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Applied and Environmental Microbiology, March 1999, p. 961-968, Vol. 65, No. 3
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Molecular Differentiation of Renibacterium salmoninarum Isolates from Worldwide Locations

Thomas H. Grayson,1,* Lynne F. Cooper,1 Franck A. Atienzar,2 Mark R. Knowles,1 and Martyn L. Gilpin1

Department of Biological Sciences,1 and Plymouth Environmental Research Centre,2 University of Plymouth, Plymouth PL4 8AA, Devon, United Kingdom

Received 30 September 1998/Accepted 8 December 1998

Renibacterium salmoninarum is a genospecies that is an obligate pathogen of salmonid fish and is capable of intracellular survival. Conventional typing systems have failed to differentiate isolates of R. salmoninarum. We used two methods to assess the extent of molecular variation which was present in isolates from different geographic locations. In one analysis we investigated possible polymorphisms in a specific region of the genome, the intergenic spacer (ITS) region between the 16S and 23S rRNA genes. In the other analysis we analyzed differences throughout the genome by using randomly amplified polymorphic DNA (RAPD). We amplified the spacer region of 74 isolates by using PCR and performed a DNA sequence analysis with 14 geographically distinct samples. The results showed that the 16S-23S ribosomal DNA spacer region of R. salmoninarum is highly conserved and suggested that only a single copy of the rRNA operon is present in this slowly growing pathogen. DNA sequencing of the spacer region showed that it was the same length in all 14 isolates examined, and the same nucleotide sequence, sequevar 1, was obtained for 11 of these isolates. Two other sequevars were found. No tRNA genes were found. We found that RAPD analysis allows reproducible differentiation between isolates of R. salmoninarum obtained from different hosts and different geographic regions. By using RAPD analysis it was possible to differentiate between isolates with identical ITS sequences.

* Corresponding author. Mailing address: Room 401A Davy Building, University of Plymouth, Plymouth PL4 8AA, United Kingdom. Phone: 44 1752 232950. Fax: 44 1752 232970. E-mail: tgrayson{at}plymouth.ac.uk.

Applied and Environmental Microbiology, March 1999, p. 961-968, Vol. 65, No. 3
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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