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SUMMARY: Non-toxic concentrations of various substances were tested for their influence on the gliding motility of Mycoplasma mobile 163K. A significant inhibitory effect on motility was observed with agents acting on nucleic acid synthesis (mitomycin), protein synthesis (puromycin, chloramphenicol), energy metabolism (p-chloromercuribenzoate, iodoacetate) and with compounds reacting with the cytoplasmic membrane or contractile elements (albumin, cholesterol, EDTA, 2-propanol, procain, CaCl2, MgCl2, colchicin and KI). The surface-active compounds Triton X-100, Tego and SDS increased the gliding velocity significantly in some concentrations and incubation periods. The results suggest that the motility of M. mobile depends on a functional cytoplasmic membrane and that cytoskeletal elements are involved in the gliding mechanism.