pH-Gated Activation of Nematodes-Secreted NUC-1 Accelerates Extracellular Antibiotic Resistance Gene Degradation in Aquatic Environments

The global dissemination of extracellular antibiotic resistance genes (eARGs) in environmental matrices necessitates urgent development of mitigation approaches. Although nematodes exhibit potential as biological agents for eARG degradation, significant research gaps exist in understanding their performance under diverse environmental conditions and strategies for enhancing degradation efficiency through systematic parameter optimization. Here, we systematically evaluated the degradation of plasmid-borne tetM by Caenorhabditis elegans across eight high nematodes-prevalent habitats, revealing a remarkable 38-fold variation in efficacy. Solution pH was identified as the pivotal regulatory parameter through controlled experiments. Acidification to pH 6 enhanced nematodes-mediated eARG degradation by 25-fold, effectively reducing the transformation efficiency below the detectable limit within 15 min. Through multidisciplinary analyses incorporating gene mutation analysis, mRNA quantification, capillary electrophoresis, and zymographic analysis, we demonstrate that environmental pH specifically modulates NUC-1 activity rather than expression. Structural modeling and pK_a calculation reveal this pH-dependent regulation operates through protonation state change in the NUC-1 catalytic center, achieving maximal enzymatic activity at pH 6. Remarkably, this pH-gated regulatory mechanism is conserved across five nematode species spanning two distinct families, highlighting its broad biological significance and biotechnological potential. Our study establishes the first comprehensive environmental assessment framework for nematodes-mediated eARG degradation and elucidates a pH-gated regulation mechanism at the molecular level, providing a novel foundation for developing biotechnologies to control AR dissemination with spatiotemporal accuracy.