Abstract
Nontypeable Haemophilus influenzae (NTHI) colonizes the human nasopharynx, but when the host immune response is dysregulated by upper respiratory tract (URT) virus infection, NTHI can gain access to more distal airway sites and cause disease. The NTHI type IV pilus (T4P) facilitates adherence, benign colonization, and infection, and its majority subunit PilA is in clinical trials as a vaccinogen. To further validate the strategy of immunization with PilA against multiple NTHI-induced diseases, it is important to demonstrate T4P expression under microenvironmental conditions that predispose to NTHI infection of the airway. Because URT infection commonly facilitates NTHI-induced diseases, we examined the influence of ongoing virus infection of respiratory tract epithelial cells on NTHI T4P expression in vitro Polarized primary human airway epithelial cells (HAEs) were sequentially inoculated with one of three common URT viruses, followed by NTHI. Use of a reporter construct revealed that NTHI upregulated pilA promoter activity when cultured with HAEs infected with adenovirus (AV), respiratory syncytial virus (RSV), or rhinovirus (RV) versus that in mock-infected HAEs. Consistent with these results, pilA expression and relative PilA/pilin abundance, as assessed by quantitative reverse transcription-PCR (qRT-PCR) and immunoblot, respectively, were also significantly increased when NTHI was cultured with virus-infected HAEs. Collectively, our data strongly suggest that under conditions of URT virus infection, PilA vaccinogen induction of T4P-directed antibodies is likely to be highly effective against multiple NTHI-induced diseases by interfering with T4P-mediated adherence. We hypothesize that this outcome could thereby limit or prevent the increased load of NTHI in the nasopharynx that characteristically precedes these coinfections.IMPORTANCE Nontypeable Haemophilus influenzae (NTHI) is the predominant bacterial causative agent of many chronic and recurrent diseases of the upper and lower respiratory tracts. NTHI-induced chronic rhinosinusitis, otitis media, and exacerbations of cystic fibrosis and chronic obstructive pulmonary disease often develop during or just after an upper respiratory tract viral infection. We have developed a vaccine candidate immunogen for NTHI-induced diseases that targets the majority subunit (PilA) of the type IV twitching pilus (T4P), which NTHI uses to adhere to respiratory tract epithelial cells and that also plays a role in disease. Here, we showed that NTHI cocultured with virus-infected respiratory tract epithelial cells express significantly more of the vaccine-targeted T4P than NTHI that encounters mock-infected (healthy) cells. These results strongly suggest that a vaccine strategy that targets the NTHI T4P will be effective under the most common predisposing condition: when the human host has a respiratory tract virus infection.
Highlights
Nontypeable Haemophilus influenzae (NTHI) colonizes the human nasopharynx, but when the host immune response is dysregulated by upper respiratory tract (URT) virus infection, NTHI can gain access to more distal airway sites and cause disease
Here, we examined T4P expression by NTHI cultured with well-differentiated primary human airway epithelial cells (HAEs) with ongoing infection due to adenovirus (AV), respiratory syncytial virus (RSV), or rhinovirus (RV), three of the most common URT viruses associated with NTHI coinfections [38, 47,48,49,50,51,52]
To first confirm that virus infection at the multiplicity of infection (MOI) used did not destroy the integrity of the epithelial barrier, we inoculated HAEs with AV, RSV, or RV as previously described [2, 55,56,57,58] and monitored transepithelial electrical resistance (TEER) as a measure of epithelial barrier integrity
Summary
Nontypeable Haemophilus influenzae (NTHI) colonizes the human nasopharynx, but when the host immune response is dysregulated by upper respiratory tract (URT) virus infection, NTHI can gain access to more distal airway sites and cause disease. Prior or concurrent upper respiratory tract virus infection dysregulates host airway epithelial defenses [1] and increases the expression of proteins that NTHI subsequently uses to adhere to and colonize the respiratory tract epithelium [2,3,4,5] In the nasopharynx, this increased colonization load allows NTHI to gain access to more distal sites of the airway, where it causes disease [6,7,8]. Here, we examined T4P expression by NTHI cultured with well-differentiated primary human airway epithelial cells (HAEs) with ongoing infection due to adenovirus (AV), respiratory syncytial virus (RSV), or rhinovirus (RV), three of the most common URT viruses associated with NTHI coinfections [38, 47,48,49,50,51,52]
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