Body Size-Related Thermal Acclimation and Underlying Gene Expression Mechanisms in Workers of the Red Fire Ant...

Body size is a critical trait shaping ectotherm thermal tolerance, yet how body size mediates thermal acclimation remains poorly understood in insects. Here, we conducted 15-day daily cold (9 °C, 2.5 h) and heat (30 °C, 2.5 h) acclimation treatments on large, medium, and small Solenopsis invicta workers, followed by extreme temperature survival assays and comparative RNA-seq transcriptome profiling to uncover size-dependent acclimation phenotypes and molecular regulatory pathways. Survival results revealed that both cold and heat acclimation significantly boosted extreme thermal resistance of medium and small workers, whereas large workers exhibited inherently low mortality under lethal temperatures with no significant acclimation-induced survival improvement. Transcriptomic analyses showed striking body-size-specific transcriptional reprogramming: medium workers produced the largest number of differentially expressed genes (DEGs) under both acclimation regimes, followed by large workers, while small workers displayed minimal transcriptional shifts. Functional enrichment highlighted core pathways including lipid and small-molecule cryoprotectant metabolism, cuticle/membrane homeostasis, ion transport, antioxidant detoxification and sensory nervous signaling. Notably, hundreds of odorant receptor genes were sharply downregulated in acclimated medium workers, suggesting potential repression of olfactory function. Unlike acute thermal stress, long-term repeated acclimation triggered only mild transcriptional fluctuations of heat shock proteins with limited fold changes. Our findings demonstrate that thermal acclimation capacity in red fire ant workers is strongly correlated with body size, and coordinated remodeling of metabolic, osmotic, and chemosensory gene networks underpins this thermal plasticity. This body-size-dependent acclimation mechanism advances our understanding of invasive ant climate adaptation and helps predict their range expansion risks under global warming.