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The degradation process was generally believed to occur only after phagosome internalization and maturation. Here, we report an early DNase activity at the nascent phagocytic cup (PC) prior to its closure. Using a fluorescent DNase sensor, we revealed rapid and ubiquitous DNase activity upon PC formation across various macrophage types. We further identified the responsible enzyme as the membrane-bound DNaseX, which is constitutively recruited to the PC during PC formation. F-actin polymerization was found to correlate with DNase activity in the PC, likely by promoting physical engagement of DNaseX with solid DNA materials. Functionally, we show that macrophages degrade extracellular DNA (eDNA) within bacterial biofilms through direct physical contact, clearing the eDNA structures without internalization. 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