Aims: This study was to investigate the methanogenic community in a biogas reactor from start-up to acidification conditions. Furthermore, reliability and accuracy of the applied quantitative real-time PCR method (Q-PCR) was briefly evaluated. Methods and Results: A mesophilic (37 degrees C), maize silage fed, continuously stirred tank reactor was surveyed. It was operated semi-continuously with increasing daily organic loading rates (OLRs) to reach acidification. Gas production and organic acid composition were measured. Methanogenic community structure was determined by 16S rDNA-based Q-PCR to estimate the abundance of key methanogenic micro-organisms. 16S rDNA of hydrogenotrophic Methanobacteriales was most abundant at OLRs of >= 3 center dot 7 g dry organic matter (DOM) l-1 day-1. By contrast, that of aceticlastic Methanosaetaceae predominated at lower OLRs but disappeared at OLRs of >= 4 center dot 1 g DOM l-1 day-1. At the same OLR, the propionate concentration increased dramatically indicating the acidification of the digester. Application of internal standards to examine Q-PCR's accuracy revealed that the detected amount of 16S rDNA may vary within one log cycle. Conclusions: These results suggest that the absence of Methanosaetaceae might be taken as biological indicator for process' instability. Inhibitory effects on Q-PCR analyses could not be determined based on the spiking experiments. Significance and Impact of the Study: In this study, reactors' microbiology was observed over time using Q-PCR. Insights into the abundance of different methanogens might be used to improve the performance of biogas reactors.