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Infection and Immunity, October 2000, p. 5742-5748, Vol. 68, No. 10
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Genetic Characterization of DNA Region Containing the trh and ure Genes of Vibrio parahaemolyticus

Kwon-Sam Park, Tetsuya Iida,^* Yoshiharu Yamaichi, Tomohito Oyagi, Koichiro Yamamoto, and Takeshi Honda

Department of Bacterial Infections, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan

Received 25 February 2000/Returned for modification 31 May 2000/Accepted 29 June 2000

We have demonstrated that possession of the gene for thermostable direct hemolysin-related hemolysin (trh) coincides with the presence of the urease gene among clinical Vibrio parahaemolyticus strains and that the location of the two genes are in close proximity on the chromosome. Here, we cloned and sequenced the 15,754-bp DNA region containing the trh gene and the gene cluster for urease production from the chromosome of clinical V. parahaemolyticus (TH3996). We found 16 open reading frames (ORFs) and a lower G+C content (41%) compared with the total genome of this bacterium (46 to 47%). The ure cluster consisted of eight genes, namely, ureDABCEFG and ureR. ureR was located 5.2 kb upstream of the other seven genes in the opposite direction. The genetic organization and sequences of the ure genes resembled those found in Proteus mirabilis. Between ureR and the other ure genes, there were five ORFs, which are homologous with the nickel transport operon (nik) of Escherichia coli. We disrupted each of the ureR, ureC, and nikD genes in TH3996 by homologous recombination and analyzed the phenotype of the mutants. In the presence of urea these mutant strains had dramatically less urease activity than the strain they were derived from. Disruption of ureR, nikD, or ureC, however, had no effect on TRH production. The DNA region containing the trh, nik, and ure genes was found in only trh-positive strains and not in Kanagawa phenomenon-positive and environmental V. parahaemolyticus strains. At the end of the region, an insertion sequence-like element existed. These results suggest that the DNA region was introduced into V. parahaemolyticus in the past through a mechanism mediated by insertion sequences. This is the first reported case that the genes for an ATP-binding cassette-type nickel transport system, which may play a role in nickel transport through bacterial cytoplasmic membrane, are located adjacent to the ure cluster on the genome of an organism.

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^* Corresponding author. Mailing address: Department of Bacterial Infections, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan. Phone: 81-6-6879-8278. Fax: 81-6-6879-8277. E-mail: iida{at}biken.osaka-u.ac.jp.

Present address: Department of Molecular Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University School of Medicine, Kumamoto 862-0976, Japan.

Present address: Faculty of Health and Welfare Science, Okayama Prefectural University, Soja, Okayama 719-1197, Japan.

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Infection and Immunity, October 2000, p. 5742-5748, Vol. 68, No. 10
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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