Food Protection Trends - February 2010 - 99

Methods: target Campylobacter and non-target microorganisms were tested for inclusivity and exclusivity by the simPlate method. 37 strains of C. jejuni and C. coli were enriched in Bolton broth, diluted and plated onto simPlate devices and 3 selective agar plates (abeyta-Hunt-Bark (aHB) agar, Campy CeFeX agar and Line agar). Finally, a field trial comparison of the performance of the simPlate C-CI method to the Campy CeFeX method was performed. Lab personnel at 3 poultry processing plants analyzed 168 BPW carcass rinse samples with both methods. Results: there was good correlation for the quantitation of Campylobacter from all three plating methods to the simPlate method; only 2 strains for aHB and 1 strain for Line agar demonstrated greater than 0.5 log difference between both methods. For exclusivity, the C-CI method detected none of the 27 non-target organisms tested. Regression analysis of the results from the field trial comparison showed a correlation of 0.96. Significance: these results indicate that the simPlate C-CI method and the reference culture method are comparable for enumeration of Campylobacter jejuni and Campylobacter coli in poultry meat and poultry meat rinses. P1-29 Detection of Shiga Toxin-producing Escherichia coli (STEC) with the Assurance GDS for STEC Assay Philip t. Feldsine, Markus t. Jucker, andrew H. Lienau and David e. Kerr, BioControl systems Inc., Bellevue, Wa, 98005–4356, Usa Introduction: Recent reports of illnesses caused by foodborne non-o157:H7 shiga toxin-producing Escherichia coli (steC) have led to increased awareness of their threat to public safety. Purpose: a rapid screening assay has been developed to detect E. coli steC isolates with the following o-serotypes: o26, o45, o103, o111, o121, o145. Methods: Immunomagnetic beads are employed to specifically isolate and concentrate bacteria that express these o-antigens during a sample preparation step. Dna from the samples is then amplified and identified using primers and probes directed against conserved, specific, virulence-associated Dna sequence targets in these bacteria. Results: the assay was able to detect 30/31 E. coli strains that expressed one of the o-antigens in question. the one undetected strain did not contain either the stx1 or stx2 gene and is not considered a steC. an additional 40 bacteria, including 15 E. coli strains that express different o-antigens, were not detected. Significance: the data show that the combination of an immunomagnetic sample preparation step and a specific Dna amplification-detection step yield a screening assay specific and sensitive for the top 6 E. coli steC strains known to cause human disease. P1-30 Development and Validation of a Real-time PCR Assay for Detection of Mycobacterium avium subsp. paraturberculosis from a Range of Dairy Matrices JOHN DONAGHy, Jane Johnston, anne Fisher, and Michael Rowe, agri-Food and Biosciences Institute (aFBI), Food Microbiology Branch, aFesD, newforge Lane, Belfast, Ireland Introduction: Mycobacterium avium subsp. paratuberculosis (Map) has concerned the dairy industry by reports of its possible association with Crohn’s disease in humans. the ‘gold standard’ for Map detection remains isolation in culture, however, due to its slow growth rate efforts have been made to use quantitative PCR methods to detect Map in dairy products, particularly milk. Rational: a key factor in the development of a PCR assay for the detection of Map from dairy matrices is the use of an effective method to recover Map from the matrix and subsequently extract Dna from the cells. the objective of this study was to develop and assess the potential of real time PCR assays coupled with a standardized magnetic bead-based Map Dna extraction method, to quantify Map in a range of dairy products including milk, yogurt and a range of cheeses. Results: Map Dna extracted from homogenized cheese and milk samples using the adiapure® kit (adiagene, France) was subjected to two independent taqMan®-based real time PCR assays targeting different gene sequences. the Is900 based real time assay was more sensitive (approx. 10-fold) than the assay targeting the F57 sequence, detecting < 4 CFU ml-1 in artificially contaminated milk and < 30 CFU g-1 in spiked cheese and milk powder samples. In an eU organized dairy products ring trial, involving 12 laboratories, average sensitivities of the prescribed adiapure/Is900 taqMan®-based qPCR method across a range of artificially contaminated dairy matrices were 85–100%. specificities for all matrices were in the range 95–100%. FEBRUARY 2010 | FOOD PROTECTION TRENDS

Food Protection Trends - February 2010

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