Fe2+ Alters Carbon and Nitrogen Metabolic Networks in a Composite Microbial Consortium: Metagenomic Insights into the Shift from Denitrification to DNRA.

Abstract

Conventional biological nitrogen removal processes are constrained by lengthy treatment trains and dependence on organic carbon sources, necessitating the development of novel enhanced nitrogen removal strategies that integrate multiple functions and ensure operational stability. In this study, a synthetic bacterial consortium was constructed, comprising the aerobic denitrifier Pseudomonas stutzeri, the facultative anaerobic denitrifier Klebsiella sp., and the heterotrophic nitrifying-aerobic denitrifying bacterium Alcaligenes sp. The effects of five iron species as well as their combined effects with polyacrylamide (PAM), on nitrogen removal performance and oxidative stress responses of the consortium were investigated, and metagenomic sequencing was employed to elucidate the regulatory mechanisms of Fe2+ on metabolic processes. The results showed that, compared with the other iron species, the Fe2+ group achieved a 20-30% increase in nitrate-N removal efficiency. The addition of PAM attenuated the specific regulatory effects of different iron species through physical mass-transfer limitation. Metagenomic analysis revealed that Fe2+ modulated the carbon and nitrogen metabolic networks: in the carbon metabolic network, enrichment of the por gene in the glycolytic pathway generated substantial reducing power in the form of reduced ferredoxin; concomitantly, the transcript abundance of the dissimilatory nitrate reduction to ammonium pathway increased from 775 to 802, whereas that of the denitrification pathway decreased from 1259 to 1222. This study elucidates the intrinsic mechanism by which Fe2+ promotes synergistic carbon and nitrogen removal, providing a theoretical foundation for the development of a multi-process coupled deep nitrogen removal system integrating bioaugmentation, chemical regulation, and physical sedimentation.