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Applied and Environmental Microbiology, November 2007, p. 7300-7307, Vol. 73, No. 22
0099-2240/07/$08.00+0     doi:10.1128/AEM.01426-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Layered Structure of Bacterial and Archaeal Communities and Their In Situ Activities in Anaerobic Granules{triangledown} ,{dagger}

Hisashi Satoh, Yuki Miura, Ikuo Tsushima, and Satoshi Okabe*

Department of Urban and Environmental Engineering, Graduate School of Engineering, Hokkaido University, North-13, West-8, Sapporo 060-8628, Japan

Received 27 June 2007/ Accepted 17 September 2007

The microbial community structure and spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by 16S rRNA gene-based molecular techniques and microsensors for CH4, H2, pH, and the oxidation-reduction potential (ORP). The 16S rRNA gene-cloning analysis revealed that the clones related to the phyla Alphaproteobacteria (detection frequency, 51%), Firmicutes (20%), Chloroflexi (9%), and Betaproteobacteria (8%) dominated the bacterial clone library, and the predominant clones in the archaeal clone library were affiliated with Methanosaeta (73%). In situ hybridization with oligonucleotide probes at the phylum level revealed that these microorganisms were numerically abundant in the granule. A layered structure of microorganisms was found in the granule, where Chloroflexi and Betaproteobacteria were present in the outer shell of the granule, Firmicutes were found in the middle layer, and aceticlastic Archaea were restricted to the inner layer. Microsensor measurements for CH4, H2, pH, and ORP revealed that acid and H2 production occurred in the upper part of the granule, below which H2 consumption and CH4 production were detected. Direct comparison of the in situ activity distribution with the spatial distribution of the microorganisms implied that Chloroflexi contributed to the degradation of complex organic compounds in the outermost layer, H2 was produced mainly by Firmicutes in the middle layer, and Methanosaeta produced CH4 in the inner layer. We determined the effective diffusion coefficient for H2 in the anaerobic granules to be 2.66 x 10–5 cm2 s–1, which was 57% in water.

* Corresponding author. Mailing address: Department of Urban and Environmental Engineering, Graduate School of Engineering, Hokkaido University, North-13, West-8, Sapporo 060-8628, Japan. Phone: 81-(0)11-706-6266. Fax: 81-(0)11-706-6266. E-mail: sokabe{at}eng.hokudai.ac.jp

{triangledown} Published ahead of print on 28 September 2007.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

Applied and Environmental Microbiology, November 2007, p. 7300-7307, Vol. 73, No. 22
0099-2240/07/$08.00+0     doi:10.1128/AEM.01426-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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