Botulism. 1 A method for determining the thermal death time of the spores of Bacillus botulinus

Abstract

In the course of a series of experiments dealing with the determination of the thermal death point of spores of Bacillus botulinus in which the method of procedure recommended by Bigelow and Estey 2 was followed with minor modifications, it was found that in the daily transplanting of several hundred specimens to the tubes in which the heated material was to be incubated, it was inevitable that a certain small percentage of the tubes became contaminated. The number of proved contaminations was not large, less than 1 per cent. in a test of approximately 2,000 tubes, but because of the fact that one could not be absolutely certain that any particular tube was free from contamination, it was impossible to draw accurate conclusions in any instance where an unusual survival time was indicated. It was therefore imperative that a method be devised in which the necessary number of tubes per day could be handled with rapidity, and, at the same time, with absolute protection against any possibility of contamination after their contents had been subjected to the heating process.

After a number of trials we have adopted the following method of procedure in our experiments, and, although it cannot be adapted to the investigation of all the problems which present themselves in thermal death time experiments, it has proved to be satisfactory for the problems under immediate investigation.

Soft glass tubes, 10 × 150 mm., are used in the experiments. Three cubic centimeters of 1 per cent. glucose peptic digest liver broth, adjusted so that the final P_H is between 7.3 and 7.5, are placed in each tube and covered with a thin layer of oil to prevent evaporation. The medium is sterilized at fifteen pounds pressure for thirty minutes.