Unraveling new therapies for diabetic foot ulcers: antimicrobial peptides and probiotics

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

Publication . Santos, Raquel; Gomes, Diana; Macedo, Hermes; Barros, Diogo; Tibério, Catarina; Veiga, Ana Salomé; Tavares, Luís; Castanho, Miguel A. R. B.; Oliveira, Manuela

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.

2016Journal article

Open access

Novel therapeutic strategies for the management of diabetic foot infections : the evaluation of selected antimicrobial peptides against clinically isolated bacterial pathogens

Publication . Santos, Tânia Raquel Martins dos; Oliveira, Maria Manuela Castilho Monteiro de

Diabetic foot infections (DFIs) are a frequent complication of Diabetes mellitus. These ulcers are prone to be colonized by Staphylococcus aureus and Pseudomonas aeruginosa, including multidrug resistant and biofilm-producing strains, possibly leading to DFI chronicity and amputation. New therapeutic strategies for DFI management are urgent and the antimicrobial peptides (AMPs) nisin and pexiganan are potential candidates. This project aimed to evaluate the activity of these AMPs, incorporated in a guar gum biogel, against selected DFI clinical isolates. Firstly, nisin’s activity against a collection of S. aureus DFI clinical isolates was determined. Results showed that nisin was able to inhibit and eradicate S. aureus planktonic and biofilm cells at concentrations below its acceptable daily intake. When incorporated in the biogel, nisin kept its antimicrobial activity. This work also evaluated the potential of nisin to complement the activity of conventional antiseptics and antibiotics against established biofilms formed by these isolates. An in vitro antimicrobial schematic protocol was developed to mimetize DFI management guidelines. Fifteen antimicrobial combinations, including nisin-biogel, chlorhexidine, clindamycin, gentamicin and vancomycin, were tested. Results showed that the higher levels of biofilm inhibitory effects were presented by therapeutic combinations that included the nisin-biogel formulation. Nisin-biogel ideal storage conditions and cytotoxicity were also evaluated. Results demonstrate that if stored at temperatures between -20 and 22ºC, nisin-biogel is able to maintain its antimicrobial activity up to 24 months. Moreover, after 24 h of exposition, the nisin-biogel presented no significant levels of toxicity regarding the human keratinocytes under study. Lastly, to cover the complex microbiota present in DFIs, a combination of AMPs with different action spectra was developed, based on the simultaneous incorporation of nisin and pexiganan in the biogel. The activity of this dual-AMPs formulation was tested against two S. aureus and P. aeruginosa strains isolated from the same DFI. Acting together, these AMPs were able to diffuse from the biogel and inhibit and eradicate biofilms formed by these DFI isolates. The effectiveness of AMPs, particularly nisin and pexiganan, as novel antimicrobial strategies for the management of DFIs is still an unknown territory that merits investigation. In vitro biofilm models are the basis of preliminary research; however, they underrepresent the complex microbiota present in DFIs and their interaction with the immune system and skin cells constituents. Further research is necessary to understand the AMPs full potential regarding the clinical management of biofilm-related diseases, such as DFIs.

2020-02-28Doctoral thesis

Open access