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To determine the effect of short-term topical therapy with 1% prednisolone acetate (PA) on normal immune adenoviral clearance in the rabbit ocular model.Thirty rabbits were topically inoculated in both eyes with adenovirus type 5 (Ad5). On day 1, the rabbits were divided into three topical treatment groups: 1% PA four times daily for 3 days, 1% PA four times daily for 5 days, control (artificial tears) four times daily for 5 days. Eyes were cultured for virus on days 0, 1, 3, 4, 5, 7, 9, 11, and 14.Compared with the control group, treatment with 1% PA for 3 or 5 days significantly increased the total and daily number of Ad5-positive cultures from days 7 to 14, prolonged the duration of Ad5 shedding, and increased the mean combined Ad5 titer from days 1 to 5. In addition, treatment with 1% PA for 5 days increased the mean combined Ad5 titer from days 7 to 14.Treatment of an ocular adenoviral infection with 1% PA for as little as four times daily for 3 days significantly enhanced adenoviral replication compared with the control group. Short-term corticosteroid treatment of acute adenoviral ocular infections with 1% PA should be used judiciously.