Variation in Confirmation Rates of Samples Screen-positive for Escherichia coli O157 in Beef Trim by Using PCR to Screen for Virulence-associated and Serotype-specific Targets

Introduction: Confirming the presence of Escherichia coli O157 in beef trim samples requires tests subsequent to screening. Because of time factors, beef processors may rely on screening methods to flag potentially positive samples and make decisions about product lot disposition. While screening methods must be sensitive, too many “false-positive” screens can lead to product disposition that is economically unfavorable for the producer.

Purpose: In this study, we determine the confirmation rate of screen-positive beef trim samples for a method to detect virulence-associated and serotype-specific genes that are found in E. coli O157 and to determine if there are factors such as processing plant, month and sample lean content that affect this rate.

Methods: From 15 different processing plants that reported O157 confirmation test results during calendar year 2010, 460,137 trim samples (375 g, 60 surface pieces) were enriched ≥ 9 h and then screened by a multiplex PCR targeting the rfb, stx and eae genes and a lateral flow (LF) immunological test for the O157 antigen.  Samples which were rfb⁺ and/or LF⁺, eae⁺ and stx⁺ (stx₁⁺ and/or stx₂⁺) were designated as O157 IR (Initially Reactive). Confirmation tests included immunomagnetic separation using anti-O157 antigen beads and PCR assays for rfb, eae, H7, γint, subtilase, and stx.

Results: Overall, 15.5% of the O157 IR samples (which signals come from mixed enrichment cultures) were confirmed positive for E. coli O157 by subsequent molecular tests.   During the year, confirmation rates ranged from 5.9-25.6% and tended to be higher from late mid-spring through fall.  For PCR signals, results ranged from 7.3-20.6% with stx₂^- the lowest and stx₁^-  the highest. Confirmation rates for processing plants ranged from 3.2-72.13%. Samples 50-69 and 70-89 percent lean had confirmation rates of 19.9 and 12.%, respectively.

Significance: Overall, 15% of screen-positive samples remained positive after confirmatory testing. Confirmation rates varied by month, by plant, individual gene tests and less so by day of the week. Sampling plans might be designed to take these variations into account, especially during the summer and fall. Variations between plants could be used to examine their processing controls.