Psychrophilic (< 20 degrees C) anaerobic digestion (AD) represents an attractive alternative to mesophilic wastewater treatment. In order to investigate the AD microbiome response to temperature change, with particular emphasis on methanogenic archaea, duplicate laboratory-scale AD bioreactors were operated at 37 degrees C followed by a temperature drop to 15 degrees C. A volatile fatty acid-based wastewater (composed of propionic acid, butyric acid, acetic acid and ethanol) was used to provide substrates representing the later stages of AD. Community structure was monitored using 16S rRNA gene clone libraries, as well as DNA and cDNA-based DGGE analysis, while the abundance of relevant methanogens was followed using qPCR. In addition, metaproteomics, microautoradiography-fluorescence in situ hybridization, and methanogenic activity measurements were employed to investigate microbial activities and functions. Methanomicrobiales abundance increased at low temperature, which correlated with an increased contribution of CH4 production from hydrogenotrophic methanogenesis at 15 degrees C. Methanosarcinales utilized acetate and H-2/CO2 as CH4 precursors at both temperatures and a partial shift from acetoclastic to hydrogenotrophic methanogenesis was observed for this archaeal population at 15 degrees C. An upregulation of protein expression was reported at low temperature as well as the detection of chaperones indicating that mesophilic communities experienced stress during long-term exposure to 15 degrees C. Overall, changes in microbial community structure and function were found to underpin the adaptation of mesophilic sludge to psychrophilic AD.