Mycoplasma pneumoniae, Acholeplasma laidlawii, M. arthritidis, and M. pulmonis were shown to induce interferon in the lymphocyte fraction of ovine peripheral blood leukocytes, but not in the polymorphonuclear leukocyte fraction. Human peripheral blood lymphocytes produced significant levels of interferon in response to infection with M. pneumoniae and M. synoviae. The antiviral substance induced by the mycoplasmas in human lymphocytes was characterized as interferon by the usual criteria.