PRESS RELEASE

FDA presents data on the use of Pathatrix for Bio-threat applications in Food at the 2005 FDA Science Forum

Isolation of Yersinia pestis from Food using Immunomagnetic Capture.

D. E. Hanes, M. L. Saylor, M. H. Kothary, M. E. Lorenzo, B. D. Tall, CFSAN, FDA, Laurel, MD

Heightened concerns about bioterrorism have required that the FDA strengthen their methods for the isolation and identification of pathogens not traditionally associated with foodborne disease. One such pathogen is Yersinia pestis (Yp), the causative agent of plague. This study examines the use of the Pathatrix system of immunomagnetic capture for the isolation of Yp from foods. Immunomagnetic beads were coated with rabbit polyclonal antiserum raised to whole cells of Yp strain A1122.

Food samples spiked with A1122 were placed in sterile stomacher bags and diluted 1:10 with Butterfield's phosphate dilution buffer. The samples were placed in the Pathatrix apparatus and circulating tubing placed into the stomacher bags. Antibody-coated immunobeads were introduced into the tubing and magnetically bound in place. The food samples were circulated past the immunobeads for 30 min at 28°C, after which the immunobeads were harvested and washed with 0.85% saline.

Aliquots of the immunobeads were directly plated onto Sheep Blood Agar (SBA) and Yersinia selective agar (CIN), and the cultures incubated for 48 h at 28°C. Immunobeads were also inoculated into Brain Heart Infusion Broth as an enrichment culture. Following 24 h of incubation, the enrichment cultures was plated onto CIN and SBA and the plates incubated at 28°C for 48 h. Identity of suspect colonies subcultured from direct plating and enrichments were confirmed using lysis by an Yp specific phage and by PCR. PCR analysis was also performed on aliquots of the immunobeads as a rapid screening method for the presence of Yp. Yp was recovered from both the direct plating and the enrichment cultures on CIN and SBA. Colonies of Yp were more evident on CIN however there was a 10-100 fold decrease in recovery on CIN when compared to SBA. Yp was also detected in all samples using PCR analysis.

These results indicate that immunomagnetic capture can be used to isolate and identify Yp from foods. These studies also will facilitate the development of a rapid method for the identification of Yp in food using immunocapture combined with PCR.

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