Cultures of adherent and non-adherent bovine blood mononuclear cells were infected with bluetongue virus (BTV) serotype 10. Production of BTV proteins in mononuclear cell cultures was detected by immune precipitation of viral proteins from [35S]methionine-labelled extracts of these cells, by immunofluorescence staining of cells using monoclonal antibodies (MAbs) to BTV proteins VP7 and NS2, and by flow cytometry with MAbs to VP2, VP7, NS1 and NS2. BTV-infected cells were most numerous in cultures of adherent mononuclear cells; infected cells were initially identified as monocytes on the basis of their morphology, and size and scatter characteristics as determined by analysis with a fluorescence-activated cell sorter (FACS). The majority of adherent mononuclear cells with these scatter characteristics were confirmed to be monocytes by FACS analysis with a MAb specific for bovine monocytes. Identification of BTV-infected adherent mononuclear cells as monocytes was further established by double immunofluorescent labelling, as infected adherent cells reacted with the MAb specific for bovine monocytes, and with another MAb specific for class II antigen. Infection of adherent mononuclear cells was also confirmed by transmission electron microscopy, as BTV virions and tubules were present in lysates of cultures of BTV-infected adherent mononuclear cells and within the cytoplasm of adherent cells. In contrast, BTV proteins were detected in few cells identified as lymphocytes on the basis of their scatter characteristics, and mean fluorescence of such cells was considerably less than that of BTV-infected monocytes. Viraemia persisted until 35 days after inoculation of a colostrum-deprived calf inoculated with BTV. Virus was isolated from blood mononuclear cells at 1 week after infection of the calf, but not thereafter. BTV infection of blood mononuclear cells was demonstrated until 9 days after inoculation by indirect immunofluorescence staining of mononuclear cells. In contrast, virus was consistently isolated from erythrocyte-enriched preparations throughout viraemia in titres comparable to those in whole blood. These results indicate that although bovine monocytes are readily infected in vitro with this strain of BTV serotype 10, infection of blood monocytes is unlikely to be responsible for the prolonged viraemia that consistently occurs in BTV-infected cattle.