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Applied and Environmental Microbiology, May 2007, p. 2787-2798, Vol. 73, No. 9
0099-2240/07/$08.00+0     doi:10.1128/AEM.02111-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Actively Growing Bacteria in the Inland Sea of Japan, Identified by Combined Bromodeoxyuridine Immunocapture and Denaturing Gradient Gel Electrophoresis{triangledown} ,{dagger}

Koji Hamasaki,1* Akito Taniguchi,1 Yuya Tada,1 Richard A. Long,2,{ddagger} and Farooq Azam3

Graduate School of Biosphere Science, Hiroshima University, Hiroshima, Japan,1 Department of Oceanography, College of Geoscience, Texas A&M University, College Station, Texas 77843,2 MBRD, Scripps Institution of Oceanography, University of California San Diego, La Jolla, California 920373

Received 6 September 2006/ Accepted 21 February 2007

A fundamental question in microbial oceanography concerns the relationship between prokaryote diversity and biogeochemical function in an ecosystem context. We combined bromodeoxyuridine (BrdU) magnetic bead immunocapture and PCR-denaturing gradient gel electrophoresis (BUMP-DGGE) to examine phylotype-specific growth in natural marine assemblages. We also examined a broad range of marine bacterial isolates to determine their abilities to incorporate BrdU in order to test the validity of the method for application to diverse marine assemblages. We found that 27 of 29 isolates belonging to different taxa could incorporate BrdU. BUMP-DGGE analysis revealed phylogenetic affiliations of DNA-synthesizing, presumably actively growing bacteria across a eutrophic to mesotrophic transect in the Inland Sea of Japan. We found that the BrdU-incorporating (growing) communities were substantially different from the total communities. The majority (34/56) of phylotypes incorporated BrdU and were presumably growing, and these phylotypes comprised 10 alphaproteobacteria, 1 betaproteobacterium, 11 gammaproteobacteria, 11 Cytophaga-Flavobacterium-Bacteroides group bacteria, and 1 unclassified bacterium. All BrdU-responsive alphaproteobacteria were members of the Rhodobacterales, suggesting that such bacteria were dominant in the growing alphaproteobacterial populations in our samples. The BrdU-responsive gammaproteobacteria belonged to the Oceanospirillales, the SAR86 cluster, the Pseudomonadales, the Alteromonadales, and the Vibrionales. Thus, contemporaneous cooccurrence of diverse actively growing bacterial taxa was a consistent pattern in our biogeochemically varied study area.

* Corresponding author. Present address: Department of Marine Ecosystem Dynamics, Ocean Research Institute, The University of Tokyo, 1-15-1 Minamidai, Nakano-ku, Tokyo 164-8639, Japan. Phone: 81-3-5351-6337. Fax: 81-3-5351-6482. E-mail: hamasaki{at}ori.u-tokyo.ac.jp

{triangledown} Published ahead of print on 2 March 2007.

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

{ddagger} Present address: Department of Biological Sciences, University of South Carolina, Columbia, SC 29208.

Applied and Environmental Microbiology, May 2007, p. 2787-2798, Vol. 73, No. 9
0099-2240/07/$08.00+0     doi:10.1128/AEM.02111-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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