Investigator: Matthew Moreau, Salve Regina University
Theme: Environmental Health Sciences
Title: Analyzing the mechanisms of unique adherence phenotypes of super shedder E. coli 
Award: SURF PUI Training Award (2022-2024)

Abstract: Super shedding (SS) of Shiga-toxin producing Escherichia coli (STEC) from its cattle reservoir is responsible for producing most of the environmental burden of this bacterium. More importantly, this phenomenon has been previously described in the most pathogenic variant of STEC, enterohemorrhagic E. coli (EHEC). Many of these isolates have been previously linked to many of the outbreaks associated with this bacterium that causes bloody diarrhea and in a smaller population, life-threatening hemolytic uremic syndrome. This phenomenon has been previously linked to a unique strong and aggregative adherence phenotype on bovine recto-anal junction squamous epithelia (RSE) cells. Though the deletion of several non-fimbrial adhesins did not result in the alteration or ablation of this phenotype in an SS isolate, some of these same deletions resulted in the ability of a non-SS isolate to have this same adherent phenotype. Thus, the goal of this study is to determine the transcriptional changes due to these deletions in the non-SS isolate and compare it to the global transcriptome of the SS isolate when adhered to these RSE cells. To do this, prokaryotic and eukaryotic RNA samples will be separated after infection of media control or RSE cells with either the SS isolate, the non-SS isolate, and/or the non-SS isolate mutants that confer the strong, aggregative adherence phenotype. RNA-Seq libraries will be constructed and sequenced followed by comparison of the fold change of RNA expression by RPKM comparisons. The specific aims of this present proposal are; 1) Examine the fold-change of genes under differential regulation during different adherence phenotypes from both the prokaryotic and eukaryotic perspective and any potential underlying regulatory and/or partnership changes that may be responsible for the change of cell-to-cell interface
between human/cattle cells and SS isolates. 2) Use locus-specific knockout technologies to perform molecular Koch’s postulates on genes and pathways identified in 1) and to test their importance/role in the SS-adherence phenotype and possibly to the SS phenotype as a whole.

Relevance: The majority of E. coli outbreaks that cause bloody diarrhea and other gastrointestinal disease states can be traced back to super shedder isolates. Our work will identify the factors important for the ability for E. coli to colonize and subsequently shed from cattle reservoirs. In doing so, we will be able to develop interventive strategies in hopes to prevent future outbreaks of E. coli and thusly future E. coli infections.