COMPLEMENTATION ANALYSIS OF THE DICHELOBACTER-NADOSUS FIMN, FIMO, ANDFIMP GENES IN PSEUDOMONAS-AERUGINOSA AND TRANSCRIPTIONAL ANALYSIS OF THE FIMNOP GENE REGION

The causative agent of ovine footrot, the gram-negative anaerobe Diche lobacter nodosus, produces polar type IV fimbriae, which are the major protective antigens. The D. nodosus genes fimN, fimO, and fimP are ho mologs of the Pseudomonas aeruginosa fimbrial assembly genes, pilB, pi lC, and pilD, respectively. Both the pilD and fimP genes encode prepil in peptidases that are responsible for cleavage of the leader sequence from the immature fimbrial subunit. To investigate the functional sim ilarity of the fimbrial biogenesis systems from these organisms, the D . nodosus genes were introduced into P. aeruginosa strains carrying mu tations in the homologous genes, Analysis of the resultant derivatives showed that the fimP gene complemented a pilD mutant of P. aeruginosa for both fimbrial assembly and protein secretion. However, the fimN a nd fimO genes did not complement pilB or pilC mutants, respectively. T hese results suggest that although the PilD prepilin peptidase can be functionally replaced by the heterologous FimP protein, the function o f the PilB and PilC proteins may require binding or catalytic domains specific for the P. aeruginosa fimbrial assembly system, The transcrip tional organization and regulation of the fimNOP gene region were also examined, The results of reverse transcriptase PCR and primer extensi on analysis suggested that these genes form an operon transcribed from two sigma(70)-type promoters located upstream of ORFM, an open readin g frame proximal to fimN. Transcription of the D. nodosus fimbrial sub unit was found to increase in cells grown on solid media, and it was p ostulated that this regulatory effect may be of significance in the in fected footrot lesion.