|This thesis is on the Archaea and Bacteria from a hypersaline hyperalkaline lake sediment of a meteorite impact crater named Lonar. We have surveyed the active and the total biodiversity of the sediment from Lonar. We have detected biogeochemically related important taxa and functional genes from the sediment. We have constructed and screened metagenomic libraries for industrially relevant enzymes. We have also investigated this communities’ ability to engineer its microenvironment in terms of pH.
The study site, Lonar crater lake represents a unique environment. As per its origin, Lonar is a meteorite impact crater and as per physicochemical parameters, it is hypersaline and hyperalkaline. This meteorite crater soda lake is located in the southern peninsula of Indian subcontinent known as Lonar lake. Main objectives were to investigate the total (DNA-based) and the active (RNA-based) biodiversity of Archaea and Bacteria, the presence of different taxa and genes (metagenome-based) with their role in biogeochemical cycles, and the ability of the microbial community to engineer their microenvironmental pH.
A total of 85,668 high-quality partial 16S rRNA gene sequences of archaeal and 182,137 sequences of bacterial origin were recovered and analyzed. In Archaea, the total and the active community diversity, a coverage of 74.21 % and 84.07% was observed. The total and the active community diversity of Bacteria showed a coverage of 59.78% and 88.98% respectively. Among the Archaea at the order level, most dominant taxa was Halobacteriales. Halobacteriales is mostly represented by Natronococcus, which was also the most dominant genera both in the total and the active community diversity. In the case of Bacteria most dominant phyla was Firmicutes and the genera were Alkaliphilus and Bacillus. Both of these genera represents Firmicutes. Upon comparison of all the previous studies on Lonar lake and this investigation, more than 67 % of all bacterial and archaeal genera are unique to this study and were not observed in previous investigations.
In our study, we have observed 24 genera, for example, Methanosaeta and Methylobacterium, which may have been involved in methane cycle. In Archaea, they contribute to an average of 39.24 % relative abundance in the active community. In the case of Bacteria, they contribute to an average of 0.50 % relative abundance in the active community. A total of 16 genera, for example, Ammonifex and Nitratireductor, were found which may have been involved in the nitrogen cycle. Among Archaea, they contribute to an average of 1.37 % relative abundance in the active community. In the case of Bacteria, they contribute to an average of 2.87 % relative abundance in the active community. A significantly high diversity of bacterial genera, totaling 36, involved in the sulfur cycle were recorded, for example, Desulfococcus and Thioalkalivibrio. They represent an average relative abundance of 0.93 % relative abundance in the active community.
A total of 32 million paired-end reads were obtained from direct metagenome sequencing. Analysis of the metagenome resulted in 588,668 contigs, with a total number of base of 371 Mb (371,120,372 bases). Several ORFs involved in these biogeochemical cycles were detected. The predicted relative abundance of ORFs in relation to methane metabolism, nitrogen metabolism, and sulfur metabolism pathways were found to be 1.49 %, 0.50 %, and 0.68 %. Experimental data mapped on the reference pathways provides a comprehensive overall view of methane, nitrogen and sulfur metabolism in the sediments of Lonar crater lake.
A total of 235,943 archaeal and 1,657,168 bacterial partial 16S rRNA gene sequences were recovered from the different time point of the nonoptimal pH exposure of the sediments. The microbial community, from the Lonar meteorite crater soda lake sediments, was exposed to suboptimal and superoptimal pH conditions. The change of pH of the culture filtrate was monitored. Community dynamics was also measured at a resolution of 5 days for a total of 25 days using high-throughput 16S rRNA gene analysis. We observed an average coverage of 71.04 % in Archaea and 85.56 % in Bacteria. We have seen a 10-fold change in the initial hydrogen ion concentration difference to a point between suboptimal and superoptimal pH. Several archaeal and bacterial taxa at phylum (Bacteria) or order (Archaea) level and genus (both Archaea and Bacteria) level have been identified to modulate significantly upon exposure to nonoptimal pH. Several of them regained their original or extremely close to their original relative abundance with the progression of time. Also, from HPLC analysis, it is evident that metabolism of ammonia and hydroxyproline have a function in this community dynamics and eventual microenvironmental pH homeostasis. However, we were not able to confirm if this observation is due to correlation or causality. It was also observed, that the dynamics of several archaeal and bacterial genera can be grouped in to different types of dynamic groups based on their changing relative abundances. We found two types of dynamic groups in Archaea and four types of dynamic groups in Bacteria. Considering, all these observations, it might be safe to speculate that this microbial community can change their microenvironment to a more favorable (hypothetical optimal) one in terms of pH at the same time resisting permanent change in its community structure.