GENERATION OF ESCHERICHIA-COLI INTIMIN DERIVATIVES WITH DIFFERING BIOLOGICAL-ACTIVITIES USING SITE-DIRECTED MUTAGENESIS OF THE INTIMIN C-TERMINUS DOMAIN

Intimins, encoded by eae genes, are outer membrane proteins involved i n attaching-effacing (A/E) lesion formation and host cell invasion by pathogenic bacteria, including enteropathogenic Escherichia coli (EPEC ) and Citrobacter rodentium, A series of intimins, harbouring specific mutations close to the C-terminus, were constructed using pCVD438, wh ich encodes the eae gene from EPEC strain E2348/69. These mutant plasm ids were introduced into EPEC strain CVD206 and C, rodentium strain DB S255, which both contain deletion mutations in their eae genes. CVD206 , CVD206(pCVD438) and CVD206(pCVD438) derivatives were assessed for th eir ability to promote A/E lesion formation or invasion of HEp-2 cells and io induce A/E lesions on fresh human intestinal In vitro organ cu ltures (IVOC), The pathogenicity of C, rodentium DBS255 harbouring the se plasmid derivatives was also studied in mice. Here, we report that intimin-mediated A/E lesion formation can be segregated from intimin-m ediated HEp-2 cell invasion, Moreover, adherence to IVOC, EPEC-induced microvillus elongation and colonization of the murine intestine by C. rodentium were also modulated by the modified intimins.