Sprouts and E coli O157 EcO157 and Se

Numerous outbreaks of EcO157 and Se associated with contaminated sprouts have occurred since 1995, and a number of the outbreaks have been traced to seeds contaminated with relatively low levels of pathogen [146]. Although calcium hypochlorite at 20 mg/ml has been recommended for sanitizing seeds [147], it does not remove the entire natural microbial flora on the seed, suggesting that bacteria are attached in sites inaccessible to chemical treatments [13]. A comparison of the growth of multiple EcO157 and Se strains on alfalfa sprouts revealed major differences in attachment among the strains [27]. Six strains of EcO157 grew an average of 1.5 log10 less on sprouts compared to five strains of Se (Table 2.3). An EcO157-GFP strain attached poorly to sprout roots and shoots, whereas individual cells and aggregates of Se Newport-GFP were observed adhering to sprout seed coat edges and root hairs (Figure 2.3D). The 10- to 1000-fold difference in attachment to sprout tissues by EcO157 compared to Se strains was confirmed

FIGURE 2.3 (Color insert follows page 594) Confocal micrographs of bacteria on plant leaf, stem, and root tissues, and bacteria bound to material extracted from leaves. (A) Natural microorganisms, mostly bacteria, bound to junction of epidermal cells on a lettuce leaf. The bacteria were stained with LIVE BacLight Gram stain (Molecular Probes, OR). (B) GFP-labeled S. enterica and dsRed-labeled P. agglomerans cells bound singly and in aggregates after their inoculation and incubation on the leaves of cilantro

plants. Natural epiphytic bacteria were stained with SYTO 62 (Molecular Probes) and were detected in the close vicinity of the inoculated strains. The SYTO 62 signal was assigned the pseudocolor blue. (C) GFP-labeled EcO157:H7 bound in the region of a lateral root emerging from an Arabidopsis thaliana plant. The arrow points to a region where the EcO157 cells have become internalized. (D) GFP-labeled S. enterica bound to the root hairs (Rh) of an alfalfa sprout. (E) A thick biofilm of natural microorganisms colonizing the root of an alfalfa sprout and stained with LIVE BacLight Gram stain. (F) GFP-labeled S. enterica cells attached to a dried compound extracted from cilantro leaves and identified as stigmasterol. (Brandl and Mandrell, unpublished data.).

in subsequent studies [148]. Interestingly, a Se Newport strain appeared to be as fit in sprouts up to at least three days as three epiphytic strains isolated from sprouts. In addition, four nonpathogenic E. coli strains isolated from field-grown cabbage (Table 2.3) attached as well as the epiphytic strains, but less than Se Newport. The results of these and other studies of Se on sprouts [149] indicated that specific interactions occur between human pathogens and plants, and suggested that enteric bacteria and human pathogens isolated recently from plant surfaces may retain fitness and attachment capability for plants.

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