THE EFFECT OF CARBONYLCYANIDE-M-CHLOROPHENYLHYDRAZONE (CCCP) AND ACETATE ON ESCHERICHIA COLI O157:H7: UNCOUPLING VERSUS ANION ACCUMULATION

Authors: DIEZ-GONZALEZ, F - CORNELL UNIVERSITY; Russell, James

Submitted to: Federation of European Microbiological Societies Microbiology Reviews
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/15/1997
Publication Date: N/A
Citation: N/A

Interpretive Summary: Escherichia coli O157:H7 is a highly toxic strain of the E. coli bacteria that kills more than 500 people per year. The growth of E. coli in the rumen (the first of four stomachs in ruminant animals such as cattle, sheep and goats) and in the intestines is inhibited by compounds known as volatile fatty acids. The volatile fatty acids are low molecular weight compounds containing carbon, hydrogen and oxygen. The primary inhibitor of E. coli is acetate, a volatile fatty acid containing two carbon atoms. Other workers found that E. coli O157:H7 had a greater resistance to acetate than other strains of E. coli, but the mechanism of this resistance was not known. It had generally been assumed that acetate facilitated a futile cycling of protons (hydrogen atoms) through the cell membrane through a process known as uncoupling. We found that acetate did not act as an uncoupler in E. coli O157:H7, thus uncoupling is not a satisfactory explanation for the resistance of E. coli 0157:H7 to acetate. This, and subsequent work aimed at better understanding the acid resistance mechanism of 0157:H7, may provide means of inhibiting the growth of E. coli in the animals and man. In turn, this would reduce the incidence of illness and/or death resulting from exposure to this bacterium.

Technical Abstract: Non-growing cells of E. coli O157:H7 that were incubated anaerobically in sodium phosphate buffer at pH 6.5 consumed glucose at a rate of 8 umol [mg protein]-1 h-1, produced acetate, formate and ethanol, and had an intracellular pH of 7.3. The uncoupler, carbonylcyanide -m- chlorophenylhydrazone (CCCP), caused a decrease in intracellular pH, glucose consumption rate, intracellular ATP and potassium. CCCP had no effect on the ratio of fermentation end-products. Acetate addition also caused a decrease in intracellular pH and glucose consumption rate, but it did not cause a decrease in intracellular ATP or potassium. Acetate addition caused a marked increased in D-lactate production. Differences between acetate and CCCP indicated that acetate was not acting as a simple uncoupler. Acetate addition caused an increase in intracellular acetate, but intracellular acetate concentrations were similar to amounts predicted by the transmembrane pH gradient (delta pH), acetate diffusion and dissociation in the more alkaline interior. If acetate were acting as an uncoupler, one would have expected lower concentrations of intracellular acetate than were predicted by delta pH.