Antimicrobial Photodynamic Therapy with Chlorin e6 Is Bactericidal against Biofilms of the Primary Human Otopathogens.

Moraxella catarrhalis, Streptococcus pneumoniae, and nontypeable Haemophilus influenzae (NTHi) are ubiquitous upper respiratory opportunistic pathogens. Together, these three microbes are the most common causative bacterial agents of pediatric otitis media (OM) and have therefore been characterized as the primary human otopathogens. OM is the most prevalent bacterial infection in children and the primary reason for antibiotic administration in this population. Moreover, biofilm formation has been confirmed as a primary mechanism of chronic and recurrent OM disease. As bacterial biofilms are inherently metabolically recalcitrant to most antibiotics and these complex structures also present a significant challenge to the immune system, there is a clear need to identify novel antimicrobial approaches to treat OM infections. In this study, we evaluated the potential efficacy of antibacterial photodynamic therapy (aPDT) with the photosensitizer chlorin e6 (Ce6) against planktonic as well as biofilm-associated M. catarrhalis, S. pneumoniae, and NTHi. Our data indicate aPDT with Ce6 elicits significant bactericidal activity against both planktonic cultures and established biofilms formed by the three major otopathogens (with an efficacy of ≥99.9% loss of viability). Notably, the implementation of a novel, dual-treatment aPDT protocol resulted in this disinfectant effect on biofilm-associated bacteria and, importantly, inhibited bacterial regrowth 24 h posttreatment. Taken together, these data suggest this novel Ce6-aPDT treatment may be a powerful and innovative therapeutic strategy to effectively treat and eradicate bacterial OM infections and, significantly, prevent the development of recurrent disease.IMPORTANCE Otitis media (OM), or middle ear disease, is the most prevalent bacterial infection in children and the primary reason for antibiotic use and surgical intervention in the pediatric population. Biofilm formation by the major bacterial otopathogens, Moraxella catarrhalis, Streptococcus pneumoniae, and nontypeable Haemophilus influenzae, has been shown to occur within the middle ears of OM patients and is a key factor in the development of recurrent disease, which may result in hearing impairment and developmental delays. Bacterial biofilms are inherently impervious to most antibiotics and present a significant challenge to the immune system. In this study, we demonstrate that antimicrobial photodynamic therapy (aPDT) using the photosensitizer chlorin e6 elicits significant bactericidal activity versus planktonic and biofilm-associated otopathogens and supports further analyses of this novel, efficacious, and promising technology as an adjunctive treatment for acute and recurrent OM.

Antibiotics Enhance Prevention and Eradication Efficacy of Cathodic-Voltage-Controlled Electrical Stimulation against Titanium-Associated Methicillin-Resistant Staphylococcus aureus and Pseudomonas aeruginosa Biofilms.

Periprosthetic joint infection (PJI) develops clinically, even with antibiotic treatment, and methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa are predominant causes of these infections. Due to biofilm formation, antibiotic treatment for patients with PJI can perpetuate resistance, further complicating the use of noninvasive treatments. This study evaluated cathodic-voltage-controlled electrical stimulation (CVCES) of titanium, in combination with a clinically relevant antibiotic, to synergistically prevent MRSA and P. aeruginosa PJIs by inhibiting bacterial adherence or as a treatment for eradicating established biofilms. CVCES of -1.0 V, -1.5 V, or -1.8 V (versus Ag/AgCl), with or without vancomycin for MRSA or gentamicin for P. aeruginosa, was applied to sterile titanium incubated with cultures to evaluate prevention of attachment or eradication of preestablished biofilms. Treatments were 24 h long and included open-circuit potential controls, antibiotic alone, CVCES, and CVCES plus antibiotic. Biofilm-associated and planktonic CFU were enumerated. In general, CVCES at -1.8 V alone or with antibiotic completely eradicated biofilm-associated CFU for both strains, and these parameters were also highly effective against planktonic bacteria, resulting in a >6-log reduction in MRSA and no detectable planktonic P. aeruginosa All CFU were reduced ∼3 to 5 logs from controls for prevention CVCES plus antibiotics at -1.0 V and -1.5 V against MRSA. Remarkably, there were no detectable P. aeruginosa CFU following prevention CVCES at -1.0 V or -1.5 V with gentamicin. Our results suggest that CVCES in combination with antibiotics may be an effective approach for prevention and treatment of PJI.IMPORTANCE Periprosthetic joint infections (PJIs) develop clinically in the presence of antibiotic therapies and are responsible for increased patient morbidity and rising health care costs. Many of these infections involve bacterial biofilm formation on orthopedic hardware, and it has been well established that these biofilms are refractory to most antibiotic treatments. Recent studies have focused on novel methods to prevent and eradicate infection. Cathodic-voltage-controlled electrical stimulation (CVCES) has previously been shown to be effective as a method for prevention and eradication of Gram-positive and Gram-negative infections. The present study revealed that the utility of CVCES for prevention and eradication of methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa is enhanced in the presence of clinically relevant antibiotics. The synergistic effects of CVCES and antibiotics are effective in a magnitude-dependent manner. The results of this study indicate a promising alternative method to current PJI mitigation techniques.

Magnesium alloy AZ91 exhibits antimicrobial properties in vitro but not in vivo.

Magnesium alloys hold great promise for developing orthopedic implants that are biocompatible, biodegradable, and mechanically similar to bone tissue. This study evaluated the in vitro and in vivo antimicrobial properties of magnesium-9%aluminum-1%zinc (AZ91) and commercially pure titanium (cpTi) against Acinetobacter baumannii (Ab307). The in vitro results showed that as compared to cpTi, incubation with AZ91 significantly reduced both the planktonic (cpTi = 3.45e8, AZ91 = 8.97e7, p < 0.001) colony forming units (CFU) and biofilm-associated (cpTi = 3.89e8, AZ91 = 1.78e7, p = 0.01) CFU of Ab307. However, in vivo results showed no significant differences in the CFU enumerated from the cpTi and AZ91 implants following a 1-week implantation in an established rodent model of Ab307 implant associated infection (cpTi = 5.23e3, AZ91 = 2.46e3, p = 0.29). It is proposed that the in vitro results were associated with an increased pH in the bacterial culture as a result of the AZ91 corrosion process. The robust in vivo buffering capacity likely diminished this corrosion associated pH antimicrobial effect. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 221-227, 2018.

Cathodic voltage-controlled electrical stimulation of titanium implants as treatment for methicillin-resistant Staphylococcus aureus periprosthetic infections.

Effective treatment options are often limited for implant-associated orthopedic infections. In this study we evaluated the antimicrobial effects of applying cathodic voltage-controlled electrical stimulation (CVCES) of -1.8 V (vs. Ag/AgCl) to commercially pure titanium (cpTi) substrates with preformed biofilm-like structures of methicillin-resistant Staphylococcus aureus (MRSA). The in vitro studies showed that as compared to the open circuit potential (OCP) conditions, CVCES of -1.8 V for 1 h significantly reduced the colony-forming units (CFU) of MRSA enumerated from the cpTi by 97% (1.89 × 106 vs 6.45 × 104 CFU/ml) and from the surrounding solution by 92% (6.63 × 105 vs. 5.15 × 104 CFU/ml). The in vivo studies, utilizing a rodent periprosthetic infection model, showed that as compared to the OCP conditions, CVCES at -1.8 V for 1 h significantly reduced MRSA CFUs in the bone tissue by 87% (1.15 × 105 vs. 1.48 × 104 CFU/ml) and reduced CFU on the cpTi implant by 98% (5.48 × 104 vs 1.16 × 103 CFU/ml). The stimulation was not associated with histological changes in the host tissue surrounding the implant. As compared to the OCP conditions, the -1.8 V stimulation significantly increased the interfacial capacitance (18.93 vs. 98.25 µF/cm(2)) and decreased polarization resistance (868,250 vs. 108 Ω-cm(2)) of the cpTi. The antimicrobial effects are thought to be associated with these voltage-dependent electrochemical surface properties of the cpTi.

Moraxella catarrhalis is susceptible to antimicrobial photodynamic therapy with Photofrin.

BACKGROUND AND OBJECTIVE: Moraxella catarrhalis is a significant cause of pediatric otitis media (OM), which is the most prevalent bacterial infection in children and primary reason for antibiotic administration in this population. Moreover, biofilm formation has been implicated as a primary mechanism of chronic or recurrent OM disease. As bacterial biofilms are inherently resistant to most antibiotics and these complex structures also present a significant challenge to the immune system, there is a clear need to identify novel antimicrobial approaches to treat OM infections. In this study, we evaluated the potential efficacy of antibacterial photodynamic therapy (aPDT) with porfimer sodium (Photofrin (PF)) against planktonic as well as biofilm-associated M. catarrhalis. MATERIALS AND METHODS: The bactericidal activity of aPDT with PF was assessed against multiple recent clinical isolates of M. catarrhalis grown planktonically as well as in biofilms. The bactericidal activity of PF-aPDT was quantified by enumeration of colony forming units post-treatment. The effect of aPDT on M. catarrhalis biofilms was further investigated with scanning electron microscopy (SEM) imaging. RESULTS: aPDT with PF significantly reduced M. catarrhalis viability. Although PF-aPDT caused higher killing in planktonic grown organisms (5-6 log kill), biofilm grown bacteria also demonstrated a statistically significant reduction in viable organisms (3-4 log decrease in recoverable bacteria) following treatment as compared to saline only controls (P < 0.01). SEM studies indicated the PF-aPDT treated bacteria exhibited prominent morphological changes with visibly distorted cell membranes. CONCLUSIONS: aPDT with PF elicits significant bactericidal activity against both planktonic and biofilm-associated M. catarrhalis, suggesting this technology warrants further analysis as a potential novel antimicrobial treatment for acute or recurrent OM.