IAFP 2009 Program Book and Abstract Book - AB108

Results: Aerobic plates showed that counts for the level of background flora were lower in boiled pork than in broccoli sprouts, less than 100 CFU/g and 6.7 × 107 (± 1.5) CFU/g, respectively. In boiled pork, the culture method and real-time PCR detected Salmonella equally well, and a statistically significant difference was not observed between the two methods. In broccoli sprouts, the culture method identified 16 out of 80 samples (20%) as positives, and real-time PCR similarly identified 39 out of 80 samples (49%) as positives. With regard to this result, obtained from 4 trials, an extremely statistically significant difference was observed between the two methods. Significance: The real-time PCR assay was less affected by background flora and more sensitive in detecting low numbers of Salmonella than the culture method, particularly in the sample with a high degree of background flora. For further improvement of the culture method and real-time PCR, additional measures will be needed to increase the number of Salmonella to detectable levels in samples with high levels of background flora to minimize the effects of background flora. P2-13 Washing and Enrichment of Jalapeño Peppers Using Small Volumes of Non-selective Broth Facilitates Rapid Cytometric Detection of Salmonella Saintpaul BLEDAR BISHA and Byron F. Brehm-Stecher Iowa State University, Food Science & Human Nutrition, Ames, IA, USA Introduction: The 2008 outbreak of salmonellosis associated with the consumption of jalapeño and Serrano peppers imported from Mexico highlights the need for rapid methods for detection of this pathogen on this and other types of fresh produce. Although enrichment of produce represents a bottleneck, it may be possible to reduce overall assay time through the use of improved methods for sampling and sample preparation prior to enrichment. Purpose: The purpose of this study was to develop a rapid, streamlined method for detection of Salmonella ser. Saintpaul on jalapeño peppers. Elements evaluated included use of a larger analytical unit and a one-step approach for wash-based removal of bacteria with delivery into a concentrated enrichment volume. Methods: Jalapeño peppers were spot inoculated with stationary phase cells of S. Saintpaul ATCC 9712. After drying (~3 h), cells were detached via pulsification into 10 ml Trypticase Soy Broth (TSB) for non-selective, low-volume enrichment at 37°C. One-milliliter aliquots were taken at regular intervals during enrichment and examined via combined fluorescence in situ hybridization (FISH) and flow cytometry. Use of a larger analytical unit (4 peppers, ~100 g), and use of low-volume washing with 0.1% peptone water (PW) for direct detection of S. Saintpaul were also examined. Results: Pulsification enabled rapid, vigorous and non-destructive washing of jalapeño surfaces, facilitating removal and concentration of attached bacteria into small volumes of TSB or PW. Using FISH and flow cytometry, we were able to detect 103 CFU g-1 S. Saintpaul directly in PW, or as few as 10 CFU g-1 after only 6 h of enrichment in TSB. Instead of being limited to the testing of individual peppers, we found that we could also analyze a larger analytical unit (4 peppers, ~100 g) using this method. Significance: We have developed a simple “FISH and flow” method for rapid and sensitive detection of S. Saintpaul from jalapeño peppers. The method takes advantage of the thorough but non-destructive nature of Pulsifier-based washing for removal of bacteria from pepper surfaces, combined with dual use of small volumes of non-selective media as both a wash and as a growth medium. Because total assay time (wash-based removal of cells, cell fixation, hybridization and flow cytometry) was ~2 h, our method may enable the detection of low numbers of salmonellae on jalapeño surfaces within a single eight hour work shift. This approach provides the fresh produce industry with a rapid tool for monitoring jalapeño peppers for contamination with Salmonella, and may also be useful for rapid detection of this pathogen in other types of fresh produce. P2-14 Evaluation of PCR Detection of Salmonella in Alfalfa Sprouts and Spent Irrigation Water Collected during Sprouting of Naturally Contaminated Seeds NICOLE D. MAKS and Tong-Jen Fu National Center for Food Safety and Technology/Illinois Institute of Technology, Summit-Argo, IL, USA Introduction: Traditional Salmonella confirmation methods can take up to four days, and high levels of background flora, if present, often interfere with the isolatation of typical colonies from selective agar plates. PCR-based detection is a more sensitive method that produces results in shorter time. Our previous work revealed that sprout irrigation water spiked with low numbers (1 to 100 CFU) of Salmonella enriched overnight in tetrathionate (TT) broth and buffered peptone water with novobiocin (BPW+n) was able to detect Salmonella using a PCR detection system, but few studies using PCR for detection of Salmonella in naturally contaminated products have been published. Purpose: This study evaluated the efficacy of a PCR-based system (DuPont Qualicon BAX) using two enrichment protocols for detection of Salmonella in sprouts and spent irrigation water collected during sprouting of seeds naturally contaminated with Salmonella. Methods: Alfalfa seeds naturally contaminated with Salmonella were grown in jars at 20°C and 30°C for three days. Afterwards, five samples of spent irrigation water and five samples of sprouts were enriched overnight individually in TT and BPW+n at 42°C and in lactose broth (FDA BAM) at 37°C. Levels of Salmonella present in the spent irrigation water and sprouts were determined by MPN analysis, and both were plated on PCA to determine background flora levels. After enrichment in TT and BPW+n, the samples were run in the BAX system and results compared to the FDA BAM method. Three trials for each growth temperature were done. Results: Salmonella levels were lower at 20°C than at 30°C for some trials, and background flora levels ranged from 107 to 108 CFU/g or ml at 20°C and 108 to 109 CFU/g or ml at 30°C. In trials with a Salmonella level >1.1 MPN/g or ml, both the BAX and FDA BAM methods were able to detect positives in all samples. In trials with lower levels of Salmonella, BAX was able to detect more positive samples than FDA BAM. For one trial with < 0.003 MPN/g or ml of Salmonella, the presence of the pathogen was not indicated by either the BAX or the FDA BAM method. 108 — Abstract Book Poster

IAFP 2009 Program Book and Abstract Book

Table of Contents for the Digital Edition of IAFP 2009 Program Book and Abstract Book

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