This study was done to develop a method to inactivate Escherichia coli O157:H7 on radish and cabbage seeds using simultaneous treatments with gaseous chlorine dioxide (ClO₂) and heat at high relative humidity (RH) without decreasing seeds’ viability. Gaseous ClO₂ was spontaneously vaporized from a solution containing hydrochloric acid (HCl, 1 N) and sodium chlorite (NaClO2, 100,000 ppm). Using a sealed container (1.8 L), an equation (y = 5687×, R² = 0.9948) based on the amount of gaseous ClO₂ generated from HCl–NaClO₂ solution at 60 ^◦C and 85% RH was developed. When radish or cabbage seeds were exposed to gaseous ClO₂ at concentrations up to 3,000 ppm for 120 min, germination rates did not significantly decrease (P > 0.05). When seeds inoculated with E. coli O157:H7 were treated with 2,000 or 3,000 ppm of gaseous ClO₂ in an atmosphere with 85% RH at 60 ^◦C, populations (6.8–6.9 log CFU/g) on both types of seeds were decreased to below the detection limit for enrichment (− 0.5 log CFU/g) within 90 min. This study provides useful information for developing a decontamination method to control E. coli O157:H7 and perhaps other foodborne pathogens on plant seeds by simultaneous treatment with gaseous ClO₂ and heat at high RH.