A team of researchers in Wyoming has developed a test that could greatly aid in the detection of brucellosis in cattle, elk and bison.

“This new diagnostic test allows us to re-evaluate the risk factors for bacterial infection in both livestock and wildlife, helping to control the disease in the Greater Yellowstone Area,” says veterinarian Brant Schumaker, an associate professor in the University of Wyoming’s Department of Veterinary Sciences.

Schumaker and colleagues developed a polymerase chain reaction (PCR) assay that has the ability to detect the bacteria that causes bovine brucellosis (Brucella abortus) in almost twice as many animals as the current diagnostic test, which is a bacterial culture.

“The potential to replace the current gold-standard diagnostic test with a more sensitive assay could decrease the cost of an outbreak in livestock,” Schumaker says. “Additionally, PCR testing of a suspect animal is about one-quarter the cost of culture and can be completed in a few hours, in comparison to 10 to 14 days for bacterial culture.”

This means livestock producers, wildlife managers, veterinarians, livestock boards and others can have results within the same day of samples arriving at a lab.

Schumaker emphasizes that the PCR assay test is still in the research phase, with validation expected within a year — but early results are promising.

Checking the test
The Wyoming researchers were able to acquire samples from 87 suspect animals, including 18 cattle and 69 bison, from within the brucellosis endemic area of northwest Wyoming, southern Montana and eastern Idaho.

Of these, only 42 (or about 48%) were culture-positive, despite all of the animals being suspect or positive on the standard blood test for B. abortus exposure.

Schumaker says that with these same animals, the top PCR candidate set that his team developed detected Brucella DNA on 77 (or nearly 90%) of the animals tested.

In the endemic area in and around Yellowstone National Park, elk and bison carrying the B. abortus bacterium can lead to infections in cattle and domesticated bison. The bacterium can cause the animals to abort their calves. And if cattle are determined to be infected, that can create negative ripple effects for other ranchers. For example, specified cattle from within a “designated surveillance area” must be tested for brucellosis before changing ownership or being moved to other counties and states.

“It is important for producers in the endemic areas of Wyoming, Montana and Idaho to understand their options to reduce disease risk,” Schumaker says. “Working with your state livestock board to develop herd plans and potentially vaccinating adult animals can help to reduce an individual producer’s risks. Additionally, proactively testing animals on an annual basis can limit the potential cost of an outbreak.”

Schumaker and his colleagues remain confident that the new PCR assay will greatly improve testing for brucellosis, which will help both the livestock industry and those who manage wildlife in the greater Yellowstone area.

“Thus far, on seronegative cattle, elk and bison located outside of this endemic area, our assay has been 100% specific,” Schumaker says. “This means that we are able to correctly identify disease-free animals, with no animals testing falsely positive.”

Additionally, he says, “With this PCR assay, we are now able to re-evaluate the cutoff values for serologic tests. Now that we have a better measure of true disease status, we can set values of screening blood tests to better identify an animal’s health profile.”

Brucellosis-caused quarantines costly
Bovine brucellosis is one of the world’s most widespread human diseases and still causes problems to domestic livestock producers in Wyoming, Idaho and Montana.

In the U.S., the Cooperative State-Federal Brucellosis Eradication Plan, initiated in 1934, was successful in eradicating brucellosis from cattle populations.

The disease, however, still has a reservoir in elk and bison in the greater Yellowstone area (northwest Wyoming, southern Montana and eastern Idaho), with multiple instances of spillover into cattle and domesticated bison herds on private and public lands within the GYA in the last decade.

Positive cases in livestock lead to quarantines that can cost the producer upward of $254,000 based on an analysis of a “typical” Wyoming beef cattle operation, according to a paper that a team of Wyoming researchers wrote for the 2017 Field Days Bulletin, which is published annually by the Wyoming Agricultural Experiment Station.

Additionally, producers may elect — and in some cases, be required — to cull their herd and submit the culls to the imperfect and time-consuming diagnostic testing that is currently available.

Producers located in the designated surveillance areas of Wyoming, Idaho and Montana are also required to undertake increased testing requirements prior to importation to some specific states. This applies to producers not under quarantine, but who conduct their livestock operations within the region.

The team that is developing a new test to better detect for brucellosis is composed of researchers from the University of Wyoming’s Department of Veterinary Sciences, Wyoming State Veterinary Laboratory and Wyoming Game and Fish Department.

Members include Brant Schumaker, Noah Hull, Sierra Amundson, Jacob Berg, Jonathan Miller, David Berry, William Laegreid, Hank Edwards and Gerard Andrews.

How did the Wyoming team develop the new test?
How did the Wyoming research team develop the new polymerase chain reaction assay to better detect brucellosis in the tissues of affected animals?

Principal investigator Brant Schumaker says that he and his colleagues conducted the most in-depth computer analysis of Brucella spp. ever performed, using genetic sequences acquired from USDA’s National Veterinary Services Laboratory in Ames, Iowa.

This analysis revealed potential targets for diagnostic testing, and these targets, in turn, went through an extensive screening process, which ultimately left eight candidates for validation, explains Noah Hull, a doctoral candidate in the University of Wyoming’s Department of Veterinary Sciences.

“We evaluated each candidate on its ability to correctly identify target pieces of bacterial DNA in known positive animals,” Hull says. “Additionally, these candidates can differentiate between a true infection and vaccinal reactions in animals.”

Hull says the team also made sure that the successful candidates would not cross-react with known organisms, which can produce false-positive results.

“Our goal was to design an assay that could give us the clearest picture of an animal’s disease status,” Hull adds.

Parallel to this process, the team wanted to make sure that the methods used to extract Brucella DNA from tissue samples produced the highest yield, as the bacterium is presumed to exist in small numbers in these samples, Schumaker says. They screened six different commercial extraction kits against blood and its fractions, and all standard tissue types.

The kits used on field samples were taken directly from this analysis.

“Ultimately, we had to perfect the way we conduct the assay, as that has been almost as important as the genetic target we are using,” Schumaker says. “We are very excited to have a new tool to help us control this disease.”

Brucellosis testing research receives broad support
A far-reaching constituency of local, state and federal officials, along with those who manage livestock and wildlife, is interested in developing improved tests to detect brucellosis in cattle, elk and bison.

That is very evident by the long list of state and federal agencies and institutions that are financially supporting brucellosis testing research being conducted at the University of Wyoming’s Department of Veterinary Sciences.

Among the financial supporters are the U.S. Department of Homeland Security’s Institute for Infectious Animal Diseases, USDA National Institute of Food and Agriculture, USDA Western Sustainable Agriculture Research and Education, Wyoming NASA Space Grant Consortium, Wyoming Department of Agriculture’s Agriculture Producer Research Grant Program, Wyoming Wildlife/Livestock Disease Research Partnership, Wyoming IDeA Networks for Biomedical Research Excellence, National Science Foundation, Wyoming Experimental Program to Stimulate Competitive Research, University of Wyoming College of Agriculture and Natural Resources, and Wyoming Agricultural Experiment Station.

Waggener writes from Laramie, Wyo.