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

This research presents the development and validation of a real-time PCR assay for the detection of a specific subspecies of Streptococcus equi bacteria in horses, which is more efficient than traditional culture techniques.

Introduction

• The study’s main focus is on the Streptococcus equi subspecies equi (S. equi), an infection endemic in UK horses.
• A population of horses serve as long-term carriers of this infection and are a primary source of its spread.
• The detection of the infection in these persistently infected horses can be challenging due to the low sensitivity of standard culture techniques and overgrowth of contaminating bacteria.
• This subspecies is closely related to S. equi zooepidemicus, making accurate diagnostic tests harder to devise.

Objective

• The research objective was to devise and validate a specific and sensitive real-time PCR (polymerase chain reaction) assay for detecting S. equi and to compare those results with traditional culture methods.

Methods

• The research approach involved a retrospective cross-sectional study that used 92 samples drawn from horses suspected to have ‘strangles’ disease caused by S. equi.
• These samples were cultured using microbial procedures and then tested with the newly created S. equi real-time PCR.
• The team compared the results of PCR tests with traditional methods and computed the diagnostic sensitivity and specificity of the real-time PCR.
• The researchers also checked the real-time PCR for cross-reactivity with horse commensal bacteria, and established the efficiencies and detection limits of the PCR test.

Results

• The real-time PCR assay demonstrated a diagnostic sensitivity of 95% and specificity of 86%, indicating it’s both reliable and precise.
• No cross-reactivity was detected with any bacterial species tested, including the closely resembling S. equi zooepidemicus.
• The assay even detected as few as 3 gene copies, signifying a high sensitivity.

Conclusions

• The real-time PCR assay was concluded to be swift, highly sensitive, and specific and can detect S. equi DNA directly from a minor clinical extract on a swab.
• An additional conclusion was that this real-time PCR assay can facilitate rapid detection of S. equi in symptomless carriers, enabling swift treatment and helping control the spread of strangles in horse populations.