locations. Upon incubation of aerated soil slurries under 7% CH4, rates of CH4 oxidation increased after a lag phase of 1-4 days and reached values of 3-10 micromol d(-1) g-dw(-1) soil. The methanotrophic community was assayed by retrieval of the pmoA gene which encodes the a subunit of the particulate methane monooxygenase. After extraction of DNA from actively CH4-oxidizing soil samples and PCR-amplification of the pmoA, the community was analyzed by Denaturant Gradient Gel Electrophoresis (DGGE) and Terminal Restriction Fragment Length Polymorphism (T-RFLP). DGGE bands were excised, the pmoA re-amplified, sequenced and the encoded amino acid sequence comparatively analyzed by phylogenetic treeing. The analyses allowed the detection of pmoA sequences related to the following methanotrophic genera: the type-I methanotrophs Methylobacter, Methylomicrobium, Methylococcus and Methylocaldum, and the type-II methanotrophs Methylocystis and Methylosinus. T-RFLP analysis detected a similar diversity, but type-II pmoA more frequently than DGGE. All soils but one contained type-II in addition to type-I methanotrophs. Type-I Methylomonas was not detected at all. Different combinations of methanotrophic genera were detected in the different soils. However, there was no obvious geographic pattern of the distribution of methanotrophs.     

Environmental factors have a strong impact on the composition and diversity of the gut bacterial community of Chinese black honeybees

The Chinese black honeybees is an ecotype of the European honeybee that is formed by the natural hybridization of Apis mellifera mellifera and A. m. carnica. It is distributed in nature reserves in North China and has been an important breeding resource for disease resistance and other desirable traits. Compared to the areas outside of reserves, the nature reserves offer significant biodiversity benefits not only to the Chinese black honey bees but also to the other valuable plants and animals. In recent years there has been growing evidence that environmental factors including food choices play an important role in shaping the composition and activity of gut microbiota, which in turn can impact host health. In the previous studies on Chinse black honeybees, little attention has been paid to the diverse population of microbes in the gut that play a vital role in host health. In order to achieve a better understanding on the role of environmental factors in diversity and composition of gut microbiota of honey bees, in the present study, we analyzed the gut bacterial communities of Chinese black honeybees using terminal restriction fragment length polymorphism (T-RFLP). The results showed that the samples from the national nature reserves that are protected and managed so as to preserve and enrich their natural condition and resources for Chinese black honeybees had higher variety and richness of gut bacteria than that collected from unreserved regions that also harbor populations of Chinese black honeybees. The four terminal restriction fragments (T-RFs), 201, 223, 247 and 320 bp, were identified to be the dominant bacteria of Chinese black honeybees. Of which 247 and 320 bp had greater differences between bee groups sampled in different regions and therefore could be used as genetic markers to separate samples collected from the national nature reserves to samples collected from unreserved regions. The results clearly indicate that national nature reserve protects biological diversity and ecological and evolutionary processes which have had a significant influence on the diversity of gut bacteria of Chinese black honeybees. The ubiquity of gut symbiotic bacteria identified in Chinese black honeybee suggests that environmental factors could play an important role in diversity and composition of gut bacteria and warrant further investigation into the functional significance of these gut bacteria for the honeybee health.     

The relationship between microbiota and polyamine concentration in the human intestine: a pilot study

The fecal microbiota of 10 hospitalized elderly subjects and 14 healthy adults were analyzed by terminal-restriction fragment length polymorphism (T-RFLP) analysis using Hha I, Msp I, Hae III, and Alu I, as well as fecal polyamine (PA) concentration. The T-RFLP profiles of the fecal microbiota of the subjects were roughly divided into 2 clusters-I (9 out of 11 were derived from hospitalized elderly subjects) and II (12 out of 13 were derived from healthy adults). The average concentration of putrescine in Cluster II was 5.8 times higher than that of putrescine in Cluster I (P=0.0015). Using a phylogenetic assignment database for T-RFLP analysis of human colonic microbiota, the terminal-restriction fragments (T-RFs) characteristically detected in the case of subjects with high fecal PA concentration were predicted to be derived from bacterial species and phylotypes belonging to Clostridium subcluster XIVa, particularly including Clostridium xylanolyticum, Clostridium saccharolyticum, the uncultured human intestinal bacterium clone JW1H4 (a relative of Desulfotomaculum guttoideum), Roseburia intestinalis, the uncultured bacterium clone 41F10 (a relative of Eubacterium ramulus), Roseburia cecicola, Ruminococcus obeum and its relatives. From these results, we concluded that fecal microbiota may be linked with fecal PA concentration and that some bacterial species belonging to Clostridium subcluster XIVa may play a major role in the control of intestinal PA concentration in humans.     

Community structure of Archaea and Bacteria in a profundal lake sediment Lake Kinneret (Israel)

The microbial community structure of an anoxic profundal lake sediment, i.e., subtropical Lake Kinneret, was analysed with respect to its composition by culture-independent molecular methods including terminal restriction fragment length polymorphism (T-RFLP) analysis, comparative sequence analysis, and quantitative real-time PCR. In particular we were interested in the structure, species composition, and relative abundance of the overall microbial community in the methanogenic sediment layer (0-10 cm depth). Pairwise comparison of archaeal and bacterial 16S rRNA gene T-RFLP profiles obtained from three independent samplings indicated stability of the microbial community. The numbers of Archaea and Bacteria, quantified by real-time PCR, amounted to about 10(8) and 10(10) 16S rRNA gene copies cm(-3) sediment, respectively, suggesting that Archaea may account for only a minor fraction (approximately 1%) of the total prokaryotic community. Hydrogenotrophic Methanomicrobiales and acetoclastic Methanosaeta spp. dominated T-RFLP profiles of the archaeal community. T-RFLP profiles of the bacterial community were dominated by Deltaproteobacteria, sulphate reducers and syntrophs in particular. The second most abundant group was assigned to the Bacteroidetes-Chlorobi-group. Only one bacterial group, which was affiliated with halorespiring bacteria of subphylum II of the Chloroflexi, showed variation in abundance within the sediment samples investigated. Our study gives a comprehensive insight into the structure of the bacterial and archaeal community of a profundal lake sediment, indicating that sulphate reducers, syntrophs, bacteroidetes, halorespirers and methanogens are of particular importance in Lake Kinneret sediment.     

Diversity of fungi in hair roots of Ericaceae varies along a vegetation gradient

Ericaceous dwarf shrubs including Calluna vulgaris and Vaccinium spp. occur both in open heathland communities and in forest ecosystems as understory vegetation. Ericaceous shrubs were once thought to form ericoid mycorrhizal associations with a relatively narrow range of ascomycetous fungi closely related to, and including, Rhizoscyphus ericae. However, perceptions have recently changed since the realization that a broader range of ascomycete fungi, and in some cases basidiomycete fungi, can also form associations with the roots of ericaceous plants. We used a combination of molecular approaches, including denaturing gradient gel electrophoresis, terminal restriction fragment length polymorphism, cloning and sequencing, to investigate the diversity of fungi associated with C. vulgaris roots collected across a heathland/native Scots pine forest vegetation gradient. We also determined differences in fungal community composition between roots of co-occurring C. vulgaris and Vaccinium myrtillus in the forest understory. Collectively, the data show that a large diversity of potentially ericoid mycorrhizal fungal taxa associate with roots of C. vulgaris and V. myrtillus, and that ascomycetes were about 2.5 times more frequent than basidiomycetes. The assemblages of fungi associated with C. vulgaris and V. myrtillus were different. In addition, the community of fungi associated with C. vulgaris hair roots was different for samples collected from the forest, open heathland and a transition zone between the two. This separation was partly, but not entirely, due to the occurrence of typical ectomycorrhizal basidiomycetes associated with the hair roots of C. vulgaris in the forest understory. These data demonstrate that forest understory ericaceous shrubs associate with a diverse range of ascomycete and basidiomycete taxa, including typical ectomycorrhizal basidiomycetes.     

长白山红松针阔叶混交林主要树种树干呼吸速率

采用土壤呼吸气室,于2003年4—10月原位测定了长白山红松针阔叶混交林主要树种红松、蒙古栎、水曲柳和紫椴的树干呼吸, 监测了树干温度及林内温度. 结果表明: 4个树种的树干呼吸速率均有明显的季节变化, 其中7月的呼吸速率最高, 10月最低, 呈单峰曲线. 各树种的树干呼吸速率日变化均为“S”型曲线, 并在4:00达到最低,而呼吸速率峰值出现时间有所不同,红松、蒙古栎、水曲柳和紫椴的呼吸速率峰值分别在18:00、20:00、16:00和14:00.不同树种的树干呼吸对温度变化的响应也不相同, 其树干呼吸Q₁₀值在2.24～2.91之间变化,为水曲柳＞蒙古栎＞红松＞紫椴.     

微生物生态学一种新研究方法-T-RFLP技术

T RFLP是建立在PCR基础之上一种新的微生物生态学研究方法。该方法克服了传统微生物培养方法的限制、应用快速、灵敏度高且输出定量的数据结果 ,被广泛应用到菌种鉴定、群落对比分析、群落中系统发育种群多样性的评估等领域。目前我国仍没有关于此方法应用的相关报道 ,但作为一种研究微生物群落特征的理想方法 ,T RFLP已经越来越受到相关研究人员的重视。该文主要介绍了该方法的基本原理、阐述了该方法的关键技术及其应用发展现状。