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Antimicrobial photodynamic therapy offers significant benefits over standard antibiotic therapy in terms of rapid sterilization and resistance reduction. In this paper, a series of ruthenium complexes were designed and synthesized for antibacterial studies. [Ru(tpy)(dppz-R)Cl][PF6] (tpy = 2,2′:6′,2′′-terpyridine; dppz = dipyrido [3,2-a:2′,3′-c] phenazine; R = H, CH3, C(CH3)3, F, CF3, NO2, NH2 or CN), Ru1–Ru8 all showed superior in vitro antimicrobial effects under blue light irradiation. Ru2 has the best antimicrobial activity, with its inherent dark toxicity and photodynamic inactivation rapidly killing S. aureus without developing drug tolerance. Binding and damaging DNA as well as generating ROS may be its potential antimicrobial targets. Ru2 lacks a hemolytic effect on rabbit erythrocytes and has low phototoxicity against G. mellonella Larvae. In addition, Ru2 has been successfully used to treat skin wounds in mice infected with S. aureus. Therefore, Ru2 has shown great potential as a photodynamic antimicrobial agent against the threat of S. aureus infection. [ABSTRACT FROM AUTHOR]