Differences in Growth of Salmonella enterica and Escherichia coli O157:H7 on Alfalfa Sprouts

Abstract

Sprout producers have recently been faced with several Salmonella enterica and Escherichia coli O157:H7 outbreaks. Many of the outbreaks have been traced to sprout seeds contaminated with low levels of human pathogens. Alfalfa seeds were inoculated with S. enterica and E. coli O157:H7 strains isolated from alfalfa seeds or other environmental sources and sprouted to examine growth of these human pathogens in association with sprouting seeds. S. enterica strains grew an average of 3.7 log 10 on sprouting seeds over 2 days, while E. coli O157:H7 strains grew significantly less, an average of 2.3 log 10 . The initial S. enterica or E. coli O157:H7 inoculum dose and seed-sprouting temperature significantly affected the levels of both S. enterica and E. coli O157:H7 on the sprouts and in the irrigation water, while the frequency of irrigation water replacement affected only the levels of E. coli O157:H7. Colonization of sprouting alfalfa seeds by S. enterica serovar Newport and E. coli O157:H7 strains transformed with a plasmid encoding the green fluorescent protein was examined with fluorescence microscopy. Salmonella serovar Newport colonized both seed coats and sprout roots as aggregates, while E. coli O157:H7 colonized only sprout roots.Sprout producers in North America, Asia, and Europe have faced numerous Salmonella enterica and Escherichia coli O157:H7 outbreaks since 1995 (2, 17). Several of the outbreaks were traced to sprouts grown from seeds contaminated with low levels of human pathogens (2,15,19,20,27).As a result of the many recent outbreaks traced to sprouts and guidance by a U.S. Food and Drug Administration document (2), many producers now sanitize their sprout seeds with 20,000 ppm of calcium hypochlorite before sprouting and test each crop of sprouts for S. enterica and E. coli O157:H7. Epidemiological evidence suggests that this sanitation protocol may prevent some, but not all, outbreaks (2, 5), and experimental sanitation of naturally contaminated alfalfa seeds did not eliminate S. enterica from the seeds (23). To date, no seed sanitation method has been shown to eliminate S. enterica or E. coli O157:H7 from laboratory-contaminated seeds (4,13,14,19,24,25). In addition, the recommended calcium hypochlorite method does not kill or remove all naturally occurring nonpathogenic bacteria from alfalfa seeds, suggesting that bacteria on seeds may be in locations inaccessible to calcium hypochlorite treatment (7).Sprouts present an unusual food safety predicament compared to other fresh produce because bacteria, including S. enterica and E. coli O157:H7, may multiply by several logs on the sprouting seeds during the first few days of germination (1,22,23). Experiments with radish and alfalfa sprouts grown from laboratory-contaminated seeds have demonstrated that human pathogens may be present between plant cells inside sprouts, where they can resist decontamination treatments (9, 12). Washing sprouts in water only slightly reduces the number of bacteria found on the sprouts, and green sprouts are usuall...

Microbiological quality of organic vegetables produced in soil treated with different types of manure and mineral fertilizer

“…The ability to colonize vegetable roots and shoots varies for different strains of a bacterial species (4,61) and for different bacterial species (4,14,19). Bacterial motility (17) and the ability to use seed exsudates as carbon sources (52) are related to the extent of colonization.…”

De Novo Amino Acid Biosynthesis Contributes to Salmonella enterica Growth in Alfalfa Seedling Exudates

“…The very low bacterial populations on contaminated seeds make detection by routine seed lot screening difficult (13), but during seed germination, the S. enterica population can grow 1 million-fold (to 10 9 CFU/ml) within 48 h (24). The rapid exponential growth of S. enterica on germinating seeds (24,25) shows that they provide a very favorable environment for enteric human pathogen growth.…”

Flow cytometry for rapid detection of Salmonella spp. in seed sprouts

“…Components of the assay included a set of cooperatively binding probes, a chemical blocking treatment intended to reduce non-specific background, and sample concentration via tangential flow filtration (TFF). We were able to detect S. Typhimurium in sprout wash at levels as low as 10 3 CFU ml 1 sprout wash (10 4 CFU g 1 sprouts) against high microbial backgrounds (∼10 8 CFU g 1 sprouts).…”

“…The most common pairing has been Salmonella in alfalfa sprouts. The high incidence of Salmonella contamination of alfalfa sprouts may be rooted in ecological factors, such as the capacity of this pathogen to adhere strongly to sprout surfaces so that it is retained at high levels in the finished product, even after periodic irrigation during the sprouting process [7,8]. Over the years, various serovars of S. enterica have been involved in sprout-borne outbreaks, including Bareilly, Cubana, Enteritidis, Kottbus, Mbandaka, Muenchen, Newport, Saintpaul, Stanley, Weltevreden and others [9,10,11,12].…”