Sliver nanoparticles (NPs) possess wonderful antibacterial activity while ZnO is a promising photocatalyst. Although it has been reported that the photocatalytic activity of Ag/ZnO heterostructure nanoparticles (HNPs) was better than that of pure ZnO NPs, the existing comparisons on their antibacterial activity were all taken between Ag-doped ZnO HNPs and pure Ag NPs (or pure ZnO NPs), and were still controversial. Ag islands on ZnO, another type of Ag/ZnO HNPs, were synthesized by a two-step preparation and characterized in this work. The antibacterial activity of as-synthesized Ag/ZnO HNPs, especially with 5.0wt.% of Ag, against Escherichia coli (ATCC 25922) and Staphylococcus aureus (ATCC 25923) was enhanced greatly in contrast with that of the simple mixture of Ag and ZnO NPs. The enhancement was found to be mainly due to the obvious increase in the reactive oxidative species (especially superoxide) and the increased damage to plasmid DNA induced by Ag/ZnO HNPs.