In this study, the interactions between the microbial populations in an anaerobic digester leading to the acidification of the fermentation medium were investigated using numerical simulation in the Anaerobic Digestion Model No. 1 (ADM1). The anaerobic digestion of cattle manure at 10 g volatile solids (VSs)/L and 35 degrees C was used as the case study. The numerical runs were performed using a factorial experimental approach with two levels. The minimum pH reached in the digestion was considered the response variable, whereas the initial concentrations of the main microbial groups included in the ADM1 were considered the factors. The statistical analysis demonstrated that the initial concentration of acetoclastic bacteria was the most critical variable in the acidification of the medium. It was estimated that permanent acidification occurred when the initial concentration of this group of microorganisms was less than or equal to 0.0838 g chemical oxygen demand (COD)/L, regardless of the population density of the other microbial groups. (C) 2013 Elsevier B.V. All rights reserved.