Three 16S rRNA gene clone libraries (P1L, P4L and P8L) were constructed using three soil samples (P1S, P4S and P8S) collected near Pindari glacier, Himalayas. The three libraries yielded a total of 703 clones. Actinobacteria, Firmicutes and Proteobacteria were common to the three libraries. In addition to the above P1L and P8L shared the phyla Acidobacteria, Bacteroidetes, Gemmatimonadetes and Planctomycetes. Phyla Chlamydiae, Chlorobi, Chloroflexi, Dictyoglomi, Fibrobacteres, Nitrospirae, Verrucomicrobia, candidate division SPAM and candidate TM7s TM7a phylum were present only in P1L. Rarefaction analysis indicated that the bacterial diversity in P4S and P8S soil samples was representative of the sample. Principal component analysis (PCA) revealed that P1S and P8S were different from P4S soil sample. PCA also indicated that arsenic content, pH, Cr and altitude influence the observed differences in the percentage of specific OTUs in the three 16S rRNA gene clone libraries. The observed bacterial diversity was similar to that observed for other Himalayan and non-polar cold habitats. A total of 40 strains of bacteria were isolated from the above three soil samples and based on the morphology 20 bacterial strains were selected for further characterization. The 20 bacteria belonged to 12 different genera. All the isolates were psychro-, halo- and alkalitolerant. Amylase and urease activities were detected in majority of the strains but lipase and protease activities were not detected. Long chain, saturated, unsaturated and branched fatty acids were predominant in the psychrotolerant bacteria.

Strain PN5 T is a Gram-positive, aerobic, motile, rod-shaped, peritrichously flagellated bacterium that was isolated from the Pindari glacier using nutrient agar medium. Cells of PN5 T are catalase-positive and oxidase-negative and contain lysine, glutamic acid and alanine in the peptidoglycan (peptidoglycan type A4a). Further, the cells are characterized by the presence of iso-C 15 : 0 and iso-C 16 : 1 as the predominant fatty acids and MK-7 as the isoprenoid quinone. Based on the above characteristics, strain PN5 T was assigned to the genus Bacillus. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain PN5 T clustered with the type strain of Bacillus silvestris with a sequence similarity of 97.2 %. DNA-DNA hybridization between PN5 T and B. silvestris DSM 12223 T resulted in a relatedness of only 15 %, clearly indicating that strain PN5 T represents a novel species. Further, PN5 T was different from B. silvestris with respect to various phenotypic and chemotaxonomic characteristics. Therefore, strain PN5 T is identified as a representative of a novel species of the genus Bacillus, for which the name Bacillus cecembensis sp. nov. is proposed. Bacillus cecembensis is unique among psychrotolerant Bacillus species in containing L-Lys-D-Glu in the cell-wall peptidoglycan. The type strain is PN5 T (5LMG 23935 T 5MTCC9127 T 5JCM 15113 T ).The genus Bacillus was created by Cohn (1872), with Bacillus subtilis as the type species. Members of the genus Bacillus are distinguished from other endospore-forming bacteria on the basis of being strict or facultative aerobes, rod-shaped, (usually) catalase-positive and they contain meso-diaminopimelic acid (meso-Dap) in the peptidoglycan, MK-7 as the major menaquinone and iso-C 15 : 0 and anteiso-C 15 : 0 as major fatty acids and have DNA G+C contents of 32-69 mol%. Interestingly, Bacillus is the only genus that contains organisms with L-lysine (Lee et al., 2006;Rheims et al., 1999) (Bae et al., 2005). Although Bacillus species have been isolated from a wide range of habitats, only one Bacillus species with a validly published name, Bacillus lehensis (Ghosh et al., 2007), has been isolated from the Himalayan region. In the present study, a unique psychrotolerant Bacillus strain (PN5 T ) containing L-lysine as the diamino acid in the peptidoglycan was isolated from a soil sample of the Pindari glacier of the Indian Himalayas; based on polyphasic taxonomic characterization, it is identified as representing a novel species.Strain PN5 T was isolated from a soil sample collected close to the Pindari glacier at an altitude of approximately 3500 m. Approximately 200 mg of the soil sample was suspended in 0.9 % saline, subjected to shaking for 1 h and allowed to settle and 100 ml was plated on nutrient agar plates [0.3 % beef extract, 0.5 % peptone, 0.8 % NaCl and 1.5 % agar (all w/v)]. The viable bacterial cell count was approximately 2.1610 5 c.f.u. and, based on colony morphology, 40 different morphotypes were observed. In the present study, one of the morp...