Evaluation of the Field Application of PCR in the Eradication of Contagious Equine Metritis from Japan.

Abstract

ABSTRACT. The effectiveness of the polymerase chain reaction (PCR) as a field application test for the eradication of contagious equine metritis (CEM) was evaluated. Seven-thousands five-hundred and thirty-four genital swabs were collected from 4,026 Thoroughbred broodmares and stallions in Japan to test "high risk" horses as well as for general surveillance testing from 1998 to 2001. Bac terial isolation as well as PCR testing of original specimens and cultured specimens was performed for detection of Taylorella equigenitalis from genital swabs. As a result, T. equigenitalis was detected in 12 mares and 1 stallion by PCR, although the bacteria were isolated from only 2 of the PCR-positive mares. CEM-infected and carrier horses were treated by a combination of chemotherapy and surgery. Subsequent follow-up testing over a 3-year period did not detect T. equigenitalis. It was demonstrated that PCR testing was more sensitive than isolation as a method for the detection of T. equigenitalis from genital swabs of horses in the field. It was therefore suggested that a combination of PCR testing and treatment were useful measures in the eradication of CEM from Japan.

Microarray-Based Identification of Bacteria in Clinical Samples by Solid-Phase PCR Amplification of 23S Ribosomal DNA Sequences

“…T. equigenitalis is a fastidious and slowly growing bacterium, with the time to detection being 6 days or more by conventional methods. In order to prevent long-term infections and possible sterility, it is extremely important to detect this bacterium as early as possible (7,8,44).…”

“…Such examinations are time-consuming, and the results are usually not available until 24 to 48 h later. For the fastidious, slowly growing species T. equigenitalis, visible colony formation even takes 4 to 6 days and is often obscured by the growth of other quick-growing bacteria (8). The principal treatment for infertility from infectious causes can be directed at reducing the number of offending organisms by exposing them to chemotherapeutic agents (9).…”

Construction of a Recombinant Plasmid as Reaction Control in Routine PCR for Detection of Contagious Equine Metritis (CEM-PCR)

“…A single-step PCR method for detection of T. equigenitalis was developed by Anzai et al [2]. Recently, this method was modified to a semi-nested PCR method (CEM-PCR) for dealing with a large number of routine diagnosis examinations [3]. This method targets the unique sequence motif of T. equigenitalis and is regarded as a far more sensitive diagnostic method for CEM than the culture method [1].…”

“…Another is that the size of the product from the plasmid should be significantly different from the true positive PCR product; this will be able to prevent false positive results by contamination of the plasmid to samples. To satisfy these conditions, we used a recombinant plasmid (CEM-1P) from the CEM-1 strain, which is a clone of the genomic library of T. equigenitalis constructed by Anzai et al [3]. CEM-1P, a recombinant plasmid constructed from pUC119 by insertion of a T. equigenitalis-specific fragment, was used to develop CEM-PCR.…”

“…The selected transformants were stored at -80°C. CEM-PCR was performed as described in a previous report [3]. After TA cloning, 34 transformed E. coli strains were examined.…”