Salinity Stress Does Not Affect Root Uptake, Dissemination and Persistence of Salmonella in Sweet-basil (Ocimum basilicum)

Abstract

Crop produce can be contaminated in the field during cultivation by bacterial human pathogens originating from contaminated soil or irrigation water. The bacterial pathogens interact with the plant, can penetrate the plant via the root system and translocate and survive in above-ground tissues. The present study is first to investigate effects of an abiotic stress, salinity, on the interaction of plants with a bacterial human pathogen. The main sources of human bacterial contamination of plants are manures and marginal irrigation waters such as treated or un-treated wastewater. These are often saline and induce morphological, chemical and physiological changes in plants that might affect the interaction between the pathogens and the plant and thereby the potential for plant contamination. This research studied effects of salinity on the internalization of the bacterial human pathogen Salmonella enterica serovar Newport via the root system of sweet-basil plants, dissemination of the bacteria in the plant, and kinetics of survival in planta. Irrigation with 30 mM NaCl-salinity induced typical salt-stress effects on the plant: growth was reduced, Na and Cl concentrations increased, K and Ca concentrations reduced, osmotic potential and anti-oxidative activity were increased by 30%, stomatal conductance was reduced, and concentrations of essential-oils in the plants increased by 26%. Despite these physical, chemical and morphological changes in the plants, root internalization of the bacteria and its translocation to the shoot were not affected, and neither was the die-off rate of Salmonella in planta. The results demonstrate that the salinity-induced changes in the sweet-basil plants did not affect the interaction between Salmonella and the plant and thereby the potential for crop contamination.

Keywords: Salmonella; basil; contamination; human pathogens; internalization; persistence; root; salt stress.

FIGURE 1

Effect of irrigation with saline water on the concentrations of the salts Na and Cl (A), and the macronutrients K, Ca, and Mg (B), in leaves of plants cultivated under non-saline (Control) and saline irrigation (Salt). The data are means ± SE. Asterisks represent a significant difference between the concentration of a mineral element in control vs. salt plants (P < 0.05).

FIGURE 2

Effect of irrigation with saline water on osmotic potential and relative water content of leaves from plants cultivated under non-saline (control) and saline irrigation (salt). The data are means ± SE. Asterisks represent a significant difference between the control and salt plants (P < 0.05).

FIGURE 3

Effect of irrigation with saline water on the survival of Salmonella within the leaf. The presence of Salmonella in the leaves was determined at the end of the pathogen loading period (time 0), and 6, 22, and 30 h thereafter. ‘Control’ represents leaves from plants cultivated under irrigation with tap water; ‘salt’ represents leaves from plants irrigated with 30 mM NaCl salinity. The data are means (±SE). Means in a single sampling day, marked by same letters, are not significantly different according to Tukey test at α = 0.05. Asterisks represent that no bacteria (< 1 CFU/g^-1) were detected in the leaves even after an enrichment test.

FIGURE 4

Growth kinetics of Salmonella in plant sap expressed from shoots of sweet basil plants cultivated with non-saline (control) or saline irrigation (salt). Changes in Salmonella concentration (main figure), and rate of concentration change (insert) over time following spiking of the sap with 10³ CFU ml^-1 Salmonella. In the main figure, time 0 present results immediately following sap inoculation; and the lines represent a sigmoidal curve-fit to the data. The data are means ± SE. In the insert, the averaged rates and SE were calculated from the sigmoidal curve-fit data for the individual biological repeats. The difference in concentration and rate between the control and the salt treatment was not statistically different (P < 0.05) for any of the sampling times.

FIGURE 5

pH, electrical conductivity (EC), and osmotic potential of the plant sap used for the in vitro experiments described in Figure 4. The sap was expressed from shoots of sweet basil cultivated under non-saline (Control) and saline irrigation (Salt). The data are means ± SE. Asterisks represent a significant difference between the control and salt plants (P < 0.05).

FIGURE 6

Effect of irrigation with saline water on survival of Salmonella (A), and total aerobic bacteria (B), in the potting medium 24 h, and 5 days following inoculation. Data are averages ± SE. The soil was inoculated with 10⁷ CFU Salmonella g soil^-1. Asterisks represent significant difference of the measurement on the 1st vs. 5th day after inoculation (P < 0.05) at any salinity level.