Metabolic injury to bacteria. II. Metabolic injury induced by distilled water or Cu++ in the plating diluent.

When distilled water from a tin-lined still served as the plating diluent, cells of Aerobacter aerogenes developed symptoms of metabolic injury as evidenced by increased counts on supplemented, as compared with minimal, plating medium. Cysteine was as effective as yeast extract as a supplement to the minimal medium in increasing the viable count. Mg(++) and, to a lesser extent, phosphate buffer at the concentrations tested protected unfrozen cells, but not cells which had been frozen and stored, against the loss of capacity to grow on minimal medium. When the plating diluent consisted of distilled water redistilled in an all-glass still, the symptoms of metabolic injury did not appear. Spectrographic analysis revealed the presence of 10(-7)m Cu(++) in the distilled water, and Cu(++) added to redistilled water serving as the plating diluent reproduced the metabolic injury effects induced by distilled water. It was concluded that freezing and storage damaged the cell membrane, rendering it more penetrable by toxic elements which were thereby enabled to act at sites in the cell where Mg(++) and other solutes in the plating diluent could not serve as effective antagonists. Increased recovery of cells on supplemented medium could be ascribed to the capacity of the supplements to remove toxic elements which had become bound to the cells during suspension in the plating diluent.