Antimicrobial assessment of phage therapy using a porcine model of biofilm infection

Antibiotic resistant bacterial communities persist in many types of wounds, chronic wounds in particular, in the form of biofilms. Biofilm formation is a major cause of severe infections and the main reason for a negative treatment outcome and slow healing progression. Chronic wounds are a silent epidemic essentially affecting people with co-morbid conditions such as diabetes and obesity and elderly persons particularly those with movement limitations. The development of complementary and alternative effective strategies to antibiotics for the treatment of chronic wounds is highly desired. Phage therapy constitutes a very promising approach in the control of topical microbial populations. In this work newly isolated phages were tested for their efficacy to control bacterial species that predominate in chronic wounds. Phage effectiveness was studied on 24-h old biofilms formed in polystyrene microplates and in porcine skin explants using two treatment approaches: individual phage and a cocktail of phages against four main pathogens commonly isolated from chronic wounds. The two models produced variations in the surface colonization ability, assessed by viable bacterial counts and microscopy visualization after using peptide nucleic acid (PNA) or locked nucleic acid probes (LNA) and 2-O-methyl (2-OMe) in fluorescence in situ hybridization (FISH), and in the phage-host interactions. Phages alone and combined caused greater reductions in the number of viable cells when biofilms had been formed on porcine skins and with greater variations detected at 4 h and 24 h of sampling. These results suggest that porcine skin models should be preferentially used to assess the use of phages and phage cocktails intended for topical use in order to understand the fate, throughout treatment time, of the population when dealing with biofilm-related infections.This work was supported by Portuguese Foundation for Science and Technology under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE2020(POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020–Programa Operacional Regional do Norte and the Project RECI/BBB-EBI/0179/2012 (FCOMP-01-0124-FEDER-027462). CM ac- knowledges the Portuguese Foundation for Science and Technology (FCT) grant SFRH/BD/94434/2013. SS is an Investigador FCT (IF/ 01413/2013).