Mucus polymer concentration andin vivoadaptation converge to define the antibiotic response ofPseudomonas aeruginosaduring chronic lung infection

Abstract

AbstractThe airway milieu of individuals with muco-obstructive airway diseases (MADs) is defined by the accumulation of dehydrated mucus due to hyperabsorption of airway surface liquid and defective mucociliary clearance. Pathological mucus becomes progressively more viscous with age and disease severity due to the concentration and overproduction of mucin and accumulation of host-derived extracellular DNA (eDNA). Respiratory mucus of MADs provides a niche for recurrent and persistent colonization by respiratory pathogens, includingPseudomonas aeruginosa, which is responsible for the majority of morbidity and mortality in MADs. Despite high concentration inhaled antibiotic therapies and the absence of antibiotic resistance, antipseudomonal treatment failure in MADs remains a significant clinical challenge. Understanding the drivers of antibiotic recalcitrance is essential for developing more effective treatments that eradicate persistent infections. The complex and dynamic environment of diseased airways makes it difficult to model antibiotic efficacyin vitro. We aimed to understand how mucin and eDNA concentrations, the two dominant polymers in respiratory mucus, alter the antibiotic tolerance ofP. aeruginosa. Our results demonstrate that polymer concentration and molecular weight affectP. aeruginosasurvival post antibiotic challenge. Polymer-driven antibiotic tolerance was not explicitly associated with reduced antibiotic diffusion. Lastly, we established a robust and standardizedin vitromodel for recapitulating theex vivoantibiotic tolerance ofP. aeruginosaobserved in expectorated sputum across age, underlying MAD etiology, and disease severity, which revealed the inherent variability in intrinsic antibiotic tolerance of host-evolvedP. aeruginosapopulations.ImportanceAntibiotic treatment failure inPseudomonas aeruginosachronic lung infections is associated with increased morbidity and mortality, illustrating the clinical challenge of bacterial infection control. Understanding the underlying infection environment, as well as the host and bacterial factors driving antibiotic tolerance and the ability to accurately recapitulate these factorsin vitro, is crucial for improving antibiotic treatment outcomes. Here, we demonstrate that increasing concentration and molecular weight of mucin and host eDNA drive increased antibiotic tolerance to tobramycin. Through systematic testing and modeling, we identified a biologically relevantin vitrocondition that recapitulates antibiotic tolerance observed inex vivotreated sputum. Ultimately, this study revealed a dominant effect ofin vivoevolved bacterial populations in defining inter-subjectex vivoantibiotic tolerance and establishes a robust and translatablein vitromodel for therapeutic development.