Polyacrylamide hydrogel-immobilized Escherichia coli cell lysate for efficient removal and reduction in transformability of extracellular antibiotic resistance genes in water

The dissemination of extracellular antibiotic resistance genes (eARGs) through horizontal gene transfer poses a serious threat to global water safety. While Escherichia coli MG1655 is known to employ intracellular nucleases to defend against foreign DNA, the potential of its cell lysate to enzymatically degrade plasmid-borne eARGs and disrupt their environmental propagation remains unexplored. This study demonstrated that E. coli MG1655 cell lysate can degrade the plasmid-borne ampR gene by 99.9% within 48 h, and the maximum first-order degradation rate constant measured by long amplicon qPCR was 0.20 h−1 at the selected dosage state. This degradation eliminated the ability of the plasmid pUC19 for transformation, reducing it to below detectable levels (<10−6 transformants/recipients). The DNase I, produced by the chromosomal endA gene, was identified as the likely nuclease driving eARG cleavage. Environmental conditions critically modulated degradation efficiency: humic acid (50 mg/L) decreased the rate by 41.3%, while divalent cations (e.g., Mg2+) increased the rate by 1.65-fold. To transmit laboratory findings into real-world application, a polyacrylamide hydrogel-immobilized lysate (PAM-cell lysate) was engineered, which outperformed free lysate in treating plasmid-spiked real wastewater, sustaining 99.9% of ampR removal across three treatment cycles. This research demonstrated that PAM-cell lysate, without genetic modification, can be a cost-effective and scalable solution for attenuating the spread of eARGs in water, advancing the development of complementary enzymatic strategies for sustainable water remediation.