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Research Call

DAFM Reference

Lead(Collaborating)Institution

DAFM Award

DAFM National Call 2011 11FP401 UL €92,673

Project Title:

Enhancing consumer safety by development of a rapid flow cytometric assay for early detection of coagulase positive Staphylococcus aureus in chilled ready meals.

Project Coordinator:

Dr Martin Wilkinson

Project Abstract

Currently, coagulase positive S. aureus are enumerated and identified by plate counting on Baird Parker selective agar using the standard method EN ISO 6888-1 which takes 48 h for a result. Thereafter, colonies are subjected to further identification procedures such as growth in BHI medium and plasma coagulation which may take a further 4-5 days. The initial “plate and wait” methodology to detect the presence of potential coagulase positive S. aureus colonies is completely unsuited to provision of rapid information to the producer for early process intervention to ensure consumer safety. This 48 h step also places a constraint on the optimal release time for the product to the market with implications for shorter shelf life and limitations on export market exploitation. This project seeks to exploit novel fluorescent activated cell sorting (FACS) based methods for microbiological enumeration of S. aureus using methodology developed in a previously funded DAFF project, 06RDUL413. The new assay will allow enhanced total time to result (TTR) of ~ 4h compared with the selective agar plating method enabling food producers to rapidly identify the presence of potential coagulase positive S. aureus in samples originating from the production process, personnel and in the final product. Thus the proposal will significantly enhance consumer safety and provide an extra degree of security for the producer. In the FIRM project, FACS with immunological tagging enabled detection of S. aureus in foods at levels > 103 within an analytical window of ~4 hours including time for sample preparation and data analysis. The assay featured: (a) selective labelling of S. aureus in a mixture of bacteria using commercial antibodies, (b) simultaneous enumeration and calculation of the labelled S. aureus cells using fluorescent bead standards, and (c) confirmatory testing by sorting of labelled cells by FACS onto traditional selective Baird-Parker media. These outputs represented a potentially significant advance in time saving and analytical labour costs for the food industry especially the chilled ready meals sector where shelf life ranges from 10 – 14 days. This proposal now seeks to improve the sensitivity of the antibody labelling technique, ensuring that it only labels “live” coagulase positive Staphylococcus aureus cells and validate its performance in comparison with ISO/EN plate count methodology with a leading Public Health Microbiology Laboratory, (PHML, Cork) and an industry partner Dawn Fresh Foods. Currently, the prototype method is operated on expensive FACS technology and a key part of this proposal will be to translate and simplify the assay methodology onto affordable and operator friendly benchtop laboratory cytometers suitable for routine quality control analysis in the food and process industries.

Final Report:

Not available yet.