Project seeks test for 'viable but non-culturable' pathogens

Project seeks test for 'viable but non-culturable' pathogens

A handful of pathogens - including Salmonella and Shiga toxigenic E. coli (STEC) - can go into a dormant state where they remain virulent but can't be detected through traditional culturing methods.

Dr. Xiaonan Lu, an associate professor of food science at the University of British Columbia in Vancouver, B.C., Canada, wants to change that. He is leading a group of researchers in an effort to develop a quick, easy-to-use and economical assay to detect these dormant pathogens - referred to as viable but non-culturable or VBNC - on fresh and processed produce.

"VBNC is a relatively new concept to the fresh produce industry," Lu said. "That's why gaining a better understanding of VBNC in fresh produce is extremely important. There's very, very limited studies that have been done to prove VBNC is on fresh produce, so this is a pretty new topic."

Depending on the project's outcome, Lu said growers and processors may need to rethink the types and levels of antimicrobials they use to treat water to account for VBNC pathogens.

Joining him are co-investigators Dr. Pascal Delaquis and Dr. Susan Bach, both research scientists with Agriculture and Agri-Food Canada based at the Summerland Research and Development Center in Summerland, B.C. Dr. Jeff Farber, professor and director of the Canadian Research Institute for Food Safety at the University of Guelph, Guelph, Ontario, also is a co-investigator.

The two-year project, titled "Detection, validation and assessment of risks implied by the viable but non-culturable state of enteric bacterial pathogens in fresh produce," has three pillars.

First, the researchers plan to develop a new molecular detection method for VBNC that is user friendly. "The current detection methods for VBNC bacteria have some limitations," Lu said. "During the past six months, after some optimization and after a lot of hard work, we've already finished proof of concept that the new detection method is working." The researchers still have a few parameters of the new assay to fine-tune, but Lu said they expect to have the work completed by this fall.

During the second part, the researchers plan to test the new detection method in lettuce fields at the Summerland Station. They also plan to spike lettuce, tomatoes and spinach with the VBNC pathogens to ensure the assay works with plants. Their earlier work was done in vitro in the laboratory. "We want to know how robust the test is," Lu said. Known technically as an isothermal PCR, it uses a warm water bath to keep the bacteria at 63 degrees Celsius (145 degrees Fahrenheit) as the population is amplified for better detection.

"This means we don't need a special machine, such as a thermal cycler for the convectional PCR, so this method is very suitable for in-field studies. You don't need to carry a very heavy thermal cycler to go to the field," he said. The entire test can be completed in less than an hour, speeding decision-making for growers and processors. "The easier the technique and the quicker the technique, the better," Lu said.

In addition, the researchers plan to survey lettuce fields to determine the survival of VBNC both in the field and during simulated processing. "We're trying to do the simulated processing and prepare the fresh produce on different cutting boards and solid surfaces so we can see the (VBNC) survival," he said.

The third pillar involves Dr. Farber, who will develop a risk assessment of VBNC using data from the field trials and simulated processing.
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