Plants can act as hosts for a wide range of microbes with colonisation occurring on all tissue types to varying extents. There have been a number of foodborne outbreaks from consumption of contaminated fresh produce, including the E.coli O157:H7 PT8 outbreak in 2011 associated with soil-contaminated vegetables, which was one of the largest we have seen in the UK. This proposal aims to investigate the ability of E.coli O157:H7 to internalise into a variety of plant species, under commercially relevant conditions. Then the pathogenic potential of internalised bacteria and the subsequent risk posed to human health will be assessed. A comparative analysis of internalisation using microscopy to visualise internalised bacteria, and microbiology and molecular techniques to quantify internalised bacteria, will be performed. The overall aim of the project is to provide growers and producers with robust data, and to provide information during outbreak investigations.
The objectives of the study are to:
- determine the how different factors affect the internalisation phenotype for VTEC
- determine the internalisation capability of into plants
- assess the viability of internalised bacteria
- determine the pathogenic potential of internalised bacteria
There are several points at which internalisation can occur, from growth through to processing and packaging. In this study it is proposed that the potential of VTEC to internalise in leafy vegetables in conditions representative of pre-harvest growth will be determined. The research will assess the impact of different factors on the ability of VTEC to internalise, by carrying out a comparative analysis to a baseline model dataset. The second part of the research will investigate whether internalisation in leafy green vegetables alters the pathogenic potential of the bacteria for humans. The data generated will be of use to producers of leafy vegetables and help to inform in the event of a produce-associated outbreak.
A combination of approaches will be used, including microbiology, microscopy and cell culture. The leafy vegetables selected and reported by the FAO/WHO seem to present the greatest concern in terms of microbiological hazards. The plants used in this study are spinach, lettuce, alfalfa, sweet basil and tomato. The plants are grown under hydroponic conditions in sterile conditions before being transferred to pots containing agar or soil, depending on the plant. Light microscopy is used to visualise and localise the bacteria internalised in the plant tissue. To enumerate the internalised population and viable population of bacteria, quantitative PCR will be used. Bacterial gene expression will also be investigated as a measure of the pathogen’s adaptability to the internalised plant tissue environment.