Terrestrial molluscs living in temperate and polar environments must contend with cold winter temperatures. However, the physiological mechanisms underlying the survival of terrestrial molluscs in cold environments and the strategies employed by them are poorly understood. Here we investigated the cold tolerance of Ambigolimax valentianus, an invasive, terrestrial slug that has established populations in Japan, Canada, and Europe. To do this, we acclimated A. valentianus to different environmental conditions (differing day lengths and temperatures), then exposed them to sub-zero temperatures and measured overall survival. Then, we measured low molecular weight metabolites using 1H NMR to see if they play a role in their cold tolerance as they do in other invertebrate species. We found that A. valentianus is not strongly freeze tolerant but does become more cold-hardy after acclimation to shorter day lengths. We also found that no metabolites were strongly upregulated in response to winter conditions despite the change in cold hardiness, and instead saw evidence of metabolic suppression leading up to winter such as formate and L-glutamine being suppressed in winter conditions.
