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LED-Based In Vitro Screening for Assessing Photoactivable Molecules in Bacterial Photodynamic Inactivation
Published on: January 24, 2025
Targeted photodynamic therapy of established soft-tissue infections in mice
Faten Gad1, Touqir Zahra, Kevin P Francis
1Wellman Laboratories of Photomedicine, Masschusetts General Hospital, Boston 02114, USA.
Abstract:
The worldwide rise in antibiotic resistance necessitates the development of novel antimicrobial strategies. Although many workers have used photodynamic therapy (PDT) to kill bacteria in vitro, the use of this approach has seldom been reported in vivo in animal models of infection. We have previously described the first use of PDT to treat excisional wound infections by Gram-(-) bacteria in living mice. However, these infected wound models involved a short timespan between infection (30 min) and treatment by PDT. We now report on the use of PDT to treat an established soft-tissue infection in mice. We used Staphylococcus aureus stably transformed with a Photorhabdus luminescenslux operon (luxABCDE) that was genetically modified to be functional in Gram-(+) bacteria. These engineered bacteria emitted bioluminescence, allowing the progress of the infection to be monitored in both space and time with a low light imaging charge-coupled device (CCD) camera. One million cells were injected into one or both thigh muscles of mice that had previously been rendered neutropenic by cyclophosphamide administration. Twenty-four hours later, the bacteria had multiplied more than one hundredfold; poly-L-lysine chlorin e6 conjugate or free chlorin e6 was injected into one area of infected muscle and imaged with the CCD camera. Thirty minutes later, red light from a diode laser was delivered as a surface spot or by interstitial fiber into the infection. There was a light dose dependent loss of bioluminescence (to <5% of that seen in control infections) not seen in untreated infections or those treated with light alone, but in some cases, the infection recurred. Treatment with conjugate alone led to a lesser reduction in bioluminescence. Infections treated with free chlorin e6 responded less well and the infection subsequently increased over the succeeding days, probably due to PDT-mediated tissue damage. PDT-treated infected legs healed better than legs with untreated infections. This data shows that PDT may have applications in drug-resistant soft-tissue infections.
Insights
Photodynamic therapy (PDT) effectively treated established Staphylococcus aureus soft-tissue infections in mice, reducing bacterial bioluminescence and improving wound healing. This shows promise for combating antibiotic-resistant infections.
Area of Science:
- Microbiology
- Biomedical Engineering
- Photomedicine
Background:
- Antibiotic resistance is a growing global health threat, driving the need for alternative antimicrobial strategies.
- Photodynamic therapy (PDT) has shown efficacy against bacteria in vitro, but in vivo applications, especially for established infections, are less explored.
- Previous studies demonstrated PDT for early-stage Gram-negative wound infections in mice.
Purpose of the Study:
- To evaluate the efficacy of photodynamic therapy (PDT) in treating established Staphylococcus aureus soft-tissue infections in a murine model.
- To assess the impact of PDT on bacterial load and infection progression in vivo.
- To investigate the potential of PDT as a therapeutic option for drug-resistant bacterial infections.
Main Methods:
- Staphylococcus aureus engineered to express bioluminescence (lux operon) were inoculated into neutropenic mouse thigh muscles.
- Infections were monitored over 24 hours using a low-light imaging charge-coupled device (CCD) camera.
- Established infections were treated with poly-L-lysine chlorin e6 conjugate or free chlorin e6, followed by diode laser irradiation.
- Bioluminescence reduction and wound healing were assessed to determine treatment efficacy.
Main Results:
- PDT treatment, particularly with the chlorin e6 conjugate, significantly reduced bacterial bioluminescence (to <5%) in a light-dose-dependent manner.
- Untreated infections or those treated with light alone showed no significant reduction in bioluminescence.
- While PDT demonstrated efficacy, some infections recurred, and free chlorin e6 treatment resulted in less favorable outcomes possibly due to tissue damage.
- PDT-treated infected legs exhibited improved healing compared to untreated controls.
Conclusions:
- Photodynamic therapy (PDT) is a viable strategy for treating established Staphylococcus aureus soft-tissue infections in vivo.
- The use of bioluminescent bacteria allows for real-time monitoring of infection progression and treatment response.
- PDT, especially with targeted conjugates, shows potential for managing antibiotic-resistant bacterial infections, although recurrence and tissue damage need further consideration.
