Microbial communities in hot springs at high elevations have been extensively studied worldwide. In this sense, the Indian Himalaya regions is valuable ecosystems for providing both the extreme 'cold' and 'hot' sites for exploring microbial diversity. In the present study, a total of 140 thermophilic bacteria were isolated from 12 samples collected from Manikaran and Yumthang hot springs of Indian Himalayas. The bacterial isolates were studied for phylogenetic profiling, growth properties at varying conditions and potential sources of extracellular thermostable hydrolytic enzymes such as protease, amylase, xylanase and cellulase. Based on production of extracellular hydrolases, 51 isolates from Manikaran (28) and Yumthang thermal springs (23) were selected and identified using 16S rRNA gene sequencing which included 37 distinct species of 14 different genera namely Anoxybacillus, Bacillus, Brevibacillus, Brevundimonas, Burkholderia, Geobacillus, Paenibacillus, Planococcus, Pseudomonas, Rhodanobacter, Thermoactinomyces, Thermobacillus, Thermonema and Thiobacillus. Out of 51 hydrolase producing bacteria, 24 isolates showed stability at wide range of temperature and pH treatments. In present investigation, three thermotolerant bacteria namely, Thermobacillus sp NBM6, Paenibacillus ehimensis NBM24 and Paenibacillus popilliae NBM68 were found to produced cellulase-free xylanase. These potential extracellular thermostable hydrolytic enzymes producing thermophilic bacteria have a great commercial prospect in various industrial, medical and agriculture applications.

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Microorganisms, native to the cold environments have successfully acclimatized their physiological, metabolic, and biological features, exhibiting uniqueness in their enzymes, proteins, and membrane structures. These cold-active enzymes have immense biotechnological potential. The diversity of culturable bacteria in two different water lakes (the sub-glacial freshwater and the brackish) of Himalayas was analyzed using SYBR green staining and cultural methods. A total of 140 bacteria were isolated and were grouped as psychrophiles, psychrotrophs, and psychrotolerant organisms, based on their optimal temperature for growth. The amplified ribosomal DNA restriction analysis using three restriction enzymes facilitated the grouping of these isolates into 96 genotypes at ≥85% polymorphism. Phylogenetic analysis using 16S rRNA gene sequences revealed that the bacterial strains from both lakes belonged to Firmicutes, Proteobacteria (α, β, and γ) or Actinobacteria. Screening of the germplasm for the activity of different cold-active hydrolases such as protease, amylase, xylanase, and cellulase, revealed that about 16 isolates were positive, and exhibiting a wide range of stability at various temperature and pH. Our results suggest that the distinctly different ecosystems of sub-glacial freshwater and brackish water lakes have diverse groups of bacteria, which can be an excellent source of extracellular hydrolases with a wide range of thermal stability.

One hundred ten alkalo-tolerant thermophilic bacteria were isolated from 17 samples (water and sediment) collected from Manikaran. Of 110 isolates, 70 showed the production of xylanases and were further screened for growth and production of xylanases at different temperature ranging from 40 to 75°C. Eleven isolates that showed growth and xylanase production at temperatures ≥50°C were selected for quantitative estimation in modified Reese mineral liquid medium containing wheat bran. Maximum xylanase activity was produced by isolate H-7 followed by H-9 and R-9 and was statistically superior to other isolates. The microscopic observation showed that the isolates possessed the typical rod with endospore, characteristic of genus Bacillus. The isolates were found to be oxidase and catalase positive. Using BIOLOG Microlog 3 software, the isolates H7, H9 and R9 were identified as Paenibacillus ehemensis, Bacillus cereus/B. thuringiensis and B. subtilis respectively, based on utilization of 95 carbon sources. PCR-RFLP analysis of 16S rDNA indicated that the isolates were genetically different from each other. DNA sequencing of the three isolates and phylogenetic analysis revealed that all the isolates obtained from Manikaran thermal springs showed 97 to 100% similarity with the sequences within the GenBank. The closest phylogenetic neighbours according to the 16S rRNA gene sequence data for the three isolates H-7, H-9, and R-9 were Paenibacillus ehemensis, Bacillus cereus and Bacillus subtilis, respectively.

Culture dependent phenotypic characterization and 16S rDNA based phylogenetic analyses were applied to study the aerobic halophilic bacterial population present in the Pulicat brackish-water Lake of India. Five different media were employed for isolation of bacteria. A total of 198 morphotypes were recovered, purified and screened for salt tolerance in nutrient agar medium amended with 5-25% NaCl. Based on 16S rDNA restriction fragment length polymorphism analysis with three restriction endonucleases, 51 isolates tolerant to 5% or more NaCl were grouped into 29 clusters. Phylogenetic analysis using 16S rRNA gene sequences revealed that 29 strains could further be allocated into two clades: 19 to Firmicutes and 10 to γ-Proteobacteria. Firmicutes included low G+C Gram-positive bacteria related to family Bacillaceae, which included five genera Bacillus, Virgibacillus, Rummelibacillus, Alkalibacillus and Halobacillus. Another genera included in Firmicutes was Salimicrobium halophilum. In the γ-Proteobacteria group, all the isolates belonged to one genus Halomonas, represented by six different species Halomonas salina, H. shengliensis, H. salifodinae, H. pacifica, H. aquamarina and H. halophila. Most of the isolates exhibited cellulase, xylanase, amylase and protease activities.