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Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of  Escherichia coli  O157:H7. While the amounts of culturable  E. coli  O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for  E. coli  O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA) real-time PCR assay to quantify viable (with PMA) and total (without PMA)  E. coli  O157:H7 cells on growth chamber and field-grown lettuce.  E. coli  O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 10 6  CFU/plant. In the growth chamber at low relative humidity (30%), culturable amounts of the nontoxigenic  E. coli  O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer  E. coli  O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 10 4  greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable  E. coli  ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of  E. coli  O157:H7 cells either die or are no longer culturable.

Assessments of Total and Viable Escherichia coli O157:H7 on Field and Laboratory Grown Lettuce