Behavior of Shiga Toxin-producing Escherichia coli (STEC) Strains during Lettuce Washing

Introduction: To validate washing processes for sanitizing leafy greens, identification of microbial surrogates for Shiga toxin-producing Eschericha coli (STEC) may be useful.  Because an ideal surrogate would imitate STEC, it is important to characterize the behavior of various STEC strains in washing processes, e.g. in transfer, attachment, and survival.

Purpose: To compare STEC serotypes and non-pathogenic E. coli in the extent of transfer and attachment to lettuce during washing.

Methods: Transfer of eight STEC strains (O157:H7, O104:H4, and the “Big Six” non-O157 STEC) and non-pathogenic E. coli Nissle 1917 (EcN) was compared in 30-ml washes of lettuce with or without chlorine. E. colipopulations remaining on inoculated leaves, in wash water, and transferred to un-inoculated leaves after a 1-min wash were enumerated by plate counts on selective agar in triplicate trials.  An attachment assay was performed to determine the number of attached cells after repeated washes. 

Results: During an un-chlorinated water wash, > 90% of inoculated cells was transferred (all strains). Transfer of EcN, O111, and O145 did not differ from that of O157:H7 (P > 0.05); but the other STEC strains showed less transfer than O157:H7 (P < 0.05).  Approximately 2 - 3 log CFU transferred onto un-inoculated leaves.  In a 1-ppm chlorine wash, EcN showed significantly more tolerance to the chlorine wash (P < 0.0001); more cross-contamination in wash water was detected for O111 (P = 0.0027), compared to O157:H7. 

Preliminary attachment data showed that O157:H7 and O145 attached less than the other STEC strains, and EcN demonstrated stronger attachment than O157:H7 (P = 0.0057).

Significance: Data of STEC transfer and attachment during washing will be valuable for assessing cross-contamination risks in post-harvest washing of lettuce. These methods also will be useful for identifying potential STEC surrogates for process validation.