The effect of hydrothermal (150 degrees C for 10 min and 70 degrees C for 40 mm), enzymatic (Accelerase 1500, 0.07 g/g volatile solids (VS)), ultrasound (45 kHz for 30 min) and chemical pretreatments (HNO3 at pH 3 and NaOH at pH 12) alone or in combination on the chemical composition and methane yield of the pulp and paper mill secondary sludge was studied in batch assays at 55 degrees C. In total, 12 different pretreatment combinations were compared. Chemical analyses showed that all pretreatments except for HNO3 and ultrasound pretreatments improved the organic matter solubilization. Among the studied pretreatments, hydrothermal (150 degrees C, 10 mm) pretreatment alone or in combination with enzymatic and/or ultrasound pretreatment had the highest impact on sludge solubilization and methane yield. The increase in methane yield was 31% (from 108 ml/g VSoriginal to 141 ml/g VSoriginal). In addition, enzymatic pretreatment also improved the methane yields but only when combined with hydrothermal pretreatment at 150 degrees C or ultrasound + hydrothermal pretreatment at 150 degrees C. On the other hand, ultrasound pretreatment did not improve the methane yields while acid and alkaline pretreatments resulted in lower methane yields than control. Improved hydrolysis and higher methane production rates noticed in assays subjected to hydrothermal pretreatment alone or in combination with enzymes and/or ultrasound could make these treatments more attractive in reducing the retention times required during full-scale anaerobic digestion of pulp and paper mill wastewater sludges. (C) 2013 Elsevier B.V. All rights reserved.