Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/29/2009
Publication Date: 10/13/2009
Citation: Smith, D.J., Oliver, C.E., Shelver, W.L., Caesar, T., Anderson, R.C. 2009. Clorate Metabolism in Pure Cultures of Escherichia Coli 0157:H7 Pretreated with Either Nitrate or Chlorate. Journal of Agricultural and Food Chemistry. 57:10216-10224. DOI:10.1021/jf901513f
Interpretive Summary: Thousands of food-born illnesses are caused each year by pathogens such as E. coli O157:H7 and(or) Salmonella that inhabit the gastrointestinal tracts of food animals. At slaughter, these bacteria may contaminate carcasses and retail products subsequent to carcass processing. One way to reduce the probability harmful bacteria will contaminate food-animal carcasses is to provide feed or water additives that will selectively kill pathogens. One such additive, sodium chlorate, has been identified that consistently kills both Salmonella and E. coli O157:H7 when fed to live animals. The exact manner in which sodium chlorate exerts its effects on these bacteria is unknown, but it is thought that chlorate acts at a specific site within the bacterial cell that allows the cell to use nitrate in energy assimilation. We conducted this study to determine the effects that nitrate might have on the action of chlorate in E. coli and to determine whether bacteria metabolize chlorate to products proposed to be bactericidal to pathogens. We found that nitrate does not increase the effect of chlorate in E. coli, and that E. coli metabolize chlorate to chlorite, an unstable metabolite. These results suggest that the effect of chlorate is mediated through the production of the active chlorate metabolite, chlorite.
Technical Abstract: Experiments were conducted to determine the effects of 5, 7.5, and 10 mM nitrate, and 5, 10, or 20 mM chlorate on total E. coli counts, chlorate metabolism, and volatile fatty acid (VFA) concentrations in anaerobic ruminal fluid cultures. Nitrate did not affect total E. coli counts (P = 0.05), chlorate metabolism (P = 0.05), or VFA concentration (P = 0.05). Chlorate (5 mM) decreased total E. coli counts relative to controls (P < 0.05) at h 3 and 6, and 20 mM chlorate decreased E. coli counts (P < 0.05) at 3, 6, and 24 h of incubation. Chlorate and nitrate did not generally affect (P = 0.05) VFA concentrations. Consistent evidence for measurable reduction of 5, 10, or 20 mM chlorate in ruminal fluid incubations was not produced. The response of pure cultures of E. coli O157:H7 to chlorate mirrored experiments with mixed ruminal cultures. That is, nitrate did not exacerbate the effects of chlorate on the pathogen. Experiments investigating chlorate biotransformation by pure cultures of E. coli O157:H7 using [36Cl]chlorate also indicated that rates of chlorate metabolism were low when chlorate concentrations exceeded 5 mM. Measurable chlorate reduction occurred in incubations containing 1 or 5 mM [36Cl]chlorate. Pure cultures of E. coli O157:H7 that were preconditioned with 10 mM chlorate had an attenuated ability to transform [36Cl]chlorate to [36Cl]chloride. The hypothesis that E. coli O157:H7 is sensitive to chlorate by virtue of the reduction of chlorate to chlorite ion was supported by the direct measurement of low concentrations of [36Cl]ClO2- in incubation media containing 0.5 mM [36Cl]ClO3-. Collectively these results indicate that concentrations of both generic E. coli in mixed culture, and E. coli O157:H7 in pure culture, will be reduced about 2 log units in the presence of 5 mM or greater concentrations of sodium chlorate.