Assessing the pressure resistance of Escherichia coli O157:H7, Listeria monocytogenes and Salmonella enterica to high pressure processing (HPP) in citric acid model solutions for process validation

Despite the commercial success of high pressure processing (HPP) in the juice industry, some regulatory agencies still require process validation. However, there is a lack of consensus on various aspects regarding validation protocols, including the selection of representative strains to be used in challenge tests. This study characterized the variable response of Escherichia coli O157:H7 (34 strains), Listeria monocytogenes (44 strains) and Salmonella enterica (45 strains) to HPP, and identified potential candidates to use in process validation. Stationary phase cells were submitted to 500 MPa for 1 min at 10 °C in model solutions consisting of tryptic soy broth + 0.6% yeast extract (TSBYE) adjusted to pH 4.5 and 6.0 with citric acid. At pH 6.0, pressure resistance widely varied between species and within strains of the same species. E. coli O157:H7 and L. monocytogenes were the most pressure resistant and showed high variability at strain level, as the total count range given by minimum and maximum counts spread between 2.0 and 6.5 log₁₀ CFU/ml. S. enterica was the least resistant pathogen with more than 82% of the isolates displaying non-detectable counts after HPP. Recovery through storage at 12 °C was also variable for all pathogens, but eventually most strains recovered with median counts on day 14 between 8.3 and 8.9 log₁₀ CFU/ml. For pH 4.5 solutions, 26 E. coli O157:H7 strains displayed survivors after HPP but did not adapt, registering non-detectable counts in the next sampling dates. None of the L. monocytogenes and S. enterica strains survived HPP or incubation at pH 4.5 (<2.0 log₁₀ CFU/ml), suggesting that citric acid at 4.16 g/l is a safe barrier for pathogen control under moderate HPP conditions. Principal component and cluster analyses served to propose strain cocktails for each species based on their pressure resistant and adaptation phenotypes. Additionally, S. enterica was identified as less pressure resistant and less prone to recover following HPP than E. coli O157:H7 and L. monocytogenes, so its relevance in process validation for juices should be questioned. Future work will validate the proposed strain cocktails on real food systems.