Development of a PCR test for rapid diagnosis of contagious equine metritis.
In order to establish a rapid diagnostic method for contagious equine metritis (CEM), we developed and evaluated a polymerase chain reaction (PCR) test. Species-specific PCR primer sets were derived from the DNA sequence of a cloned DNA fragment of Taylorella equigenitalis that did not hybridize with the genome of a taxomonically related species, Oligella urethralis. Single step PCR with primer set P1-N2 and two-step semi-nested PCR with primer sets P1-N2 and P2-N2 detected as low as 100 and 10 CFU of the bacteria, respectively. Single-step PCR detected T. equigenitalis from genital swabs of experimentally infected mares with sensitivity comparable to that of bacterial isolation. Furthermore, two-step PCR was more sensitive than the culture method. Upon examination of field samples, 12 out of 3,123 samples were positive by single-step PCR while only 2 were positive by bacterial culture. The 12 PCR-positive samples originated from 5 mares, of which 3 animals were considered to be carriers based on previous bacteriologic and serologic diagnoses for CEM. The PCR test described in this study would provide a specific and highly sensitive tool for the rapid diagnosis of CEM.
Anzai
T
T
Epizootic Research Station, Equine Research Institute, Japan Racing Association, Tochigi, Japan.
Eguchi
M
M
Sekizaki
T
T
Kamada
M
M
Yamamoto
K
K
Okuda
T
T
D000818:Animals; D001483:Base Sequence; D002584:Cervix Uteri / Q000382:microbiology; D002987:Clitoris / Q000382:microbiology; D017931:DNA Primers / Q000737:chemistry; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel / Q000662:veterinary; D004716:Endometritis / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D005260:Female; D015723:Gene Library; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis / Q000662:veterinary*; D006734:Horse Diseases / Q000175:diagnosis* / Q000382:microbiology; D006736:Horses; D008297:Male; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010413:Penis / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D020625:Taylorella equigenitalis / Q000235:genetics* / Q000302:isolation & purification
D017931:DNA Primers; D004269:DNA, Bacterial