The study evaluated antiviral efficacy of antibiotics- Doxycycline and Kanamycinagainst Japanese encephalitis virus (JEV) infection in vivo. Adult Swiss albino mice (4- 6weeks) were used. Mice were distributed in four groups- control group (A), only drug group(B), JEV infected group (C) and JEV + drug treated group (D). Mice were given intravenousinoculation of JEV strain P20778. Doxycycline was given via intra peritoneal (i. p.) route at 50mg/kg dose. Kanamycin was given to mice via subcutaneous (s. c.) route at 20 mg/kg dose. Alldrug dosages were administered at 24 hr, 48 hr and 96 hr post infection (p.i.) twice a day (BID)for upto 14 days. The mice were monitored for 21 days. The viral load was determined byplaque assay. Viral RNA load and cytokine levels were determined. The infected mice died by8 days of infection. Doxycycline treatment at 50 mg/kg dose after 24 hr p.i. lowered diseaseprogression, prolonging the survival of the animals by a week. Antiviral effect was evidentwith reduction of progeny plaque formation. The plaque formation was reduced at 24 hr p.i.compared to virus group. Doxycycline inhibited viral RNA replication. Doxycycline was ableto moderately modulate proinflammatory cytokines. Kanamycin administration was lesseffective. Thus, the studies demonstrated encouraging results in treatment of JEV infectionby Doxycycline. It indicated that Doxycycline delayed the disease progression. Thus, thefindings suggest that Doxycycline could be used as an adjuvant treatment against JE.