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Applied and Environmental Microbiology, May 2000, p. 2211-2215, Vol. 66, No. 5
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Simultaneous Direct Counting of Total and Specific Microbial Cells in Seawater, Using a Deep-Sea Microbe as Target

Akihiko Maruyama1,* and Michinari Sunamura1,2

Bioconsortium Analysis Program, National Institute of Bioscience and Human Technology, Higashi, Tsukuba, Ibaraki 305-8566,1 and Mitsubishi Kasei Institute of Life Science, Machida, Tokyo 194-8511,2 Japan

Received 1 November 1999/Accepted 2 March 2000

To rapidly and accurately enumerate total and specific microbes in aquatic samples, fluorescent in situ hybridization was combined with direct counting via direct immobilization of cells on a polymer-coated Nuclepore filter. The technique, named FISH-DC, achieved almost complete recovery of total cells and reproducibility of Psychrobacter pacificensis cells of deep-sea origin (error, <= 3%) in a mixed culture and in natural seawater. Target cells immobilized on the filter were also successfully enumerated after stringent 3-cycle hybridization and even after a 16-month preservation at -30°C.

* Corresponding author. Mailing address: Marine Microbiology and Ecology Group, National Institute of Bioscience and Human Technology, Higashi, Tsukuba, Ibaraki 305-8566, Japan. Phone: 81-298-61-6062. Fax: 81-298-61-6412. E-mail: maruyama{at}nibh.go.jp.

Applied and Environmental Microbiology, May 2000, p. 2211-2215, Vol. 66, No. 5
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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