Metabolic Injury to Bacteria: I. Effect of Freezing and Storage on the Requirements of Aerobacter Aerogenes and Escherichia Coli for Growth

Various complex supplements including Bacto-casamino acids when added to a minimal salts glucose medium increased the bacterial count on suspensions of cells of both Aerobader aerogenes and Escherichia coli frozen and stored for 3 weeks. Replica plating procedures established that no auxotrophic mutants had been formed. Attempts to fractionate Casamino acids revealed that the active material could be retained by and eluted from both acidic and basic ion exchange resins. A mixture of 18 crystalline amino acids was equally as good as Casamino acids as a supplement to the minimal medium for A. aerogenes. Cystine was the most effective of the amino acids in the mixture tested singly, though other amino acids singly and in combination also increased the bacterial count. Cysteine was even more active than cystine and as effective as the Casamino acids for both organisms. Ethylenediaminetetraacetic acid had some activity for A. aerogenes but little for E. coli. Cells frozen and stored were more sensitive to added phosphate in the medium than unfrozen cells. It is postulated that freezing and storage damage the cytoplasmic membrane of a proportion of the cells rendering them more penetrable by toxic metal ions present as trace contaminants. The enriched medium would permit growth of damaged cells by providing compounds capable of chelating the toxic ions.