Development of a real-time PCR to detect Streptococcus equi subspecies equi

Reasons for performing studyInfection with Streptococcus equi subspecies equi (S. equi) is endemic in the UK. A proportion of horses serve as long-term carriers and act as a reservoir of infection. Detection of these persistently infected horses is difficult using standard culture techniques owing to a lack of sensitivity and overgrowth by contaminating bacteria. In addition, differentiation of this causative bacterium from the closely related S. equi zooepidemicus has made the development of reliable and accurate diagnostic tests difficult. ObjectiveTo develop and validate a sensitive and specific real-time PCR assay to detect S. equi and to compare the results with traditional culture techniques. Study designRetrospective cross-sectional study. MethodsThe assay was validated using a panel of 92 samples from suspected clinical cases of strangles. These were cultured using microbial techniques and tested using the S. equi real-time PCR. The results of the 2 methods were compared, and the diagnostic sensitivity and specificity of the real-time PCR were calculated. The real-time PCR was tested for cross-reactivity with horse commensal bacteria, and the efficiencies and limits of detection were established. ResultsThe assay had a diagnostic sensitivity of 95% and specificity of 86%. No cross-reactivity was observed with any of the bacterial species tested, including S. equi zooepidemicus. The assay detected as few as 3 gene copies. ConclusionThe assay is fast, sensitive and specific and will detect S. equiDNA directly from a crude extract of clinical material on a swab. Potential relevanceThis assay could aid in the rapid detection of subclinical shedders of S. equi, enabling quicker treatment and helping to limit the spread of strangles in equine populations.