The clone library analysis this website showed that Firmicutes and Bacteroidetes are the dominant phyla present in human

gut flora in our subjects and also confirmed the results of DGGE analysis showing that different bacterial genera are dominating the gut flora in different aged individuals as shown in Figure  3. The clone library analysis with Sanger sequencing has limitations of having low depth of sequencing as compared to Next generation sequencing technologies AG-120 clinical trial like pyrosequencing, however longer read length obtained by Sanger sequencing are beneficial when mapping the sequence to the species level [40]. Fewer than 100 sequences are enough to detect the pattern of variation among the microbial communities in gut of diverse hosts [40–42]. Although clone library analysis

would not yield total bacterial diversity, it would give the variation in major bacterial groups within the samples. Recently Zupancic et al. reported bacterial genera which forms the core gut microbiota of Amish subjects [43]. We retrieved the sequences for almost all the genera defined as core microbiota by Zupancic et al. in our study. This further supports the fact that clone library analysis could be useful in determining the variation in major bacterial phyla in a sample. A study by Mariat et al. on European Population showed that the Firmicutes /Bacteroidetes ratio being 0.4 in Infants which increases to 10.9 in Carnitine palmitoyltransferase II adults and decreases to see more 0.6 in elderly [16]. Somewhat different results were observed by Biagi et al. in Italian population, the Firmicutes /Bacteroidetes ratio for adults 3.9 which increased to 5.1 for elderly and decreased to 3.6 for centenarians respectively [44]. Moving from young to elderly the Firmicutes /Bacteroidetes ratio was observed to be decreased in Mariat et al. study while it increased in Biagi et al. study [16, 44]. In contrast, in our study we observed a consistent decrease in Firmicutes number and increase in Bacteroidetes number with increasing age. This was observed

in the clone library analysis and then validated by qPCR. The decrease in Firmicutes number and increase in Bacteroidetes suggest that there would be a gradual decrease in Firmicutes /Bacteroidetes ratio in our subjects with increasing age which further implies that our subjects do not follow the same trend of change in Firmicutes /Bacteroidetes ratio with age as to what has been reported earlier in European population. Isolation of strict anaerobes from one of the family showed age related differences in the culturable anaerobic diversity. To the best of our knowledge this is the first study focusing on age related changes in culturable anaerobic diversity from Indian subcontinent.