Lethality of a Heat- and Phosphate-Catalyzed Glucose By-Product to Escherichia coli O157:H7 and Partial Protection Conferred by the rpoS Regulon

Abstract

A by-product of glucose produced during sterilization (121°C, 15 lb/in2, 15 min) at neutral pH and in the presence of phosphate (i.e., phosphate-buffered saline) was bactericidal toEscherichia coli O157:H7 (ATCC 43895). Other six-carbon (fructose and galactose) and five-carbon (arabinose, ribose, and xylose) reducing sugars also produced a toxic by-product under the same conditions. Fructose and the five-carbon sugars yielded the most bactericidal activity. Glucose concentrations of 1% (wt/vol) resulted in a 99.9% decline in the CFU of stationary-phase cells per milliliter in 2 days at 25°C. An rpoS mutant (pRR10::rpoS) of strain 43895 (FRIK 816-3) was significantly (P < 0.001) more sensitive to the glucose-phosphate by-product than the parent strain, as glucose concentrations from 0.05 to 0.25% resulted in a 2- to 3-log10 reduction in CFU per milliliter in 2 days at 25°C. Likewise, log-phase cells of the wild-type strain, 43895, were significantly more sensitive (P < 0.001) to the glucose-phosphate by-product than were stationary-phase cells, which is consistent with the stability of rpoS and the regulation ofrpoS-regulated genes. The bactericidal effect of the glucose-phosphate by-product was reduced when strains ATCC 43895 and FRIK 816-3 were incubated at a low temperature (4°C). Also, growth in glucose-free medium (i.e., nutrient broth) did not alleviate the sensitivity to the glucose-phosphate by-product and excludes the possibility of substrate-accelerated death as the cause of the bactericidal effect observed. The glucose-phosphate by-product was also bactericidal to Salmonella typhimurium, Shigella dysenteriae, and a Klebsiella sp. Attempts to identify the glucose-phosphate by-product were unsuccessful. These studies demonstrate the production of a glucose-phosphate by-product bactericidal to E. coli O157:H7 and the protective effects afforded by rpoS-regulated gene products. Additionally, the detection of sublethally injured bacteria may be compromised by the presence of this by-product in recovery media.

Factors Affecting Thermally Induced Furan Formation

“…There are studies showing interactions of phosphate with sugar in chemical reactions. For example, it has been found that sugar solutions became bactericidal after being heated in the presence of phosphate , . Recent studies , have shown that when sugars and ascorbic acid were heated in the presence of phosphate (without realizing the phosphate effect), many compounds besides furan were produced including acetic acid, formic acid, acetaldehyde, and glycolaldehyde.…”