Gel microbead cultivation with a subenrichment procedure can yield better bacterial cultivability from a seawater sample than standard plating method |
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Authors: | Shiqi Ji Rui Zhao Qi Yin Yuan Zhao Chenguang Liu Tian Xiao Xiaohua Zhang |
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Institution: | JI Shiqi1), ZHAO Rui1), YIN Qi1), ZHAO Yuan2), LIU Chenguang1), XIAO Tian2), and ZHANG Xiaohua1), 1) College of Marine Life Sciences, Ocean University of China, Qingdao 266003, P. R. China 2) Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, P. R. China |
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Abstract: | A gel microbead (GMD) cultivation method was employed to cultivate microorganisms from an amphioxus breeding zone in Qingdao,
P. R. China. The culture results were compared with those by standard plating method. In the GMD-based method, the microcolony-forming
GMDs were sorted by fluorescence-activated cell sorting (FACS). To further get pure cultures, a subsequent enrichment culture
and a streaking purification procedure were conducted on marine R2A medium. Eighty bacterial strains isolated by the GMD-based
method were randomly selected for sequencing. These isolates belonged to Alphaproteobacteria (33%), Gammaproteobacteria (44%),
Bacteroidetes (11%), Actinobacteria (5%), Firmicutes (5%), Epsilonproteobacteria (1%), and Verrucomicrobia (1%), the last
two groups being usually difficult to culture. The 16S rRNA gene sequences revealed a diverse community with 91.1%–100% of
the bacterial rRNAs similarities. Thirteen strains were sharing 16S rRNA gene sequence which was less than 97% similar to
any other rRNA genes currently deposited in TYP16S database. Seventy isolates derived from the standard plating method fell
into 4 different taxonomic groups: Alphaproteobacteria (9%), Gammaproteobacteria (81%), Bacteroidetes (7%) and Firmicutes
(3%) with a 16S rRNA gene sequence similarities between 95.8%–100%, in which only 3 strains were sharing 16S rRNA gene sequence
of less than 97%. The results indicated that the GMD-based method with subenrichment culture yielded more taxonomic groups
and more novel microbial strains, including members of previously rarely cultured groups, when compared with the standard
plating method, and that this method markedly improved the bacterial cultivability. |
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Keywords: | GMD cultivation marine microorganism bacterial cultivability amphioxus |
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