Formulation, stabilisation and encapsulation of bacteriophage for phage therapy

Against a backdrop of global antibiotic resistance and increasing awareness of the importance of the human microbiota, there has been resurgent interest in the potential use of bacteriophages for therapeutic purposes, known as phage therapy. A number of phage therapy phase I and II clinical trials have concluded, and shown phages don’t present significant adverse safety concerns. These clinical trials used simple phage suspensions without any formulation and phage stability was of secondary concern. Phages have a limited stability in solution, and undergo a significant drop in phage titre during processing and storage which is unacceptable if phages are to become regulated pharmaceuticals, where stable dosage and well defined pharmacokinetics and pharmacodynamics are de rigueur. Animal studies have shown that the efficacy of phage therapy outcomes depend on the phage concentration (i.e. the dose) delivered at the site of infection, and their ability to target and kill bacteria, arresting bacterial growth and clearing the infection. In addition, in vitro and animal studies have shown the importance of using phage cocktails rather than single phage preparations to achieve better therapy outcomes. The in vivo reduction of phage concentration due to interactions with host antibodies or other clearance mechanisms may necessitate repeated dosing of phages, or sustained release approaches. Modelling of phage-bacterium population dynamics reinforces these points. Surprisingly little attention has been devoted to the effect of formulation on phage therapy outcomes, given the need for phage cocktails, where each phage within a cocktail may require significantly different formulation to retain a high enough infective dose. This review firstly looks at the clinical needs and challenges (informed through a review of key animal studies evaluating phage therapy) associated with treatment of acute and chronic infections and the drivers for phage encapsulation. An important driver for formulation and encapsulation is shelf life and storage of phage to ensure reproducible dosages. Other drivers include formulation of phage for encapsulation in micro- and nanoparticles for effective delivery, encapsulation in stimuli responsive systems for triggered controlled or sustained release at the targeted site of infection. Encapsulation of phage (e.g. in liposomes) may also be used to increase the circulation time of phage for treating systemic infections, for prophylactic treatment or to treat intracellular infections. We then proceed to document approaches used in the published literature on the formulation and stabilisation of phage for storage and encapsulation of bacteriophage in micro- and nanostructured materials using freeze drying (lyophilization), spray drying, in emulsions e.g. ointments, polymeric microparticles, nanoparticles and liposomes. As phage therapy moves forward towards Phase III clinical trials, the review concludes by looking at promising new approaches for micro- and nanoencapsulation of phages and how these may address gaps in the field

Phage-Bacterium War on Polymeric Surfaces: Can Surface-Anchored Bacteriophages Eliminate Microbial Infections?

These studies illustrate synthetic paths to covalently attach T1 and φ11 bacteriophages (phages) to inert polymeric surfaces while maintaining bacteriophage’s biological activities capable of killing deadly human pathogens. The first step involved formation of acid (COOH) groups on polyethylene (PE) and polytetrafluoroethylene (PTFE) surfaces using microwave plasma reactions in the presence of maleic anhydride, followed by covalent attachment of T1 and φ11 species via primary amine groups. The phages effectively retain their biological activity manifested by a rapid infection with their own DNA and effective destruction of Escherichia coli and Staphylococcus aureus human pathogens. These studies show that simultaneous covalent attachment of two biologically active phages effectively destroy both bacterial colonies and eliminate biofilm formation, thus offering an opportunity for an effective combat against multi-bacterial colonies as well as surface detections of other pathogens. [Image: see text

Chestnut honey and bacteriophage application to control Pseudomonas aeruginosa and Escherichia coli biofilms: evaluation in an ex vivo wound model

Chronic skin wounds represent a major burn both economically and socially. Pseudomonas aeruginosa and Escherichia coli are among the most common colonizers of infected wounds and are prolific biofilm formers. Biofilms are a major problem in infections due to their increasingly difficult control and eradication, and tolerance to multiple prescribed drugs. As so, alternative methods are necessary. Bacteriophages (phages) and honey are both seen as a promising approach for biofilm related infections. Phages have specificity towards a bacterial genus, species or even strain, self-replicating nature, and avoid dysbiosis. Honey has gained acknowledgment due to its antibacterial, antioxidant and anti-inflammatory and wound healing properties. In this work, the effect E. coli and P. aeruginosa phages vB_EcoS_CEB_EC3a and vB_PaeP_PAO1-D and chestnut honey alone, and combined were tested using in vitro (polystyrene) and ex vivo (porcine skin) models and against mono and dual-species biofilms of these bacteria. In general, colonization was higher in the porcine skins and the presence of a second microorganism in a consortium of species did not affect the effectiveness of the treatments. The antibacterial effect of combined therapy against dual-species biofilms led to bacterial reductions that were greater for biofilms formed on polystyrene than on skin. Monospecies biofilms of E. coli were better destroyed with phages and honey than P. aeruginosa monospecies biofilms. Overall, the combined phage-honey formulations resulted in higher efficacies possibly due to honeys capacity to damage the bacterial cell membrane and also to its ability to penetrate the biofilm matrix, promoting and enhancing the subsequent phage infection.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (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-01-0124FEDER-027462). AO acknowledge financial support from the Portuguese Foundation for Science and Technology (FCT) through the project PTDC/CVT-EPI/4008/2014 (POCI-01-0145-FEDER-016598). SS is an Investigador FCT (IF/01413/2013).info:eu-repo/semantics/publishedVersio

Bacteriophages in the Experimental Treatment of Pseudomonas aeruginosa Infections in Mice

International audienceThe regular increase of drug-resistant pathogens has been a major force in the renewed interest in the use of bacteriophages as therapeutics. In addition to experience acquired in eastern Europe where bacteriophages have been used to treat bacterial infections in humans, in Western countries only experimental models have been developed until recently. The Gram-negative bacterium Pseudomonas aeruginosa is an opportunistic pathogen causing particularly severe infections in cystic fibrosis patients. Several experimental models in mice have yielded encouraging results for the use of bacteriophages to treat or prevent septicemia, skin and lungs infections caused by P. aeruginosa. Now, a phase II clinical trial conducted in the United Kingdom provides evidence for the efficacy of bacteriophage treat- ments in chronic otitis due to antibiotic-resistant P. aeruginosa strains. Together with experimental models, these results provide an incen- tive to develop more research and clinical studies to fully appreciatethe benefits of the use of bacteriophages in medicine