Ingress of Salmonella enterica Typhimurium into tomato leaves through hydathodes

Abstract

Internal contamination of Salmonella in plants is attracting increasing attention for food safety reasons. In this study, three different tomato cultivars "Florida Lanai", "Crown Jewel", "Ailsa Craig" and the transgenic line Sp5 of "Ailsa Craig" were inoculated with 1 µl GFP-labeled Salmonella Typhimurium through guttation droplets at concentrations of 10(9) or 10(7) CFU/ml. Survival of Salmonella on/in tomato leaves was detected by both direct plating and enrichment methods. Salmonella cells survived best on/in the inoculated leaves of cultivar "Ailsa Craig" and decreased fastest on/in "Florida Lanai" leaves. Increased guttation in the abscisic acid over-expressing Sp5 plants may have facilitated the entrance of Salmonella into leaves and the colonization on the surface of tomato leaves. Internalization of Salmonella Typhimurium in tomato leaves through guttation drop inoculation was confirmed by confocal laser microscopy. For the first time, convincing evidence is presented that S. enterica can enter tomato leaves through hydathodes and move into the vascular system, which may result in the internal translocation of the bacteria inside plants.

Figure 1. Survival of Salmonella Typhimurium on/in tomato leaves after inoculation (10⁹ CFU/ml) into guttation droplets.

Populations of Salmonella (log(CFU/g)) in inoculated tomato leaves of tomato cultivar: Florida Lanai (A), Crown Jewel (B), Ailsa Craig (C) and the transgenic line (Sp5) of Ailsa Craig (D).

Figure 2. Survival of Salmonella Typhimurium on/in tomato leaves after inoculation (10⁷ CFU/ml) into guttation droplets.

Populations of Salmonella (log(CFU/g)) in inoculated tomato leaves of tomato cultivar: Florida Lanai (A), Crown Jewel (B), Ailsa Craig (C) and the transgenic line (Sp5) of Ailsa Craig (D).

Figure 3. Confocal laser microscope images of tomato leaf tissue sections colonized by Salmonella Typhimurium after inoculation (10⁹ CFU/ml) through guttation droplets.

White arrows point out the GFP-tagged Salmonella cells (green) which entered into the vascular system of tomato leaves. Red fluorescence is the autofluorescence of plant chloroplasts. Images A2, B2, C2 and D2 are merged images under GFP and TRITC filters obtained by projecting 20 Z section overlaid fluorescence images of different layers with 1 µm interval into one combined image .