Metatranscriptome analysis unveils the mechanisms of zero-valent iron enhancing reactivation of starvation hydrolysis acidification sludge by inducing high-level gene expression.

Hydrolysis acidification (HA) is a promising method for wastewater treatment and resource recovery. However, the extended time required for bacterial reactivation after starvation or a change in living conditions often poses a challenge to the efficient operation of the system. Although the addition of zero-valent iron (ZVI) could enhance HA performance, its effects on sludge reactivation in the HA process are not fully understood. In this study, ZVI was employed to accelerate sludge reactivation and its involved genetic mechanisms were unveiled. The results demonstrated that ZVI addition activated the sludge within 35 days with stable HA performance. Sludge characteristics revealed that ZVI improved active biomass, enzyme activity (by 11.4 % ∼ 26.7 %), ETS activity (by 566 %), and cell viability, with a higher concentration of MLVSS, live cells, more microbial byproducts in EPS, and relative abundance of HA bacteria (63.41 %). Moreover, metatranscriptome analysis showed that ZVI upregulated the expression of genes related to key enzymes in carbohydrate degradation metabolism, biosynthesis of electron transfer media such as heme and ubiquinone, and biosynthesis of vital cofactors like vitamin B12 and folate during microbial growth and metabolism. These findings suggest that ZVI enhanced electron transfer, bacterial growth, and metabolism, resulting in effective starch conversion and VFAs generation. Overall, these results deepen our understanding of the mechanism by which ZVI enhanced HA sludge reactivation, providing valuable information for addressing sludge starvation issues in HA systems.