Phage therapy presents an alternative approach against the emerging methicillin-resistant Staphylococcus aureus (MRSA) threat. Some of the problems encountered during isolation of MRSA phages include the high prevalence of enteric phages in natural sources, nonspecific absorption of viable phage, and the formation of pinpoint or tiny plaques. The phage isolated in this study, MR-5, also formed tiny plaques against its host S. aureus ATCC 43300 (MRSA), making its detection and enumeration difficult. An improved method of increasing the plaque size of MRSA phage by incorporating sublethal concentrations of three different classes of antibiotics (inhibitors of protein synthesis) in the classical double-layer agar (DLA) method was investigated. The -lactam and quinolone antibiotics commonly employed in earlier studies for increasing the plaque size did not show any significant effect on the plaque size of isolated MR-5 phage. Linezolid (oxazolidinone class), tetracycline, and ketolide antibiotics brought significant enhancements (3 times the original size) in the plaque size of MR-5 phage. Prior treatment with these antibiotics resulted in significant reductions in the time of adsorption and the latent period of MR-5 phage. To rule out whether the action of linezolid (which brought the maximum increase in plaque size) was specific for a single phage only, its effect on the plaque size of seven other S. aureus-specific phages was also assessed. Significant enhancements in the plaque size of these phages were observed. These results indicate that this modification can therefore safely be incorporated in the traditional DLA overlay method to search for new MRSA-virulent phages.
Staphylococcus aureus is a leading cause of both communityand hospital-acquired infections, including skin and soft tissue infections, wound infections, endocarditis, osteomyelitis, and lifethreatening bacteremia (6, 23). The emergence of high levels of penicillin resistance, followed by the development and spread of strains resistant to the action of nafcillin, oxacillin, methicillin, and now, even vancomycin has made therapy of staphylococcal disease a global challenge (23,24,34). More people in the United States now die from methicillin-resistant S. aureus (MRSA) infection than from AIDS (33). Hence, there is an unmet medical urgency and pressing need to develop novel antibacterials to tackle the staphylococcal menace. Phage therapy can serve as one of the stand-alone approaches for treating staphylococcal infections refractory to the action of commonly deployed antibiotics (6).The first step in investigating the potential of therapy using phage or its products against MRSA infections involves the isolation and selection of a lytic bacteriophage covering a maximum number of MRSA strains, both from community and hospital sources. The difficulty encountered in the isolation of phages specific to staphylococcal strains is partly attributed to the prevalence of enteric phages in sewage samples and partly to the high level of nonspecific adsorpti...