Fimbriae-mediated enhanced attachment of nontypable Haemophilus Influenzae (NTHI) to respiratory syncytial virus (RSV)-infected epithelial cells

NTHI infections occur soon after respiratory viral infections. Adherence of NTHI to normal respiratory epithelial cells is mediated in part by the bacterial surface fimbriae. In the present study, the role of NTHI fimbrae in attachment to RSV-infected A549, a human Type II-like alveolar epithelial cell line, was analyzed. Confluent monolayers of A549 cells were infected with RSV (multiplicity of infection = 0.1) or control media. After 24 h, the cells were detached with IM EDTA and mixed with suspensions of PKH (red fluorescence) dye-labeled NTHI or its funbriae-deficient isogenic mutant (NTHI-minusF) al ratio A549:NTHI = 1:100). The mixtures were incubated for In at 37°C on a shaker. Attachment of the PKH-labeled NTHI by flow cytomi try were: Ratio Mean (n = 4) A No. NTHI attached to RSV⁺ A549 1.81 No. NTHI attached to control A549 B No. NTHI-minusF attached to RSV⁺ A549 0.89 No NTHI-minusF attached to control A549 A + B 2.16 The results show that RSV increases the attachment of NTHI to A549 cells by 181% (P<0.05), and it is 216% higher when compared with NTHI-minusF mutant (P<0.05). Furthermore, by transmission microscopy it was shown that NTHI attachment to control A549 cells was rare, whereas it was significantly increased to RSV-infected A549 cells. The NTHI-minusF mutants rarely attached to the normal or RSV-infected A549 cells. In summary, RSV infection enhances the attachment of NTHI to respiratory epithelial cells which is primarily mediated by the bacterial fimbriae. Studies on RSV-induced expression of the fimbriae receptors on the epithelial cells are ongoing.