Anaerobic digestion is a process that can produce renewable energy through the fermentation of biodegradable biomass. Industrial anaerobic digestion tanks are usually made of concrete but the production of various aggressive compounds (CO2, NH4+ and volatile fatty acids) during the microbial fermentation leads to deterioration of the concrete structure. In addition, the formation of a microbial biofilm on the cementitious material surface could generate even more intense biodeterioration. The objective of this study is to gain a better understanding of the involvement of biofilm in the biodeterioration of cementitious materials during an anaerobic digestion process. More specifically, the study focuses on the heterogeneity of microbial populations within the biofilm and the reactive medium in anaerobic digestion. Laboratory scale anaerobic bioreactors mimicking the industrial anaerobic digestion medium were constructed and CEM I cement pastes were immersed in this medium for 2, 3, 4, 5, 10 and 15 weeks. The biodeterioration of the cement pastes was evaluated by determining the deteriorated thickness. The aggressive compounds in the medium were quantified. The biofilm attached to the surface of the cement pastes was analyzed using 16 s rRNA gene sequencing. To evaluate the heterogeneity of the biofilm, the growth of biofilm layers was successively caused to stall by using two distinct biofilm removal techniques. Three microbial fractions were defined: planktonic microorganisms, and the microorganisms within the biofilm that were loosely and strongly attached. The results showed that the planktonic lifestyle was more associated with microorganisms producing methane and consuming volatile fatty acids, while the biofilm was more associated with bacteria producing acids, mainly members of the Clostridium genus. A microbial community shift due to a reversible propionic acid accumulation during the first 5 weeks was also observed. In addition, no major differences were spotted between the loosely and strongly attached biomass, indicating homogeneity in the two layers of the biofilm. These results suggest that the biofilm could increase the biodeterioration of concrete since volatile fatty acids could be produced in massive quantities near the surface of the cement samples by the acidogenic microbial population more present within the biofilm.