Adaptive Microbial Dynamics in Hydrochar-Enhanced Chain Elongation under Loading Rate Regulation Explored through Mechanistic Network Analysis

Resource recovery from sewage sludge via chain elongation (CE) is often constrained by declining efficiency under high-rate conditions. This study developed a hydrochar-assisted anaerobic fermentation strategy to enhance the CE performance under reduced sludge retention times (SRTs). Under equivalent conditions, hydrochar significantly improved the fermentation performance toward medium-chain fatty acids (MCFAs). At an SRT of 7.5 days, the system maintained a yield comparable to the 15 day control (14.0 g of COD/L), demonstrating hydrochar's capacity-enhancing effect under elevated loading. Mechanistic analyses showed that hydrochar promoted a reduced redox environment, stimulating acidogenesis and electron donor resynthesis. Humic acids further facilitated CE, enhancing the caproate yield and selectivity. Dynamic time-series and path analyses under SRT regulation revealed adaptive responses, with a strong negative path coefficient from SRT to caprylate (β=-95.3), and positive coefficients from acetate to butyrate (β = +6.96) and valerate to heptanoate (β = +0.85). Microbial analysis identified <i>Clostridium_sensu_stricto_12</i> as a species showing marked variation under hydrochar-mediated SRT regulation, exhibiting strong correlations (<i>r</i> > 0.9) with even-chain fatty acid accumulation. These findings demonstrate the dual role of hydrochar in enhancing CE performance and fermentation capacity. Moreover, integrating mechanistic modeling with network-level microbial and metabolite analyses provides a robust framework for elucidating adaptive responses in product-oriented anaerobic systems.