Heterogeneity Governs 3D-Cultures of Clinically Relevant Microbial Communities

The intrinsic heterogeneity of bacterial niches should be retained in in vitrocultures to represent the complex microbial ecology. As a case study,mucin-containing hydrogels -CF-Mu3Gel - are generated by diffusion-inducedgelation, bioinspired on cystic fibrosis (CF) mucus, and a microbial nichechallenging current therapeutic strategies. At breathing frequency, CF-Mu3Gelexhibits aG′andG′′equal to 24 and 3.2 Pa, respectively. Notably, CF-Mu3Gelexhibits structural gradients with a gradual reduction of oxygen tensionacross its thickness (280–194μmol L−1). Over the culture period, a steepdecline in oxygen concentration occurs just a few millimeters below theair–mucus interface in CF-Mu3Gel, similar to those of CF airway mucus.Importantly, the distinctive features of CF-Mu3Gel significantly influencebacterial organization and antimicrobial tolerance in mono- and co-cultures ofStaphylococcus aureusandPseudomonas aeruginosathat standard culturesare unable to emulate. The antimicrobial susceptibility determined inCF-Mu3Gel corroborates the mismatch on the efficacy of antimicrobialtreatment between planktonically cultured bacteria and those in patients.With this example-based research, new light is shed on the understanding ofhow the substrate influences microbial behavior, paving the way for improvedfundamental microbiology studies and more effective drug testing anddevelopment

Cystic Fibrosis Mucus Model to Design More Efficient Drug Therapies

Mucus represents a strong barrier to tackle for oral or pulmonary administered drugs, especially in mucus-related disorders. This study uses a pathological cystic fibrosis (CF) mucus model to investigate how mucus impacts the passive diffusion of 45 ad hoc commercial drugs selected to maximize physicochemical variability. An in vitro mucosal surface was recreated by coupling the mucus model to a 96-well permeable support precoated with structured layers of phospholipids (parallel artificial membrane permeability assay, PAMPA). Results show that the mucus model was not a mere physical barrier but it behaves like an interactive filter. In nearly one-half of the investigated compounds, the diffusion was reduced by mucus, while other drugs were not sensitive to the mucus barriers. We also found that permeability can be enhanced when drug-calcium salts are formed. This was confirmed with cystic fibrosis sputum as a rough ex vivo model of CF mucus. Since the drug discovery process is characterized by a high rate of failure, the mucus platform is expected to provide an efficient support to early reduce the number of poor-performing drug candidates