Tips for successful oral-fluid sampling for PRRSV

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Oral-fluid sampling is an easy way to detect porcine reproductive and respiratory syndrome virus (PRRSV) or track changes in PRRSV infections in populations — and it can optimize your survey efforts by keeping a few simple tips in mind, said Jeff Zimmerman, DVM, PhD, a professor at Iowa State University.

Oral fluids contain local and serum-derived antibodies and pathogens from the pig, but they also contain whatever other pathogens happen to be in the pen environment. This combination makes them a good diagnostic specimen, Zimmerman said.

Oral fluid is more efficient than bleeding pigs for detecting PRRSV and other pathogens. For example, if 20% of the pigs in a pen were PRRSV-positive, the probability of detecting the infection using one oral-fluid sample is over 90% — actually, 98% by polymerase chain reaction (PCR) and 94% by ELISA. To match that probability of detection, you would need to bleed at least 10 pigs (Table 1), he said.

SAMPLING PROCEDURE

Zimmerman advised obtaining oral-fluid samples first thing in the morning, when pigs are most active. It’s important to use cotton rope, which gives better PCR performance than ropes made with synthetic fibers, and to cut the length of the rope so that the pigs need to raise their heads just a little to reach it.

After fluids are extracted from the rope and poured into a tube, they should be chilled immediately and kept cool until they get to the lab. If there will be more than a 72-hour delay, it may be better to freeze the samples, he added.

“Fixed” spatial sampling — that is, sampling pens throughout the barn (or air space) and at equal distances from each other — is easier than trying to randomly select pens and is actually as good as or better than random sampling or risk-based sampling. Sampling the same pens the next time and every time is best.

“This is a new idea for us, but it works well for both detection and monitoring,” he said.

*Assumption is that pigs are both viremic and antibody positive.

If only one sample is going to be collected, he recommended sampling pens in the middle of the barn. “That’s the spot where you’ll have the highest likelihood of detection,” Zimmerman said.

SAMPLE SAME PENS

For surveillance or monitoring, sampling the same pens each time will make it easier to interpret results. “If you mix different pens, they’re going to be at different stages of infection, and you have a more difficult time understanding what the results are telling you over time,” the veterinarian explained.

Repeated sampling over time is critical. “One sample is a picture that’s static and historic, while sampling over time is a movie that’s dynamic and predictive,” Zimmerman explained.

One of the biggest challenges, he noted, is determining the best strategy for monitoring presumed-negative populations. In this case, Zimmerman recommended collecting six, eight or 10 samples initially and then one or two more samples every week or two. Collecting from all barns exponentially increases the probability of detection.

“I’d rather see fewer samples — even one sample — collected every week or 2 weeks from all or most barns versus more samples collected less often,” he said.

DON’T ABANDON ANTIBODY TESTS

Testing of oral-fluid samples for PRRSV can be done by PCR. Test results can vary depending on the PCR protocols and kits used. “That’s something to bear in mind when you look at your data. There are differences between labs and between kits,” Zimmerman cautioned.

A problem with PCR testing for PRRSV is that not all labs are running the same protocol. Results can therefore vary with each lab. “There’s still some work to be done to find the best protocol and get all the labs to run the same protocol,” he said.

Samples also can be tested for PRRSV antibodies, Zimmerman said, adding that the antibody response is more definitive than PCR results for a number of different pathogens. “We’ve had a tendency to abandon antibody tests, but it may be premature. Antibodies are detectable even after nucleic acid disappears. For effective surveillance and monitoring, we need both nucleic acid [PCR] and antibody tests,” he emphasized.

One problem with antibody testing occurs when maternal antibody (IgG) is present. IgG declines but can be detected for a long time. If there is doubt about a positive antibody result, a PRRSV IgM/IgA ELISA can be used to determine whether the antibody is from the sow (IgG) or from the pig (IgM and/or IgA), Zimmerman said.

1 Olsen C, et al. Probability of detecting porcine reproductive and respiratory syndrome virus infection using pen-based swine oral fluid specimens as a function of within-pen prevalence. J Vet Diagn Invest. 2013;25(3): 328-335.