Nisin Z Potential for the Control of Diabetic Foot Infections Promoted by Pseudomonas aeruginosa Persisters.

Diabetic foot ulcers (DFU) are a major complication of diabetes mellitus and a public health concern worldwide. The ability of P. aeruginosa to form biofilms is a key factor responsible for the chronicity of diabetic foot infections (DFIs) and frequently associated with the presence of persister cells. These are a subpopulation of phenotypic variants highly tolerant to antibiotics for which new therapeutic alternatives are urgently needed, such as those based on antimicrobial peptides. This study aimed to evaluate the inhibitory effect of nisin Z on P. aeruginosa DFI persisters. To induce the development of a persister state in both planktonic suspensions and biofilms, P. aeruginosa DFI isolates were exposed to carbonyl cyanide m-chlorophenylhydrazone (CCCP) and ciprofloxacin, respectively. After RNA extraction from CCCP-induced persisters, transcriptome analysis was performed to evaluate the differential gene expression between the control, persisters, and persister cells exposed to nisin Z. Nisin Z presented a high inhibitory effect against P. aeruginosa persister cells but was unable to eradicate them when present in established biofilms. Transcriptome analysis revealed that persistence was associated with downregulation of genes related to metabolic processes, cell wall synthesis, and dysregulation of stress response and biofilm formation. After nisin Z treatment, some of the transcriptomic changes induced by persistence were reversed. In conclusion, nisin Z could be considered as a potential complementary therapy for treating P. aeruginosa DFI, but it should be applied as an early treatment or after wound debridement.

Pexiganan in Combination with Nisin to Control Polymicrobial Diabetic Foot Infections.

Diabetic foot ulcers (DFUs) are major complications of Diabetes mellitus being responsible for significant morbidity and mortality. DFUs frequently become chronically infected by a complex community of bacteria, including multidrug-resistant and biofilm-producing strains of Staphylococcus aureus and Pseudomonas aeruginosa. Diabetic foot infections (DFI) are often recalcitrant to conventional antibiotics and alternative treatment strategies are urgently needed. Antimicrobial Peptides (AMPs), such as pexiganan and nisin, have been increasingly investigated and reported as effective antimicrobial agents. Here, we evaluated the antibacterial potential of pexiganan and nisin used in combination (dual-AMP) to control the growth of planktonic and biofilm co-cultures of S. aureus and P. aeruginosa clinical strains, co-isolated from a DFU. A DFU collagen three-dimensional (3D) model was used to evaluate the distribution and efficacy of AMPs locally delivered into the model. The concentration of pexiganan required to inhibit and eradicate both planktonic and biofilm-based bacterial cells was substantially reduced when used in combination with nisin. Moreover, incorporation of both AMPs in a guar gum delivery system (dual-AMP biogel) did not affect the dual-AMP antimicrobial activity. Importantly, the application of the dual-AMP biogel resulted in the eradication of the S. aureus strain from the model. In conclusion, data suggest that the local application of the dual-AMPs biogel constitutes a potential complementary therapy for the treatment of infected DFU.

Potential of two delivery systems for nisin topical application to dental plaque biofilms in dogs.

BACKGROUND: Periodontal disease (PD) is caused by the development of a microbial biofilm (dental plaque) in the periodontium, affecting approximately 80% of dogs. Several bacterial species present in the canine oral cavity can be implicated in the development of this disease, including Enterococcus spp. To decrease antibiotic administration, a possible control strategy for dog's enterococcal PD may involve the use of the antimicrobial peptide (AMP) nisin. Nisin's inhibitory activity was evaluated against a collection of previously characterized enterococci obtained from the oral cavity of dogs with PD (n = 20), as well as the potential of a guar-gum gel and a veterinary toothpaste as topical delivery systems for this AMP. The Minimum Inhibitory (MIC) and Bactericidal Concentrations (MBC) and the Minimum Biofilm Eradication (MBEC) and Inhibitory Concentrations (MBIC) were determined for nisin and for the supplemented guar-gum gel. For the supplemented veterinary toothpaste an agar-well diffusion assay was used to evaluate its inhibitory potential. RESULTS: Nisin was effective against all isolates. Independently of being or not incorporated in the guar-gum gel, its inhibitory activity on biofilms was higher, with MBIC (12.46 ± 5.16 and 13.60 ± 4.31 µg/mL, respectively) and MBEC values (21.87 ± 11.33 and 42.34 ± 16.61 µg/mL) being lower than MIC (24.61 ± 4.64 and 14.90 ± 4.10 µg/mL) and MBC (63.09 ± 13.22 and 66.63 ± 19.55 µg/mL) values. The supplemented toothpaste was also effective, showing inhibitory activity against 95% of the isolates. CONCLUSIONS: The inhibitory ability of nisin when incorporated in the two delivery systems was maintained or increased, demonstrating the potential of these supplemented vehicles to be applied to PD control in dogs.

Guar gum as a new antimicrobial peptide delivery system against diabetic foot ulcers Staphylococcus aureus isolates.

Diabetic patients frequently develop diabetic foot ulcers (DFUs), particularly those patients vulnerable to Staphylococcus aureus opportunistic infections. It is urgent to find new treatments for bacterial infections. The antimicrobial peptide (AMP) nisin is a potential candidate, mainly due to its broad spectrum of action against pathogens. Considering that AMP can be degraded or inactivated before reaching its target at therapeutic concentrations, it is mandatory to establish effective AMP delivery systems, with the natural polysaccharide guar gum being one of the most promising. We analysed the antimicrobial potential of nisin against 23 S. aureus DFU biofilm-producing isolates. Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), minimum biofilm inhibitory concentration (MBIC) and minimum biofilm eradication concentration (MBEC) were determined for nisin diluted in HCl and incorporated in guar gum gel. Statistical analysis was performed using the Wilcoxon matched-pair test. Nisin was effective against all isolates, including some multidrug-resistant clinical isolates, independent of whether it is incorporated in guar gum. While differences among MIC, MBC and MBIC values were observed for HCl- and guar gum- nisin, no significant differences were found between MBEC values. Inhibitory activity of both systems seems to differ only twofold, which does not compromise guar gum gel efficiency as a delivery system. Our results highlight the potential of nisin as a substitute for or complementary therapy to current antibiotics used for treating DFU infections, which is extremely relevant considering the increase in multidrug-resistant bacteria dissemination. The guar gum gel represents an alternative, practical and safe delivery system for AMPs, allowing the development of novel topical therapies as treatments for bacterial skin infections.