Observation of Injured E. coli Population Resulting from the Application of High-Pressure Throttling Treatments

This study used flow cytometry (FC), epifluorescent microscopy (EM), and conventional culture media (PC) to evaluate the potential for high-pressure throttling (HPT) to produce injury in E. coli. E. coli cells suspended at a concentration of approximately 8 log (CFU/mL) in Butterfield's phosphate buffer and UHT skimmed milk, were treated with HPT at pressures ranging from 35 to 283 MPa. Cells were stained with SYTO 9 and propidium iodide (Live/Dead Baclight kit) to assess their membrane integrity. MacConkey and Tryptone Soy agars and a modification of the thin agar layer method were used to determine injured and non-injured cells. PC results indicated a reduction in E. coli counts as pressure increased but no significant injured population was detected in either matrix. However, FC and EM observations indicated that the membrane integrity of a portion of the bacterial population was affected by HPT, producing different degrees of cell injury that could be sublethal. The percentage of this heterogeneous population increased with applied pressure. These results reassert the importance of understanding the potential of new processing treatments to produce sublethally-injured bacteria, and developing appropriate detection techniques.