Browsing by Author "Saurabh Kumar"
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ThesisItem Open Access Documentation of the bacterial and diazotrophic diversity from Garhwal Himalaya through culturable and unculturable approaches(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2018-07) Saurabh Kumar; Goel, ReetaTo study the bacterial diversity and soil physiochemical properties of Garhwal Himalaya, total five soil samples were collected from different locations of Garhwal Himalaya namely Harsil, Gangotri, Uttarkashi, Dhanaulti and Kandakhal. Based on the primary screening through quantification of 16SrDNA and nifH genes and 16SrDNA DGGE analysis, Gangotri and Dhanaulti were selected for further study. Comparative soil physiochemical characterization of the both soil samples revealed that the Kandakhal soil was neutral in pH, poor in soil organic matter (SOM) and total Kjeldal nitrogen (TKN) and rich in micronutrient. However, Gangotri soil was found alkaline, rich in TKN and SOM and poor in mineral nutrients. Thus, it was found that the soil physicochemical properties vary significantly with the altitude. Metagenomics of Kandakhal soil revealed that the Proteobacteria (31%) was the major phylum followed by Bacteroidetes (20%), Fermicutes (9%), Acidobacteria (7%), Actinobacteria (6%) and Chloroflexi (5%). However, in phylum Proteobacteria, Alphaproteobacteria (8%), Betaproteobacteria (8%), Gammaproteobacteria (8%) and Deltaproteobacteria (7%) were uniformly distributed. On the other hand in Gangotri, total 31 bacterial phyla were observed where, Proteobacteria (38.49 %), Acidobacteria (17.88 %), Actinobacteria (14.48 %), Bacteroidetes (7.89 %), Gemmatimonadetes (7.87 %) and Chloroflexi (5.94 %) were dominant bacterial phyla. However, in phylum Proteobacteri, Alphaproteobacteria (16.88%) was the most abundant bacterial class followed by Betaproteobacteria (9.44%). Presence of large numbers of unidentified OTU in Gangotri soil suggests that it has unique bacteria diversity. Further, an indigenous diffusion chamber was designed and incubated in-situ for cultivation of readily not cultivable bacterial from Gangotri soil which revealed the potential of this device for isolation of bacteria which were previously difficult to isolate from conventional methods. Moreover, six psychrophilic diazotrophic bacteria were isolated form Gangotri soil and identified as Pseudomonas helmanticensis, Arthrobacter humicola, Brevibacillus invocatus and Pseudomonas mandelii etc. However, no psychrophilic diazotrophic were isolated from the Kandakhal soil. Further investigation was carried out to study the cold adaptation in Pseudomonas helmanticensis at molecular level. This was the first time when proteomics of psychrophilic diazotrophs Pseudomonas helmanticensis was reported under the cold stress. Differential proteomics of P. helmanticensis under cold stress through LC-MS analysis revealed that majority of upregulated and unique proteins expressed under cold stress were functionally involved in RNA metabolism, translation and energy production. Moreover, major upregulated proteins were molecular chaperones which assist in the proper protein folding and RNA stability at low temperature. Therefore, both culture dependent and independent studies revealed that the Kandakhal and Gangotri soil harbor rich bacterial diversity. However, diversified culturable psychrophilic nitrogen fixers were only present in the Gangotri soil. Thus, Garhwal region of the western Indian Himalaya is rich in cold adapted bacterial diversity which could be explored for basic understanding of cold adaptation besides its applied significance.