Direct and rapid detection of Erysipelothrix rhusiopathiae DNA in animals by PCR.
Erysipelothrix rhusiopathiae is a gram-positive rod capable of causing erysipelas in swine. To establish a method for specifically detecting E. rhusiopathiae for practical applications, such as for the inspection of slaughterhouses, the feasibility of using primers derived from the DNA sequence coding for 16S rRNA in a PCR-specific detection system was investigated. Oligonucleotide primers were designed to amplify a 407-bp DNA fragment by PCR. The amplification was specific to the Erysipelothrix DNA but not to that of other bacterial genera tested. This PCR-based method efficiently and specifically detected the Erysipelothrix DNA sequence in joint and spleen samples from mice within 6 h, and application of the 407-bp DNA segment from samples containing very low numbers of bacteria (< 20 bacteria per spleen from mice) was possible. Although this PCR amplification is specific for the Erysipelothrix genus, which contains at least two species, E. rhusiopathiae and E. tonsillarum, it can be concluded that all Erysipelothrix strains detected by this PCR system in diseased pigs are E. rhusiopathiae because only E. rhusiopathiae is virulent for pigs. These results show that this PCR amplification system using the DNA sequence coding for 16S rRNA is very rapid and reliable and avoids cumbersome and lengthy cultivation steps, demonstrating that this system could be used for practical applications.
Makino
S
S
Department of Veterinary Public Health, Institute of Public Health, Tokyo, Japan.
Okada
Y
Y
Maruyama
T
T
Ishikawa
K
K
Takahashi
T
T
Nakamura
M
M
Ezaki
T
T
Morita
H
H
D000818:Animals; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics; D004888:Erysipelothrix / Q000235:genetics / Q000302:isolation & purification*; D004889:Erysipelothrix Infections / Q000175:diagnosis*; D005240:Feasibility Studies; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013047:Specific Pathogen-Free Organisms; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis*; D013997:Time Factors
D004269:DNA, Bacterial; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S