LLNL biologists James Thissen and Crystal Jaing along with researchers from Kansas State University found that the Microbial Detection Array could help identify diseases in the commercial swine industry.Photo courtesy of Lawrence Livermore National LaboratoryA study by LLNL and Kansas State University scientists found that the Lawrence Livermore Microbial Detection Array (LLMDA) could help identify diseases in the commercial swine industry.

The research paper will be carried in the May edition of the Journal of Veterinary Diagnostic Investigation, which is published by the American Association of Veterinary Laboratory Diagnosticians.

Many of the diseases affecting the commercial swine industry involve complex syndromes caused by multiple pathogens, including emerging viruses and bacteria.

One pivotal advantage of the Livermore-developed LLMDA over other detection technologies is that it can detect within 24 hours any bacteria or virus that has been previously sequenced.

Using 180,000 probes, the current version of the LLMDA has the capability to detect approximately 8,100 microorganisms, including 3,855 bacteria and 3,856 viruses.

“I call the LLMDA the ‘everything test,’ ” said Raymond “Bob” Rowland, professor of diagnostic medicine and pathobiology at Kansas State’s College of Veterinary Medicine. “It’s really the future of diagnostics for both humans and animals.”

New infectious diseases in animal food production systems can create enormous impacts that can affect domestic consumption and exports, as well as public health in the case of diseases that can move from animals to humans, the paper’s authors wrote.

Two examples of new diseases introduced into the swine industry include theInfluenza A virus subtype H1N1 and Porcine epidemic diarrhea virus. Two other foreign diseases, African swine fever and classical swine fever, remain constant threats to the U.S. industry.

“The best assurance for the timely identification of known and unknown threats is to employ techniques that can detect endemic, emerging and foreign animal diseases using a single test,” Rowland said.

Currently, polymerase chain reaction (PCR) assays represent one technology widely used for pathogen detection, but typically only a handful of microorganisms can be identified in a single test.        

Another method of detecting pathogens, DNA sequencing, greatly expands the number of microorganisms that can be identified, but the technique can be lengthy with costly procedures and requires significant expertise.

“The LLMDA can identify co-infections from a single sample,” said LLNL biologist Crystal Jaing, who oversees LLNL’s microbial detection array collaborations. “A PCR test cannot. The array also can identify co-infections faster and cheaper than DNA sequencing.”  

In their paper, the authors noted that as the LLMDA technology’s cost decreases and throughput increases, it becomes feasible to look at microarrays as everyday tools for use in the diagnostic laboratory.

“The beauty of the LLMDA is that it lets you identify unknown diseases that the researcher isn’t looking for,” Jaing said.

Rowland compared the LLNL-Kansas State University collaboration to mixing peanut butter and chocolate.

“They bring the high-end technology and we bring the practical, real-world application. Typically, these tools are used to identify new organisms, but we recognize in food animal health that many of our problems are complex and polymicrobial.

“These multiple bacteria and viruses end up in a disease syndrome. We’re looking at a complex situation and we need the tools that can give us a comprehensive look at the disease factors involved.”

The Kansas State University-Livermore collaboration is viewed as a strategic partnership by Rowland, who sees the LLMDA as potentially being useful for cattle and poultry diagnostic tests, as well as for pets, such as dogs and cats.

“The most interesting thing that we’ve found in our work is that we’ve been able to pick up not only what is in the animal, but outside the animal. We’ve been able to find out what’s in the water, the feed and the air,” he said.

During their yearlong study, LLNL and Kansas State researchers evaluated known and unknown microbes in blood, oral fluid and tonsils from pigs that have co-infections of porcine reproductive and respiratory syndrome virus (PRRSV) and Porcine circovirus-2 (PCV-2).

The LLMDA easily identified PRRSV and PCV-2, but at higher concentrations compared to gold-standard PCR detection methods. 

The oral fluid samples worked well, with the LLMDA detecting other signatures for several swine-associated bacteria, including Streptococcus, Clostridium and Staphylococcus.

“The use of the microarray technology could help the U.S. detect the emergence of foreign animal diseases at their outset to prevent major disease outbreaks,” Jaing said.

Since its introduction in 2010, the LLMDA has had successful proof of concept demonstrations for a number of areas, including clinical medicine, food safety testing, environmental monitoring and biodefense.

In addition to Jaing and Rowland, other co-authors on the paper are biologist James Thissen, bioinformaticists Shea Gardner and Kevin McLoughlin, all of LLNL; veterinarian Pam Hullinger, of LLNL and the University of California, Davis School of Veterinary Medicine as its Large Animal Clinic director; and biologist Nicholas Monday and veterinarian Megan Niederwerder, both of Kansas State.

The collaboration with Kansas State started in the fall of 2013 under a LLNL Laboratory Directed Research and Development project to develop next-generation diagnostics for detecting agricultural pathogens.