Aggregative adherence fimbria I expression in enteroaggregative Escherichia coli requires two unlinked plasmid...

Adherence to HEp-2 cells by many enteroaggregative Escherichia coli (EAggEC) strains is associated with the expression of flexible, bundle-forming fimbriae 2 to 3 nm in diameter, designated aggregative adherence fimbriae I (AAF/I). We have previously reported the molecular cloning and TnphoA mutagenesis of AAF/I genes from the large plasmid of prototype EAggEC strain 17-2 (J. P. Nataro, Y. Deng, D. R. Maneval, A. L. German, W. C. Martin, and M. M. Levine, Infect. Immun. 60:2297-2304, 1992). Here, we report that further mapping and subcloning of AAF/I regions suggest that expression of the fimbriae requires two separate plasmid regions (designated regions 1 and 2). Approximately 9 kb of DNA unnecessary for fimbrial expression separates the two regions; this intervening segment encodes the EAggEC heat-stable enterotoxin (EAST1). Neither region was capable of conferring aggregative HEp-2 adherence (AA) when cloned individually; when the two regions were cloned as a single fragment or when each was cloned into a different vector and introduced into the same E. coli HB101 cell, AA was restored. AA-positive constructs also expressed human erythrocyte hemagglutination, autoagglutination in broth cultures, and the production of AAF/I as detected by immunogold electron microscopy.