Bacterial motility is essential for navigating heterogeneous environments like soil, where it plays a key role in nutrient cycling, bioremediation, and overall soil health. Despite its importance, the interplay between bacterial motility and soil microstructures—such as the effects of physical confinement and interfacial interactions—remains underexplored. In this study, we investigated the motility of Escherichia coli bacteria in aqueous micro-environments with three different natural soil samples and examined how the particle size, void fraction, and proximity to soil particles affect bacterial motility and movement patterns by quantitatively analyzing bacterial trajectories, velocities, and directional changes. We observed that bacterial velocity decreased significantly in soil micro-environments, showing a strong positive correlation with the soil particle size and a negative correlation with the void fraction of the soil samples. Additionally, bacteria in soil micro-environments showed rapid and dramatic directional changes, and the rate of directional changes of bacteria was negatively correlated with the particle size. These results were further validated with synthetic micro-environments with glass microspheres. As the density of microspheres increased, the translational velocity of bacteria decreased while the directional changes increased. This study enhances our understanding of how the soil type, porosity, and particle proximity impact bacterial movement and is expected to contribute to a better understanding of bacterial activities on soil health and management.

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