Microfluidic droplet application for bacterial surveillance in fresh-cut produce wash waters

Foodborne contamination and associated illness in the United States is
responsible for an estimated 48 million cases per year. Increased food
demand, global commerce of perishable foods, and the growing threat of
antibiotic resistance are driving factors elevating concern for food
safety. Foodborne illness is often associated with fresh-cut, ready-to-eat
produce commodities due to the perishable nature of the product and
relatively minimal processing from farm to the consumer. The research
presented here optimizes and evaluates the utility of microfluidic
droplets, also termed ultraminiaturized bioreactors, for rapid detection
of viable Salmonella enterica ser. Typhimurium in a shredded lettuce wash
water acquired from a major Mid-Atlantic produce processing facility
(denoted as Producer) in the U.S. Using a fluorescentlylabeled anti- S.
Typhimurium antibody and relative fluorescence intensities, paired with
in-droplet incubation, S. Typhimurium was detected and identified with
100% specificity in less than 5 h. In initial optimization experiments
using S. Typhimuriumspiked sterile water, the relative fluorescence
intensity of S. Typhimurium was approximately two times that of the
observed relative intensities of five non- S. Typhimurium negative
controls at 4-h incubation in droplets containing Rappaport-Vasiliadis
(RV) broth at 37 ° C: relative fluorescence intensity for S. Typhimurium =
2.36 (95% CI: 2.15-2.58), Enterobacter aerogens 1.12 (95% CI: 1.09-1.16),
Escherichia coli 700609 = 1.13 (95% CI: 1.09-1.17), E. coli 13706 1.13
(95% CI: 1.07-1.19), E. coli 700891 1.05 (95% CI: 1.03-1.07) and
Citrobacter freundii 1.04 (95% CI: 1.03-1.05). S. Typhimurium - and E.
aerogens -spiked shredded lettuce wash waters acquired from the Producer
were then incubated 4 h in-droplet at 37 ° C with RV broth. The observed
relative fluorescence of S. Typhimurium was significantly higher than that
of E. aerogens , 1.56 (95% CI: 1.42-1.71) and 1.10 (95% CI: 1.08-1.12),
respectively. While further optimization focusing on compatible
concentration methodologies for highly-dilute produce water samples is
needed, this application of droplet microfluidics shows great promise in
dramatically shortening the time necessary – from days to hours – to
confirm viable bacterial contamination in ready-to-eat produce wash waters
used throughout the domestic and international food industry.