Purpose: The objective of this study is to compare the levels of DnaK protein under heat-shock and acid-shock conditions among different STEC strains.
Methods: Overnight cultures of six non-O157 STEC, one E. coli O157:H7 (EDL 933) and one non-pathogenic E. coli K12 were grown in tryptic soy broth (TSB). For the heat shock treatment, cells were held at 46°C for 25 min prior to protein extraction. For the acid shock treatment, cells were held in acidified TSB with acetic acid 0.50M (pH 6.5) for 1 h at 37°C. Cells were disrupted using a commercial enzyme cocktail followed by the addition of phenymethylsulfonyl fluoride to inhibit proteases and extract DnaK. A competitive ELISA was used to determine the intracellular DnaK concentration using absorbance at 450 nm. Each experiment was replicated thrice and data were statistically analyzed.
Results: Significant differences (P>0.05) in the intracellular concentration of DnaK were found between the tested strains exponentially grown at 37°C with values ranging from 1,821±120 to 6,204±423 molecules per cell. The values observed for heat shock and acid shock ranged from 6,867±340 to 18,182±460 and 5,032±475 to 15,919±549 molecules/cell, respectively.
Significance: The monitoring of the internal concentration of DnaK proteins shows that varying DnaK concentrations are detected during sub-lethal injury with either heat or acid stress. The elevated concentration of DnaK is associated with the ability of the cells to adapt to stress. Therefore heat shock proteins can potentially be used in addition to cell counts to determine the susceptibility of STEC to adapt to or survive different food processing conditions.