Concentration of Large Volumes of Irrigation Water Facilitates Sensitive Detection of Foodborne Pathogens II

Fecally contaminated irrigation water is frequently implicated as either a source of contamination of fresh produce with foodborne pathogens. Since the concentrations of pathogenic microorganisms in irrigation water are usually low, sampling of larger volumes of water should be performed prior to detection and quantification. The objective of this study was to evaluate the use of Modified Moore swabs and continuous centrifugation to concentrate Escherichia coli O157:H7 and Salmonella spp. from large volumes of irrigation water. Three strains each of produce isolates of E. coli O157:H7 and Salmonella spp. were independently transformed with plasmids expressing one of red, green, or cyan fluorescent proteins. The strains were grown overnight and combined in two cocktails (one for E. coli, and another for Salmonella), and suspended in 10 liters of water at final concentrations of 10-1, 100, 101, and 102 CFU/ml. Concentration via centrifugation was performed in a CFC-200 continuous flow centrifuge (Scientific Methods Inc., Granger, In) at a flow rate of 300 rpm and 2,500 x g. For Modified Moore swab filtration, concentration was performed at a flow rate of 300 rpm, using a peristaltic pump attached to a filter housing in which the swabs were placed. Following concentration, samples were surface plated onto tryptic soy agar supplemented with 50 µg/ml of ampicillin, followed by incubation at 37oC for 24h. For samples with lower contamination levels (10-1 and 100/ml), 500 µl of concentrate was transferred to 5 ml of tryptic soy broth with ampicillin and enriched overnight at 37oC with shaking, followed by testing via lateral flow assay (Neogen Corp., Lansing, Mi). The results indicated that both E. coli O157:H7 and Salmonella spp. were concentrated by 1.5 to 2 orders of magnitude within 35 minutes, when either the Modified Moore swab, or continuous centrifugation was performed. Centrifugation was more sensitive that the Modified Moore swab, although this difference was statistically insignificant. At lower bacterial concentrations, the samples were enriched both pre and post-concentration, since the numbers of bacteria were below the levels of detection by plating. The results showed that E. coli O157:H7 and Salmonella were not detected in the 10-1 and 100 CFU/ml before concentration by either method, but were detected following concentration in the initial 100 CFU/ml sample (Moore swab) and 10-1 and 100 CFU/ml samples (centrifugation). Both Modified Moore Swabs and continuous centrifugation represent rapid methods to concentrate bacterial pathogens in irrigation water prior to detection.