In vitro model of infectious crystalline keratopathy: Tissue architecture determines pattern of microbial spread

PURPOSE. TO develop an in vitro model of infectious crystalline keratopathy using human corneal buttons and to test the hypothesis that the compactnes s of the corneal stroma determines the pattern of microbial spread. METHODS. Twenty human corneal buttons obtained after penetrating keratoplas ty for keratoconus (KC) and eight human corneal buttons obtained from eye b ank (EB) donor eyes were maintained in organ culture. Fourteen buttons (10 KC and 4 EB donors) were maintained ill a turgid state (swollen, edematous) and 14 in a nonturgid state (compact, normal state of deturgescence) by th e omission or addition of 5% dextran to the culture medium. Eight KC and fo ur EB nonturgid buttons and eight KC and four EB turgid buttons were inocul ated with Streptococcus viridans (Lancefield group Cr, gram-positive) organ isms. Two KC nonturgid and two KC turgid buttons were inoculated with Klebs iella oxytoca (gram-negative) organisms. Bacterial migration and spread in the tissue were observed by light and electron microscopy. RESULTS. Of the nonturgid buttons, six KC buttons and all four EB buttons i noculated with S. viridans and both KC buttons inoculated with K. oxytoca d emonstrated an arborizing, crystallike pattern of bacterial spread. In the turgid buttons, five KC and all four EB buttons inoculated with S. viridian s and both KC buttons inoculated with K. oxytoca demonstrated globular, amo rphous colonies. This was in complete contrast to the needlelike branching appearance seen in nonturgid corneal buttons. Electron microscopy confirmed an interlamellar spread of the bacterial colonies. CONCLUSIONS. This is the fil:st in vitro model of bacterial keratitis. It d emonstrates that the pattern of spread of bacteria within corneal tissue is largely determined by the compactness of the corneal stroma. Altering tiss ue architecture changed the pattern of bacterial migration and spread. This model has considerable potential in further understanding host-microbe int eractions and microbial spread that occurs during infection.