Studies are presented on the effect of cytochalasin B (CB) on the growth of five Mycoplasma species, three Acholeplasma species, and one Spiroplasma species. The three gliding mycoplasma species (M gallisepticum, M pneumoniae and M pulmonis are the only mycoplasmas inhibited by CB. These are the only prokaryotes reported to be inhibited by CB. This suggested that these three mycoplasmas might have some sort of cytoskeletal structure. A protein fraction has been isolated from M gallisepticum which polymerizes in 0.6 M KCl and depolymerizes when KCl is removed. This fraction contains a major 58,000-dalton protein, a 46,000-dalton protein, and a minor 87,000-dalton protein.