Multilevel modelling as a tool to include variability and uncertainty in quantitative microbiology and risk assessment. Thermal inactivation of Listeria monocytogenes as proof of concept.

Variability is inherent in biology and also substantial for microbial populations. In the context of food safety risk assessment, it refers to differences in the response of different bacterial strains (between-strain variability) and different cells (within-strain variability) to the same condition (e.g. inactivation treatment). However, its quantification based on empirical observations and its incorporation in predictive models is a challenge for both experimental design and (statistical) analysis. In this article we propose the use of multilevel models to quantify (different levels of) variability and uncertainty and include them in the predictions. As proof of concept, we analyse the microbial inactivation of Listeria monocytogenes to thermal treatments including different levels of variability (between-strain and within-strain) and uncertainty. The relationship between the microbial count and time was expressed using a (non-linear) Weibullian model. Moreover, we defined stochastic hypotheses to describe the different types of variation at the level of the kinetic parameters, as well as in the observations (microbial counts). The model parameters (kinetic parameters and variances) are estimated using Bayesian statistics. The multilevel approach was compared against an analogous, single-level model. The multilevel methodology shrinks extreme parameter estimates towards the mean according to uncertainty, thus mitigating overfitting. In addition, this approach enables to easily incorporate different levels of variation (between-strain and/or within-strain variability and/or uncertainty) in the predictions. On the other hand, multilevel (Bayesian) models are more complex to define, implement, analyse and communicate than single-level models. Nevertheless, their ability to incorporate different sources of variability in predictions make them very suitable for Quantitative Microbial Risk Assessment.