ROLE OF FIMBRIAE EXPRESSED BY NONTYPABLE HAEMOPHILUS-INFLUENZAE IN PATHOGENESIS OF AND PROTECTION AGAINST OTITIS-MEDIA AND RELATEDNESS OF THE FIMBRIN SUBUNIT TO OUTER-MEMBRANE PROTEIN-A

Notypeable Haemophilus influenzae is a primary pathogen in both acute otitis media (OM) and chronic OM, yet the pathogenesis of this disease is not fully understood. Although fimbriae have been observed on all clinical OM isolates examined to date, their role in pathogenesis rema ins unclear. Therefore, the gene which codes for the fimbrial subunit protein (fimbrin) in nontypeable H. influenzae 1128 was isolated, clon ed, and sequenced. The nucleotide sequence of the fimbrin gene was fou nd to contain an open reading frame of 1,077 bp which would encode a m ature fimbrin protein consisting of 338 amino acids with a calculated molecular mass of 36.4 kDa. The translated amino acid sequence was fou nd to be homologous with various OmpA proteins of other gram-negative bacteria, and algorithmic analysis predicted that this protein is orga nized as a coiled coil. To directly test whether fimbriae are involved in pathogenesis, the fimbrin gene was disrupted, and the biological c onsequences of disruption were absence of both expression of the fimbr ial appendage and the specific immunogold labeling thereof with antise ra directed against isolated fimbrial protein, reduced adherence to hu man oropharyngeal cells in vitro, augmented clearance from the tympanu m post-transbullar inoculation, and significantly reduced induction of OM post-intranasal inoculation in a chinchilla model compared with th e fimbriated parent strain. We additionally find that either passive i mmunization or active immunization against isolated fimbrial protein c onfers partial protection against transbullar challenge. A Western blo t (immunoblot) indicated a degree of serological relatedness among fim brin proteins of 15 nontypeable and type b isolates. These data sugges t that fimbrin could be useful as a component of a vaccine to protect against OM.