Evaluation of Commercial Test Kits for Detection of Salmonella in Alfalfa Sprout Spent Irrigation Water

Introduction: Salmonella contamination accounts for most of the reported sprout outbreaks and alfalfa sprouts are the most frequently linked vehicle. Microbiological testing of spent irrigation water has been recommended as part of an overall strategy to reduce sprout-associated outbreaks. An increasing number of pathogen screening kits based on different assay formats are commercially available and many of these kits have been certified by the AOAC. However, very few of them have been validated for detection in sprouts or spent irrigation water.

Purpose: Evaluate the performance of a number of AOAC certified rapid methods in comparison with the FDA BAM method for detection of Salmonella in spiked alfalfa sprout spent irrigation water. The methods selected include three lateral flow tests (Reveal, RapidChek, Singlepath), three enzyme immunoassays (Assurance Gold EIA, TECRA VIA, VIDAS UP), and five DNA-based assays (BAX, iQ-Check, BAX, MicroSEQ, GeneDisc, Atlas).   

Methods: Six strains of Salmonella (Cubana, Mbandanka, Muenchen, Newport, Tennessee and St. Paul) previously associated with sprout outbreaks were used for preparation of the inoculum. Alfalfa sprout spent irrigation water samples were collected from a commercial sprouting facility at approx. 48 ± 12 h of sprouting. Twenty-five ml of spent irrigation water was inoculated with 0, 1, or 10 CFU of the Salmonella cocktail and was subjected to the enrichment and assay protocols recommended by each method. Regardless of the assay results, the presence of Salmonella in the enrichments was determined following procedures outlined in the FDA BAM. 

Results: All test kits except VIDAS UP were able to detect the presence of Salmonella at a level of 10 CFU/25 ml. For samples spiked at 1 CFU/25 ml level, the Assurance Gold and Atlas tests performed the best, probably due to the use of better enrichment media. RapidChek was the most sensitive test among the three lateral flow tests. Real-time PCR based tests required a shorter period of culture enrichment and were easy to perform with the automated systems. False positive results were observed for the BAX test due to sample interference. The performance of VIDAS UP could be improved by adjusting the pre-enrichment temperature from 42ºC to 37ºC, but false negative results were still observed in samples with the low inoculation level.

Significance: Most of the methods evaluated were able to detect low levels of Salmonella in spiked alfalfa sprout spent irrigation water. The availability of additional validated methods will improve the microbial testing programs for sprouts.