Distinctin

Amphibian skin secretions are an important treasure house of bioactive antimicrobial peptides (AMPs). Despite having been the focus of decades of research in this context, investigations of phyllomedusine frogs continue to identify new AMPs from their skin secretions. In this study, the prototype of a novel family of AMP distinctin-like-peptide-PH (DLP-PH) was identified from the skin secretion of the otherwise well-studied Tiger-Legged Tree Frog Phyllomedusa hypochondrialis through cloning of its precursor-encoding cDNA from a skin secretion-derived cDNA library by a 3'-rapid amplification of cDNA ends (RACE) strategy. Subsequently, the mature peptide was isolated and characterized using reverse-phase HPLC and MS/MS fragmentation sequencing. DLP-PH adopted an α-helical conformation in membrane mimetic solution and demonstrated unique structural features with two distinct domains that differed markedly in their physiochemical properties. Chemically synthesized replicates of DLP-PH showed antimicrobial activity against planktonic bacterial and yeast cells, but more potent against Escherichia coli at 32 μg/mL. However, DLP-PH showed much weaker inhibitory activity against the growth of sessile cells in biofilms. In addition, DLP-PH exhibited anti-proliferative activity against human cancer cell lines, H157, and PC3, but with no major toxicity against normal human cell, HMEC-1. These combined properties make DLP-PH deserving further study as an antimicrobial agent and further investigations of its structure-activity relationship could provide valuable new insights into drug lead candidates for antimicrobial and/or anti-cancer purposes.

The aim of this study was to evaluate the efficacy of distinctin in the management of cutaneous methicillin-resistant Staphylococcus aureus (MRSA) wound infections in an experimental mouse model. Wounds, made in the panniculus carnosus of BALB/c mice, were inoculated with 5 × 10(7) colony-forming units (CFU) of MRSA. Mice were treated with topical distinctin (1 mg/kg of body weight), topical teicoplanin (7 mg/kg of body weight), intraperitoneal teicoplanin (7 mg/kg of body weight); topical teicoplanin and daily intraperitoneal teicoplanin; topical distinctin and daily intraperitoneal teicoplanin. Bacterial cultures of excised tissues and histological examination of microvessel density and of vascular endothelial growth factor (VEGF) expression were studied. It was found that topical distinctin combined with parenteral teicoplanin inhibited bacterial growth to levels comparable with those observed in uninfected animals. Wounded areas of animals treated with distinctin were characterized by a more mature granulation tissue, with a more organized and denser type of connective tissue, compared to mice treated only with teicoplanin. Treatment with topical distinctin had a significant impact on VEGF expression and microvessel density. The combined use of distinctin with teicoplanin may be useful in the management of infected wounds by significantly inhibiting bacterial growth and accelerating the repair process.

The ability of microorganisms to adhere to medical implants is a problem of high visibility and has been focused in numerous investigations. To assess the efficacy of distinctin and conventional antibiotics in the treatment of central venous catheter in vitro and in vivo studies were performed. The in vitro susceptibility assay was performed against S. aureus biofilms developed on 96-well polystyrene tissue culture plates. Efficacy studies were performed in a rat model of CVC infection. Twenty-four hours after implantation, the catheters were filled with distinctin. Thirty minutes later, rats were challenged via the CVC with S. aureus. Administration of antibiotics into the CVC at a concentration equal to the MBC for adherent cells, or at 1024 microg/mL began 24 h later. The killing activities of all antibiotics against adherent bacteria were at least four- to eightfold lower than against freely growing cells. When antibiotics were used in distinctin pretreated wells, they showed a significant increase of activity. The in vivo studies showed that when CVCs were pretreated with distinctin biofilm bacterial load was further decreased to 10(1) CFU/mL and bacteremia was not detected. Distinctin displays potential as an adjunctive agent to antibiotics in the treatment of CVC-related infections.