Background: Controlled human infection models are widely used to study infectious diseases. Since the 1960s, these studies have primarily relied on intranasal inoculation, as earlier aerosol-based methods raised safety concerns and were largely abandoned despite better mimicking the natural route of infection.

Methods: Utilizing modern advances in aerosol delivery, we conducted a dose-escalation study to evaluate the safety and feasibility of aerosol inoculation for human influenza challenge. Healthy adults aged 18-49 years were inoculated with influenza A/Perth/16/2009 (H3N2) using either a flow-focusing monodisperse aerosol generator (FMAG) delivering coarse particles or a medical nebulizer delivering fine particles. Participants were monitored in a controlled inpatient setting with symptom tracking, virologic sampling, and serology. Doses were escalated according to predefined safety and attack rate thresholds.

Results: Fourteen participants were enrolled. Flow-focusing monodisperse aerosol generator delivery at higher dose levels resulted in infection in 75% of participants (3/4), while nebulizer delivery produced infection in 50% (2/4). Illnesses were mild and self-limited. Viral shedding was detected at multiple respiratory sites, and adverse events were infrequent and generally mild. No serious adverse events occurred. Antibody responses were observed in a subset of infected participants.

Conclusions: In this small pilot, aerosolized challenge using modern delivery systems was feasible under controlled conditions; no safety concerns were identified, and MMID was induced in a subset of participants. These data establish a methodological framework for future studies evaluating pathogenesis and mucosal immune responses to a variety of respiratory pathogens.