Development of a real time PCR for the detection of Taylorella equigenitalis directly from genital swabs and discrimination from Taylorella asinigenitalis

• Published in: Veterinary microbiology 2006-12, Vol.118 (3), p.247-254
• Main Authors: Wakeley, P.R., Errington, J., Hannon, S., Roest, H.I.J., Carson, T., Hunt, B., Sawyer, J., Heath, P.
• Format: Article
• Language: English
• Subjects: analytical kits; Animals; Bacteriology; Base Sequence; Biological and medical sciences; Colony Count, Microbial - veterinary; Contagious equine metritis; Diagnosis, Differential; Direct PCR amplification; disease detection; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; DNA, Ribosomal - chemistry; DNA, Ribosomal - genetics; Endometritis - diagnosis; Endometritis - microbiology; Endometritis - veterinary; Female; Fundamental and applied biological sciences. Psychology; Gene Amplification; genetic techniques and protocols; genitalia; Gram-Negative Bacterial Infections - diagnosis; Gram-Negative Bacterial Infections - microbiology; Gram-Negative Bacterial Infections - veterinary; Horse Diseases - diagnosis; Horse Diseases - microbiology; Horses; Microbiology; Miscellaneous; new methods; pathogen identification; Phylogeny; polymerase chain reaction; Polymerase Chain Reaction - methods; Polymerase Chain Reaction - veterinary; rapid methods; reliability; Reproducibility of Results; ribosomal DNA; sampling; screening; Sensitivity and Specificity; Sequence Alignment - veterinary; sexually transmitted diseases; Species Specificity; Taylorella - classification; Taylorella - genetics; Taylorella - isolation & purification; Taylorella asinigenitalis; Taylorella equigenitalis; Taylorella equigenitalis - classification; Taylorella equigenitalis - genetics; Taylorella equigenitalis - isolation & purification; validity
• ISSN: 0378-1135; 1873-2542
• DOI: 10.1016/j.vetmic.2006.08.007

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 112 bp 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.