Iron content of commercial mucin contributes to compositional stability of a cystic fibrosis airway synthetic microbiota community

In vitro culture models of mucosal environments are used to elucidate the mechanistic roles of the microbiota in human health. These models often include commercial mucins to reflect the in-situ role of mucins as an attachment site and nutrient source for the microbiota. Two types of mucins are commercially available: porcine gastric mucin (PGM) and bovine submaxillary mucin (BSM). These commercial mucins have been shown to contain iron, an essential element required by the microbiota as a co-factor for a variety of metabolic functions. In these mucin preparations, the concentration of available iron can exceed physiological concentrations present in the native environment. This unexpected source of iron influences experimental outcomes, including shaping the interactions between co-existing microbes in synthetic microbial communities used to elucidate the multispecies interactions within native microbiota. In this work, we leveraged the well-characterized iron-dependent production of secondary metabolites by the opportunistic pathogen Pseudomonas aeruginosa to aid in the development of a simple, low-cost, reproducible workflow to remove iron from commercial mucins. Using the mucosal environment of the cystic fibrosis (CF) airway as a model system, we show that P. aeruginosa is canonically responsive to iron concentration in the chemically defined synthetic CF medium complemented with semi-purified PGM, and community composition of a clinically relevant, synthetic CF airway microbial community is modulated, in part, by iron concentration in PGM. IMPORTANCEMucins are critical components of in vitro systems used to model mucosal microbiota. However, crude commercial mucin preparations contain high concentrations of iron, which impacts interactions between members of the microbiota and influences interpretation of experimental results. Therefore, we developed and applied a simple, reproducible method to semi-purify commercial porcine gastric mucin as an affordable, low-iron mucin source. The development of this simplified workflow for semi-purification of commercial mucin enables researchers to remove confounding iron from a critical nutrient source when modeling clinically relevant microbial communities in vitro.