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The Himalayan Mountains are placed among the globally recognized biodiversity hot spots. While the Indian Himalayan Region (IHR) has been subjected to extensive studies on plant and animal biodiversity, microbial diversity is now being studied for its bioprospection. The present paper deals with the evaluation of bacterial diversity in high-altitude soil samples from IHR following polyphasic approach including comparison between the MALDI-TOF mass spectrometry and 16S rRNA gene sequencing for species-level identification. Initially, a culture collection of large number of bacterial isolates was established in the laboratory. Performing morphological and biochemical screenings, sixty-one representative isolates were selected for mass spectrometry and gene sequencing. Both the methods emerged with bacterial identification showing maximum number of Bacillus followed by Pseudomonas species. The other frequently isolated strains belonged to the genera Alcaligenes, Carnobacterium, Lysinibacillus, Microbacterium, Paenarthrobacter, Rhodococcus, Serratia and Stenotrophomonas. Although the MALDI-TOF technique appeared to be advantageous as less time-consuming in comparison with 16S rRNA-based method, the discrepancies at species level indicated the limited database of MALDI Biotyper and species complexity in the genera. The remarkable characteristics of the bacterial isolates were their tolerance to wide range of pH and temperature. Their potential to produce industrially valuable enzymes indicated their importance in bioprospection. Accessioning of these bacterial isolates in microbial culture collections is a cautious effort for their availability to conduct advanced research on these cold-adapted bacteria in future.

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