Effects of Escherichia coli spheroplast formation on assays of H2 and adenosine triphosphate based ampicillin susceptibility tests.

The present study examined the effects of ampicillin on one strain of Escherichia coli in lactose peptone broth with an osmolality of 342 mosm/L under anaerobic conditions. Spheroplast formation occurred at 10 X MIC of ampicillin. The metabolic changes that took place during spheroplast formation disfavored the production of molecular hydrogen. The intracellular bacterial adenosine triphosphate (ATP) level remained normal or slightly elevated during spheroplast formation while viability (cfu/ml) decreased. Thus spheroplast formation did not interfere significantly with ampicillin susceptibility as interpreted by assaying molecular hydrogen and viability. The effect on the ATP assay was, however, pronounced. It was found that the reversion of spheroplasts to bacterial cells for this particular strain (as recorded by cfu/ml) did not occur in quantitative numbers. The ATP assay thus indicated an approximate of the density of cells, while viability studies reported a lower cell density. When using a broth with lower osmolality (50 mosm/L) no spheroplast formation occurred and a close relation between viability and intracellular ATP was observed.