Purpose: The goals were to characterize the physiological state of EHEC on pre-harvest lettuce, and determine if the stresses experienced by EHEC on pre-harvest lettuce impact its ability to survive subsequent decontamination treatments.
Methods: Greenhouse-grown Romaine lettuce was spray inoculated with one of 4 different EHEC strains (O157:H7, n = 2; O26:H11, n = 2) and incubated in the greenhouse for 5 days. On days 1, 3, and 5, lettuce leaves were washed for 2 min in sterile water or 50 ppm chlorine. EHEC were enumerated after washing by plating on MacConkey agar. Changes in EHEC gene expression were measured over time by extracting RNA from inoculated lettuce and using RNA-seq.
Results: One day post-inoculation, all 4 strains had an average 1.0 ± 0.3 log CFU/g lower recovery from chlorine-washed lettuce vs. water-washed lettuce. Three and five days post-inoculation, the O157:H7 spinach outbreak strain had an average of 0.4 ± 0.2 log CFU/g lower recovery from chlorine washed lettuce, while the other three strains remained at ~ 1 log CFU/g. Analysis of RNA-seq data indicated significantly higher transcript levels for genes encoding proteins involved in responding to oxidative stress for the O157:H7 spinach strain on day 3 post-inoculation (P > 0.05).
Significance: Adaptation of EHEC to stresses on pre-harvest lettuce can lead to cross-protection to subsequent stresses. Assessing physiological changes that occur during this adaptation can provide a greater understanding of stresses the pathogen is experiencing, and may be used to develop control measures for this pathogen in the food supply.