title: Recombinant anti-EspA antibodies block Escherichia coli O157:H7-induced attaching and effacing lesions in vitro.
abstract: Intimin and EspA proteins are virulence factors expressed by attaching and effacing Escherichia coli (AEEC) such as enteropathogenic and enterohaemorrhagic E. coli. The EspA protein makes up a filament structure forming part of the type III secretion system (TTSS) that delivers effector proteins to the host epithelial cell. Bacterial surface displayed intimin interacts with translocated intimin receptor in the host cell membrane leading to intimate attachment of the bacterium and subsequent attaching and effacing lesions. Here, we have assessed the use of recombinant monoclonal antibodies against E. coli O157:H7 EspA and intimin for the disruption of AEEC interaction with the host cell. Anti-gamma intimin antibodies did not reduce either adhesion of E. coli O157:H7 to host cell mono-layers or subsequent host cell actin rearrangement. Anti-EspA antibodies similarly had no effect on bacterial adhesion however they had a marked effect upon E. coli O157:H7-induced host cell actin rearrangement, where both monoclonal and polyclonal antibodies completely blocked cytoskeletal changes within the host cell. Furthermore, these anti-EspA antibodies were shown to reduce actin rearrangement induced by some but not all other AEEC serotypes tested. Both polyclonal and monoclonal antibodies could be used to label E. coli O157 EspA filaments and these immunoreagents did not inhibit the formation of such filaments. This is the first report of monoclonal antibodies to EspA capable of disrupting the TTSS function of E. coli O157:H7.
authors.affiliation: Veterinary Laboratories Agency, Department of Food and Environmental Safety, Woodham Lane, New Haw, Addlestone, Surrey KT15 3NB, UK.
authors.forename: Roberto M
authors.initials: RM
authors.lastname: La Ragione
authors.forename: Shila
authors.initials: S
authors.lastname: Patel
authors.forename: Ben
authors.initials: B
authors.lastname: Maddison
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Woodward
authors.forename: Angus
authors.initials: A
authors.lastname: Best
authors.forename: Garry C
authors.initials: GC
authors.lastname: Whitelam
authors.forename: Kevin C
authors.initials: KC
authors.lastname: Gough
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology*; D001422:Bacterial Adhesion / Q000276:immunology*; D002460:Cell Line; D018604:Epitope Mapping; D019453:Escherichia coli O157 / Q000276:immunology / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000276:immunology* / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology; D010590:Phalloidine; D011817:Rabbits; D011994:Recombinant Proteins / Q000276:immunology*; D013194:Staining and Labeling / Q000379:methods
chemical_list: D000199:Actins; D000907:Antibodies, Bacterial; D029968:Escherichia coli Proteins; C102051:EspA protein, E coli; D007128:Immunoglobulin Fragments; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D010590:Phalloidine; D016650:Fluorescein-5-isothiocyanate