INCREASED LEVELS OF INTRACELLULAR CALCIUM ARE NOT REQUIRED FOR THE FORMATION OF ATTACHING AND EFFACING LESIONS BY ENTEROPATHOGENIC AND ENTEROHEMORRHAGIC ESCHERICHIA-COLI

Elevated concentrations of intracellular calcium ([Ca](i)) have been i mplicated as an important signalling event during attaching and effaci ng (A/E) lesion formation by enteropathogenic Escherichia coli (EPEC), The highly localized nature of the cytoskeletal and cell surface alte rations occurring during A/E lesion formation suggests that there shou ld be equally localized EPEC-induced signalling events. To analyze fur ther the calcium responses to infection of HEp-2 cells by EPEC, we emp loyed calcium-imaging fluorescence microscopy, which allows both tempo ral and spatial measurements of [Ca](i) in live cells. Using this imag ing technique, not only were we unable to detect any significant eleva tion in [Ca](i) at sites of A/E EPEC adhesion, but, with several diffe rent classical EPEC and enterohemorrhagic E. coli (EHEC) strains and t hree different infection procedures, each of which resulted in extensi ve A/E bacterial adhesion, we were unable to detect any significant al terations in [Ca](i) in infected cells compared to uninfected cells, I n addition, chelation of intracellular free calcium with bis(aminophen oxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA) did not, as previously reported, prevent A/E lesion formation. We conclude that increased [Ca ](i) are not required for A/E lesion formation by EPEC and EHEC.