RSV & rhinovirus coinfection order dictates outcome via type III interferon-mediated interference in infant respiratory epithelium.

UNLABELLED: Coinfection with respiratory syncytial virus (RSV) and rhinovirus (RV) is the most frequently detected coinfection in infants, yet mechanisms governing viral interference and disease outcome remain poorly understood. Although infection order is a key determinant of coinfection outcome involving RSV, influenza, rhinovirus, and SARS-CoV-2, its role in RSV/RV coinfections has not been defined. Using infant-derived human nasal organoids at air-liquid interface, we demonstrate that coinfection order fundamentally determines the outcome of RSV/RV interactions. An initial RSV infection almost completely inhibits secondary RV coinfection, whereas primary RV infection only partially impairs a subsequent RSV coinfection. These distinct outcomes are accompanied by divergent temporal antiviral and pro-inflammatory responses. Mechanistically, we identify type III interferons as a sufficient mediator of RSV-induced restriction of RV, establishing a host-driven mechanism of viral interference. In contrast, partial impairment of RSV following RV infection occurs through an interferon-independent mechanism. Despite marked differences, RSV/RV coinfections do not exacerbate epithelial injury or innate immune responses beyond that induced by RSV alone, indicating RSV is the dominant driver of pathology in coinfections. Collectively, our data establishes coinfection order as a critical determinant of RSV/RV outcome and reveals novel and mechanistic insight into critical viral-host interactions in the infant nasal epithelium.

IMPORTANCE: This study uncovers fundamental differences in infant epithelial control of two major pediatric viral pathogens. We demonstrate for the first time that the order of coinfection fundamentally determines the outcome of RSV-RV interactions within infant respiratory epithelium. By identifying type III interferon as an important regulator of these interactions, our study suggests that differences in interferon susceptibility among respiratory viruses can heavily influence coinfection outcomes. Importantly, despite clear differences in viral interference, RSV/RV coinfections did not produce greater epithelial injury or pro-inflammatory responses than RSV infection alone. Instead, RSV consistently emerged as the dominant driver of epithelial damage and inflammation, providing a mechanistic explanation for clinical observations that RSV/RV coinfections are often not more severe than RSV infection alone.