Multi‐dimensional tolerance to low temperature for high‐latitude invasion success by the rapidly expanding golden mussel

Many invasive species are capable of rapidly advancing invasion forefronts, often into areas with challenging or extreme environmental conditions. Understanding the mechanisms driving these invasions is essential for predicting their future spread and developing effective conservation strategies. A notable example is the recent range expansion of the golden mussel (Limnoperna fortunei) from low- to high-latitude regions, which poses significant threats to the integrity of global freshwater ecosystems and socio-economic sustainability. Successful invaders often utilize a variety of physiological, behavioural and ecological strategies to survive and thrive in harsh environments. To investigate the multi-dimensional mechanisms underlying low-temperature tolerance for high-latitude invasion success, we collected golden mussels from their northernmost invasive range. The mussels were exposed to a control temperature of 25°C and low temperatures of 15°C and 5°C. We conducted multi-level analyses, including behavioural (survival and valve activity), morphological (filament ultrastructure), biochemical (enzyme content) and molecular (transcriptome and metabolome) changes in the gill, a key organ involved in low-temperature response. We observed low mortality rates (<30%) across all stressed groups, indicating the golden mussel's tolerance to low temperatures. Upon initial exposure, the golden mussels significantly reduced their valve-opening rate, effectively blocking cold water outside their bodies. Microscopic observations and staining analyses revealed significant changes in the gill filaments, including alterations in cilia, filament cells and junction distances, suggesting functional morphological adjustments following the behavioural response of valve closure. The energy conserved through these morphological changes, along with actively generated energy, was utilized to enhance tolerance at the biochemical and molecular levels. This energy-intensive tolerance mechanism involved apoptosis inhibition, membrane fluidity enhancement, improved antioxidant capacity and elevated immune regulation, as evidenced by enzyme assays and integrated transcriptomic and metabolomic analyses. These adjustments interacted and coordinated, demonstrating highly systematic and complementary responses to the low temperature stress. Our study elucidates the multi-dimensional mechanisms employed by golden mussels to cope with extreme temperature conditions in high-latitude regions, highlighting the integrated strategies that facilitate their survival in harsh environments. These findings offer valuable insights for developing management strategies for regions characterized by extreme environmental conditions for invasive species.