Development and evaluation of PCR test for detection of Taylorella equigenitalis.

The researchers developed a PCR (Polymerase Chain Reaction) test to efficiently detect Taylorella equigenitalis, the bacterium responsible for contagious equine metritis in horses. The results showed that the PCR test is more sensitive and accurate than traditional culture methods, suggesting that the latter might have led to an underestimation of the spread of this bacterium among horses.

Development of a PCR Test

• The researchers developed a PCR test aimed to detect Taylorella equigenitalis, a bacterium causing contagious equine metritis, a severe reproductive infection in horses.
• A genus-specific primer-probe set tailored for the bacterium was created from the 16S ribosomal DNA sequences. A primer-probe set is a pair of short synthetic oligonucleotide sequences that bind to the DNA segment to be amplified in the PCR process.
• The PCR test was found to be specific, and it amplified a 585-base pair product from all 64 available T. equigenitalis isolates.

Evaluation of the PCR Test

• The PCR product was then hybridized with a specific probe in a dot spot assay.
• A variety of microorganisms from the genital tracts of horses or those with a close phylogenetic relationship to T. equigenitalis did not yield a visible PCR product and were all negative in the dot spot hybridization assay.

Comparison with Traditional Culture Methods

• The effectiveness of the PCR assay was then benchmarked against traditional culture methods. Using 191 genital swabs from horses of different breeds, it was found that the PCR assay surpassed the culture method in terms of sensitivity.
• The assay was found to be most sensitive when using DNA from culture plates incubated for at least two days.

Implications of the PCR Test

• In the study, only 1.5% of the tested samples were positive in the culture assay, as opposed to 35% positive in the culture PCR assay. This striking difference suggests that a significant number of T. equigenitalis-carrying horses may go unnoticed with the current culturing technique, leading to incorrect assessments related to the spread and control of T. equigenitalis.

This new PCR test, therefore, represents an important advance in the diagnosis and control of contagious equine metritis since it is more sensitive, specific, and faster than the traditional culturing technique.