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An apparatus consisting of a Dewar flask and a relay system controlling the flow of liquid nitrogen permitted the irradiation of samples in tin cans or Pyrex tubes at temperatures ranging from 0 ± 1.5 C to -194 ± 2 C. An inoculated pack comprising 320 cans of ground beef containing 5 × 104 spores ofClostridium botulinum 33A per can (10 cans per radiation dose) was irradiated with Co60 at 0 and -196 C. Incubation was carried out at 30 C for 6 months. Approximately 0.9 Mrad more radiation was required to inactivate the spores at -196 C than at 0 C. Cans irradiated at -196 C showed partial spoilage at 3.6 Mrad and no spoilage at 3.9 Mrad; the corresponding spoilage-no spoilage doses at 0 C were 2.7 and 3.0, respectively. The majority of positive cans swelled in 2 to 14 days; occasional swelling occurred as late as 20 days. At progressively higher doses, swelling was delayed proportionally to the radiation dose received. The remaining nonswollen cans had no toxin after 6 months of storage, although occasional cans contained very low numbers of viable spores comprising on the average 0.1% of the original spore inoculum. TheD 10 values in phosphate buffer were 0.290 Mrad for 0 C and 0.396 Mrad for -196 C; in ground beef, the correspondingD 10 values were 0.463 Mrad and 0.680 Mrad, respectively. TheseD 10 values indicate that the lethal effect of γ rays decreased at -196 C as compared with 0 C by 13.5% in phosphate buffer, and by 47% in ground beef.

Effect of Temperature of Liquid Nitrogen on Radiation Resistance of Spores of Clostridium botulinum