Studies On The Diversity Of The Culturable Bacteria From Kongsfjorden And The Polyphasic Taxonomy Of Two Flavobacterial Strains

Kongsfjorden is an inlet on the west coast of Spitsbergen (a part of the Svalbard archipelago in the Arctic Ocean), covering about209km2and is a typical fjord developed at the edge of the high arctic continental ice. This area contains huge numbers of marine microorganisms with extremely diversities. These microorganisms have evolved unique physiological and metabolic characteristics and have important ecological functions due to the area’s special geographical position, climate and environment. In this study, bacterial strains were isolated from different samples from the Arctic Kongsfjorden including surface seawater, marine bottom and intertidal sandy sediments samples and diversity of these culturable bacteria were studied; two novel flavobacterial strains isolated were then polyphasically characterized, based on which their exact taxonomic positions were finally determined.1. Diversity of the culturable bacteria fom KongsfjordenA total of295bacterial strains were isolated from various samples from the Arctic Kongsfjorden. The16S rRNA genes of the295strains were amplified and sequenced. Analysis of these16S rRNA gene sequences revealed that the isolated bacterial strains from surface seawater samples belonged to y-Proteobacteria (Pseudoalteromonas, Vibrio, Alteromonas and Psychrobacter), a-Proteobacteria (Sphingorhabdus), Bacilli (Bacillus), Actinobacteria (Kouria and Rhodococcus) and Flavobacteriia (Maribacter, Cellulophage, Flavobacterium, Bizionia and Olleya); the isolated bacterial strains from marine bottom sediment samples belonged to y-Proteobacteria (Pseudoalteromonas, Enterobacteria, Vibrio, Photobacterium, Psychromonas, Alteromonas and Psychrobacter), a-Proteobacteria (Sphingorhabdus), Actinobacteria (Microbacterium), Bacilli (Bacillus) and Flavobacteriia (Flavobacterium, Aestuariicola, Algibacter, Formosa, Bizionia and Polaribacter); the isolated bacterial strains from the intertidal sandy samples belonged to γ-Proteobacteria (Pseudoalteromonas, Psychromonas, Shewanella, Psychrobacter, Marinomonas, Pseudomonas) and Bacilli (Planococcus).2. Polyphasic taxonomic study of strain SM1202TStrain SM1202T was isolated from marine bottom sediment collected from the Arctic Kongsfjorden. Strain SM1202T formed orange, circular and convex colonies with smooth surfaces on the TYS agar plates. Cells of strain SM1202T were gram-negative, non-flagellated, oxidase-and catalyse positive rods and able to secrete abundant extracellular polysaccharide. Strain SM1202T grew at4-35℃(optimum at25℃), in1-5%NaCl (w/v, optimum in2.0%) and at pH5.5-8.5(optimum at pH7.0). Strain SM1202T could not reduce nitrate to nitrite. It hydrolyzed casein, starch and Tween20but not agar, DNA, gelatin, Tweens40,60,80or aesculin.The major cellular fatty acids of strain SM1202T were iso-C15:0, iso-C15:03-OH, iso-C13:0, C15:0, iso-C15:1G, iso-C17:03-OH and C15:1ω6c. Its respiratory quinone was MK-6and polar lipids included phosphatidylethanolamine (PE), one unidentified lipid (L), two unidentified aminophosphalipids (APL1-2) and one aminolipid (AL). The genomic DNA G+C content of strain SM1202T was36.4mol%.Strain SM1202T was found to have the highest16S rRNA gene sequence similarity with type strain of P. dokdonensis (94.2%) and92.7%～93.9%sequence similarities to type strains of other recognized Polaribacter species. In the phylogenetic trees based on the16S rRNA gene sequences, strain SM1202T clustered with members of the genus Polaribacter and formed a distinct phylogenetic branch at the periphery of the cluster.On the basis of the polyphasic data from this study, strain SM1202T represents a novel species in the genus Polaribacter of the family Flavobacteriaceae, for which the name Polaribacter huanghezhanensis sp. nov. is proposed. The type strain of Polaribacter huanghezhanensis is SM1202T (=CCTCC AB2013148T=KCTC32516T).3. Polyphasic taxonomic study of strain SM1203TStrain SM1203T was isolated from marine surface seawater of Kongsfjorden. The bacterium formed yellow-colored, circular and convex colonies with smooth surfaces on the TYS agar plates. Cells of strain SM1203T were Gram-negative, aerobic, non-flagellated, oxidase-negative and catalyse-positive rods which were able to secrete extracellular polysaccharide. The strain grew at4-30℃(optimum at20℃), with1.0-5.0%(w/v) NaCl (w/v, optimum in2.0%) and at pH5.5-8.5(optimum at pH7.0). It could not reduce nitrate to nitrite. The strain hydrolyzed casein, starch, Tweens20, 40,80and gelatin but not agar, DNA, Tween60or aesculin.The major fatty acids of strain SM1203T were iso-C15:0, iso-C15:1, anteiso-C15.0and C15:0and the main polar lipids were phosphatidylethanolamine (PE), two unidentified lipids (L1-2), one unidentified phospholipid (PL), one unidentified aminophospholipids (APL) and three unidentified aminolipids (AL1-AL3).Comparative analysis of the16S rRNA gene sequence showed that strain SM1203T shared the highest16S rRNA gene sequence similarities (>96%) with Formosa spongicola (96.8%) and Bizionia paragorgiae (96.3%). Sequence similarities with other recognized Bizionia species were95.4～96.3%. The phylogenetic trees based on16S rRNA gene sequences showed that strain SM1203T was grouped into the genus Bizionia. The genomic DNA G+C content of strain SM1203T was34.8mol%.On the basis of the polyphasic data from this study, strain SM1203T represents a novel species in the genus Bizionia, for which the name Bizionia arctica sp. nov. is proposed. The type strain of Bizionia arctica is SM1203T (=CGMCC1.12751T=CCTCC AB2013360T).