The ability of wild-type and mutant strains to adhere to scarified corneal epithelium was determined essentially as described by Gupta et al.
44 All animals used in this study were treated in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. Female 10- to 12-week-old 129/Sv mice
45 were anesthetized with isoflurane (Mallinckrodt Veterinary, Inc., Mundelein, IL) and killed by cervical dislocation. Corneas of each eye were scarified with a sterile 26-gauge needle. Eyes were then enucleated and placed into organ culture, as previously described.
46 Eyes (three per strain) were inoculated with 5 μL 10
7 colony-forming units (CFU) of bacteria on the corneal surface. After incubation for 1 hour at 37°C, eyes were rinsed to remove nonadherent bacteria and prepared for scanning electron microscopy, as previously described.
44 Eyes were examined with a scanning electron microscope (JSM-840A; JEOL, Tokyo, Japan), and bacteria adhering to corneal wounds were quantitated as described by Hazlett et al.
30 In brief, five representative fields were photographed from each group of three eyes at a magnification of ×3000. Negatives from these five photographs were enlarged to ×6000 m and three fields (80 mm
2 each) were counted on each photograph. Counts are presented as the mean number of bacteria bound per field ± SEM. Statistical comparisons between strains were conducted by an unpaired, two-tailed Student’s
t-test (
n = 15/strain) at
P ≤ 0.05 significance.