Title: A bioluminescence ATP assay for estimating surface hydrophobicity and membrane damage of Escherichia coli cells treated with pulsed electric fields Authors
Submitted to: Foodborne Pathogens and Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 3, 2011
Publication Date: September 1, 2011
Citation: Ukuku, D.O., Yuk, H., Zhang, H.Q. 2011. A bioluminescence ATP assay for estimating surface hydrophobicity and membrane damage of Escherichia coli cells treated with pulsed electric fields. Foodborne Pathogens and Disease. Volume 8, Number 10, 2011, Pages 1103-1109. Interpretive Summary: Pulse Electric Field (PEF) treatments, a non-thermal process have been reported to injure and kill bacteria in liquid foods. This technology can be used to kill Escherichia coli in apple juice. However, some E. coli may survive the PEF process as injured cells. There is no information available regarding the effect of PEF treatments on E. coli cells surface charge and how it relates to injury and death. In this study, we investigated the effect of PEF on E. coli cell surface charge and hydrophobicity leading to injury with subsequent inactivation of the treated E. coli bacteria in apple juice. The two E. coli O157:H7 strains (SEA13B88 and Oklahoma) studied here were implicated in apple juice cider-related outbreaks. Their surface charge and hydrophobicity determined before the PEF treatments was 32.10 and 0.462, respectively. The PEF treatments decreased these numbers significantly to 6.9 and 0.337, respectively. The decrease in surface charge and hydrophobicity by PEF treatment led to E. coli cells cellular injury. And the injury to the bacterial membrane led to the leakage of intracellular adenosine tryphosphate (ATP), the energy needed for maintaining normal physiological activity and, this led to their death. Understanding how this happens will enable processors to improve the PEF process for enhancing the safety of apple juice.
Technical Abstract: Pulse Electric Field (PEF) treatments, a non-thermal process have been reported to injure and inactivate bacteria in liquid foods. However, the effect of this treatment on bacterial cell surface charge and hydrophobicity has not been investigated. Apple juice (AJ, pH 3.8) purchased from a wholesale distributor was inoculated with Escherichia coli O157:H7 at 6.8 log CFU/ml, processed with a PEF at 32.2 kv/cm, 18.4 A with pulse width of 2.6 µs, at 35 C and 45C and a flow rate of 120 ml/min. Treatment condition was periodically adjusted to achieve outlet temperatures of 47.5 and 57C. Treated samples were plated (0.1 ml) on Sorbitol MacConkey Agar (SMAC) and Tryptic Soy Agar (TSA) plates to determine injured populations and viability loss. Bacterial cell surface charge and hydrophobicity of untreated and PEF treated E. coli O157:H7 were determined using hydrophobic and electrostatic interaction chromatography. Immediately after treatment at 47.5C, the average cell populations determined on TSA plates and SMAC plates was 4.9 log and 4.7 log, respectively. At 57C treatment, E. coli cell populations averaged 3.9 and 2.4 log on the TSA and SMAC plates, respectively. The relative E. coli cell surface charge and hydrophobicity before PEF treatment averaged 32.10+/-8.12 and 0.4620+/-0.112, respectively. PEF treatment at 47.5C and 57C decreased the above values to 12.85+/-3.30 and 0.335+/-0.121 and 6.94+/-3.85 and 0.337+/-0.114, respectively. The results of this study indicate that damage of PEF treatment to E. coli bacteria occurs mostly at the outer cell envelope exposed to the extracellular environment as evidenced by the reduction of cell surface charge and leakage of intracellular ATP after the PEF treatment.