Chemical inhibition of alpha-toxin, a key corneal virulence factor of Staphylococcus aureus.

PURPOSE: alpha-Toxin mediates extreme corneal damage during Staphylococcus aureus keratitis. Chemical inhibition of this toxin was sought to provide relief from toxin-mediated pathology.

METHODS

Inhibition of alpha-toxin by phosphate-buffered saline (PBS), 0.1% methyl-beta-cyclodextrin (CD), or CD plus cholesterol (0.1%, CD-cholesterol) was assayed by hemolysis of rabbit erythrocytes. Pathologic changes in rabbit corneas injected with 12 hemolytic units of alpha-toxin suspended in PBS, 1% CD, or 1% CD-cholesterol were compared over time. Rabbit corneas injected with 10(2) colony forming units (CFU) of S. aureus were treated from 7 to 13 hours postinfection (PI) with a total of 15 drops of CD-cholesterol, CD, or PBS. Slit lamp examination (SLE) and measurement of erosions were performed at 13 hours PI and bacteria were quantified at 14 hours PI.

RESULTS

Toxin-mediated lysis of erythrocytes was inhibited up to 16,000-fold in the presence of CD-cholesterol compared with CD or PBS. Eyes injected with alpha-toxin mixed with CD-cholesterol had, at 7 hours postinjection, significantly smaller erosions than eyes injected with alpha-toxin in PBS or alpha-toxin mixed with CD (P = 0.0090 and P = 0.0035, respectively). Eyes infected with S. aureus and treated with CD-cholesterol had significantly lower SLE scores than eyes treated with CD or PBS (P <or= 0.0103 and P <or= 0.0017, respectively); however, there were no differences in the number of bacteria present (P >or= 0.0648).

CONCLUSIONS

CD-cholesterol is a potent inhibitor of alpha-toxin activity in vitro and an effective means to arrest corneal damage during S. aureus keratitis.