Mechanofiltration Enables High-Throughput Measurements of Bacterial Cell Mechanics

Bacteria experience diverse mechanical forces throughout their natural environments, yet quantitative measurements of bacterial biomechanics remain challenging because most existing techniques require specialized instrumentation, such as atomic force microscopy (AFM) or microfluidic devices. Here, we introduce mechanofiltration, a simple, high-throughput assay that estimates whole-cell mechanics using standard laboratory equipment. In mechanofiltration, bacterial suspensions are centrifuged through porous membranes in multiwell plates. Pressure generated during centrifugation drives cells toward pores that are smaller than cell width, requiring cells to deform to transit through the membrane. By combining recovery on the opposite side of the filter with measurements of cell size, the assay estimates maximum cell deformation, a size-corrected proxy for whole-cell stiffness. We show that mechanofiltration detects established reductions in Escherichia coli cell stiffness caused by genetic disruption of load-bearing cell envelope components, treatment with outer membrane- destabilizing agents, and sublethal exposure to antibiotics. Fold changes in maximum deformation closely agree with published measurements obtained using AFM, cell-bending assays, and osmotic shock experiments. Mechanofiltration also reproduces previously reported mechanical differences among bacterial species with distinct morphologies and envelope architectures. Together, these results establish mechanofiltration as an accessible, inexpensive, and scalable approach for identifying genetic and environmental determinants of bacterial mechanics and for high-throughput screening of bacterial biomechanical phenotypes.