Efficacy of a Topical Nitric Oxide-Releasing Gel on Polymicrobial Wound Infections

Wounds are colonized frequently by heterogeneous microflora. Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA) are two of the most isolated bacterial species from wounds, and both typically form highly organized biofilms. Nitric oxide (NO) is a short-lived, diatomic, lipophilic gas with antimicrobial activity. Recently, NO and its derivatives have been shown to exhibit broad-spectrum antimicrobial activity against bacteria, viruses, and parasites.

P. aeruginosa strain ATCC 27312 or military isolate PA09-010 were combined with methicillin-resistant S. aureus strain MRSA USA300 to demonstrate the ability of NO to reduce polymicrobial infections in a porcine wound infection model. Deep partial-thickness wounds (10 mm × 7 mm × 0.5 mm) were made on four animals using a specialized electrokeratome. Wounds were inoculated with MRSA USA300 combined with PA09-010 in three animals and MRSA USA300 combined with PA27312 in one animal, then wounds were covered with polyurethane film dressings. After 48 hours, three wounds were recovered for baseline enumeration. The remaining wounds were randomly assigned to treatment groups and treated once daily. The NO topical gels tested were combinations of two phases, ointment phases with various concentrations (2-20%) combined with hydrogels with fast or slow release kinetics. A 4-day study with microbiological recovery was conducted on day 4. A separate 7-day study was also conducted, with microbial burden assessed on day 7.

The largest efficacy against MRSA USA300 was observed for the NO formulation with 2% concentration and fast release kinetics. This treatment reduced the MRSA USA300 bacterial count by more than 99.97% and 99.95% from baseline in wounds co-infected with PA09-010 and PA 27312, respectively, at day 7. Treatments showed a minimal efficacy against PA27312 and PA09-010 strains in both assessment times. MRSA USA300 was reduced to a lesser extent when it was combined with PA27312 as compared to PA09-010.

These studies demonstrate that NO-releasing topical formulations effectively reduce the MRSA burden in established biofilms composed of multiple microorganisms. Minimal efficacy against PA was observed. It has been demonstrated that MRSA bioburden is significantly reduced when inoculated together with P. aeruginosa. A better understanding of mechanisms of host-bacteria interactions, in single or mixed species biofilms, may lead to the development of novel therapeutic approaches. Overall, NO offers a promising alternative treatment against MRSA in polymicrobial infections.