Project description:Metagenomic sequencing of mice with different treatments: Mice were randomly divided into donor control group (Donor + MRS), constipation model group (STC + MRS), or a Lactobacillus acidophilus treated group (STC + La): A humanized mouse model was established by intragastric administration of fecal bacterial liquid from healthy donors or STC patients on alternate days, followed by continuous administration of Lactobacillus acidophilus in treatment group. Finally, the feces of each group of mice were collected, and the intestinal microbial communities of the mice were analyzed through metagenomic sequencing. 16S rRNA sequencing of mice before and after the use antibiotics: Before and after treating the mice with antibiotics, the mice's feces were collected for 16s rRNA sequencing respectively.
Project description:The characterization of microbial community structure via 16S rRNA gene profiling has been greatly advanced in recent years by the application of amplicon pyrosequencing. The possibility of barcode-tagged sequencing of templates gives the opportunity to massively screen multiple samples from environmental or clinical sources for community details. However, an on-going debate questions the reproducibility and semi-quantitative rigour of pyrotag sequencing and, as in the early days of genetic community fingerprinting, pros and cons are continuously provided. In this study we investigate the reproducibility of bacterial 454 pyrotag sequencing over biological and technical replicates of natural microbiota. Moreover, via quantitatively defined template spiking to the natural community, we explore the potential for recovering specific template ratios within complex microbial communities. For this reason, we pyrotag sequenced three biological replicates of three samples, each belonging from yearly sampling campaigns of sediment from a tar oil contaminated aquifer in Düsseldorf, Germany. Furthermore, we subjected one DNA extract to replicate technical analyses as well as to increasing ratios (0, 0.2, 2 and 20%) of 16S rRNA genes from a pure culture (Aliivibrio fisheri) originally not present in the sample. Unexpectedly, taxa abundances were highly reproducible in our hands, with max standard deviation of ~3% abundance across biological and ~2% for technical replicates. Furthermore, our workflow was also capable of recovering A. fisheri amendmend ratios in reliable amounts (0, 0.29, 3.9 and 23.8%). These results highlight that pyrotag sequencing, if done and evaluated with due caution, has the potential to robustly recapture taxa template abundances within environmental microbial communities. 9 Biological and 3 technical replicates were evaluated, as well as potential to recover qPCR-defined ratios of DNA, in 454 pyrotag sequencing
Project description:The characterization of microbial community structure via 16S rRNA gene profiling has been greatly advanced in recent years by the application of amplicon pyrosequencing. The possibility of barcode-tagged sequencing of templates gives the opportunity to massively screen multiple samples from environmental or clinical sources for community details. However, an on-going debate questions the reproducibility and semi-quantitative rigour of pyrotag sequencing and, as in the early days of genetic community fingerprinting, pros and cons are continuously provided. In this study we investigate the reproducibility of bacterial 454 pyrotag sequencing over biological and technical replicates of natural microbiota. Moreover, via quantitatively defined template spiking to the natural community, we explore the potential for recovering specific template ratios within complex microbial communities. For this reason, we pyrotag sequenced three biological replicates of three samples, each belonging from yearly sampling campaigns of sediment from a tar oil contaminated aquifer in Düsseldorf, Germany. Furthermore, we subjected one DNA extract to replicate technical analyses as well as to increasing ratios (0, 0.2, 2 and 20%) of 16S rRNA genes from a pure culture (Aliivibrio fisheri) originally not present in the sample. Unexpectedly, taxa abundances were highly reproducible in our hands, with max standard deviation of ~3% abundance across biological and ~2% for technical replicates. Furthermore, our workflow was also capable of recovering A. fisheri amendmend ratios in reliable amounts (0, 0.29, 3.9 and 23.8%). These results highlight that pyrotag sequencing, if done and evaluated with due caution, has the potential to robustly recapture taxa template abundances within environmental microbial communities.
Project description:Total bacterial DNA was isolated from water and sediment samples from a local watershed and 16S rRNA sequences were analyzed using the Illumina MiSeq v3 platform in order to generate snapshots of bacterial community profiles. A total of 56 samples were collected that represent water and sediment samples from 14 sample sites over two different time points (November 18 and 25, 2011).
Project description:We investigated a contaminant-degrading microbial community by sequencing total RNA (without rRNA depletion) from microcosms containing sediment from a hypoxic contaminated aquifer fed with isotopically labeled toluene.
Project description:The impact of mono-chronic S. stercoralis infection on the gut microbiome and microbial activities in infected participants was explored. The 16S rRNA gene sequencing of a longitudinal study with 2 sets of human fecal was investigated. Set A, 42 samples were matched, and divided equally into positive (Pos) and negative (Neg) for S. stercoralis diagnoses. Set B, 20 samples of the same participant in before (Ss+PreT) and after (Ss+PostT) treatment was subjected for 16S rRNA sequences and LC-MS/MS to explore the effect of anti-helminthic treatment on microbiome proteomes.
Project description:Waste decomposition in landfills is a complex and microbe-mediated process. Understanding the microbial community composition and structure is critical for accelerating decomposition and reducing adverse impact on the environment. Here, we examined the microbial communities along with landfill depth and age (LDA) in a sanitary landfill in Beijing, China using 16s rRNA Illumina sequencing and GeoChip 4.6. We found that Clostridiales and Methanofollis were the predominant bacteria and archaea in the present landfill, respectively. Interestingly, in contrast with the decreasing trend of microbial diversity in soil, both phylogenetic and functional diversities were higher in deeper and older refuse in the landfill. Phylogenetic compositions were obviously different in the refuse with the same LDA and such difference is mainly attributed to the heterogeneity of refuse instead of random process. Nevertheless, functional structures were similar within the same LDA, indicating that microbial community assembly in the landfill may be better reflected by functional genes rather than phylogenetic identity. Mantel test and canonical correspondence analysis suggested that environmental variables had significant impacts on both phylogenetic composition and functional structure. Higher stress genes, genes for degrading toxic substances and endemic genes in deeper and older refuse indicated that they were needed for the microorganisms to survive in the more severe environments. This study suggests that landfills are a repository of stress-resistant and contaminant-degrading microorganisms, which can be used for accelerating landfill stabilization and enhancing in situ degradation.
Project description:Waste decomposition in landfills is a complex and microbe-mediated process. Understanding the microbial community composition and structure is critical for accelerating decomposition and reducing adverse impact on the environment. Here, we examined the microbial communities along with landfill depth and age (LDA) in a sanitary landfill in Beijing, China using 16s rRNA Illumina sequencing and GeoChip 4.6. We found that Clostridiales and Methanofollis were the predominant bacteria and archaea in the present landfill, respectively. Interestingly, in contrast with the decreasing trend of microbial diversity in soil, both phylogenetic and functional diversities were higher in deeper and older refuse in the landfill. Phylogenetic compositions were obviously different in the refuse with the same LDA and such difference is mainly attributed to the heterogeneity of refuse instead of random process. Nevertheless, functional structures were similar within the same LDA, indicating that microbial community assembly in the landfill may be better reflected by functional genes rather than phylogenetic identity. Mantel test and canonical correspondence analysis suggested that environmental variables had significant impacts on both phylogenetic composition and functional structure. Higher stress genes, genes for degrading toxic substances and endemic genes in deeper and older refuse indicated that they were needed for the microorganisms to survive in the more severe environments. This study suggests that landfills are a repository of stress-resistant and contaminant-degrading microorganisms, which can be used for accelerating landfill stabilization and enhancing in situ degradation. Fifteen refuse samples with five landfill depths and ages (6m/2a, 12m/4a, 18m/6a, 24m/8a and 30m/10a) were collected from a sanitary landfill in Beijing, China. Three replicates in every landfill depth and age
Project description:Total bacterial DNA was isolated from water and sediment samples from a local watershed and 16S rRNA sequences were analyzed using the Illumina MiSeq v3 platform in order to generate snapshots of bacterial community profiles.
Project description:Comparison of probe-target dissociations of probe Eub338 and Gam42a with native RNA of P. putida, in vitro transcribed 16s rRNA of P. putida, in vitro transcribed 16S rRNA of a 2,4,6-trinitrotoluene contaminated soil and an uncontaminated soil sample. Functional ANOVA revealed no significant differences in the dissociation curves of probe Eub338 when hybridised to the different samples. On the opposite, the dissociation curve of probe Gam42a with native RNA of P. putida was significantly different than the dissociation curves obtained with in vitro transcribed 16S rRNA samples. Keywords: Microbial diversity, thermal dissociation analysis, CodeLink microarray