Poster Z4 - Development of tomato xylem-mimicking microfluidic system to study Ralstonia pseudosolanacearum biofilm formation

Introduction: The bacterial wilt pathogen Ralstonia pseudosolanacearum (Rps) colonizes plant xylem vessels and blocks the flow of xylem sap by its biofilm, causing devastating wilt disease across various economical host plants including tomatoes. The technical challenges of imaging the xylem environment along with the use of artificial cell culture plates and media in existing in-vitro systems limit the understanding of Rps biofilm and its infection dynamics.

Materials and

Methods: In this study, we designed and built a microfluidic system that mimicked the physical and chemical conditions of the tomato xylem vessels, and allowed us to dissect Rps responses to different xylem-like conditions. The microfluidic system was designed with twenty parallel channels with a constant 50um in width and 7mm in length in each channel. Photolithography technique was used to fabricate a mold and replicated the chessboard-like patterns, eventually assembled to a glass slide to create a sealed system mirroring the natural xylem. The system was coated with carboxymethyl cellulose – dopamine (CMC-DOPA) via a amidation reaction to enhance the bioactivity and mimic the natural substrate. The flow profile within the system was analyzed experimentally and computationally using tracer particles and COMSOL Multiphysics®, respectively. Finally, the efficacy of the system was validated through Rps biofilms assay to examine its bacterial attachment and biofilm formation under static and dynamic conditions.

Results, Conclusions, and Discussions: The system, incorporating functional surface coatings of CMC-DOPA, provided a bioactive environment that significantly enhanced Rps attachment and biofilm formation in the presence of xylem sap. Using computational approaches, we confirmed that Rps experienced linear increasing drag forces in xylem-mimicking channels at higher flow rates. Consistently, attachment and biofilm assays conducted in our microfluidic system revealed that both seeding time and flow rates were critical for bacterial adhesion to surface and biofilm formation processes. These findings provided insights into the Rps attachment and biofilm formation processes, contributing to a better understanding of plant-pathogen interactions during wilt disease development.

Acknowledgements (Optional):