Effect of environmental parameters (temperature, pH and a(w)) on the individual cell lag phase and generation time of Listeria monocytogenes

Abstract

The effect of the individual environmental factors temperature (2-30 degrees C), pH (4.4-7.4) and a(w) (0.947-0.995) as well as the combinations of these factors on the individual cell lag phase and the generation time of Listeria monocytogenes was investigated. Individual cells were isolated using a serial dilution protocol in microtiter plates, and subsequent growth was investigated by optical density (OD) measurements at 600 nm. About 100 replicates were made for each set of environmental conditions. Part of the data were previously published in Francois et al. (Francois, K., Devlieghere, F., Smet, K., Standaert, A.R., Geeraerd, A.H., Van Impe, J.F., Debevere, J., 2005a. Modelling the individual cell lag phase: effect of temperature and pH on the individual cell lag distribution of Listeria monocytogenes. Int. J. Food Microbiol. 100, 41-53.), but were recalculated here using the calibration curves for transformation of optical density to colony forming units/ml from Francois et al. (Francois, K., Devlieghere, F., Standaert, A.R., Geeraerd, A.H., Cools, I., Van Impe, J.F., Debevere, J., 2005b. Environmental factors influencing the relationship between optical density and cell count for Listeria monocytogenes. J. Appl. Microbiol. 99, 1503-1515), as this calibration curve appeared to be dependent on the environmental parameters. The previous dataset was also extended with a factor a(w), observed individually and combinations with the above mentioned environmental factors. Individual cell lag phases and subsequent growth rates were calculated assuming an exponential growth model. The results are discussed as mean values to determine the general trends and in addition, histograms are made and statistical distributions are fitted to the different data sets. When stress levels increased, the mean values and the variability observed for the individual cell lag phases increased, resulting in broader histograms and distributions that were shifting to the right. Also the gravity point of the distributions was shifting from a skewed left type to a more symmetrical type. The best description of the data is obtained with an exponential distribution for low stress levels, a gamma distribution for intermediate stress and a Weibull distribution for severe stress levels. When only low stress levels were applied, a significant percentage of the cells showed no lag phase. In those cases, a new approach was used to obtain better fits: cells with a lag phase and those without a lag phase were separated using a binomial distribution while in a second step, a gamma or a Weibull distribution is fitted to the fraction of cells showing a lag phase. A normal distribution is used to describe the variability of the generation times. These distributions can be applied to refine the exposure assessment part of the risk assessment concerning L. monocytogenes by incorporating intercellular variability.