A group of researchers is working to develop a quick, easy and affordable molecular test to simultaneously detect and differentiate four protozoan pathogens that could potentially cause foodborne outbreaks in packaged salads.
Should the results from the multiplex PCR assay, as the test is known, be positive, a subsequent but different PCR test could determine whether the pathogens are viable.
To make it even more user friendly, the PCR assay would be designed for use alongside those routinely used to screen produce for other pathogens, such as E. coli, Salmonella or Listeria.
Leading the effort is Dr. Stefan Wuertz, a professor emeritus in environmental engineering at the University of California, Davis.
Having the ability to readily screen for these protozoan pathogens would give the produce industry another tool to better manage food safety risks, he said.
"Protozoa are increasingly seen as biological agents involved in outbreaks," Wuertz said. "They are implicated in a growing number of reports, and we know some of these protozoa are expected to have a larger presence in the United States."
The project focuses on four protozoa associated with produce-borne outbreaks: Cryptosporidium, Giardia, Toxoplasma and Cyclospora. Although the produce industry routinely tests for fecal indicator bacteria or specific bacterial pathogens, they rarely test for protozoan contamination.
Current methods used to detect protozoan oocysts - the hard-shelled protozoan life stage shed in human feces - require time-consuming techniques conducted by specialized laboratories.
"That would never be something that the producer or processor would be likely to routinely implement," Wuertz said. "So even though there are methods, they are really better used in the hands of experts. Ultimately any such method for the produce industry would only be used and interpreted if it is relatively straight forward and relatively easy to interpret."
Real-time PCR (or qPCR), or polymerase chain reaction, can quickly identify microorganisms through their DNA. Many tests have been developed to detect numerous other organisms, and Wuertz said his project's goal is to do the same for the four protozoa.
But just because the test allows you to detect DNA doesn't necessarily indicate whether the organism is alive or dead. Follow-up qPCR assays would assess the organism's viability.
Joining in the research are
Dr. Karen Shapiro
, a faculty member in the UC Davis School of Veterinary Medicine, and Dr. Minji Kim, a post doctoral researcher in the UC Davis
Department of Civil and Environmental Engineering
Shapiro has conducted research on Toxoplasma, one of the target organisms, for more than 10 years. "So it's really good to have someone who has a good understanding of the biology of the organism," Wuertz said.
Like himself, Kim is an environmental engineer who takes an analytical approach to water quality and public health. Dr. Veronica
Rajal, an associate professor at the National University of Salta in Argentina, brings an international perspective and first-hand experience with protozoa.
"She has worked many years on the detection of pathogens, and the pathogens we're working on are very commonly detected in her country," Wuertz said. "She's also an expert in PCR, so she brings that element."
In addition, Rajal teaches fundamentals of biotechnology and food preservation and understands the food processors' perspective.
Wuertz said he had hoped to work with a cooperating packaged
salad processor that would remain anonymous, and he still hopes to be able to do so. Any company interested in collaborating may contact CPS or Dr. Wuertz directly.
"The most important benefit would be to simply listen to their viewpoints," he said. "What we think is so important to them may not be so. Is it more important to have a method that gives us a 'yes' or 'no' answer? Or are we more relevant if the method is not as quick but is more quantitative?
We can complete the project without a cooperator, but having one would bring a different viewpoint and can help us set priorities and provide inputs into the writing part of it."