The biofilm matrix scaffold of Pseudomonas species consists of non-canonically base paired extracellular DNA and RNA

While the array of emergent properties assigned to biofilms is extensive (e.g. antimicrobial tolerance), the mechanisms that underpin these are largely unknown. In particular, the extracellular matrix, a defining feature of biofilms, remains poorly understood in terms of its composition and contribution to biofilm structure and function. Here we demonstrate that extracellular DNA exists in a complex with RNA that forms the main cross-linking exopolymer of Pseudomonas biofilms, and explains biofilm elasticity. The RNA has a high purine content and our solid-state NMR data indicate the formation of Hoogsteen guanine base pairs. This may suggest the presence of G-quadruplexes, which is also corroborated by the enhancement of biofilm formation in the presence of potassium. The finding that non-canonical interactions mediate networking of matrix-forming extracellular nucleic acids addresses how eDNA is organized and contributes to matrix biophysical properties. This understanding will allow for the development of more effective biofilm control strategies.