However, the storage conditions had a large impact on the taxonomic composition of the samples at the genus and species level for all subjects (figure 2B). Variations were found depending on both the storage
condition and the individual. In Table 2, we showed the effect of storage conditions on the proportion of 3 main bacterial taxa. selleck chemical As shown in this table, the abundance comparison between frozen and selleck inhibitor unfrozen samples was affected by thawing samples for 1 h and 3 h as exemplified by the significant decrease of a dominant unknown taxon from the Bacteroides genus (from an average of 19% (F) to 13% (UF1h; p = 0.044, Poisson regression model) and to 9% (UF3h; p < 0.0001, Poisson regression model)). The proportion of the two other bacterial taxa was significantly affected when thawing the
samples over 3 h (p = 0.02 and p = 0.0007 respectively, Poisson regression model). The room temperature condition was only significantly affecting the bacterial proportion after 2 weeks (p < 0.04 for all taxa, Poisson regression model) as shown in Table 3. Figure 2 Bacterial community analysis based on 16S rRNA gene survey. A) Alpha-diversity analysis of number of species observed in 6 storage conditions: Immediately frozen (F); unfrozen 1 h and 3 h (UF1h, UF3h); room temperature 3 h, 24 h, and 2 weeks check details (RT3h, RT24h, RT2w). The plot averages the number of species from the samples provided by 4 individuals in each condition. B) Taxonomy analysis at the species level of the 24 samples based on alignment performed using PyNast against Silva 108 release database and OTUs assignment using blast and the Silva 108 release taxa mapping file. Individual #1 (red), #2 (blue), #3 (green), #4 (purple). A more detailed taxonomy assignment is provided in the additional data (See Additional file 3: Table S1). C) UPGMA clustering of the 24 samples based on weighted UniFrac method. Samples Nintedanib (BIBF 1120) from the 4 individuals are colored as in B. The scale bar
represents 2% sequence divergence. Table 2 Taxonomic comparison for 3 main bacterial taxa between frozen and unfrozen samples Taxon F* UF1h* UF3h* p value F vs UF1h p value F vs UF3h Bacteroides;uncultured bacterium 19 13 9 0.044 9.68e-05 Prevotellaceae;uncultured;human gut metagenome 7 6 3 0.6804 0.0222 Bifidobacterium;uncultured bacterium 2 4 8 0.2257 0.0007 Statistical analysis was performed using Poisson regression model; p value < 0.05 is considered significant; n = 4 subjects; * Values are mean proportion of sequences (%). F = frozen; UF1h = unfrozen during 1 h; UF3h = unfrozen during 3 h; Taxonomy is indicated at the genus level and if not possible at the family level.