GROWTH-CONDITIONS MEDIATE DIFFERENTIAL TRANSCRIPTION OF FIM GENES INVOLVED IN PHASE VARIATION OF TYPE-1 PILI

Type 1 pili in Escherichia coli undergo phase variation in which indiv idual cells in a population reversibly switch between piliated (Pil+) and nonpiliated (Pil-) states. The switching process is mediated by an invertible DNA fragment which contains the promoter for fimA, the gen e encoding the major structural subunit of type 1 pili. Although type 1 pili randomly phase vary in broth cultures, many clinical isolates o f E. coli do not express type 1 pili when cultured on agar media. We i nvestigated the role of the invertible element and the upstream genes, fimB and fimE, in the agar-mediated suppression of pili in an agar-ne gative clinical isolate, strain 149. Southern hybridization and polyme rase chain reaction analyses of the fimA promoter region in broth-grow n 149 cells indicated that the invertible element was present in orien tations corresponding to both Pil+ and Pil- phenotypes. In contrast, o nly one orientation of the invertible element, corresponding to the Pi l- phenotype, was observed in strain 149 cells cultured on agar. A sec ond clinical isolate, strain 2-7, which expresses type 1 pili on agar was also examined; the invertible element was found in both the Pil+ a nd Pil- orientations during growth of this strain on agar as well as i n broth. The introduction of the fim gene cluster from strain J96 on a multicopy plasmid into agar-negative strain 149 resulted in the produ ction of both J96 and 149 pili during growth on agar. Experiments with subclones of the J96 genes indicated that the presence of an intact f imB gene allowed strain 149 pili to be produced on agar. Northern (RNA ) hybridizations showed that transcripts of fimB and fimA, as well as an antisense transcript of fimE, were present in broth-grown cells of strain 149, but not in agar-grown cells. Only fimE transcripts were pr esent in agar-grown cells, but they appeared to be degraded and were n ot consistently observed. Transcripts of fimA, fimB, fimE, and antisen se fimE were detected in both broth- and agar-grown cells of phase-var iable strain 2-7. The results suggest that in agar-negative strain 149 , the fimA promoter element is phase locked in an off orientation on a gar. Differences in pilus production between agar and broth cultures a ppear to be the result of differential transcription of fimB and fimE under the two growth conditions. In contrast, the pattern of expressio n of these genes in agar phase-variable strain 2-7 did not differ betw een broth- and agar-grown cells.