Vibrio vulnificus is a gram-negative, highly invasive bacterium responsible for human opportunistic infections. We studied the antibacterial effects of toluidine blue O (TBO)-mediated photodynamic therapy (PDT) for V. vulnificus wound infections in mice. Fifty-three percent (10 of 19) of mice treated with 100 g of TBO per ml and exposed to broad-spectrum red light (150 J/cm 2 at 80 mW/cm 2 ) survived, even though systemic septicemia had been established with a bacterial inoculum 100 times the 50% lethal dose. In vitro, the bacteria were killed after exposure to a lower light dose (100 J/cm 2 at 80 mW/cm 2 ) in the presence of low-dose TBO (0.1 g/ml). PDT severely damaged the cell wall and reduced cell motility and virulence. Cell-killing effects were dependent on the TBO concentration and light doses and were mediated partly through the reactive oxygen species generated during the photodynamic reaction. Our study has demonstrated that PDT can cure mice with otherwise fatal V. vulnificus wound infections. These promising results suggest the potential of this regimen as a possible alternative to antibiotics in future clinical applications.Photodynamic therapy (PDT) is an experimental treatment which shows great potential for the treatment of neoplastic and nonneoplastic diseases (11,12). PDT involves a light-sensitive photosensitizer, light, and molecular oxygen (14). After excitation with visible light, highly cytotoxic singlet oxygen and other reactive oxygen species (ROS) are generated by either energy or electron transfer (14,30,32). The use of photosensitizers for microbial eradication can be traced back to before the age of chemotherapy (16,38). The antimicrobial effects of PDT are increasingly recognized. The technique has been shown to have effects against a range of oral pathogens and also against drug-resistant bacteria. More importantly, there have been reports of the use of PDT to treat infections in selected animal models (13,15,17) and some clinical trials (16), with encouraging results. The activities of the virulence factors of gram-negative bacteria were also reduced with PDT (24). Among the many photosensitizers, toluidine blue O (TBO), a cationic phenothiazinium photosensitizer (16,38), has been shown to be phototoxic to gram-negative bacilli with red-light irradiation (39). However, in vivo antimicrobial studies of TBO-PDT were mainly limited to oral infections (39).Vibrio vulnificus is a gram-negative, motile, curved bacillus of the family Vibrionaceae (22). Most Vibrio species are free-living in marine or brackish water (6). Many cases of V. vulnificus infection have been reported from the coastal areas of the United States (4), Asia (6), and Europe (10). V. vulnificus causes primary sepsis, wound infection, and gastrointestinal illness in humans (6,22). This organism is extremely virulent, and infections with this organism typically occur in patients with underlying liver disease 1 to 2 days after exposure. The mortality rate is up to 55% in septic patients and 25% in those with wound infect...