Diagnosing viruses in pigs easier thanks to non-invasive techniques

SDSU Extension

BROOKINGS — Over the last decade, surveillance and diagnosis of swine respiratory disease have been transformed by innovations in sampling and testing.

“In pigs, the same visual signs of disease can be caused by very different germs. It’s important to know which ones are present in the midst of a respiratory disease outbreak. This is where quick and accurate agent detection through laboratory testing comes into play,” said Russ Daly, professor, SDSU Extension veterinarian, state public health veterinarian.

Below, Daly discusses some of these recent developments in laboratory techniques.

Oral fluids sampling: This novel method of sample collection has dramatically changed testing for disease agents in pigs. Rather than restraining and obtaining blood or swab samples from individual pigs, which can be stressful to the animal, investigators simply collect viruses, such as porcine reproductive and respiratory syndrome virus (PRRSV) or influenza A virus (IAV), which are shed in saliva by animals who chew on a cotton rope hung in their pens. In recent years, this sampling method has been validated for more and more test procedures and is now widely accepted for many applications.

Polymerase chain reaction testing: While oral fluids have transformed sample collection, polymerase chain reaction (PCR) technology has revolutionized agent detection in such samples.

“The speed, accuracy and sensitivity of PCR technology is unparalleled,” Daly said. “It’s given swine veterinarians and producers the ability to confidently and quickly respond to new disease threats in the animals they care for.”

The PCR testing method is particularly valuable in swine medicine compared to other species. “Perhaps no other domestic animal species has been faced with as many emerging disease threats over the past couple decades as have swine,” Daly said, referencing PRRS, porcine epidemic diarrhea virus and porcine circovirus.

In a typical PCR test, samples submitted to diagnostic labs are processed such that nucleic acid (DNA or RNA) is extracted from the various viruses and bacteria present in the particular sample.

Then, primers and bases specific to the tested-for germ’s nucleic acid are added.

Repeated cycles of heating and cooling result in an exponential amplification of the target nucleic acid. In “real-time” PCR tests, an indicator dye fluoresces in the midst of this reaction when sufficient nucleic acid has built up in the test chamber. This signifies that the target nucleic acid is present in the sample and the test is deemed positive.

Most PCR tests run at veterinary diagnostic labs detect nucleic acid of a single pathogen. The section of nucleic acid targeted by the test must be one that’s “conserved” (the same) across multiple strains of a pathogen. In this manner, PCR tests can also be designed to target a very specific strain or portion of the germ.

Multiplex polymerase chain reaction tests: These PCR procedures utilize the same pig sample, but test for more than one germ simultaneously. This can mean savings in both testing time and cost for veterinarians and producers.

“Establishing a multiplex PCR test is not as simple as throwing two “single-plex” tests together,” Daly explained. “The reagents and time and temperature conditions for the test for germ A might be different than those for germ B.”

Tweaks and adjustments need to be made by the test developers so they work together.

Current examples of multiplex PCR tests which are run at the SDSU Animal Disease Research and Diagnostic Laboratory include porcine epidemic diarrhea virus + transmissible gastroenteritis + deltacoronavirus and the multiplex PRRSV PCR that detects both North American and European strains.

Testing services coming to SDSU

A new multiplex PCR test coming online at the SDSU Animal Disease Research and Diagnostic Laboratory will greatly help veterinarians and producers sort out clinical signs in growing pigs and other groups.

“For the first time, the influenza A PCR and PRRSV PCR tests are being combined in one multiplex procedure,” Daly said.

He explained that the multiplex PCR test has been validated on oral fluids samples, nasal swabs and tissues. Blood and serum samples will still be tested with individual single PCR tests.

Benefits of multiplex testing

“The multiplex test will be particularly valuable in light of the end of USDA support for influenza testing in swine populations,” Daly said.

He explained that its price is the same as a current single-plex test for either virus, making for more cost-effective diagnostics, whether for routine surveillance or for disease diagnostic investigations.

“The development of tests such as the multiplex influenza-PRRSV PCR test demonstrates the responsiveness of today’s veterinary diagnostic laboratories to the needs of swine veterinarians and producers,” Daly said. “Labs such as the SDSU Animal Disease Research and Diagnostic Laboratory continue to adapt even newer technologies such as whole genome sequencing of germs in order to help these professionals make better decisions for the health of their pigs.”