The handling and treatment of biological samples is critical when characterizing the composition of the intestinal microbiota between different ecological niches or diseases. Specifically, exposure of fecal samples to room temperature or long term storage in deep freezing conditions may alter the composition of the microbiota. Thus, we stored fecal samples at room temperature and monitored the stability of the microbiota over twenty four hours. We also investigated the stability of the microbiota in fecal samples during a six month storage period at −80°C. As the stability of the fecal microbiota may be affected by intestinal disease, we analyzed two healthy controls and two patients with irritable bowel syndrome (IBS). We used high-throughput pyrosequencing of the 16S rRNA gene to characterize the microbiota in fecal samples stored at room temperature or −80°C at six and seven time points, respectively. The composition of microbial communities in IBS patients and healthy controls were determined and compared using the Quantitative Insights Into Microbial Ecology (QIIME) pipeline. The composition of the microbiota in fecal samples stored for different lengths of time at room temperature or −80°C clustered strongly based on the host each sample originated from. Our data demonstrates that fecal samples exposed to room or deep freezing temperatures for up to twenty four hours and six months, respectively, exhibit a microbial composition and diversity that shares more identity with its host of origin than any other sample. 相似文献
Two novel gull-specific quantitative PCR (qPCR) assays were developed using 16S rRNA gene sequences from gull fecal clone libraries: a SYBR green assay targeting Streptococcus spp. (gull3) and a hydrolysis TaqMan assay targeting Catellicoccus marimammalium (gull4). The objectives of this study were to compare the host specificity of a previous C. marimammalium qPCR assay (gull2) with that of the new markers and to examine the presence of the three gull markers in environmental water samples from different geographic locations. Most of the gull fecal samples tested (n = 255) generated positive signals with the gull2 and gull4 assays (i.e., >86%), whereas only 28% were positive with gull3. Low prevalence and abundance of tested gull markers (0.6 to 15%) were observed in fecal samples from six nonavian species (n = 180 fecal samples), whereas the assays cross-reacted to some extent (13 to 31%) with other (nongull) avian fecal samples. The gull3 assay was positive against fecal samples from 11 of 15 avian species, including gull. Of the presumed gull-impacted water samples (n = 349), 86%, 59%, and 91% were positive with the gull2, the gull3, and the gull4 assays, respectively. Approximately 5% of 239 non-gull-impacted water samples were positive with the gull2 and the gull4 assays, whereas 21% were positive witg the gull3 assay. While the relatively high occurrence of gull2 and gull4 markers in waters impacted by gull feces suggests that these assays could be used in environmental monitoring studies, the data also suggest that multiple avian-specific assays will be needed to accurately assess the contribution of different avian sources in recreational waters. 相似文献
Inulin is a well-known fructose-based prebiotic which has been shown to stimulate the growth of bifidobacteria, a bacterial group generally considered beneficial for intestinal health. In the present study, we analyzed inulin-associated shifts in the total bacterial community of wild-type mice and mice carrying a genetically inactivated adenomatous polyposis coli tumor suppressor gene by using DNA-based approaches independent of bacterial culturability. Mice were fed a high-fat, nonfiber diet with or without inulin inclusion at a 10% (wt/wt) concentration. Cecal contents were analyzed after 0, 3, and 9 weeks on the experimental diets. Inulin inclusion significantly affected the total bacterial community structure of the cecum as determined by both a nonselective percent-guanine-plus-cytosine-based profiling analysis and a more specific 16S ribosomal DNA sequence analysis. The shifts included stimulation of bifidobacteria and suppression of clostridia, but sequence comparison revealed that the major shifts were within previously unknown bacterial taxa. Concomitantly, significantly higher bacterial densities, determined by flow cytometry, were observed with the inulin-amended diet, and the metabolism of the cecal bacterial community was altered, as indicated by higher levels of residual short-chain fatty acids, particularly lactic acid. With regard to all of the microbiological parameters measured, the wild-type mice and mice carrying a genetically inactivated adenomatous polyposis coli tumor suppressor gene were essentially identical. Studies of the implications of pre- and probiotics may need to be expanded to include careful analysis of their effects on the entire microbial community, rather than just a few well-known species. Further studies are needed to increase our understanding of the possible roles of currently unknown gastrointestinal bacteria in health and disease. 相似文献
The phylogenetic diversity of the microbial community assemblage of the carpet-like mucilaginous cyanobacterial blooms in the eutrophic Lake Taihu was investigated. 16S ribosomal DNA clone libraries produced from the DNA of cyanobacterial assemblages that had been washed to remove unattached bacteria contained only cyanobacteria. However, a further treatment which included grinding the freeze-dried material to physically detach cells followed by the removal of larger cells by filtration allowed us to detect a large variety of bacteria within the cyanobacterial bloom community. Interestingly, the dominant members of the microbial community were Planctomycetes followed by Cytophaga–Flavobacterium–Bacteroides (CFB), Betaproteobacteria, and Gammaproteobacteria. The analysis of the 16S ribosomal DNA clone libraries made from enrichment culture revealed much higher phylogenetic diversity of bacteria. Dominant bacterial groups in the enrichment system were identified as members of the Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Deltaproteobacteria subdivisions, CFB group, and Planctomycetes. In addition, the clone libraries constructed from Planctomycetes-specific 16S ribosomal RNA primers also verified that the enrichment allowed a diversity of Planctomycetes to proliferate, although the community composition was altered after enrichment. 相似文献
Root exudate composition and quantity vary in relation to plant nutritional status, but the impact of the differences on rhizosphere microbial communities is not known. To examine this question, we performed an experiment with barley (Hordeum vulgare) plants under iron-limiting and iron-sufficient growth conditions. Plants were grown in an iron-limiting soil in root box microcosms. One-half of the plants were treated with foliar iron every day to inhibit phytosiderophore production and to alter root exudate composition. After 30 days, the bacterial communities associated with different root zones, including the primary root tips, nonelongating secondary root tips, sites of lateral root emergence, and older roots distal from the tip, were characterized by using 16S ribosomal DNA (rDNA) fingerprints generated by PCR-denaturing gradient gel electrophoresis (DGGE). Our results showed that the microbial communities associated with the different root locations produced many common 16S rDNA bands but that the communities could be distinguished by using correspondence analysis. Approximately 40% of the variation between communities could be attributed to plant iron nutritional status. A sequence analysis of clones generated from a single 16S rDNA band obtained at all of the root locations revealed that there were taxonomically different species in the same band, suggesting that the resolving power of DGGE for characterization of community structure at the species level is limited. Our results suggest that the bacterial communities in the rhizosphere are substantially different in different root zones and that a rhizosphere community may be altered by changes in root exudate composition caused by changes in plant iron nutritional status. 相似文献
Inactivation (loss of culturability) by sunlight of enterococci and fecal coliforms within sewage effluent diluted in seawater was investigated in field experiments. In most experiments, 500-ml flasks of pure silica were used to confine activated sludge effluent diluted to 2% (vol/vol) in seawater. Inactivation of bacteria in these flasks (diameter, 0.1 m) was faster than in either open chambers (depth, 0.25 m) or patches of dyed effluent (depth of order, 1 m), probably because of the longer light paths in the latter two types of experiment, which caused greater attenuation of sunlight. Inactivation of 90% of enterococci generally required 2.3 times the insolation required for 90% inactivation of fecal coliforms, because of both the presence of larger initial shoulders on survival curves and a lower final inactivation rate. Two parameters are required to model inactivation of enterococci, a shoulder constant as well as a rate coefficient. The depth dependence of inactivation rate for both fecal indicators matched the attenuation profile of UV-A radiation at about 360 nm. Inactivation by UV-B radiation (290 to 320 nm), which penetrates much less into seawater, is of minor importance compared with the UV-A and visible radiation in sunlight, contrary to expectations in consideration of published action spectra for bacterial inactivation. 相似文献
The animal gastrointestinal tract contains a complex community of microbes, whose composition ultimately reflects the co-evolution of microorganisms with their animal host. An analysis of 78,619 pyrosequencing reads generated from pygmy loris fecal DNA extracts was performed to help better understand the microbial diversity and functional capacity of the pygmy loris gut microbiome. The taxonomic analysis of the metagenomic reads indicated that pygmy loris fecal microbiomes were dominated by Bacteroidetes and Proteobacteria phyla. The hierarchical clustering of several gastrointestinal metagenomes demonstrated the similarities of the microbial community structures of pygmy loris and mouse gut systems despite their differences in functional capacity. The comparative analysis of function classification revealed that the metagenome of the pygmy loris was characterized by an overrepresentation of those sequences involved in aromatic compound metabolism compared with humans and other animals. The key enzymes related to the benzoate degradation pathway were identified based on the Kyoto Encyclopedia of Genes and Genomes pathway assignment. These results would contribute to the limited body of primate metagenome studies and provide a framework for comparative metagenomic analysis between human and non-human primates, as well as a comparative understanding of the evolution of humans and their microbiome. However, future studies on the metagenome sequencing of pygmy loris and other prosimians regarding the effects of age, genetics, and environment on the composition and activity of the metagenomes are required. 相似文献
Diverse sessile organisms inhabit the coral reef ecosystems, including corals, sponges, and sea anemones. In the past decades, scleractinian corals (Cnidaria, Anthozoa, Scleractinia) and their associated microorganisms have attracted much attention. Zoanthids (Cnidaria, Anthozoa, Zoanthidea) are commonly found in coral reefs. However, little is known about the community structure of zoanthid-associated microbiota. In this study, the microbial community associated with the zoanthid Palythoa australiae in the South China Sea was investigated by 454 pyrosequencing. As a result, 2,353 bacterial, 583 archaeal, and 36 eukaryotic microbial ribotypes were detected, respectively. A total of 22 bacterial phyla (16 formally described phyla and six candidate phyla) were recovered. Proteobacteria was the most abundant group, followed by Chloroflexi and Actinobacteria. High-abundance Rhizobiales and diverse Chloroflexi were observed in the bacterial community. The archaeal population was composed of Crenarchaeota and Euryarchaeota, with Marine Group I as the dominant lineage. In particular, Candidatus Nitrosopumilus dominated the archaeal community. Besides bacteria and archaea, the zoanthid harbored eukaryotic microorganisms including fungi and algae though their diversity was very low. This study provided the first insights into the microbial community associated with P. australiae by 454 pyrosequencing, consequently laid a basis for the understanding of the association of P. australiae–microbes symbioses. 相似文献
Limestone aquifers provide the main drinking water resources of southern Italy. The groundwater is often contaminated by fecal bacteria because of the interaction between rocks having high permeability and microbial pollutants introduced into the environment by grazing and/or manure spreading. The microbial contamination of springwater in picnic areas located in high mountains can cause gastrointestinal illness. This study was carried out in order to analyze the interaction between Enterococcus faecalis and the soil of a limestone aquifer and to verify the influence of this interaction on the time dependence of groundwater contamination. E. faecalis was chosen because, in the study area involved, it represents a better indicator than Escherichia coli. The research was carried out through field (springwater monitoring) and laboratory experiments (column tests with intact soil blocks). The transport of bacterial cells through soil samples was analyzed by simulating an infiltration event that was monitored in the study area. Comparison of laboratory results with data acquired in the field showed that discontinuous precipitation caused an intermittent migration of microorganisms through the soil and produced, together with dispersion in the fractured medium (unsaturated and saturated zones), an articulated breakthrough at the spring. The short distances of bacterial transport in the study area produced a significant daily variability of bacterial contamination at the field scale. 相似文献
The objective of this study was to analyze human fecal Lactobacillus community and its relationship with rheumatoid arthritis. Samples taken from rheumatoid arthritis (RA) patients and healthy individuals were analyzed by quantitative real-time PCR. Bacterial DNA was extracted from feces, and amplicons of the Lactobacillus-specific regions of 16S rRNA were analyzed by denaturing gradient gel electrophoresis. The richness, Shannon-Wiener index, and evenness of gut microbiota of both groups were analyzed to compare fecal Lactobacillus community structures. Results of this study demonstrated that fecal microbiota of RA patients contained significantly more Lactobacillus (10.62 ± 1.72 copies/g) than the control group (8.93 ± 1.60 copies/g). Significant increases were observed in RA patients in terms of the richness, Shannon-Wiener, and evenness measures, indicating more bacterial species, and increased bacterial diversity and abundance. These results suggest a potential relationship between Lactobacillus communities and the development and progression of rheumatoid arthritis. 相似文献
Kartchner Caverns in Benson, AZ, was opened for tourism in 1999 after a careful development protocol that was designed to
maintain predevelopment conditions. As a part of an ongoing effort to determine the impact of humans on this limestone cave,
samples were collected from cave rock surfaces along the cave trail traveled daily by tour groups (200,000 visitors year–1) and compared to samples taken from areas designated as having medium (30–40 visitors year–1) and low (2–3 visitors year–1) levels of human exposure. Samples were also taken from fiberglass moldings installed during cave development. Culturable
bacteria were recovered from these samples and 90 unique isolates were identified by using 16S rRNA polymerase chain reaction
and sequencing. Diversity generally decreased as human impact increased leading to the isolation of 32, 27, and 22 strains
from the low, medium, and high impact areas, respectively. The degree of human impact was also reflected in the phylogeny
of the isolates recovered. Although most isolates fell into one of three phyla: Actinobacteria, Firmicutes, or Proteobacteria,
the Proteobacteria were most abundant along the cave trail (77% of the isolates), while Firmicutes predominated in the low
(66%) and medium (52%) impact areas. Although the abundance of Proteobacteria along the cave trail seems to include microbes
of environmental rather than of anthropogenic origin, it is likely that their presence is a consequence of increased organic
matter availability due to lint and other organics brought in by cave visitors. Monitoring of the cave is still in progress
to determine whether these bacterial community changes may impact the future development of cave formations. 相似文献
Broiler chickens from eight commercial farms in Southern Finland were analyzed for the structure of their gastrointestinal microbial community by a nonselective DNA-based method, percent G+C-based profiling. The bacteriological impact of the feed source and in-farm whole-wheat amendment of the diet was assessed by percent G+C profiling. Also, a phylogenetic 16S rRNA gene (rDNA)-based study was carried out to aid in interpretation of the percent G+C profiles. This survey showed that most of the 16S rDNA sequences found could not be assigned to any previously known bacterial genus or they represented an unknown species of one of the taxonomically heterogeneous genera, such as Ruminococcus or Clostridium. The data from bacterial community profiling were analyzed by t-test, multiple linear regression, and principal-component statistical approaches. The percent G+C profiling method with appropriate statistical analyses detected microbial community differences smaller than 10% within each 5% increment of the percent G+C profiles. Diet turned out to be the strongest determinant of the cecal bacterial community structure. Both the source of feed and local feed amendment changed the bacteriological profile significantly, whereas profiles of individual farms with identical feed regimens hardly differed from each other. This suggests that the management of typical Finnish farms is relatively uniform or that hygiene on the farm, in fact, has little impact on the structure of the cecal bacterial community. Therefore, feed compounders should have a significant role in the modulation of gut microflora and consequently in prevention of gastrointestinal disorders in farm animals. 相似文献
Consistently high recoveries of enterococci as compared to the low numbers of coliforms obtained from the same samples of frozen sea foods are indirect evidence that enterococci are better indicators of contamination in such foods.
The use of azide dextrose broth, modified by the incorporation of bromthymol blue, and of ethyl violet azide broth as presumptive and confirmation tests, respectively, were found to be highly specific for the detection and enumeration of enterococci in these samples. Tetrazolium agar medium, when used as a third step after the confirmation test, provides a reliable differentiation of Streplococcus faecalis types from other group D streptococci. A simple procedure is described for further identification of S. faecalis varieties and other enterococcal species.
Incidence of biotypes within certain species is noted and relationships of these subgroups to the organisms described by other workers is discussed.
The striking resistance of all group D streptococci to dihydrostreptomycin and polymyxin B seems to offer promise for evolving a new selective medium for these organisms.