<p>Phyllite, when used as a cement subgrade, is susceptible to thermal fatigue degradation due to temperature fluctuations resulting from prolonged exposure to natural environments. However, limited research exists on the anti-seepage properties of microbial-modified phyllite. This study investigates the capillary water migration characteristics of two types of strongly weathered modified phyllite subjected to thermal fatigue cycles, focusing on their anti-seepage performance and structural integrity for use as roadbed fillers or in infrastructure applications. Through spontaneous imbibition and thermal fatigue experiments, the effects of microbial modification on phyllite’s permeability were explored. The results indicated that microbial modification reduced phyllite’s porosity, improved its internal pore structure, and enhanced its anti-seepage performance. Compared to unmodified phyllite, the modified materials exhibited greater resistance to capillary water absorption. This study highlights the potential of microbial treatment as a sustainable method to enhance the properties of weathered phyllite. By combining knowledge of phyllite and microbial ecology, we aim to foster a deeper understanding of the relationship between these fields and better assess how microorganisms affect phyllite’s permeability.</p>