A study was made of antiviral activities in vitro of 54 derivatives of N-phenyl-N′-aryl or alkylthiourea with a basic structural formula C6H5NHCSNHR against poliovirus 1 (Mahoney). Eleven active compounds were selected by the Screening gradient-plaque-inhibiting test, some of which (N-phenyl-N′-4-carboxy-5-hydroxyphenylthiourea, N-phenyl-N′-3-hydroxyphenylthiourea) in a one-step growth cycle inhibit 99.4–99.9% of the virus yield. It was noted that, along with the -HN-CS-NH- group, the presence and spatial location of the hydroxyl group in the aryl or alkyl radical at N′-atom is of importance to antiviral