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R. Kapur, E.Y. Tu, S.L. Pendland, J. Sugar; The Effect of Temperature on Microbial Activity in Optisol–GS . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4908.
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Purpose: To determine the survival of different bacteria inoculated in Optisol–GS at current storage temperature (6o C) and at room temperature (22o C). Methods: S. aureus, E. faecium, S. pneumoniae, and P. aeruginosa were chosen from stock clinical isolates for inclusion in the study. Group A consisted of 12 Optisol–GS vials, and Group B consisted of 12 Optisol–GS vials containing corneas deemed inappropriate for transplantation according to EBAA protocols. Each group was inoculated with three concentrations (106, 105, 104) of each of the bacterial species. The final inoculum was prepared via spectrophotometric methods and confirmed by viable colony counts. All 24 vials were refrigerated per EBAA protocol. A small sample was taken at 24 hours to determine viability. After 48 hours, all vials were placed at room temperature. Sampling for viable counts was performed at 48, 50, 54, 60, 72, and 96 hours. At 96 hours, Group B corneas containing 106 and 104 starting inoculi were grinded and cultured. All samples underwent serial dilution (10–1 to 10–4) to minimize antibiotic carryover. The diluted samples and undiluted samples were spiral plated onto blood agar plates. The plates were incubated at 35oC, and viable colony counts were determined at 24 hours. An uninoculated vial from each group was tracked to determine the time required to reach temperature equilibration. Results:With the exception of the lowest concentration of Pseudomonas, all isolates were viable after refrigeration. Rapid bactericidal activity was observed against Pseudomonas after 2 hours at room temperature. The rate and extent of killing against S. aureus were influenced by the initial inoculum. Of note, S. pneumoniae and Enterococcus showed significant resistance. Time–kill curves of each individual bacterium will be presented showing the viability of organisms in Optisol–GS following refrigeration for 48 hours and subsequent incubation at room temperature for 48 hours. No effect on colony counts was seen by the presence of corneal tissue. Cultures of the corneal tissues grew 1 colony of Pseudomonas and 69 colonies of S. pneumoniae from the high inoculation vials. S. aureus did not grow in the tissue cultures. However, heavy growth of Enterococcus was observed from those corneas that were inoculated with the 104 and with the 106 starting inoculum. Conclusions: The antimicrobial activity of Optisol–GS was reduced at refrigerated temperatures. Bactericidal activity was not observed against S. pneumoniae and Enterococcus at the temperatures tested. Recommendations on tissue handling with regard to temperature will be discussed.
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