Molecular evidence for algal bloom decay as a critical window for bacteria-mediated DOM transformation toward more refractory forms.

Algal blooms profoundly alter the sources and composition of dissolved organic matter (DOM) in eutrophic lakes, with bacterial processes playing a central role in DOM transformation. However, how algal bloom succession reshapes bacteria-mediated DOM transformation and further influences the molecular fate of DOM remains unclear. Here, we integrated Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), paired mass distance (PMD)-based reactomics, and high-throughput sequencing to characterize DOM transformations and their microbial driving mechanisms across the pre-outbreak, outbreak, and decay stages of a natural algal bloom in a eutrophic lake. Across bloom succession, DOM molecular richness peaked during the outbreak stage, whereas molecular phylogenetic dispersion, aromaticity, and recalcitrance increased toward the decay stage, alongside reduced potential bioavailability. Relative to other stages, bacterially active DOM during the decay stage showed the highest relative intensity of biologically refractory DOM (i.e., tannin-like, condensed aromatic-like, and lignin-like compounds), along with higher aromaticity and unsaturation. PMD-based reactomics revealed the strongest reaction signatures during the outbreak stage, dominated by categories annotated as oxygenases acting on paired donors and acyltransferases, consistent with rapid DOM turnover. By contrast, decay-enriched reaction signatures, including categories annotated as intramolecular oxidoreductases, carbon-nitrogen lyases, and racemases/epimerases, suggested selective molecular transformation of DOM, potentially favoring molecular persistence. These findings highlight the algal decay stage as a critical window for bacteria-mediated DOM transformation toward more refractory forms, providing molecular-level evidence for microbial regulation of organic carbon persistence in eutrophic lakes.