Article Figures & Data
Figures
- FIG 1
Salad extracts induce Salmonella growth in water. (A) In vitro time course of growth at 37°C of Salmonella in water with increasing levels of spinach juice; values shown are means from triplicate time points; n = 3. (B) Images showing the fluid that accumulates in bagged salads, spinach, and mixed-leaf varieties. (C) Histograms showing the in vitro growth at 4°C of 101 CFU/ml Salmonella inoculated into sterile water supplemented with 5% to 20% (vol/vol) doses of fluids from a spinach and mixed-leaf salad bag; values shown are means from triplicate time points; n = 3. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001.
- FIG 2
Salad extracts increase Salmonella motility and biofilm formation. (A) Salmonella motility at room temperature in 2% (vol/vol) salad leaf juice-supplemented DMEM relative to unsupplemented controls (P ≤ 0.01). Key: C, control (no additions); Cl, cos lettuce; Bo, baby green oak lettuce; Rr, red romaine lettuce; S, spinach; Rc, red chard; n = 5. (B) Initial in vitro attachment of Salmonella in water in the presence of salad juice relative to unsupplemented controls (P ≤ 0.001). Initial inocula were 106 CFU/ml; values shown are means of 4 data points; n = 3. Key: C, control (no additions); Cl, cos lettuce; Bo, baby green oak lettuce; Rr, red romaine lettuce; S, spinach; Rc, red chard. The various leaf juices were supplemented as 2% (vol/vol) additions; n = 3. (C) Light microscopy image showing biofilm formation of Salmonella treated for 4 and 18 h with incubation at room temperature with 2% spinach juice–DMEM; n = 6. (D) Histogram showing that 2% spinach extract in water enhances Salmonella attachment to spinach leaves after a 30-min incubation at room temperature; values shown are means of 6 data points; n = 3. **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001.
- FIG 3
Spinach juice increases Salmonella colonization of salad leaves. (A) Representative scanning electron micrographs of untreated (−) and 2% (vol/vol) spinach juice-exposed (+) Salmonella seeded at 105 CFU/ml onto 5-by-5-mm sections of spinach leaves as described in Materials and Methods. (B) Attachment profiles of the leaf microbiota in the absence (−) and presence (+) of 2% (vol/vol) spinach juice; n = 3. Scale bars are shown in white at the bottom of each set of images. (C) In vitro growth responses of the salad leaf microbiota (inoculated at 102 CFU/ml) to spinach juice (2%) and spinach bag fluid (20%); P > 0.05; n = 8.
- FIG 4
Salad juice increases colonization of salad bags by Salmonella. (A and B) Typical pictures of a salad bag microbiota. Panel A shows microbes growing from the salad bag and panel B the diversity of microbes transferred from a bag imprint onto Luria agar. (C) Light microscopy images of Salmonella (seeded at 105 CFU/ml) and endogenous microbiota colonization of salad bag plastic in water in the absence (−) and presence (+) of 2% spinach juice; images were captured using a light microscope (bright field) set at ×60 magnification; n = 8. (D) Blue light microscopy images (×60 magnification) of 105 CFU/ml GFP-Salmonella incubated in water in the absence (Control) and presence of 2% spinach juice or 20% spinach bag fluid; n = 3.
- FIG 5
Salad bag fluids enhance Salmonella growth in host-like media. (A) Histograms showing the salad juice response growth responses of Salmonella in serum-SAPI medium. Bacteria were inoculated at 102 CFU/ml and incubated for 18 h at 37°C. Values shown are means of 6 data points; relative to controls, all data values have a P value of ≤0.001; n = 3. Key: C, control (no additions); Cl, cos lettuce; Bo, baby green oak lettuce; Rr, red romaine lettuce; S, spinach; Rc, red chard. (B) Urea gel showing that salad extracts (2% vol/vol) can remove iron from holo-transferrin (Holo); n = 5. Key: C, control (no additions); Cl, cos lettuce; Bo, baby green oak lettuce; Rr, red romaine lettuce; S, spinach; Rc, red chard. (C) Histograms showing the uptake of transferrin-complexed 55Fe by Salmonella in the absence and presence of 2% spinach juice (Juice) (P ≤ 0.0001) and 20% spinach bag fluid (Bag fluid) (P ≤ 0.001; n = 3). (D) Histograms comparing the serum-SAPI growth response profile at 37°C of a Salmonella enterobactin mutant with that of the wild type in the presence of salad juice and salad leaf dietary catechols. Values shown are means from 3 data points; n = 3. Key: C, control (no additions); S, 2% (vol/vol) spinach juice; Fe, 100 μM ferric nitrate; Ta, 100 μM tannic acid; Ca, 100 μM chlorogenic acid. **, P ≤ 0.01; ****, P ≤ 0.0001.
