Transcriptome-and Metabolome-Based Mechanisms of High-Temperature Adaptation in Triploid Rainbow Trout (Oncorhynchus...

High-temperature stress poses a critical challenge to cold-water aquaculture; however, the size-associated molecular mechanisms underlying thermal adaptation in triploid rainbow trout (Oncorhynchus mykiss) remain poorly understood. Here, we integrated transcriptomic and metabolomic profiling of liver tissues, which plays central role in energy metabolism and stress integration, from three body-weight classes-small (0.8 kg ± 0.18 kg), medium (1.5 kg ± 0.22 kg), and large (2.5 kg ± 0.31 kg)-sampled under peak summer heat stress (20.5 °C). Transcriptomic analysis identified 974, 570, and 862 group-specific differentially expressed genes, respectively, revealing a non-linear, size-associated transcriptional pattern. Genes assigned to the ribosome pathway were significantly enriched in comparisons involving MLA and SLA relative to LLA, but not between SLA and MLA, whereas carbon metabolism and amino acid biosynthesis were enriched exclusively in smaller fish, indicating a higher catabolic burden under heat stress. Metabolomic profiling identified 1123 metabolites, with lipids accounting for 45.06%, and showed size-specific enrichments in biosynthesis of unsaturated fatty acids, glycerophospholipid metabolism, arachidonic acid metabolism, and pathways related to necroptosis. Integrative analysis revealed that in smaller fish, pla2g1b and gpx4a coordinately regulate the accumulation of prostaglandin H2 and 16(R)-HETE, forming a regulatory network with ferroptosis-related genes acsl4a and hmox1a; concurrently, chka, lpin1, and phospholipase A2 members drive extensive membrane phospholipid remodeling. The observed negative correlation between hsd17b3 expression and 7α-hydroxytestosterone levels suggests size-associated steroid-mediated energy repartitioning. Collectively, smaller fish undergo extensive transcriptional and metabolic reprogramming with heightened activation of cell death pathways, whereas larger fish maintain greater thermal buffering capacity. These findings provide molecular targets for size-stratified thermal management and selective breeding in rainbow trout aquaculture.