H-052. Identification of a Novel Gene Z0021 Involved in Flagella-Mediated Motility in Escherichia coli O157:H7

U. Silphaduang1, B. K. Coombes2, A. M. Kropinski3, M. Mascarenhas3, M. A. Karmali3;
1Univ. of Guelph, Guelph, ON, CANADA, 2McMaster Univ., Hamilton, ON, CANADA, 3Lab. for Foodborne Zoonoses, Pub. Hlth. Agency of Canada, Guelph, ON, CANADA.

Introduction: Verotoxin-producing Escherichia coli (VTEC) are important pathogens that can cause severe human disease, including haemorrhagic colitis and haemolytic-uremic syndrome. We previously reported that O-island 1 (OI-1), a genomic island containing a fimbrial operon, is highly specific for VTEC serotype O157 strains. The present study was undertaken to elucidate the function and pathogenic significance of OI-1 in VTEC. Methods and Results: Lambda red recombination was used to generate an OI-1 deletion mutant (ΔOI-1). Deletion of OI-1 resulted in significantly increased swimming motility compared to the wild-type strain, Escherichia coli EDL933. Further mutagenesis revealed that the specific gene responsible for this phenotype was Z0021, an OI-1 gene encoding a protein of unknown function in E. coli O157:H7. Complementation experiments with overexpression of the OI-1 genes, Z0021, Z0022 (usher protein) and Z0024 (type-1 fimbrial protein), resulted in a predictable loss of swimming motility. Elevated levels of flagellin in ΔOI-1 and ΔZ0021 compared to EDL933 were demonstrated by western blot analysis and enhanced surface expression of flagella was demonstrated by transmission electron microscopy (TEM). Hence, the increased motility in ΔOI-1 and ΔZ0021 is associated with elevated levels of the flagella filament and suggests a molecular connection between flagella and fimbrial formation in E. coli serotype O157. Overexpression of Z0021 protein using a plasmid encoding Z0021 in other Enterobacteriaceae including E. coli K12 MG1655 and Salmonella enterica serovar Typhimurium led to a non-motile phenotype and loss of surface flagella. Conclusion: We have identified a new genetic locus in OI-1 that controls swimming motility in O157:H7 VTEC. We propose that E. coli O157 use these novel genes on the OI-1 genomic island to regulate flagellum formation and swimming motility through a novel mechanism.