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Journal of Clinical Microbiology, January 1998, p. 86-89, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Use of an Enrichment Broth Cultivation-PCR Combination Assay for Rapid Diagnosis of Swine Erysipelas

Yoshihiro Shimoji,1,* Yasuyuki Mori,1 Koji Hyakutake,2 Tsutomu Sekizaki,1 and Yuichi Yokomizo1

National Institute of Animal Health, Ibaraki 305,1 and Utsunomiya City Meat Inspection Office, Tochigi 321-01,2 Japan

Received 18 April 1997/Returned for modification 24 June 1997/Accepted 10 October 1997

We have previously described the creation by Tn916 mutagenesis of avirulent transposition mutants from a highly virulent strain of Erysipelothrix rhusiopathiae, the causative agent of swine erysipelas. In this study, we cloned a 2.2-kb DNA fragment which flanked the Tn916 insertion in an avirulent mutant (strain 33H6) and evaluated the possibility that this region could be used for the specific detection of E. rhusiopathiae. According to the sequences of this region, oligonucleotide primers were designed to amplify a 937-bp fragment of the E. rhusiopathiae chromosome by PCR. The specificity of the PCR was investigated by analyzing 64 strains of Erysipelothrix species and 27 strains of other genera different from Erysipelothrix. A 937-bp DNA fragment could be amplified from all E. rhusiopathiae strains tested, and no amplification was observed by using DNAs from the other species tested. To make a rapid and definite diagnosis of swine erysipelas in slaughterhouses, we developed an enrichment broth cultivation-PCR combination assay, which used a commercially available DNA extraction kit, to identify E. rhusiopathiae in the specimens from swine with arthritis. After samples were enriched in selective broth culture, detection of E. rhusiopathiae was tested by either conventional methods or the PCR. Of 102 samples tested, 15 samples were positive by conventional methods and 12 of the 15 samples were positive by the PCR. The detection limit of the PCR was 103 CFU per reaction mixture for the PCR-positive samples. These results indicate that this PCR technique could be used as a first-line screening technique for the specific detection of E. rhusiopathiae in specimens.

* Corresponding author. Mailing address: National Institute of Animal Health, 3-1-1 Kannondai, Tsukuba, Ibaraki 305, Japan. Phone: 81-298-38-7857. Fax: 81-298-38-7880. E-mail: shimoji{at}niah.affrc.go.jp.

Journal of Clinical Microbiology, January 1998, p. 86-89, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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