In wastewater treatment plants (WWTP), anaerobic digester (AD) units are commonly operated under mesophilic and thermophilic conditions. In some cases, during the dry season, maintaining a stable temperature in the digester requires additional power to operate a conditioning system. Without proper conditioning systems, methanogens are vulnerable to temperature shifts. This study investigated the effects of temperature shifts on CH4 gas production and microbial diversity during anaerobic digestion of anaerobic sewage sludge using a metagenomic approach. The research was conducted in lab-scale AD under stepwise upshifted temperature from 42 to 48 ◦C. The results showed that significant methanogen population reduction during the temperature shift affected the CH4 production. With 70 days of incubation each, CH4 production decreased from 4.55 L·g −1 -chemical oxygen demand (COD) at 42 ◦C with methanogen/total population (M·TP−1 ) ratio of 0.041 to 1.52 L·g −1 COD (M·TP−1 ratio 0.027) and then to 0.94 L·g −1 COD ( M·TP−1 ratio 0.026) after the temperature was shifted to 45 ◦C and 48 ◦C, respectively. Methanosaeta was the most prevalent methanogen during the thermal change. This finding suggests that the Methanosaeta genus was a thermotolerant archaea. Anaerobaculum, Fervidobacterium, and Tepidanaerobacter were bacterial genera and grew well in shifted-up temperatures, implying heat-resistant characteristics.