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Biofilms are viscoelastic materials that are a prominent public health problem and a cause of most chronic bacterial infections, in large part due to their resistance to clearance by the immune system. Viscoelastic materials combine both solid-like and fluid-like mechanics, and the viscoelastic properties of biofilms are an emergent property of the intercellular cohesion characterizing the biofilm state (planktonic bacteria do not have an equivalent property). However, how the mechanical properties of biofilms are related to the recalcitrant disease that they cause, specifically to their resistance to phagocytic clearance by the immune system, remains almost entirely unstudied. We believe this is an important gap that is ripe for a large range of investigations. Here we present an overview of what is known about biofilm infections and their interactions with the immune system, biofilm mechanics and their potential relationship with phagocytosis, and we give an illustrative example of one important biofilm-pathogen ( Pseudomonas aeruginosa ) which is the most-studied in this context. We hope to inspire investment and growth in this relatively-untapped field of research, which has the potential to reveal mechanical properties of biofilms as targets for therapeutics meant to enhance the efficacy of the immune system. 

Abstract Pseudomonas aeruginosais a pathogen that forms robust biofilms which are commonly associated with chronic infections and cannot be successfully cleared by the immune system. Neutrophils, the most common white blood cells, target infections with pathogen‐killing mechanisms that are rendered largely ineffective by the protective physicochemical structure of a biofilm. Visualisation of the complex interactions between immune cells and biofilms will advance understanding of how biofilms evade the immune system and could aid in developing treatment methods that promote immune clearance with minimal harm to the host. Scanning electron microscopy (SEM) distinguishes itself as a powerful, high‐resolution tool for obtaining strikingly clear and detailed topographical images. However, taking full advantage of SEM's potential for high‐resolution imaging requires that the fixation process simultaneously preserve both intricate biofilm architecture and the morphologies and structural signatures characterising neutrophils responses at an infection site. Standard aldehyde‐based fixation techniques result in significant loss of biofilm matrix material and morphologies of responding immune cells, thereby obscuring the details of immune interactions with the biofilm matrix. Here we show an improved fixation technique using the cationic dye alcian blue to preserve and visualise neutrophil interactions with the three‐dimensional architecture ofP. aeruginosabiofilms. We also demonstrate that this technique better preserves structures of biofilms grown from two other bacterial species,Klebsiella pneumoniaeandBurkholderia thailandensis.