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Despite regulatory and technological measures, edible sprouts are still often involved in foodborne illness and are considered a high-risk food. The present study explored the potential of spore-forming Bacillus isolates to mitigate Salmonella and Escherichia coli contamination of alfalfa sprouts. Food-derived Bacillus strains were screened for antagonistic activity against S. enterica serovar Typhimurium SL1344 (STm) and enteropathogenic E. coli O55:H7. Over 4 days of sprouting, levels of STm and E. coli on contaminated seeds increased from 2.0 log CFU/g to 8.0 and 3.9 log CFU/g, respectively. Treatment of the contaminated seeds with the most active Bacillus isolate, strain BX77, at 7 log CFU/g seeds resulted in substantial reductions in the levels of STm (5.8 CFU/g) and E. coli (3.9 log CFU/g) in the sprouted seeds, compared to the control. Similarly, co-culturing STm and BX77 in sterilized sprout extract at the same ratio resulted in growth inhibition and killed the Salmonella. Confocal-microscopy experiments using seeds supplemented with mCherry-tagged Salmonella revealed massive colonization of the seed coat and the root tip of 4-day-old sprouted seeds. In contrast, very few Salmonella cells were observed in sprouted seeds grown with BX77. Ca-hypochlorite disinfection of seeds contaminated with a relatively high concentration of Salmonella (5.0 log CFU/g) or treated with BX77 revealed a mild inhibitory effect. However, disinfection followed by the addition of BX77 had a synergistic effect, with a substantial reduction in Salmonella counts (7.8 log CFU/g) as compared to untreated seeds. These results suggest that a combination of chemical and biological treatments warrants further study, toward its potential application as a multi-hurdle strategy to mitigate Salmonella contamination of sprouted alfalfa seeds.

Hydroponics is a farming technique for growing plants with mineral nutrients using a soil-free medium. The plant roots are submerged in soil-free media, such as vermiculite or perlite, or just in mineral nutrient solutions. This allows for high production yields throughout the year with less water and agro-chemical inputs. Consequently, hydroponics is considered a sustainable agriculture technology. Hydroponically grown crops are usually protected from the diseases transmitted through soil or animals in open fields. Therefore, they require fewer chemicals for pest control and are safer than conventionally grown crops in terms of possible chemical contamination. Nevertheless, hydroponics guarantees neither plant health nor the microbial safety of fresh produce. In the case of microbial contamination by human pathogens, unlike soil-grown crops, the pathogens may rapidly spread through the circulating water and simultaneously infect all the plants in the facility. This review summarizes the up-to-date knowledge regarding the microbial safety of hydroponically grown crops and discusses the role of the hydroponic system in reducing the microbial hazards for leafy and fruity crops as well as the potential risks for contamination by human pathogens. Finally, it outlines the approaches and the available science-based practices to ensure produce safety. The contamination risk in hydroponic systems may be diminished by using novel planting materials and the appropriate decontamination treatment of a recirculating liquid substrate; by modulating the microbiota interactions; and by following strict phytosanitary measures and workers’ hygienic practices. There is a timely need to adopt measures, such as the Good Agricultural Practice (GAP) guidelines, to mitigate the risks and ensure safe hydroponically grown vegetables for consumers.

In a previous study, comparing the internalization of S. enterica serovar Typhimurium in various leaves by confocal microscopy, we have demonstrated that the pathogen failed to internalize tomato leaves. Numerous reasons may account for these findings, yet one such factor might be the methodology employed to quantify leaf internalization. To this end, we have systematically studied leaf localization of a Green-fluorescent protein-labeled Salmonella strain in tomato, lettuce, and Arabidopsis leaves by surface sterilization and enumeration of the surviving bacteria, side by side, with confocal microscopy observations. Leaf sterilization was performed using either sodium hypochlorite, silver nitrate, or ethanol for 1 to 7min. The level of internalization varied according to the type of disinfectant used for surface sterilization and the treatment time. Treatment of tomato leaves with 70% ethanol for up to 7min suggested possible internalization of Salmonella, while confocal microscopy showed no internalization. In the case of in lettuce and Arabidopsis leaves, both the plate-count technique and confocal microscopy demonstrated considerable Salmonella internalization thought different sterilization conditions resulted in variations in the internalization levels. Our findings highlighted the dependency of the internalization results on the specific disinfection protocol used to determine bacterial localization. The results underscore the importance of confocal microscopy in validating a particular surface sterilization protocol whenever a new pair of bacterial strain and plant cultivar is studied.