Polymerase Chain Reaction: Benefits and Drawbacks

The application of polymerase chain reaction (PCR) assays to veterinary medicine has revolutionized the way diagnosticians detect infectious agents and genetic markers of noninfectious disease. Confusion frequently encircles this technology due to its novelty and complexity. This article will briefly review the underlying basis of PCR, describe interpretation of results, and discuss its benefits and drawbacks over traditional methods of pathogen detection.

All living organisms contain unique sequences of genetic material that consist of deoxyribonucleic acid (DNA) and/or ribonucleic acid (RNA ). These hereditary materials serve as templates (or codes) for cellular reproduction and orchestrate the construction of proteins that are essential to all biologic processes. Traditional methods of pathogen recognition (i.e., bacterial culture, fluorescent antibody tests, serologic tests, etc.) have focused on the presence of proteins for identification. Conversely, PCR tests are designed to amplify and detect segments of DNA or RNA that are very specific to a genetic sequence or organism.

Interpretation of positive and negative PCR results can be challenging. Similar to traditional pathogen detection techniques, PCR results must be strictly interpreted in conjunction with the history, clinical signs, and evidence of disease. A positive PCR result only indicates the detection of the target genetic sequence. It can not differentiate between the incidental presence of an organism, colonization without disease, transient infection, or active infection with disease.

In addition, a positive PCR test can't differentiate between living or dead organisms as genetic material can be present in both. Assays can be designed to differentiate pathogenic from nonpathogenic isolates but may not distinguish between vaccine strains and wild-type pathogens. Knowledge of the vaccine (modified live versus killed; route of inoculation; and duration of vaccine persistence) in combination with clinical disease can greatly aid in the interpretation of whether a positive PCR is due to vaccine interference or vaccine failure. Equally important, false negative PCR results may arise due to PCR inhibitors found in clinical samples. Newer technology has greatly eliminated the influence of these inhibitors on PCR results.

PCR technology can provide many advantages over traditional techniques. Many PCR tests can be rapidly performed and interpreted the same day as sample submission. Large numbers of samples can be simultaneously completed. A major advantage of PCR over traditional techniques includes the ability to rapidly identify organisms that are difficult to culture, such as Lawsonia. Differential PCR assays can determine if an isolate is nonpathogenic or contains toxigenic properties (genes) necessary to induce disease. Lastly, PCR can amplify very small amounts of genetic material, thus it can detect very low numbers of organisms in a sample. The major drawbacks are a lack of antimicrobial sensitivity data, complexity of the assay, and the price of PCR equipment and kits.

Prices and availability of different PCR tests vary based on laboratory. Please call or visit your laboratory's website to see available tests, pricing, collection procedures, and shipping guidelines prior to sample collection.

This is an excerpt from Equine Disease Quarterly, funded by underwriters at Lloyd's, London, brokers, and their Kentucky agents.

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Equine Disease Quarterly

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