Rapid and Specific Detection of Escherichia coli Serogroups O26, O45, O103, O111, O121, O145, and O157 in Ground Beef, Beef Trim, and Produce by Loop-Mediated Isothermal Amplification

摘要

Escherichia coli O157 and six additional serogroups of Shiga toxin-producing E. coli (STEC) (O26, O45, O103, O111, O121, and O145) account for the majority of STEC infections in the United States. In this study, O serogroup-specific genes ( wzx or wzy ) were used to design loop-mediated isothermal amplification (LAMP) assays for the rapid and specific detection of these leading STEC serogroups. The assays were evaluated in pure culture and spiked food samples (ground beef, beef trim, lettuce, and spinach) and compared with real-time quantitative PCR (qPCR). No false-positive or false-negative results were observed among 120 bacterial strains used to evaluate assay specificity. The limits of detection of various STEC strains belonging to these target serogroups were approximately 1 to 20 CFU/reaction mixture in pure culture and 10~(3) to 10~(4) CFU/g in spiked food samples, which were comparable to those of qPCR. Standard curves generated suggested good linear relationships between STEC cell numbers and LAMP turbidity signals. In various beef and produce samples spiked with two low levels (1 to 2 and 10 to 20 CFU/25 g) of respective STEC strains, the LAMP assays consistently achieved accurate detection after 6 to 8 h of enrichment. In conclusion, these newly developed LAMP assays may facilitate rapid and reliable detection of the seven major STEC serogroups in ground beef, beef trim, and produce during routine sample testing.