Transcriptome–metabolome uncoupling Underlies chronic phytotoxicity of glufosinate residues in rapeseed

Glufosinate-ammonium (GLA) is increasingly used worldwide, but the chronic phytotoxicity of its soil residues to subsequent crops remains poorly understood. To address this knowledge gap, we integrated physiological, ultrastructural, transcriptomic, and metabolomic analyses to investigate the responses of rapeseed (Brassica napus L.) to three soil GLA residue concentrations across five transplanting intervals with all measurements performed at 30 days after transplanting (DAT). GLA residue levels in leaves and roots increased with treatment concentration and decreased with prolonged transplanting intervals. GLA residues were associated with visible growth inhibition and reduced photosynthesis in a concentration‑ and time‑dependent manner, as evidenced by decreases in chlorophyll content and net photosynthetic rate. Chloroplasts exhibited progressively severe ultrastructural damage with increasing concentration. Glutamine synthetase (GS) activity was inhibited, leading to ammonium accumulation, decreased soluble protein content, and—notably—glutamate accumulation at 30 DAT, a pattern that contrasts with the classical acute‑toxicity paradigm of glutamate depletion. In contrast, Ribulose-1,5-bisphosphate carboxylase (Rubisco) activity was uniformly suppressed to similarly low levels across all GLA treatments. By day 30, the antioxidant system exhibited functional impairment, potentially linked to energy limitation: although antioxidant‑related genes remained transcriptionally responsive, H₂O₂ levels showed no significant change, whereas MDA accumulated and ASA–GSH cycle components declined. Transcriptomic analysis revealed 1,967 commonly upregulated differentially expressed genes (DEGs) across the three treatment groups, but metabolomics identified only 56 commonly upregulated metabolites, which were heavily concentrated in glutamate‑centric nitrogen metabolism—indicating a profound transcriptome‑metabolome uncoupling. Notably, the DEG count in M3T8 exceeded that in the acute‑stress group M2T1, indicating that an 8‑day transplant interval may be insufficient to ensure safety under high‑dose conditions in this pot experiment. Collectively, these results suggest that, at 30 DAT, GLA exposure is associated with a distinct “chronic decline” pattern—chloroplast disintegration, glutamate accumulation, antioxidant impairment, and transcriptome–metabolome uncoupling—a pattern that contrasts with the acute ammonia‑driven lethality.