Regulatory roles of extracellular polymeric substances in uranium reduction via extracellular electron transfer by Desulfovibrio vulgaris UR1

The pathway of reducing U(VI) to insoluble U(IV) using electroactive bacteria has become an effective and promising approach to address uranium-contaminated water caused by human activities. However, knowledge regarding the roles of extracellular polymeric substances (EPS) in the uranium reduction process involving in extracellular electron transfer (EET) mechanisms is limited. Here, this study isolated a novel U(VI)-reducing strain, Desulfovibrio vulgaris UR1, with a high uranium removal capacity of 2.75 mM/(g dry cell). Based on a reliable EPS extraction method (45 degrees C heating), manipulation of EPS in D. vulgaris UR1 suspensions (removal or addition of EPS) highlighted its critical role in facilitating uranium reduction efficiency. On the second day, U(VI) removal rates varied significantly across systems with different EPS contents: 60.8% in the EPS-added system, 48.5% in the pristine system, and 22.2% in the EPS-removed system. Characterization of biogenic solids confirmed the reduction of U(VI) by D. vulgaris UR1, and the main products were uraninite and UO2 (2.88-4.32 nm in diameter). As EPS formed a permeable barrier, these nanoparticles were primarily immobilized within the EPS in EPS-retained/EPS-added cells, and within the periplasm in EPS-removed cells. Multiple electroactive substances, such as tyrosine/tryptophan aromatic compounds, flavins, and quinone-like substances, were identified in EPS, which might be the reason for enhancement of uranium reduction via providing more electron shuttles. Furthermore, proteomics revealed that a large number of proteins in EPS were enriched in the subcategories of catalytic activity and electron transfer activity. Among these, iron-sulfur proteins, such as hydroxylamine reductase (P31101), pyruvate: ferredoxin oxidoreductase (A0A0H3A501), and sulfite reductase (P45574), played the most critical role in regulating EET in D. vulgaris UR1. This work highlighted the importance of EPS in the uranium reduction by D. vulgaris UR1, indicating that EPS functioned as both a reducing agent and a permeation barrier for access to heavy metal uranium.