Isolation, Identification And Biochemical Characterization Of Halomonas And Bacillus Licheniformis From Deep-sea Hydrothermal Vent In Indian Ocean

There are many extreme microorganisms live in deep sea. Halomonas and Bacillus licheniformis are two of the important groups. We have isolated Halomonas and Bacillus licheniformis from deep-sea hydrothermal vent in Indian Ocean, and studied their biochemical characteristics.Halomonas is a kind of halophilic bacteria that can live in salinity 0.1%-32%. They distribute in saline, salt field, sea ice, marine extreme environment and so on. Halomonas has unique structure and special physiological mechanism, having strong ability of acclimatization to extreme environnment. They have very special characteristics differ from other microorganism and therefore possess a high value for research and utilization.Halomonas sp. strains YD-6, YD-7 and YD-13 were isolated from sediments of the deep-sea hydrothermal vent in Indian Ocean. Results of investigation showed that their appearances were rod and Gram stain was negative. The colonies of three strains were yellow and round. Their surface were smooth and wet, and the center of colonies were orange. The diameters of three colonies were around one mm. YD-7 was chosen for observation under electron microscope. The cell was approximately 1.5-2.0μm long and 0.5-0.7μm wide. Cell growths were observed at 4-55°C, pH 4.0-10.0 and salinity 0-30% respectively. The optimum growth temperature, pH and salinity of YD-6's were 35℃, 6.0 and 6%, so did were 35℃, 7.0 and 5% for YD-7'and 40℃, 7.5 and 5% for YD-13. Three of them all belonged to facultative anaerobe. Phylogenetic analysis based on 16S rDNA gene sequences indicated that they belonged to the genus Halomonas, phylumγ-Proteobacteria. The similarity of Halomonas sp.YD-6 and Halomonas sp. YD-13 was 98.7%, that Halomonas sp.YD-6 and Halomonas meridiana (AJ306891)was 98.8%, Halomonas sp. YD-7 and Halomonas sp. ANT9086(AY167278)was 98.1% and Halomonas sp. YD-13 and Halomonas meridiana(AJ306891)or Halomonas aquamarina(AJ306888)were 98.5%.The results demonstrated that all tested strains had extensive acclimatization. Their range of adaptation to different temperature, salinity and oxygen concentration were broad. Halomonas sp.YD-6, Halomonas sp.YD-7 and Halomonas sp.YD-13 had the ability to resist and remove high Cr(Ⅵ) concentration. In addition, Halomonas sp.YD-7 could resist and remove high Mn(II) concentration,indicating a potentiality in sewage treatment and in bioremediation.Besides, ten strains of Bacillus licheniformis were isolated. Results of investigation showed that the appearances of isolated cells were rod, three strains were long-rod, four strains were short-rod and three strains were medium-rod. Gram stain of four strains were positive and six strains were negative. They belonged to mesophilic bacteria. Cell growth ranges were 10-55°C (optimum around 45°C), pH 5.0 -10.0 (optimum pH 6.0-8.0) and salinity 0.5-6% (optimum around 3%). The result of amylase experiment showed that extracellular enzyme activity of YDGW2 was the biggest, its activity was small under 60℃. When the temperature was above 60℃, enzyme activity increased dramatically. The suitable temperature were 75-90℃and the optimum temperature was 80℃. This amylase should be identified as themosotableα- amylase.