Acetylation of alginate enables the production of inks that mimic the chemical properties of P. aeruginosa biofilm

The reason why certain bacteria, e.g., Pseudomonas aeruginosa (PA), produce acetylated alginate (Alg) in their biofilms remains one of the most intriguing facts in microbiology. Being the main structural component of the secreted biofilm, like the one formed in the lungs of cystic fibrosis (CF) patients, Alg plays a crucial role in protecting the bacteria from environmental stress and potential threats. Nonetheless, to investigate the PA biofilm environment and its lack of susceptibility to antibiotic treatment, the currently developed in vitro biofilm models use native seaweed Alg, which is a non-acetylated Alg. The role of the acetyl side group on the backbone of bacterial Alg has never been elucidated, and the transposition of experimental results obtained from such systems to clinical conditions (e.g., to treat CF-infection) may be hazardous. We systematically investigated the influence of acetylation on the physico-chemical and mechanical properties of Alg in solution and Ca2+-crosslinked hydrogels. Furthermore, we assessed how the acetylation influenced the interaction of Alg with tobramycin, a common aminoglycoside antibiotic for PA. Our study revealed that the degree of acetylation directly impacts the viscosity and Young's Modulus of Alg in a pH-dependent manner. Acetylation increased the mesh size in biofilm-like Alg hydrogels, directly influencing antibiotic penetration. Our results provide essential insights to create more clinically relevant in vitro infection models to test the efficacy of new drugs or to better understand the 3D microenvironment of PA biofilms.