Quantitative transfer of Escherichia coli O157:H7 during pilot-plant production of fresh-cut leafy greens

Cross-contamination/multi-directional transfer of Escherichia coli O157:H7 during commercial shredding, conveying, flume-washing and drying of fresh-cut leafy greens has become a major public health concern. However, the extent of bacterial transfer during the various unit operations involved in post-harvest processing of leafy greens is poorly understood. Consequently, a pilot-scale leafy green processing line consisting of a commercial shredder, conveyor, flume tank, shaker table, and centrifugal dryer was assembled to E. coli O157:H7 transfer during processing of iceberg lettuce, Romaine lettuce, and baby spinach.;Using 22.7 kg of leafy greens inoculated with a 4-strain avirulent, GFP-labeled, ampicillin-resistant E. coli O157:H7 cocktail at 6, 4 and 2 log CFU/g, bacterial transfer coefficients were determined between the product, the water, and the equipment surfaces during processing, with the effect of post-inoculation hold time and shred size on transfer also evaluated. During leafy green processing, ∼90% of the E. coli O157:H7 inoculum transferred to the wash water. After processing, E. coli O157:H7 populations on equipment were highest on the shredder and conveyor. Shred size did not affect the numbers of E. coli O157:H7 transferred from contaminated leafy greens during processing. However, extending the time between inoculation and processing decreased the removal and transfer of this pathogen during processing. When 22.7 kg of product containing 10 6 and 10 4 E. coli O157:H7 CFU/g were processed followed by 22.7 kg uninoculated product, E. coli O157:H7 was quantifiable in all iceberg and romaine lettuce samples. At an inoculation level of 2 log CFU/g, E. coli O157:H7 was sporadically detected after processing 81.6 kg of uninoculated iceberg and found throughout the entire batch of uninoculated romaine lettuce.;Using Radicchio as a colored surrogate for iceberg lettuce, this study demonstrated that 9.1 kg of E. coli O157:H7-inoculated Radicchio could contaminate 907 kg of subsequently processed uncontaminated iceberg lettuce. After processing, shreds of Radicchio were found in all but one of the ∼38 bags of shredded lettuce. Hundreds of contaminated Radicchio shreds remained on the processing line, with the majority found on the conveyor, followed by the shredder, flume tank, and shaker table, showing that contaminated product can be continually spread during leafy green processing long after a contamination event.;The effect bacterial attachment time on cross-contamination was determined by processing 0.5 kg of E. coli O157:H7-inoculated Radicchio immediately and after storage for 1 d at 4°C or 5 d at 22°C, followed by 45.5 kg of uninoculated iceberg lettuce. After processing, E. coli O157:H7 was found in all uncontaminated iceberg lettuce samples, with mean counts highest for 1 h- followed by 1 d- and 5 d-held product. Scanning electron microscopy images showed greater bacterial attachment for 5 d-held product, with larger cell aggregates enveloped in a dense extracellular matrix.;When transfer between inoculated and uninoculated product was assessed during partial dewatering, faster cross-contamination was seen using an E. coli O157:H7-inoculated Radicchio to uninoculated iceberg lettuce ratio of 1:100 than 1:10,000 (w/w) until a maximum population was reached due to the higher number of E. coli O157:H7 cells available for transfer. These findings, which demonstrate E. coli O157:H7 transfer between product, wash water, and equipment surfaces, and to large quantities of uncontaminated fresh-cut leafy greens, are critical to the development of science-based transfer models for risk analysis.