Jan. 1, 2009 - Dec. 31, 2009Award Number
Beilei Ge, Ph.D.
Food and Drug Administration, Center for Veterinary Medicine
Salmonella is a major human pathogen in a variety of fresh produce items. Recent outbreaks linked to Salmonella-contaminated produce greatly undermined consumer confidence and heightened the need for applied targeted research from field to fork. Among multi-faceted strategies to prevent future outbreaks and to reduce the incidence of produce-associated illness, sensitive and specific detection methods are imperative since they serve as an important quality control tool for safe produce from field, processing, to retail. However, there are two main drawbacks of current PCR-based methods: lack of specificity and the inability to differentiate live/dead cells. This project aims to address these challenges associated with Salmonella detection in a variety of fresh produce items by incorporating two novel techniques, the loop-mediated isothermal amplification (LAMP) for increased specificity and the use of ethidium monoazide bromide for the distinction of live/dead cells. Firstly, LAMP primers will be designed that target a unique marker for more than 100 Salmonella serovars, and the assay will be fully optimized. Secondly, the assay will be evaluated for sensitivity and specificity using a collection of 50 Salmonella strains and 50 non-Salmonella strains. The ability to differentiate live/dead cells will also be evaluated. Finally, the assay will be applied in the detection of live Salmonella in experimentally contaminated produce items (shredded lettuce, baby spinach, sliced tomato, sprouts, and cantaloupe cubes) of various stages of maturity. Upon completion, this project will generate an optimized LAMP detection assay readily applicable to the produce industry for the sensitive and specific detection of live Salmonella cells. Such detection assays will provide the produce industry personnel an increased ability to control potential hazardous microbes in produce. This improvement will provide significant benefits to produce growers, harvesters, processors, retailers, and the ultimate consumers by implementing rapid mitigation methods for contaminated produce, and therefore significantly reducing the risks and illnesses associated with the consumption of fresh produce.