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This study compared the behavior of the biomass in two fixed-film anaerobic reactors operated under equivalent organic loading rates but at different temperatures, i.e., 30 °C (RMM) and 55 °C (RMT). The reactors were fed with sugarcane vinasse and molasses (both fermented) in a simulation of sequential periods of season and off-season. The dynamics of biomass growth and retention, as well as the microbial composition, were assessed throughout 171 days of continuous operation, coupled with an additional 10-day test assessing the microbial activity in the bed region. Despite the different inoculum sources used (mesophilic granular vs. thermophilic flocculent sludge types), the biomass growth yield was identical (0.036–0.038 g VSS g⁻¹COD) in both systems. The retention rates (higher in RMT) were regulated according to the initial amount of biomass provided in the inoculation, resulting in similar amounts of total retained biomass (46.8 vs. 43.3 g VSS in RMT and RMM) and biomass distribution patterns (30–35% in the feeding zone) at the end of the operation. Meanwhile, hydrogenotrophic methanogenesis mediated by Methanothermobacter coupled to syntrophic acetate oxidation prevailed in RMT, while the Methanosaeta-mediated acetoclastic pathway occurred in RMM. The results show that different anaerobic consortia can behave similarly in quantitative terms when subjected to equivalent organic loads, regardless of the prevailing methane-producing pathway. The community grows and reaches a balance (or a given cell activity level) defined by the amount of substrate available for conversion. In other words, while the metabolic pathway may differ, the endpoint (the amount of biomass) remains the same if operational stability is maintained. Full article