Attachment of Escherichia coli O157 : H7 to lettuce leaf surface and bacterial viability in response to chlorine treatment as demonstrated by using confocal scanning laser microscopy

Confocal scanning laser microscopy was used to observe the location of Esch erichia coli O157:H7 on and within lettuce leaves. Sections of leaves (ca. 0.5 by 0.5 cm) were inoculated by submersion in a suspension of E. coli O15 7:H7 (ca. 10(7) to 10(8) CFU/ml) overnight at 7 degrees C. Fluorescein isot hiocyanate-labeled antibody was used to visualize the attached bacteria. E. coli O157:H7 was found attached to the surface, trichomes, stomata, and cu t edges. Three-dimensional volume reconstruction of interior portions of le aves showed that E, coli O157:H7 was entrapped 20 to 100 mu m below the sur face in stomata and cut edges. Agar plate culturing and microscopic observa tion indicated that E. coli O157:H7 preferentially attached to cut edges, a s opposed to the intact leaf surface. Dual staining with fluorescein isothi ocyanate-labeled antibody and propidium iodide was used to determine viabil ity of cells on artificially contaminated lettuce leaves after treatment wi th 20 mg/liter chlorine solution for 5 min. Many live cells were found in s tomata and on cut edges following chlorine treatment. E. coli O157:H7 did n ot preferentially adhere to biofilm produced by Pseudomonas fluorescens on the leaf surface. In contrast to E. coli O157:H7, Pseudomonas adhered to an d grew mainly on the intact leaf surface rather than on the cut edges.