IDENTIFICATION AND CHARACTERIZATION OF A GENE-CLUSTER MEDIATING ENTEROAGGREGATIVE ESCHERICHIA-COLI AGGREGATIVE ADHERENCE FIMBRIA-I BIOGENESIS

The aggregative pattern of adherence (AA) exhibited by enteroaggregati ve Escherichia coli upon HEp-2 cells is a plasmid-associated property which correlates with aggregative adherence fimbria I (AAF/I) expressi on and human erythrocyte hemagglutination. By using cloning and mutage nesis strategies, two noncontiguous plasmid segments (designated regio ns 1 and 2) required for AA expression have previously been identified in enteroaggregative E. coli 17-2. TnphoA mutagenesis was performed o n clones containing region 1, and 16. TnphoA mutants which were negati ve for the AA phenotype were analyzed. The TnphoA insertion site for e ach mutant was determined by junctional DNA sequencing. All 16 mutatio ns occurred within a 4.6-kb span in region 1. Nucleotide sequence anal ysis of the region revealed four contiguous open reading frames, desig nated aggDCBA, in the same span. AA-negative TnphoA insertions into al l open reading frames except aggB were obtained. On the basis of mutat ional analysis and protein homology data, it is inferred that aggA, ag gC, and aggD are involved in biogenesis of AAF/I, encoding a major fim brial subunit, outer membrane usher, and periplasmic fimbrial chaperon e, respectively. By immunogold electron microscopy, polyclonal antiser um raised against the aggA gene product decorated AAF/I fimbriae, affi rming that AggA encodes an AAF/I subunit.