Real‐Time Polymerase Chain Reaction: A Novel Molecular Diagnostic Tool for Equine Infectious Diseases

Abstract

The focus of rapid diagnosis of infectious disease of horses in the last decade has shifted from the conventional laboratory techniques of antigen detection, microscopy, and culture to molecular diagnosis of infectious agents. Equine practitioners must be able to interpret the use, limitations, and results of molecular diagnostic techniques, as they are increasingly integrated into routine microbiology laboratory protocols. Polymerase chain reaction (PCR) is the best-known and most successfully implemented diagnostic molecular technology to date. It can detect slow-growing, difficult-to-cultivate, or uncultivatable microorganisms and can be used in situations in which clinical microbiology diagnostic procedures are inadequate, time-consuming, difficult, expensive, or hazardous to laboratory staff. Inherent technical limitations of PCR are present, but they are reduced in laboratories that use standardized protocols, conduct rigid validation protocols, and adhere to appropriate quality-control procedures. Improvements in PCR, especially probe-based real-time PCR, have broadened its diagnostic capabilities in clinical infectious diseases to complement and even surpass traditional methods in some situations. Furthermore, real-time PCR is capable of quantitation, allowing discrimination of clinically relevant infections characterized by pathogen replication and high pathogen loads from chronic latent infections. Automation of all components of PCR is now possible, which will decrease the risk of generating false-positive results due to contamination. The novel real-time PCR strategy and clinical applications in equine infectious diseases will be the subject of this review.Key words: Horse; Pathogen detection; TaqMan polymerase chain reaction. Equine practitioners have always considered accurate and rapid identification of infectious pathogens a priority. The ready availability of a correct etiologic diagnosis, particularly in contagious infections, enables the veterinarian to make early decisions on the patient's care and management, address appropriate treatment, and allow timely notification and discussion of management issues regarding prevention of disease spread. The last 2 decades have seen a revolution in the understanding, management, diagnosis, control, and prevention of infectious diseases.1,2 This period has encompassed the discovery of emerging equine agents, antimicrobials, vaccines, as well as a wealth of improved diagnostic tests for equine practitioners. Despite these advances, infectious diseases remain a leading cause of equine morbidity and mortality, with resurgence of certain infections (eg, West Nile virus), an increasing population of elderly, more susceptible horses, and an increasing international equine commerce, expanding the geographic distribution of pathogens. 3,4The focus of rapid diagnosis of infectious diseases also has shifted during this time. The most obvious change has been the appearance and increasing importance of nucleic acid (NA) amplification-based techniques, primarily the...

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