Extracellular DNA (eDNA) enables early detection of the phenotypic switch of Pseudomonas sp. during biofilm development

The different steps involved in biofilm formation have been the subjects of intensive researches. However, the very early cell decision-making process related to the switch from planktonic to sessile state still remains uncharacterized. Based on the use of Pseudomonas putida KT2440 and derivatives with varying biofilm-forming capabilities, we observed a subpopulation of cells bound to extracellular DNA (eDNA) in the planktonic phase, as indicated by propidium iodide (PI) staining. Strikingly, the size of this eDNA-bound/PI-positive subpopulation correlated with the overall biofilm forming capability of the bacterial population. This finding challenges the conventional view of phenotypic switching and suggests that, in Pseudomonas, biofilm switching is determined collectively based on the quantity of eDNA released in the supernatant. The whole process can be followed based on automated flow cytometry, and the appearance of PI-positive cells was considered as an early-warning indicator for biofilm formation. For this purpose, automated glucose pulsing was used successfully to interfere with the proliferation of PI-positive cells, resulting in a reduction of biofilm formation. This study provides insights into the collective determinants of biofilm switching in Pseudomonas species and introduces a potential strategy for controlling biofilm formation.