The investigation of an outbreak of SeT associated with cilantro suggested that the cause of the outbreak was a result of preharvest contamination of imported cilantro . The outbreak strain of SeT increased on cilantro leaves ~1.0 log10 and 2.0 log10 at 24 and 30°C, respectively, 18 hours after inoculation . Observation by confocal microscopy of SeT-GFP incubated on the leaves of cilantro plants revealed that SeT cells localized to the vein area of leaves. Small microcolonies of cells were observed on the leaf veins, but larger concentrations of both individual and aggregated cells were observed on senescent portions of the leaf and in lesions, suggesting that the release of nutrients from leaky or damaged plant cells enhanced growth of the pathogen (Figure 2.2L) . Co-inoculation of cilantro with SeT-GFP and P. agglomerans, a plant epiphyte isolated from cilantro and containing a red fluorescent protein, revealed that SeT cells (Figure 2.3B, green cells) were attached to the leaf in aggregates with P. agglomerans (red/pink cells) and other natural epiphytic bacteria (purple cells) . These results suggest that SeT interacted with the plant and native bacteria after prolonged exposure to the leaf surface.
In an attempt to assess the mechanism of attachment of SeT to cilantro leaves, a chloroform-methanol (2:1) extract of cilantro leaf surfaces was prepared and fractions obtained by separation by thin-layer chromatography (TLC). Multiple TLC-purified bands were applied to glass slides, exposed to a suspension of SeT-GFP cells and the slides were incubated. Unbound cells were washed from the slide and the slide was observed by confocal microscopy. Figure 2.3F is representative of the results observed with one of the samples that bound SeT cells most effectively. This sample was analyzed by mass spectrometry and shown to be composed of >90% stigmasterol, a sterol compound that is present in the cuticle and has been detected in other leaf extracts . Interaction with cuticular waxes or sterols in regions of the leaf where nutrients are more available is a reasonable strategy for bacteria. A recent study reported the growth of epiphytic Pseudomonas spp. on apple cuticle membranes without disrupting the membrane, and the release of a variety of bacterial proteins (e.g., flagellin, porin, ABC transporter binding component) through the membrane .
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