Biodiversity Of Culturable Bacteria Associated With A Atrina Pectinata Collected From Of The Tidal Flat Of Naozhou Island

To understand the diversity of cultivable bacteria associated with Atrina pectinatacollected from the tidal flat of Naozhou Island (20°52′–20°56′N,110°33′–110°38′E) inSouth China Sea, bacteria strains were isolated from homogenates of the sample byusing conventional culture-dependent method. And the antimicrobial activities of thefermentation broth extracts from the isolates were detected to find some isolatesexhibiting strong bioactivity.We isolated125bacteria strains from the sample on MA (marine agar), MH(moderate halophilic medium agar) and NA (nutrient agar) supplemented with0–25%(w/v) NaCl. Out of them,90strains were selected and subjected to a molecularsystematic study based on16S rRNA gene sequences. The results showed that the90isolates were members of ten genera (Marinobacter, Chromohalobacter, Halomonas,Salinivibrio, Vibrio, Bacillus, Halobacillus, Virgibacillus, Staphylococcus, Micrococcus)of six families(Alteromonadaceae, Halomonadaceae, Vibrionaceae, Bacillaceae,Staphylococcaceae, Micrococcaceae) in three phylogenetic groups (Gamma-Proteobacteria, Firmicutes, Actinobacteria). The most abundant and diverse isolateswere within the phylum Firmicutes (56strains,62.2%) and the subphylum Gamma-Proteobacteria (31strains,34.5%). The phylogenetic analysis results suggested thatthere were obvious genetic divergences between most isolates and their closestly relatedtype strains (16S rRNA gene sequence similarities ranged from95.7%–99.9%), and that,out of90isolates at least five JSM strains (112019,112024,114045,114058,114083)should represent potential new taxa. Strain JSM112024could represent a novel genusin the family Halomonadaceae, and strains JSM112019, JSM114045, JSM114058andJSM114083could represent four novel species with in three characterized generaSalinivibrio, Bacillus, Halomonas and Virgibacillus, respectively.The antimicrobial activity of the125bacteria strains were tested using the agardiffusion method employing eight indicator strains. The results showed that out of125test strains, there were18strains exhibiting antibacterial activity (14.4%), in which6strains exhibited strong antibacterial activity. Then, these6strains were submitted forbiochemical and physiological tests. The results suggested that JSM112006belonged tothe genera Marinobacter, and JSM114095belonged to the genera Halomonas, and JSM114060and JSM112019belonged to the genera Salinivibrio, and JSM114054and JSM114093belonged to the genera Staphylococcus. The results of phylogenetic analysis, morphological and chemotaxonomic studiessupported the view that strain JSM114083Tshould be assigned to the genusVirgibacillus (Heyndrickx et al.,1998; Heyrman et al.,2003). To establish the precisetaxonomic position of strain JSM114083T, DNA–DNA hybridizations were performedbetween the novel isolate and the type strains of Virgibacillus halodenitrificans,Virgibacillus necropolis and Virgibacillus carmonensis, and the levels of DNA–DNArelatedness between them were19.5±3.2,10.3±2.1and11.8±2.5%, respectively, whichwere far below the threshold value of70%recommended by Wayne et al.(1987) forassigning strains to the same species. Therefore, on the basis of the polyphasictaxonomic data presented here, strain JSM114083Tshould be placed in the genusVirgibacillus as a member of a novel species, for which we propose the nameVirgibacillus marinus sp. nov.In conclusion, the results presented above showed that there are abundant bacterialspecies diversity and phylogenetic diversity in the Atrina pectinata.