Antimicrobial photodynamic inactivation (aPDI), using photosensitisers in combination with antibiotics, is a promising multi-target strategy to address antibiotic resistance, particularly in wound infections. This study aimed to elucidate the antibacterial mode of action of combinations of berberine (Ber) or curcumin (Cur) with selected antibiotics (Ber-Ab or Cur-Ab) under blue light irradiation (420 nm) against Staphylococcus aureus, including methicillin-resistant (MRSA) and methicillin-susceptible (MSSA) strains. Multiple physiological parameters were assessed using complementary assays (fluorometry, epifluorescence microscopy, flame emission and atomic absorption spectroscopy, zeta potential, flow cytometry, and the plate agar method) to examine the effect on ROS production, membrane integrity, DNA damage, motility and virulence factors of S. aureus. Results indicated that blue light photoactivated Ber-Ab and Cur-Ab combinations led to substantial ROS generation, even at low concentrations, causing oxidative stress that severely impacted bacterial membrane integrity (approximately 90 % in MRSA and 40 % in MSSA). Membrane destabilization was further confirmed by elevated intercellular potassium release (≈ 2.00 and 2.40 µg/mL in MRSA and MSSA, respectively). Furthermore, significant DNA damage was observed in both strains (≈ 50 %). aPDI treatment with blue light also reduced S. aureus pathogenicity by impairing motility and inhibiting key virulence factors such as proteases, lipases, and gelatinases, all of which play key roles in the infectious process. Overall, Ber-Ab combinations demonstrated the highest efficacy across all parameters tested, highlighting for the first time the multi-target therapeutic potential of this phytochemical-based aPDI strategy to combat antibiotic-resistant S. aureus infections and improve wound infection treatment outcomes.