LOCALIZATION OF HIGH-MOLECULAR-WEIGHT ADHESION PROTEINS OF NONTYPABLEHAEMOPHILUS-INFLUENZAE BY IMMUNOELECTRON MICROSCOPY

A family of high-molecular-weight (HMW) surface-exposed proteins impor tant in the attachment of nontypeable Haemophilus influenzae (NTHi) to human epithelial cells was previously identified (J. W. St. Geme III, S. Falkow, and S. J. Barenkamp, Proc. Natl. Acad. Sci. USA 90:2875-28 79, 1993). In the present investigation, indirect immunogold labeling and electron microscopy were used to localize these proteins on three clinical isolates of NTHi,, mutants deficient in expression of one or both HMW proteins, and embedded sections of human oropharyngeal cells after incubation with NTHi strain 12. The filamentous material compris ing the proteins was labeled with monoclonal antibodies directed again st two prototype HMW proteins (HMW1 and HMW2) of prototype NTHi strain 12. Gold labeling was observed as a cap or discrete aggregate off one pole or centrally along one long axis of the bacterial cell. Heavily labeled, non-bacterial-cell-associated, disk-like aggregates of the HM W proteins were frequently noted in both bacterial preparations as wel l as in association with the oropharyngeal cell surface and intracellu larly. Mutants demonstrated diminished labeling or an absence thereof, respectively; which correlated well with their previously demonstrate d reduced ability or inability to adhere to Chang conjunctival epithel ial cells in vitro. The Haemophilus HMW proteins share antigenic deter minants with and demonstrate amino acid sequence similarity to the fil amentous hemagglutinin protein of Bordetella pertussis, a critical adh esin of that organism. The studies presented here demonstrate that the Haemophilus proteins and B. pertussis filamentous hemagglutinin show impressive morphologic and perhaps additional functional similarity.