Evaluating Bacterial Pathogenesis Using a Model of Human Airway Organoids Infected with Pseudomonas aeruginosa Biofilms.

Pseudomonas aeruginosa is one of the leading invasive agents of human pulmonary infection, especially in patients with compromised immunity. Prior studies have used various models to establish P. aeruginosa infection and to analyze transcriptomic profiles of either the host or pathogen, and yet how much those works are relevant to the genuine human airway still raises doubts. In this study, we cultured and differentiated human airway organoids (HAOs) that recapitulate, to a large extent, the histological and physiological features of the native human mucociliary epithelium. HAOs were then employed as a host model to monitor P. aeruginosa biofilm development. Through dual-species transcriptome sequencing (RNA-seq) analyses, we found that quorum sensing (QS) and several associated protein secretion systems were significantly upregulated in HAO-associated bacteria. Cocultures of HAOs and QS-defective mutants further validated the role of QS in the maintenance of a robust biofilm and disruption of host tissue. Simultaneously, the expression magnitude of multiple inflammation-associated signaling pathways was higher in the QS mutant-infected HAOs, suggesting that QS promotes immune evasion at the transcriptional level. Altogether, modeling infection of HAOs by P. aeruginosa captured several crucial facets in host responses and bacterial pathogenesis, with QS being the most dominant virulence pathway showing profound effects on both bacterial biofilm and host immune responses. Our results revealed that HAOs are an optimal model for studying the interaction between the airway epithelium and bacterial pathogens. Human airway organoids (HAOs) are an organotypic model of human airway mucociliary epithelium. The HAOs can closely resemble their origin organ in terms of epithelium architecture and physiological function. Accumulating studies have revealed the great values of the HAO cultures in host-pathogen interaction research. In this study, HAOs were used as a host model to grow Pseudomonas aeruginosa biofilm, which is one of the most common pathogens found in pulmonary infection cases. Dual transcriptome sequencing (RNA-seq) analyses showed that the cocultures have changed the gene expression pattern of both sides significantly and simultaneously. Bacterial quorum sensing (QS), the most upregulated pathway, contributed greatly to biofilm formation, disruption of barrier function, and subversion of host immune responses. Our study therefore provides a global insight into the transcriptomic responses of both P. aeruginosa and human airway epithelium.