Studies on the Death of Bacteria at Low Temperatures

Abstract

Suspensions of E. coli in 1% peptone and in peptone-buffer mixture at pH 7 were subjected to various low temp. treatments and extensive quantitative data on mortality following such treatments obtained. Comparison of death rates at different stages in the process of freezing indicated that death by freezing involves a rapid-acting or "immediate death" due to freezing and thawing per se and a "storage death" which continues after ice formation is completed and is a direct function of time and temp. Mortality resulting from "immediate death" by freezing was marked but did not vary with the intensity of the freezing temp. " Immediate death" apparently occurs at a brief stage in the freezing process during which extracellular ice formation is being completed. The rate of "storage death" at the higher freezing temps. was found to be very rapid. It was much greater at temps. above -30[degree] C. than at temps. of -30[degree] C. and below. This difference was regarded as being due to eutectic change. It was observed that repeated freezing is more lethal than a single freezing or storage in the frozen state for a similar interval of time. Freezing proved to be profoundly more lethal than supercooling. Repeated fluctuation of temp. of frozen suspensions through various low temp. ranges from[long dash]1.5[degree] C. to[long dash]195[degree] C. did not exert a lethal action additional to that of storage. However, it was found that repeated fluctuation of temp. of frozen suspensions between -30[degree] C. and -78[degree] C. resulted in a lower mortality than storage at either temp. The authors suggest that this lower mortality may be the result of physical effects such as gas exchange, which promote survival of the cell. "Storage death" at[long dash]195[degree] C. either does not take place or is so slow that it was not detected within the storage period of 10 hrs. studied. The data presented strongly support the theory that death due to freezing results from the formation of extracellular ice. No evidence was found to indicate that intra-cellular ice is formed or exerts lethal action. The death incident to the formation of extracellular ice is apparently the result of both direct and indirect effects. The direct action ("immediate death") is regarded as being mechanical and the indirect action ("storage death") largely the result of concentration of solutes in the intercrystallic film.