The antiviral activities of 9-(2-hydroxyethoxymethyl)guanine (acyclovir; ACV) either alone or combined with recombinant human leukocyte (alpha) A/D interferon (rHuIFN-alpha) against feline herpesvirus type 1 (FHV-1) were evaluated in feline embryo cell cultures, using an infectivity-inhibition assay. In ACV-treated cultures, the 50% inhibitory dose (ID50) was approximately 10 to 20 micrograms of ACV/ml. Maximal inhibition of FHV-1 infectivity (range, 3.4 to 4.2 log10 TCID50) was observed when high test doses of ACV (125 or 250 micrograms/ml) were given 1 to 6 hours after infection. Although mild inhibition (range, 0.3 to 1.6 log10 TCID50) of virus was observed at lower drug doses (10 to 62.5 micrograms/ml), FHV-1 was relatively resistant to ACV and required higher minimal inhibitory doses than those reported for other herpesviruses. However, when ACV was combined with 10 or 100 U of rHuIFN-alpha/ml, synergistic antiviral effects were associated with ACV dosage of 10 to 62.5 micrograms/ml. Antiviral activities resulting from use of the combined drugs permitted nearly eightfold reduction in the dose of ACV required to achieve maximal inhibition of FHV-1. Significant (P less than 0.01) synergistic interactions with ACV resulted when the rHuIFN-alpha was given before or after infection; at the lower doses of ACV, however, rHuIFN-alpha pretreatment was more effective. Although dosages of either greater than or equal to 62.5 micrograms of ACV/ml or 100 U of rHuIFN-alpha/ml were cytosuppressive in control cell cultures, additive anticellular effects were not observed at synergistic combinations of ACV and 10 U of rHuIFN-alpha/ml.