Development of a real time PCR for the detection of Taylorella equigenitalis directly from genital swabs and discrimination from Taylorella asinigenitalis.
A discriminatory real time PCR for the detection of Taylorella equigenitalis, the causative agent of contagious equine metritis (CEM), and the related species T. asinigenitalis was developed for the direct examination of genital swabs. The 112bp amplicons produced from the two species were discriminated from each other using TaqMan probes labelled with different fluorophores. The TaqMan PCR was shown to be specific for the 16S ribosomal DNA of the two species of taylorella and did not cross-hybridise with the 16S ribosomal DNA of other bacteria tested. Direct amplification from genital swabs was shown to be equally sensitive to that of culture methods. Prevalence in a sample set from The Netherlands was shown to be equivalent to that demonstrated by culture. A companion real time PCR that amplified a fragment of the 16S rDNA gene of equine commensal bacteria was developed to ensure bacterial DNA was extracted from swab material supplied for testing. The use of a rapid and reliable real time PCR for the organism causing CEM should aid the control of this disease.
Wakeley
P R
PR
Technology Transfer Unit, Biotechnology Department, Veterinary Laboratories Agency, New Haw, Addlestone, Surrey, UK. p.wakeley@vla.defra.gsi.gov.uk
Errington
J
J
Hannon
S
S
Roest
H I J
HI
Carson
T
T
Hunt
B
B
Sawyer
J
J
Heath
P
P
D000818:Animals; D001483:Base Sequence; D015169:Colony Count, Microbial / Q000662:veterinary; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D003937:Diagnosis, Differential; D004716:Endometritis / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D005260:Female; D005784:Gene Amplification; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D006734:Horse Diseases / Q000175:diagnosis* / Q000382:microbiology; D006736:Horses; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016415:Sequence Alignment / Q000662:veterinary; D013045:Species Specificity; D042502:Taylorella / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D020625:Taylorella equigenitalis / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*
D004269:DNA, Bacterial; D004275:DNA, Ribosomal