Pseudomonas aeruginosa is considered one of the most important pathogens that represent life-threatening risk in nosocomial environments, mainly in patients with severe burns. Antimicrobial photodynamic therapy (aPDT) has been effective to kill bacteria. The purpose of this study was to develop a burn wound and bloodstream infection model and verify aPDT effects on it. In vitro, we tested two wavelengths (blue and red LEDs) on a clinical isolate of P. aeruginosa strain with resistance to multiple antibiotics using HB:La+3 as photosensitizer. Verapamil® associated to aPDT was also studied. In vivo, P. aeruginosa-infected burned mice were submitted to aPDT. Bacterial counting was performed on local infection and bloodstream. Survival time of animals was also monitored. In this study, aPDT was effective to reduce P. aeruginosa in vitro. In addition, Verapamil® assay showed that HB:La+3 is not recognized by ATP-binding cassete (ABC) efflux pump mechanism. In the in vivo study, aPDT was able to reduce bacterial load in burn wounds, delay bacteremia and keep the bacterial levels in blood 2–3 logs lower compared with an untreated group. Mice survival was increased on 24 h. Thus, this result suggests that aPDT may also be a novel prophylactic treatment in the care of burned patients.