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Uranium is a risk-driving radionuclide in both radioactive waste disposal and contaminated land scenarios. In these environments, a range of biogeochemical processes can occur, including sulfate reduction, which can induce sulfidation of iron (oxyhydr)oxide mineral phases. During sulfidation, labile U(VI) is known to reduce to relatively immobile U(IV); however, the detailed mechanisms of the changes in U speciation during these biogeochemical reactions are poorly constrained. Here, we performed highly controlled sulfidation experiments at pH 7 and pH on U(VI) adsorbed to ferrihydrite and investigated the system using geochemical analyses, X-ray absorption spectroscopy (XAS), and computational modeling. Analysis of the XAS data indicated the formation of a novel, transient U(VI)–persulfide complex as an intermediate species during the sulfidation reaction, concomitant with the transient release of uranium to the solution. Extended X-ray absorption fine structure (EXAFS) modeling showed that a persulfide ligand was coordinated in the equatorial plane of the uranyl moiety, and formation of this species was supported by computational modeling. The final speciation of U was nanoparticulate U(IV) uraninite, and this phase was evident at 2 days at pH 7 and 1 year at pH Our identification of a new, labile U(VI)-persulfide species under environmentally relevant conditions may have implications for U mobility in sulfidic environments pertinent to radioactive waste disposal and contaminated land scenarios.