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Research articleEnvironmental Microbiology and Engineering
Microbial Community Analysis of a Methane-Oxidizing Biofilm Using Ribosomal Tag Pyrosequencing
Department of Environmental Science and Engineering, Ewha Womans University, 11-1 Daehyun-dong, Seodaemun-gu, Seoul 120-750, KoreaReceived: September 21, 2011; Accepted: November 4, 2011
J. Microbiol. Biotechnol. 2012; 22(3): 360-370
Published March 28, 2012
Copyright © The Korean Society for Microbiology and Biotechnology.
AbstractCurrent ecological knowledge of methanotrophic biofilms is incomplete, although they have been broadly studied in biotechnological processes. Four individual DNA samples were prepared from a methanotrophic biofilm, and a multiplex 16S rDNA pyrosequencing was performed. A complete library (before being de-multiplexed) contained 33,639 sequences (average length, 415 nt). Interestingly, methanotrophs were not dominant, only making up 23% of the community. Methylosinus, Methylomonas, and Methylosarcina were the dominant methanotrophs. Type II methanotrophs were more abundant than type I (56 vs. 44%), but less richer and diverse. Dominant non-methanotrophic genera included Hydrogenophaga, Flavobacterium, and Hyphomicrobium. The library was de-multiplexed into four libraries, with different sequencing efforts (3,915 - 20,133 sequences). Sørrenson abundance similarity results showed that the four libraries were almost identical (indices > 0.97), and phylogenetic comparisons using UniFrac test and P-test revealed the same results. It was demonstrated that the pyrosequencing was highly reproducible. These survey results can provide an insight into the management and/or manipulation of methanotrophic biofilms.
Keywordsmethanotrophs, biofilm, community analysis, pyrosequencing, quantitative real-time PCR