The preservation of bacteriophage H1 of Corynebacterium ulcerans U103 by freeze-drying.

Abstract

SUMMARYThis paper describes an investigation into the successful preservation at room temperature of the bacteriophage H1 ofCorynebacterium ulceransU 103 which was extremely labile when in suspension at 4° C.Cooling at a rate of 1° C./min. showed that the survival decreased logarithmically at temperatures between – 14 and – 45° C. Survival of broth suspensions of the corynebacteriophage were found to increase proportionally with an increase in the rate of cooling though there was a marked drop in survival at rates of approximately 900° C./min. The addition of peptone solutions was found to increase the survival over the range studied, whereas the addition of sucrose solutions had only a slight effect.By avoiding freezing damage by cooling at rates of 450° C./min. in (a) 20% peptone solution, (b) 20% peptone and 10% sucrose, and (c) 20% peptone, 10% sucrose and 2% sodium glutamate, a study was made of the drying stage of the freeze-drying process. On drying at controlled temperatures it was found that there was no damage on rewarming to temperatures below – 21° C. after cooling to – 196° C., but that the survival immediately after drying in the absence of glutamate, showed a logarithmic relationship with the temperature of drying, lower temperatures giving better survival.On storage for a period of 3 months at room temperaturein vacuoand darkness, there was no appreciable loss in survival in the mixtures though suspensions in peptone alone showed a slight decrease. At higher temperatures this decrease in survival could be differentiated into two types of damage, each of which could be influenced by the presence of sucrose or glutamate.We are grateful to Professor H. R. Carne for providing the initial sample of bacteriophage H 1 ofCorynebacterium ulceransU 103, and to Professor R. I. N. Greaves for his interest and encouragement during the course of this investigation.