Developing effective therapies to fight against biofilm-associated infection is extremely urgent. The complex environment of biofilm forces the bacteria to evade the elimination of antibiotics, resulting in recalcitrant chronic infections. To address this issue, a cationic antibacterial agent based on phosphindole oxide (beta-PM-PIO) is designed and prepared. The unique molecular structure endows beta-PM-PIO with aggregation-induced emission feature and efficient singlet oxygen generation ability. beta-PM-PIO shows excellent visual diagnostic function to planktonic bacteria and biofilm. In addition, owing to the synergistic effect of phototoxicity and dark toxicity, beta-PM-PIO can achieve superb antibacterial and antibiofilm performance against Gram-positive bacteria with less potential of developing drug resistance. Notably, beta-PM-PIO also holds excellent anti-infection capacity against drug-resistant bacteria in vivo with negligible side effects. This work offers a promising platform to develop advanced antibacterial agents against multidrug-resistant bacterial infection.