Temperature-metabolism relationships above and below the body fluid freezing point in the freeze-tolerant white worm, Enchytraeus albidus.

Enchytraeus albidus is a freeze-tolerant oligochaete with potential for long-term cryopreservation storage as live feed in aquaculture. In this study, we measured the metabolic rate (MR) of E. albidus across a range of temperatures (+20 to -14 °C) in both unfrozen and frozen states using intermittent flow-through CO2 respirometry. Our results demonstrate an exponential decrease in MR with declining temperature, and a markedly steeper decline below the freezing point of body fluids. Frozen worms exhibited a Q10 of 24.1, in contrast to 2.2 in unfrozen worms, indicating significant metabolic depression associated with freezing. We argue that this depression is strongly correlated with intracellular osmolality, which increases due to freeze-induced dehydration. Theoretical calculations based on MR data suggest that glycogen stores could support frozen aerobic metabolism for up to 3-4 months at -14 °C, aligning with overwinter survival observed in Arctic populations. These findings indicate that E. albidus primarily relies on aerobic metabolism when frozen and highlight its suitability for long-term storage as live feed under moderate freezing conditions. Our results provide valuable physiological knowledge for optimizing industrial cryopreservation strategies to balance survival and shelf-life in freeze-tolerant invertebrates.