The poxvirus Cotia was studied by electron microscopy and by serological and biochemical analyses. Thin-sectioned preparations of infected Vero cells indicated that Cotia virus morphogenesis was similar to other mammalian poxviruses; unique filamentous structures and inclusion matrices were apparent in the cytoplasm. Complement fixation tests that included purified Cotia virions showed a reciprocal cross-reaction with rabbit myxoma virus and no cross-reaction with vaccinia virus. Serological results coupled with gradient polyacrylamide gel electropherograms of the structural proteins of purified Cotia, vaccinia, myxoma and fibroma viruses suggested that Cotia virus was similar to the latter two viruses. Agarose gel electropherograms of cleavage fragments of each of these virus DNAs digested with three separate restriction endonucleases showed that each of these viruses had a unique DNA gel profile.