Contributions of EspA Filaments and Curli Fimbriae in Cellular Adherence and Biofilm Formation of Enterohemorrhagic Escherichia coli O157:H7.

Abstract

In Escherichia coli O157:H7 (O157), the filamentous structure of the type III secretion system is produced from the polymerization of the EspA protein. EspA filaments are essential for O157 adherence to epithelial cells. In previous studies, we demonstrated that O157 hha deletion mutants showed increased adherence to HEp-2 cells and produced abundant biofilms. Transcriptional analysis revealed increased expression of espA as well as the csgA gene, which encodes curli fimbriae that are essential for biofilm formation. In the present study, we constructed hha espA, hha csgA, and hha csgA espA deletion mutants to determine the relative importance of EspA and CsgA in O157 adherence to HEp-2 cells and biofilm formation. In vitro adherence assays, conducted at 37°C in a tissue culture medium containing 0.1% glucose, showed that HEp-2 cell adherence required EspA because hha espA and hha csgA espA mutants adhered to HEp-2 cells at higher levels only when complemented with an espA-expressing plasmid. Biofilm assays performed at 28°C in a medium lacking glucose showed dependency of biofilm formation on CsgA; however EspA was not produced under these conditions. Despite production of detectable levels of EspA at 37°C in media supplemented with 0.1% glucose, the biofilm formation occurred independent of EspA. These results indicate dependency of O157 adherence to epithelial cells on EspA filaments, while CsgA promoted biofilm formation under conditions mimicking those found in the environment (low temperature with nutrient limitations) and in the digestive tract of an host animal (higher temperature and low levels of glucose).