Scanning Electron Microscopy Visualization of Methicillin-Resistant Staphylococcus aureus After Contact With Gatifloxacin With and Without Preservative

Abstract

To test a visual model by looking at the differences in effect of Zymar((R)) (gatifloxacin plus benzalkonium chloride [BAK]) when compared to gatifloxacin and a normal saline (NS) control upon a methicillin and gatifloxacin-resistant Staphylococcus aureus (MRSA) species. An ocular isolate of gatifloxacin-resistant (minimal inhibitory concentration >2 to 4 microg/mL) MRSA was grown to confluency. Chambered slides were prepared with bacterial culture smears, and then incubated with either gatifloxacin at the concentrations of 1 and 10 microg/mL, Zymar containing equivalent concentrations of gatifloxacin, or NS. Bacterial cultures were fixed after 10, 30, and 60 min. Fixed slides were coated in gold sputter for examination. Bacteria were visually evaluated with scanning electron microscopy (SEM) at 50,000x. Blinded review of SEM images compared structural changes and mitotic activity across samples. MRSA exposed to 10 microg Zymar for 60 min showed significantly greater pleomorphism and cell wall surface changes when compared to gatifloxacin (P < 0.0001) and NS (P = 0.001), and significantly less mitotic activity than NS (P = 0.002). Using SEM, the topical formulation of gatifloxacin 0.3% (Zymar), which contains BAK, had greater antibacterial activity than did gatifloxacin alone in gatifloxacin and methicillin-resistant S. aureus, thereby illustrating potential advantages of the preservative in the commercial formulation. We further show that these effects can be visualized and quantified.