Study On The Psychrophiles Diversity And Phylogenetic Analysis In Zoige Plateau Wetland

Zoige Plateau Wetland is located at the northeastern Tibetan Plateau with global climate change most sensitive. Zoige soil microbes were affected by high altitude, cold, hypoxia, precipitation, sunshine length and radiation intensity, so it has Special biodiversity. The study of Psychrophiles Domestic is concentrated on the Oceans, the polar environment, Glacier. The report on Psychrophiles of Zoige Plateau Wetland, the particular geographical environment, has not been found. We selected soil sample of eight different sampling sites of Zoige Plateau Wetland as experimental material, pure culture and culture-independent methods were used to analyze the soil physical and chemical properties, microbial biomass carbon and nitrogen, the enzyme activity of Psychrophilic bacteria, phylogeny and diversity of psychrophilic bacteria.The main Results were as follows:1. The results suggest that the soil nutrients in different sampling sites of Ruoergai plateau with different impact on soil microbial quantity and activity. The value of soil physical and chemical, Colony-Forming Units, microbial biomass are different. Correlation analysis revealed that microbial biomass carbon was significant relationship with shannon index and colony-forming units, it has no relationship with soil physical and chemical properties, microbial biomass nitrogen was significant relationship with shannon index and soil organic matter, Microbial carbon and nitrogen reflect an effective indicator of microbial activity. E011has the highest value of microbial biomass carbon and nitrogen,it also has the highest value of shannon index,it indicated that the soil samples of Zoige Wetland which has richer bacterial populations the higher microbial activity it has.2. In this study, All the16psychrophile strains has been found can produced cold-adapted enzyme,after the Enzyme Activity Assay aboutβ-galactosidase, Lecithinase, Chitinase, Amylase,Celluase,Caseinase,Esterase, and all can produce cellulase enzyme.is shows that cellulose play an important role in growth and development as one of nutrients in microorganism. The ability of better produce Celluase is REGP1, REGP3and REGP13have the better ability of produce Caseinase, REGP2and REGP16have the better ability of produce Esterase, The ability of better produce Chitinase is REGP1.Therefore, there are abundant and various microbial resources which can produce cold-adapted enzyme in bog soil of the zoige wetland plateau as a perfect enzyme-producing place3.37of psychrotrophic bacteria obtained by growth temperature tests from87bacteria cultured at4℃. The definition of psychrophilic bacteria is that it must live in low temperature conditions, growth and reproduction at0℃, the optimum temperature does not exceed15℃,the maximum temperature does not exceed20℃There is only42%of the low-temperature-resistant bacteria called psychrophilic bacteria from isolated strains. four enzyme digestion analysis was used by37strains, only16strains have different restriction bands. It shows that43%of isolated strains with different DNA subsequent, then DNA sequencing.4. Through purification and phylogeny research for psychrophile,we found that coating plate after enrichment is better for psychrophile isolating and getting more strain number of psychrophile than coating plate directly.6of the16strains of culture psychrophile isolated from plate are Gram-positive bacteria,10strains are Gram-negative bacteria whom is dominant group.the optimum temperature of the16strains of psychrophile is range from10℃to15℃6strains of culture psychrophile are isolated from E011,2strains are isolated from BO11,3strains are isolated from DO11,1strain is isolated from G011and3strains are isolated from H011,all have higher Shannon Wiener diversity index by DGGE analysis.through phylogeny analysis after sequencing the16s rDNA of the16strains of culture psychrophile,the Gram-positive culture psychrophile were clustered into three categories,belongs to Arthrobacter,Exiguobacterium and Sporosarcina respectively.Gram-negative culture psychrophile were clustered into four categories,respectively belongs to Variovorax,Janthinobacterium,Pseudomonas and Stenotrophomonas.β-,γ-proteobacteria are dominant groups.Pseudomonas sp. is greatest contrubite to biodiversity of wetland of Ruoergai plateau,following is Arthrobacter.The present study stated indicated that the zoige wetland plateau is the natural place for culture preservation and abundant in biodiversity of the culture cold adapted microorganism.5. Study found that the most of bacterial is belong to uncultured bacteria in this eare,through Shannon Wiener diversity index.Band X7-1is similar to Uncultured Oxalobacteraceae,but band X7-3is similar to Sphingobacteriales, it show that Sphingobacteriales and Uncultured Oxalobacteraceae may be the dominant bacteria in Zoige Plateau Wetland which has special topography and geomorphology. Band X7-4is similar to Janthinobacterium sp. belong to (3-proteobacteria, Janthinobacterium sp. has also been Isolated from psychrophile medium.The results show that the DGGE play some roles in Culture and isolation of psychrophile.