Phage therapy for treatment of methicillin-resistant Staphylococcus aureus biofilm: administration in vivo in a Galleria mellonella model of implant-associated infection

The placement of indwelling medical devices, such as pacemakers, coronary stents, orthopedic articular prostheses, has led to a significant improvement in the quality of life, restoring lost biological functions in patients undergoing these operations. However, the surgical procedures required for the insertion of the devices may be associated with a moderate risk of contracting bacterial infections mainly due to Staphylococcus aureus. The surgical site, in fact, is defined as the main entrance door of the microorganisms. Furthermore, the affinity for biomaterials of which medical devices are made favors bacterial adhesion and subsequent colonization of surfaces, with the formation of biofilms. Biofilms are sessile communities of metabolically heterogeneous bacterial cells, encased in an extracellular matrix that they produce. The presence of an extracellular polymer matrix and less metabolically active cells makes biofilms-embedded cells extremely tolerant to antibiotics. Furthermore, if the strain responsible for the infection is resistant to antibiotics commonly used in the clinical practice, the therapeutic options are further reduced. Recently, bacteriophages, viruses that have bacteria as their only target, are at the center of a renewed interest of the scientific community, representing an alternative antimicrobial strategy potentially capable of eradicating even persistent infections sustained by multi-antibiotic-resistant microorganisms. In this context, aim of this thesis was to evaluate the ability of Sb-1 bacteriophage, commercially available in Georgia (the only country in which phage therapy is approved), to control the infection associated with biofilm formation due to methicillin-resistant S. aureus in vivo, in a Galleria mellonella larval model of implant-associated infection. In particular, short fragments of Kirschner wires (K-wires), thin steel pins that are used in orthopedic hand surgery, were inserted in the larva pro-leg. Subsequently, the larvae were infected with the methicillin-resistant laboratory strain of S. aureus (ATCC43300). This model was useful for assessing the ability of Sb-1 (tested alone and/or in combination with antibiotics) to prevent colonization and treat the infection of the implant by S. aureus showing its efficacy in rescuing larvae from infection. In order to assess the potential intravenously administration in humans, the ability to lyse bacteria by Sb-1 has also been assessed in decomplemented human serum. The loss of lytic capacity of phages in serum suggested the development of a controlled phage release system located at the site of infection. For this reason, release studies of Sb-1 phages from injectable and thermosensitive hydrogels have been carried out, since the latter are capable of jelling at human body temperature, which could, therefore, favor a prolonged release of viruses to therapeutic viral titers at the infection site. In conclusion, a G. mellonella model of implant-associated infection was developed and it might be used in the future to test the antibiofilm efficacy of different compounds and antimicrobial strategies. Moreover, the ability to form biofilm by other human pathogens can be also tested. Finally, the antibiofilm activity of Sb-1 previously observed in vitro was also confirmed in vivo, suggesting the possibility to use it alone or in combination with antibiotics for either the treatment or prevention of staphylococcal infection of medical implants