SubPopulations of Shiga Toxin Producing Escherichia coli: Persistence, Resistance and Pathogenicity

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Experimental microbiology typically assumes bacterial populations are homogenous with respect to resistance to stress or gene expression. However, in reality pure strain cultures are heterogeneous with a proportion of cells existing in a dormant (persistent) state that have enhanced resistance compared to the main body of the population. Moreover, dormant cells are missed during culturing as they either fail or experience delayed growth. In evolutionary terms the persistent state is a kind of “bet hedging” whereby a proportion of cells will survive over extended periods should conditions for the population become adverse. To date, the majority of studies relating to persistence has been with respect to residual populations remaining after antibiotic treatment. Yet, persistence is commonly encountered in the wider environment such as soil, and food systems, such as foods treated with high pressure processing. The significance of residual populations with respect to food safety is unknown although the potential for the re-growth of pathogens or enhanced virulence are both possibilities. In the proposed research program we will look at the dormant (persistent, VBNC and tolerant) state in Shiga Toxin producing Escherichia coli (STEC) a class of significant clinical and foodborne pathogens. For the first time, the persistent state will be investigated in STEC and the triggers that both induce or break the dormant state identified. Of relevance will be the regulators of persistence derived from soil microbiota and rhizosphere that ultimately control pathogen:plant interactions. For the first time, it will be demonstrated how prophage influences the persistent state leading to mutual long term survival of phage and host. Studies of the persistent state will use state-of-the-art imaging techniques in combination with microfluidics that enable manipulation/ observation at the single cell level. Cells in the persistent state will be generated and harvested to facilitate studies on the role of dormancy in resistance to high pressure and lytic phages, in addition to environmental persistence. The impact of the research is potentially wide reaching. From an academic standpoint greater insights into prophage host interactions will be obtained. On an application side the research will provide data to assess the food safety risks represented by persister cells in the environment and food systems. To date, risk analysis has frequently designated a specified log reduction to verify the efficacy of interventions (for example, pasteurization) without taking into account residual survivors. Although present at low levels, residual survivors could grow during storage or become more virulent within the host. The research will provide data to assess such risks and moreover lead to more effective pathogen control strategies through breaking of the dormant state.