The Effect of Temperature on the Antimicrobial Activity of Optisol-GS

Abstract

To determine the survival of different bacteria inoculated in Optisol-GS at refrigerated storage temperature (6 degrees C) and after subsequent warming to room temperature (19-22 degrees C). Staphylococcus aureus, Enterococcus faecium, Streptococcus pneumoniae, and Pseudomonas aeruginosa were chosen from stock clinical isolates for inclusion in the study. The first group consisted of 12 Optisol-GS vials. The second group consisted of 12 Optisol-GS vials containing corneas inappropriate for transplantation according to the Eye Bank Association of America (EBAA) protocols. Each group was inoculated with 3 concentrations of approximately 10, 10, and 10 colony-forming units (CFU)/mL of each bacterial species and then refrigerated per EBAA protocol. After 48 hours of refrigeration, all vials were placed at room temperature (RT) and counts were performed at 48, 50 (2 hour RT), 54 (6 hour RT), 60 (12 hour RT), 72 (24 hour RT), and 96 (48 hour RT) hours. At 96 hours, the corneal tissue from 10 and 10 inocula were cultured. All samples underwent serial dilution, spiral plating on blood agar plates, and incubation at 35 degrees C. Viable colony counts were determined at 24 hours. Except for the 10 CFU/mL inocula of P. aeruginosa, all isolates were viable after 48 hours of refrigeration. Rapid bactericidal activity was observed against P. aeruginosa after 2 hours at RT, with complete sterilization by 6 hours. The rate and extent of killing against S. aureus were influenced by the initial inoculum. Bactericidal activity was achieved after 2 hours at RT with 10 CFU/mL of S. aureus versus 24 hours with the 10 inoculum. Of note, bactericidal activity was not observed against S. pneumoniae and E. faecium following 24 hours of storage at RT. The presence of corneal tissue did not affect viable counts, with counts from corneal tissue cultures reflecting the counts seen from Optisol-GS after 48 hours at RT. The antimicrobial activity of Optisol-GS was reduced at refrigerated temperature and enhanced at RT. Bactericidal activity was not observed against E. faecium at either refrigerated temperature or RT.