A Study Of The Microbial Diversity In The Cold Seep Of South China Sea And Description Of A Novel Bacterial Species

Deep sea is the largest habitat with rich biological resource,accounting for about two-thirds of the earth's surface.However,due to the difficulty of research,knowledge about this ecological environment and its biological feature is very limited.The cold seep system,a fluid formed by the leakage of hydrocarbon from the seabed,is an"oasis"in the barren environment of the deep sea,widely existing in the world.Most of cold seeps are characterized by high concentrations of methane and hydrogen sulfide.Microorganisms,such as sulfur-oxidizing bacteria,sulfate-reducing bacteria and methane-oxidizing archaea,play extremely important roles in sulfur and methane cycling process.In this study,both culture-independent and culture-dependent methods were used to analyze the microbial diversity in the water and sediment samples collected in 2017 from the cold seep located at southwest Taiwan of the South China Sea.Through 16S rDNA high-throughput sequencing technology,the microbial structure in the water and sediment samples from the cold deep,non-leakage area and non-cold seep area of the South China Sea were analyzed.The results showed that there was a unique microbial structure in the cold seep area.In the cold seep area,the relative abundance of microorganisms involved in sulfur and methane cycle were much higher than that in non-leakage zone and non-cold seep zone and varied with depth.To be specific,the water samples and sediment samples of the cold seep with depth of 0-8 cm were dominated by sulfur-oxidizing bacteria Sulfurovum,accounting for 40.78%-77.76%of the total OTUs?operational taxonomic unit?of the sample,and decreased with the depth.The sediment samples of the cold seep with depth of8-22 cm contained plenty of sulfate reducing bacteria SEEP-SRB1.The sediment samples of the cold seep with the depth of 10-26 cm contained abundant methane-oxidizing archaea,of which,ANME-2a-2b and ANME-3 were detected in the depth of 10-22 cm,which was similar to the distribution of SEEP-SRB1,while ANME-1 appeared in the depth of 22-26 cm.The microbes in the water samples of non-leakage zone were mainly?-Proteobacteria,and the microbes in non-cold seep samples were mainly Acidimicrobiia.Several cultural media and methods were used to cultivate microorganisms isolated from the cold seep sediment sample of the South China Sea.A total of 161strains were isolated and identified.According to 16S rDNA sequences analysis,2strains were speculated to be new species.The 155 aerobic isolated strains belonged to 3 phyla,5 classes,13 orders,23 families,26 genera and 60?sub?species.Gammaproteobacteria occupied the largest proportion,containing 88 strains belonging to 13 genera,followed in abundance by Alphaproteobacteria?41 stains,5genera?,Bacilli?23 stains,5 genera?,Cytophagia?2 strains,1genus?and Flavobacteria?1 strains,1 genus?.The 2 anaerobic strains belonged to Pseudomonas and Sphingomonas.All 4 microaerophilic strains belonged to Alteromonas.In this study,we also characterized strain L2^T,a novel species of the genus Muricauda isolated from the deep sea sediments.Analysis of 16S rDNA sequence?1423bp?indicated that strain L2^T exhibited the highest sequence identity to M.marina H19-56^T?96.4%?.Phylogenetic analysis showed that strain L2^T fell within the clade comprising various Muricauda species.Within the clade,strain L2^T formed a group with four other Muricauda species(M.marina H19-56^T,M.antarctica Ar-22^T,M.taeanensis 105^T and M.flavescens SW-62^T).The chemotaxonomic profiles of strain L2^T were generally similar to those of the genus Muricauda.However,phenotypic characteristics,including the temperature range for growth,hydrolysis of some macromolecules,some enzyme activities and antibiotic susceptibilities,distinguished strain L2^T from other related Muricauda species.Therefore,strain L2^T represents a novel species of the genus Muricauda,for which the name Muricauda iocasae sp.nov.is proposed with the type species L2^T?=CCTCC AB 2017193 ^T=KCTC 62196 ^T?.Through this study,we have learned more about the diversity of microbial communities in the cold seep ecosystem.The unique microbial population of the leak zone was explored.The results promote the understanding the function of microbes in the process of material and energy cycle in the cold seep.Meanwhile,novel species and other bacteria isolated and identified in this study will be useful to the development and utilization of marine microbial resources.