Isolation and In Vitro Evaluation of Bacteriophage against Methicillin-Resistant Staphylococcus aureus (MRSA) from Burn wounds

Abstract

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is a major threat to human health. It is considered to be a Multi-Drug Resistant (MDR) pathogen capable of causing a variety of diseases. MRSA is also one of the most important nosocomial pathogens in burn infection. As a treatment strategy against MRSA infections phage therapy has the potential of becoming alternative remedy. Thus, the aim of present study was to isolate and characterize lytic bacteriophage from hospital sewage effective against burn wounds infecting MRSA isolates. Materials and methods: Staphylococcus aureus strains were isolated from hospitalized burnt patients. Strains were confirmed as MRSA by Kirby-Bauer disk diffusion tests to penicillin, methicillin, and oxacillin and also PCR assay for mecA gene. Phage was isolated from hospital sewage sample and tittered by Double Layer Agar (DLA) method. Spot test was used for host range determination. The latent period and burst size were estimated from one step growth curve. Phage morphology was observed by electron microscopy. The nature of the nucleic acid of isolated bacteriophage was confirmed by using Rnase A, Dnase I and 6 restriction enzymes. Results: The titer, latent period and burst size of the isolated phage was determined to be 1×109 PFU/ml, 20 min and 190 PFU per infected cell, respectively. It displayed a wide host range for MRSA bacteria by spot test (27 isolate from 30).Electron microscopy observation demonstrated that the phage belonged myoviridea family. Digestion profiles of Rnase A, Dnase I and XbaI restriction enzyme on 1% agarose gel Electrophoresis, show that the genome of isolated phage was a double-stranded DNA with a size of <∼43 kbp. Conclusion: The isolated phage was active against a wide range of MRSA strains recovered from burnt patients. Therefore it can be considered as a suitable candidate for use in prophylaxis and treatment of these clinical infections and for the preparation of anti-staphylococcal cocktails.