Single-Cell Force Spectroscopy Uncovers Root Zone- and Bacteria-Specific Interactions

Understanding root-bacteria interactions with plant growth-promoting rhizobacteria (PGPR) is key to developing effective biofertilizers for sustainable agriculture. We performed single-cell force spectroscopy using the atomic force microscope (AFM) to study the primary attachment of two PGPR, Bacillus velezensis and Pseudomonas defensor, to different regions of Arabidopsis thaliana roots. Force measurements with individual cells uncovered distinct attachment strategies by each strain, involving binding via micrometer-long polymers from both bacteria and root surfaces. Flagella differentially affected the binding interactions of each PGPR; their removal altered binding characteristics differently for each strain, highlighting the importance of flagella in early root colonization. Using silica beads to mimic the negatively charged bacteria, we demonstrated the influence of electrostatic forces on root-bacteria interactions. We also examined interactions with abiotic surfaces of varying surface energies, revealing the roles of hydrophilic and hydrophobic forces in initial binding. Our measurements show that differences in the physicochemical properties of bacteria and roots are responsible for variations in primary attachment strategies between PGPR strains and root regions. Parallel fluorescence measurements corroborated our AFM single-cell analysis. Overall, our results provide a nanoscale view of bacterial attachment to roots, offering key insights into how beneficial bacteria colonize roots, crucial for enhancing biofertilizer effectiveness.