(330e) Elucidating a New Extracellular Function of Degp Inhibiting Biofilm Formation of Enterohemorrhagic Escherichia coli O157:H7

Enterohemorrhagic Escherichia coli O157:H7 (EHEC) is a pathogen causing serious outbreaks of hemorrhagic colitis. Conventional antibiotic treatment is not recommended for EHEC infection as antibiotics trigger Shiga-toxin production of EHEC and aggravate hemolytic-uremic syndrome. Biofilm formation of EHEC is closely associated with its virulence expression. Previously, we identified that probiotic Escherichia coli Nissle 1917 (EcN) secretes DegP and thereby inhibits EHEC biofilm formation. However, the extracellular function of DegP inhibiting EHEC biofilms is unclear. DegP is known as a serine protease exhibiting both proteolytic and chaperone functions and binds to outer membrane proteins (OMPs) of target cells. We hypothesized that DegP interacting with OMPs of EHEC may inhibit EHEC biofilm formation. We constructed EHEC mutants lacking ompA, ompC, or ompF individually or in combination and assessed their biofilm formation in the presence of DegP-secreting EcN in the co-culture or by adding purified DegP. When the omp were deleted in EHEC strains, their biofilm inhibition by DegP-secreting EcN was lessened, indicating that DegP may interact with OMPs to repress EHEC biofilm. Similar results were obtained when purified DegP was added into EHEC culture to form single biofilms. Next, we examined whether the EHEC biofilm inhibition is caused by protease activity or chaperone activity of DegP. A DegP variant (DegP S210A) with protease function disabled still inhibited EHEC biofilms similarly to the wild-type DegP activity, implying that protease-activity of DegP is not required to repress EHEC biofilms. In addition, biofilm related gene expressions (e.g., curli, fimbriae) of EHEC cells were affected upon DegP interacting with OMPs on EHEC cells. This study elucidates a new extracellular function of DegP in controlling EHEC biofilms and provides us with an insight to develop an alternative strategy to control biofilm-related infections.