The
structure of sojourners gut microbiota was analyzed at different
heights, measuring the abundance of bacteria by WGS, representing known
sequences at phyla level more than 1% using Wilcoxon rank test (odds
ratio at 95% confidence intervals). There were eight phyla present in
the fecal samples at different heights (Figure 3). Phyla Bacteriodetes
was predominantly present, accounting for 80% of the identified taxa at
H1 time point. The second richest phyla was Fermicutes with 18%
presence. The next detectable phyla was 2% Proteobacteria (Figure 3 A).
After ascending to and six months stay at H2, there was a significant
decrease in Bacteriodetes to 64%, though still remaining the richest
phyla, increase in Fermicutes contributing 35% with Proteobacteria
remaining the same. The microbial composition varied immensely,
depicting an interesting phenomenon when sojourners descended to H3 and
stayed for two months. There was a sudden 5% appearance of Fusobacteria
and increase in Proteobacteria to 6%. Moreover, the predominance of
Bacteriodetes was reversed and increased to 76%, whereas Fermicutes
reduced to 8%. At the end when sojourners ascended to H4 and stayed for
four months, the newly appeared phyla Fusobacteria at H3, suddenly
disappeared remaining with Bacteriodetes, Fermicutes and Proteobacteria
77%, 13% and 3% respectively. Other bacterial members from
Actinobacteria, Verrucomicrobia, Lentisphaereae, Euryarchaeota
were also detected with lower abundance.
Further, confirmation of the variation in the composition of fecal
microbiota of sojourners at different heights was achieved by linear
discriminant analyses (LEfSe) using the Greengene software
(http://greengenes.lbl.gov). FDR correction of the p value was to
determine statistical significance (p < 0.05/number of tests)
and considered p < 0.05 statistically significant for
reproducibility. Figure 3D indicates the association between the
relative abundance of presence or absence of phyla and the significant
change in the microbiota mainly with abundance of Bacteriodetes
(p= 0.05), Fermicutes (p= 0.05) and Proteobacteria (p=0.05). LEfse analysis also showed concordant result with Wilcoxon rank
test (Supplementary FigureS3). The results thus indicate the most
important contributing factor to the diversity could be altitude or
hypoxia as diet and ethnicity of all the subjects were the same.
The presence of eight phyla by WGS analysis in limited three subjects
were reproduced by 16s RNA analysis in another 16 subjects from the same
cohort at H1 and H2 heights (p<0.05 Wilcoxon rank test) with
odds ratio of 95% (Figure 3B). The second method reconfirmed the
significant changes in the abundance of phyla Bacteriodetes and
Fermicutes (Figure3C).