Diversity Of Thermophiles In The Biotopes Of Terrestrial Hot Springs And Deep-sea Hydrothermal Sediment

Diverse thermophiles are mostly isolated from various thermal environments on earth,among which terrestrial hot springs and deep-sea hydrothermal vents are most typical.Aiming at the species resource of thermophiles,prokaryotic communities exist in such two kinds of geothermal biotopes were analyzed in this paper.A set of samples were collected from several hot springs which are respectively located in different geothermal areas of China.Thermophilic bacteria were recovered by normal culture-dependant technique.Finally,totally 138 strains,which were moderately to extremely thermophilic,were isolated and preserved properly.Phylogenetic analysis based on 16S rRNA gene sequences indicated that all strains were respectively affiliated to genus Geobacillus,Anoxybacillus,Brevibacillus, Thermus or Meiothermus.Bacterial community of each hot spring was dominated by either Geobacillus or Anoxybacillus which are both members of Bacillus sensu lato. However,interspecific diversity of each hot spring was different because of the geographic interval.Moreover,the results of enzymological tests showed highly intraspecific diversity of thermophilic bacilli,some of which had potential in biotechnological application,such as industrial biocatalysis(thermostable enzymes) and biodegradation(environmental treatment).Two gram-negative novel species were found in hot springs that respectively located in northeastern and southestern geothermal areas of China.Based on the polyphasic analysis,they were described as Meiothermus cateniformans sp.nov.(type strain LY1~T= CGMCC 1.6990~T= JCM15151~T,reference strain L462 = CGMCC 1.6989 = JCM 15150) and Thermus arciformis sp.nov.(type strain TH92~T= CGMCC 1.6992~T= JCM15153~T,reference strain TH91 = CGMCC 1.6991 = JCM 15152). Another potential novel species represented by thermophilic and spore-forming strain 3nP-4 was isolated from hot spring in western Sichuan.Phylogenetic analysis based on complete sequences of 16S rRNA genes indicated that strain 3nP-4 diverged from the lineage of Anoxybacillus by forming the deepest branch.A sediment sample collected at EPR 13°N hydrothermal field(12°42′40″N, 103°54′25″W) was used for prokaryotic community analysis with culture-independent method.Diverse 16S rRNA genes were cloned directly from megagenome.Totally 52 bacteial clones and 50 archaeal ones were selected randomly to establish the 16S rRNA gene library.All 16S rRNA gene sequences were checked for chimera or anomaly before used for aligning and phylogenetic analysis.As a phylotype was defined as a group of clones which have sequence similarity higher than 97%and simultaneously cluster together in phylogenetic tree,all the 102 clones were classified into 26 bacterial phylotypes and 14 archaeal ones,which were respectively affiliated to Crenarchaeote (including marine groupⅠ),Euryarchaeote(including marine groupⅡ),α-Proteobacteria,γ-Proteobacteria,Firmicutes,Planctomycetes,Gemmatimonadetes, Actinobacteria,Acidobacteria and an uncultured bacterial group named candidate division OP11.Both Euryarchaeote andγ-Proteobacteria were the dominant groups of the prokaryotic community.Gemmatimonadetes-related sequences were found for the first time in the deep-sea hydrothermal sediment.As for the potential novel phylotypes existed in the hydrothermal sediment, complete sequences of their typical clones were determined and then rechecked for chimera or anomaly.As a result,all sequences were authentic.Besides,further sequence analysis indicated that clones EPR7-21 and EPRJ22 should respectively represent a novel unknown lineage of archaea and of bacteria while clones EPR7-8 and EPR1-9 should represent two novel euryarchaeal phylotypes.