Multifaceted roles of methylisothiazolinone intervention in sludge disintegration and acidogenic and methanogenic pathways for efficient carboxylate production during anaerobic fermentation

Abstract

Anaerobic fermentation is a promising approach for sludge treatment, although it is frequently hindered by the tight structure of extracellular polymeric substances (EPS) and limited metabolic activities. This work firstly discovered that methylisothiazolinone (MIT, a commonly used antimicrobial agent in water treatment) effectively disintegrated the sludge structure and stimulated metabolic pathways, leading to efficient volatile fatty acids (VFAs) production. The presence of MIT (ranging from 10 to 40 mg/g TSS) contributed to the acidogenic process but strongly restrained the methanogenesis, and the distinct interfering effect is highly associated with the MIT dose. The VFAs accumulation reached a maximum of 1776.7 mg COD/L (5.2 times of control) but faced a complete inhibition of methanogenic activity at 40 mg/g TSS. Spectroscopic and molecular docking analysis revealed that MIT interacted with and disrupted the EPS (mainly proteins) structure by forming hydrogen bonds with N-H groups. This disruption facilitated the release of biodegradable organics, which served as fermentation substrates. Furthermore, the MIT-mediated improvement selectively enriched the hydrolytic-acidifying bacteria (i.e., Clostridium and Caloramator) and stimulated the associated metabolic functions, particularly in the uptake, hydrolysis, and metabolism of amino acids considering the high expressions of encoding genes (e.g., oppA, pepP, gdhA). However, methanogens' reproductive and metabolic abilities were suppressed due to their inefficient response and adaptation to MIT. This suppression was evident through downregulated genetic expressions involved in DNA replication and aminoacyl-tRNA synthesis. Consequently, a decline in methanogens abundance (e.g., Methanothrix, and Methanosarcina) and methanogenic activity occurred, impeding the consumption of VFAs. This work provided an innovative niche to promote resource recovery in the sludge anaerobic system and deciphered the underlying mechanisms at both biochemical interactive steps and microbial genetic traits.