title: Bacterial interactions with human respiratory mucosa in vitro
Creators: name: Tsang, Kenneth Wah Tak
Publisher: name: University of Glasgow
Year: 1995

description: The theme of this thesis is to study the interactions of non-typable Haemophilus influenzae (NTHi) and Pseudomonas aeruginosa (PA) with intact human respiratory mucosa in vitro. Recent evidence suggests that bacteria are mainly associated with respiratory mucus during exacerbation of chronic bronchitis but penetration of antibiotics into respiratory mucus is generally poor. A study was therefore performed to evaluate the effects of 0.25 and 0.5 minimal inhibitory concentrations of amoxycillin, loracarbef (a new carbacephem) and ciprofloxacin on NTHi infection of adenoid organ cultures in an agar-embedded model in which only the intact respiratory mucosa was exposed to bacteria-containing culture medium. The results from this study may help explain the clinical efficacy of antibiotics in treatment of bronchial infection despite poor antibiotic penetration into respiratory secretions. By using the same organ culture model the effects of NTHi infection of intact human bronchial mucosa was also studied. NTHi infection of bronchial organ cultures was associated with ultrastructural damage compared with uninfected organ cultures after 24h incubation. This damage was similar to the pattern observed in adenoid organ cultures described earlier. Similar experiments using nasal turbinate tissue showed virtually no adherence of NTHi to nasal respiratory mucosa suggesting that there may be a difference in epithelial surface receptors for NTHi between adenoid and nasal turbinate mucosa. Infection of adenoid organ cultures with an air-mucosal interface by PA caused significant ultrastructural damage (mitochondrial damage, loss of cilia, cytoplasmic blebbing and extrusion of cells from the epithelial surface) and slowing of ciliary beat when assessed by transmission electron and light microscopy respectively after 8h incubation. PA was found to cause disruption of epithelial tight junctions and adhere to basement membrane collagen. A matrix-like material was probably produced by PA which bridged PA with respiratory mucosa and might therefore be a PA adhesin. PA formed bacterial biofilms on the surface of respiratory mucosa that might have hindered its removal by the mucociliary clearance mechanism. These findings might help explain the difficulty in eradicating PA from the lower respiratory tract of patients with cystic fibrosis and bronchiectasis. The organ culture model with an air-mucosal interface was also used to study the effects of a bacterial toxin on intact human respiratory mucosa. An exotoxin of PA, pyocyanin was found to cause significant mucosal damage to adenoid organ cultures when assessed by transmission electron microscopy. Moreover, by using a newly developed transmission electron microscopy method to assess orientation of central microtubules of cilia and foot processes, pyocyanin was found to cause significant disorientation of the central microtubules of respiratory cilia but not the foot processes. PA pyocyanin may therefore have a role in the pathogenesis of PA infection in vivo. By using these organ culture models of intact human respiratory mucosa, bacteria interactions with human respiratory mucosa can be studied using the transmission, scanning electron, and light microscopy methods described in this thesis. Potential virulent factors for NTHi and PA can be tested and the mechanisms of bacterial pathogenesis can be studied further to advance current understanding of bacterial interactions with the human respiratory tract mucosa that may lead to the development of novel therapies. (Abstract shortened by ProQuest.)