Jpn. J. Infect. Dis., 64 (1), 19-27, 2011
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Evaluation of a Cytolethal Distending Toxin (cdt) Gene-Based Species-Specific Multiplex PCR Assay for the Identification of Campylobacter Strains Isolated from Diarrheal Patients in Japan
S. M. Lutful Kabir, Ken Kikuchi1, Masahiro Asakura, Sachi Shiramaru, Naoki Tsuruoka2, Aeko Goto2, Atsushi Hinenoya, and Shinji Yamasaki*
Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka 598-8531; 1Department of Infection Control Science, Faculty of Medicine, Juntendo University, Tokyo 113-8421; and 2Clinical Laboratory, Tokyo Women's Medical University Hospital, Tokyo 162-8666, Japan
(Received November 1, 2010. Accepted November 23, 2010)
*Corresponding author: Mailing address: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1, Rinku Ourai-Kita, Izumisano, Osaka 598-8531, Japan. Tel & Fax: +81-72-463-5653, E-mail: firstname.lastname@example.org
SUMMARY: We have developed a cytolethal distending toxin (cdt) gene-based species-specific multiplex PCR assay for the detection and identification of Campylobacter jejuni, C. coli, and C. fetus. The applicability of this assay was evaluated with 325 Campylobacter strains isolated from diarrheal patients in Japan and the results were compared with those obtained by other genetic methods, including hipO gene detection and 16S rRNA gene sequencing. Of the 325 strains analyzed, 314 and 11 were identified as C. jejuni and C. coli, respectively, by combination of hipO gene detection and 16S rRNA gene sequencing. When the multiplex PCR assay was employed, 309, 310, and 314 strains were identified as C. jejuni on the basis of cdtA, cdtB, and cdtC gene-specific primers, respectively. Similarly, 11, 11, and 10 strains were identified as C. coli on the basis of cdtA, cdtB, and cdtC gene-specific primers, respectively. Sequence analysis of the cdt gene region of 6 strains (5 C. jejuni and 1 C. coli) which did not yield specific PCR products in any of the cdt gene-based multiplex PCR assays revealed deletions or mutations of the cdt genes. Pulsed-field gel electrophoresis indicated that C. jejuni and C. coli strains were genetically diverse. Taken together, these findings suggest that the cdtC gene-based multiplex PCR seems to be a particularly simple and rapid method for differentiating between species of Campylobacter strains, such as C. jejuni and C. coli. However, combination of these multiplex PCR assays will allow more accurate identification.
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