1, which showed a similar diversity but distinct abundance of Operational Taxonomic Units (OTUs) (Figure 1, Additional file 1: Table S1). This observation was confirmed by an Unweighted Selleckchem MLN2238 Pair Group Method with Arithmetic Mean (UPGMA) clustering analysis based on unweighted and weighted UniFrac distances (Figure 2). Sample LO4.1 from subject #4 was the only one that clustered far from the other find more samples from the same stool when both microbial composition and abundance were considered (weighted UniFrac
analysis, Figure 2B). Figure 1 Spatial organization of the microbial community (species level) in stool specimens. 250 mg of stool (N = 8) was collected in the outer (LO) and inner area (LI) layer and once the stool had been homogenised (LH). Stools were collected in duplicates for each condition. Figure 2 UPGMA clustering based on weighted (A) and unweighted UniFrac (B) distance analysis. 250 mg of stool (N = 8) was collected from the outer (LO) and inner (LI) layers and after the stool
had been homogenised (LH). Stools were collected in duplicates for each condition (48 samples in total). Unweighted UniFrac allows clustering by taking into account only the microbial composition, while weighted UniFrac considers both composition and abundance of OTUs. Effect GSK2399872A clinical trial of stool water content To evaluate how stool water content affects the microbial community, we analysed the 46 samples from four out of the eight participants, as described in the experimental design section above. After the extraction procedure, genomic DNA was loaded in an Agilent 2100 Bioanalyzer chip in order to evaluate integrity. A comparison of the DNA extracted from DL1 samples (presence
of beads and PBS) with those of DL1B’s (presence of beads but not PBS) showed that the addition of PBS caused greater genomic DNA degradation (Figure 3A). This finding was confirmed by a decrease in DNA size to lower than 10 Mbp with 125 mg of stool (sample DL1.50, Table 1) and 50% PBS. In contrast, in the absence of PBS this degradation was also observed but only when the stool weighed 62.5 mg (DL1B.75). Interestingly, we observed a double effect of stool water content and bead-beating when dealing with a small amount of stool matter. Figure 3 Effect of water content on genomic DNA integrity. (A) Gel electrophoresis CHIR-99021 supplier analysis. For each sample, genomic DNA equivalent to 1 mg of faecal sample was loaded on an Agilent 2100 Bioanalyzer chip using the Agilent 12000 kit. DL1 corresponds to participant L1 from the homogenisation evaluation. (B) Microbial diversity at the species level. The taxonomic analysis was performed using a cut-off of 97% similarity. The “#” followed by a number indicates an arbitrary identifier for an unknown OTUs. Although the presence of PBS could increase the degradation of genomic DNA, the microbial community profile was not affected at the species level (Figure 3B).