The roles of eDNA and Alginate in biofilm mechanics and mechanical resistance to the immune system

Biofilm bacteria are embedded in a matrix of extracellular polymeric substances (EPS). Our goal is to understand the importance of different EPS types that Pseudomonas aeruginosa produce (specifically extracellular DNA (eDNA) and alginate) in the biofilm mechanics and how this relates to the success of neutrophils (phagocytic white blood cells) in engulfing biofilm bacteria. We apply neutrophils to biofilms treated with varying amounts of alginate lyase, which enzymatically breaks down alginate. We then use antibiotics to kill any bacteria in our samples that haven’t been engulfed by neutrophils. Then, we lyse neutrophils to release bacteria and count the number of bacteria remaining that were protected from antibiotics by being enclosed in neutrophils. We use the same process when assessing eDNA’s role, with DNase instead alginate lyase to break apart alginate structures. We also study the role of calcium ions at physiological concentrations; calcium ions can electrostatically crosslink alginate and eDNA, altering the mechanics of biofilms and their response to enzymatic treatments.