During the course of frog virus 3 (FV3) infection in baby hamster kidney 21 (BHK) cells, vimentin-type intermediate filaments reorganize to surround the virus's cytoplasmic assembly sites. To determine whether the association between vimentin filaments and viral assembly sites has a functional role in the virus life-cycle, we treated cells with the antimicrotubule drugs taxol or colchicine, or injected them with monoclonal antivimentin antibodies prior to FV3 infection. Each of these reagents caused the collapse of the normally extended BHK intermediate filament system. In the case of taxol-treated or antivimentin-injected cells, the collapsed vimentin filaments were unable to reorganize around the newly forming viral assembly sites. The viral assembly sites that did form were aberrant and there was a significant reduction in the number of mature virions present. Colchicine, which also caused the collapse of vimentin filament organization, did not block the reorganization of vimentin filaments in response to viral infection and viral assembly sites appeared normal. These results suggest that intermediate filaments play an important role in maintaining the structural and functional integrity of FV3 assembly sites.