Aims: The effect of temperature (2-30 degrees C), pH (4(.)8-7(.)4) and water activity (0(.)946-0(.)995) on the relationship between optical density (OD) at 600 nm and the plate count (CFU ml(-1)) was investigated for Listeria monocytogenes. Methods and Results: Calibration curves, relating OD with plate counts, were collected by measuring the OD of consecutive one-half dilution series, before determining the cell density by classic plate count methods. The calibration curves were observed to be shifting in a parallel way, with increasing stress levels. Especially pH influenced the curve in a great extent, while the other variables were showing more synergetic effects. The reason for the shift was investigated by a microscopic viability test, showing a viability decrease with increasing stress levels, causing the shift of the calibration curve. In a last step a model was made describing the effect of environmental factors on the calibration curve, with different data transformations being tested. A polynomial equation was fitted to the data, taking into account a set of constraints to incorporate microbiological knowledge in the black box model. Hence, illogical interpolation results and overfitting of the data could be avoided. Conclusions: Different stress factors are affecting the relationship between the OD and the cell count of L. monocytogenes by lowering the cell viability. These effects could be modelled using a constrained polynomial model. Significance and Impact of the Study: The observed phenomena are important when calculating growth parameters, like growth rate and lag phase, based on OD data.