Metagenomic analysis reveals the size effect of magnetite on anaerobic digestion of waste activated sludge after thermal hydrolysis pretreatment

Although magnetite has been widely investigated in anaerobic digestion (AD), its role in the practical AD of waste-activated sludge (WAS) after thermal hydrolysis pretreatment (THP) and its size effect remain unclear. In this study, magnetite with four different particle sizes was added during the AD of WAS after THP. With the reduction of magnetite particle size, cumulative methane production was increased, while the optimal dosage of magnetite de-creased, with 0.1 mu m magnetite at an optimal dosage of 2 g/L achieving the highest cumulative methane production increase of 111.97 % compared with the blank group (without magnetite). Smaller magnetite particles increased alpha-glucosidase and protease activities, coenzyme F420 concentration, and electron-transport system activity (20.30 %, 173.02 %, 60.39 % and 158.08 % higher respectively than the blank group). The size of magnetite also influenced the establishment of direct interspecies electron transfer (DIET) during AD. Based on the analysis of the pilA gene abun-dance, magnetite with a large particle size could promote the formation of e-pili in syntrophic electroactive bacteria (Clostridium, Syntrophomonas, and Pseudomonas) and methanogens (Methanospirillum), thereby completing electron transfer. However, small-sized magnetite particles stimulated DIET by enhancing the secretion of conductive proteins in extracellular polymeric substances and membrane-bound enzymes (Fpo) in Methanosarcina.