Use of gaseous ozone to prevent post-harvest microbial spoilage of leafy produce

Abstract

Fresh leafy salads suffer from post-harvest microbial contamination and decay. Due to increasing pesticide resistance and consumer pressures, residue free alternatives, such as ozone, are being actively explored and encouraged to reduce microbial loads of crops in storage/transit. Previous work has demonstrated that long-term exposure to low concentrations of ozone can be effective in retarding the degradation of some fruit and vegetable. Much less is known about the potential of ozone-exposure to protect leafy produce. The first goal of this project was to determine ozone exposure levels that did not damage produce, but reduced microbial loads significantly. Different produce types exhibited varying abilities to resist ozone damage, e.g. coriander and rocket were relatively resistant to ozone (10 ppm for 10 min); while spinach, watercress and lettuce were more sensitive (1 ppm for 10 min). These ozone exposure levels reduced bacterial loads by at least 1-log. Confocal microscopy confirmed that some bacterial cells (1−10%) survived ozone treatment. These visual observations demonstrated heterogeneity in the resistance of the leaf surface microflora to ozone treatment. It was tested if colony age and/or stress (e.g. cold) may be responsible for the variation in ozone resistance observed. Stressed cells of Pseudomonas sp. isolated from coriander exhibited greater resistance to ozone than control cells. Subsequent gene expression analysis using RNA-Seq technology of stressed cells showed significant changes in the expression of genes related to stress resistance compared to controls. In particular, it was observed that in aged colonies, about 90% of the changes in gene expression mapped to one gene, a non-coding RNA that is part of RNase P. Many of the genes showing differential expression were involved in energy production and transport, motility or cell wall/membrane integrity. This improved mechanistic understanding of ozone resistance may lead to novel anti-microbial treatments. As there are growing concerns about the contamination of leafy products with pathogens resulting in food poisoning the final part of this work focused on the potential of ozone to inactivate food pathogens on leafy produce. Results showed that this treatment significantly reduced E. coli and Listeria spp. on spinach, and the pathogens did not re-grow after treatment over a 9-day storage period. v It was concluded that gaseous ozone treatment is worthy of further exploration as a potential commercial tool to improve the safety of fresh leafy salads and herbs, and reduce microbial spoilage.