Infection by Bluetongue virus is acid activated and can be inhibited by raising endosomal pH. Consistent with these observations, Forzan et al [45] concluded that BTV-10 infection of Vero and Hela cells is mediated by CME, with capsid disassembly and membrane penetration occurring from within early-endosomes. In this report, we have confirmed that active endosomal acidification is required for BTV-1 infection of BHK cells. However, in contrast to previous studies we found that the clathrin pathway is not the major entry route used by BTV-1 to infect BHK cells. This conclusion is based on a number of observations, including; (1) although a low endosomal pH was shown to be essential for infection, the kinetics of BTV-1 delivery to acidic compartments was slower than expected for uptake by the clathrin pathway [53] ; (2) during entry, BTV-1 did not colocalise with transferrin, a commonly used marker for CME; and (3) cell-entry and infection was not inhibited by expression of three different DN inhibitors of CME (AP180C, DN-Eps15 and DNdynamin-2). Furthermore, both EIPA and cytochalasin-D were shown to inhibit uptake of BTV-1 but not uptake of transferrin indicating that BTV-1 and transferrin are internalised by different endocytic pathways.