Geographic variation of freeze-tolerance in the earthworm Dendrobaena octaedra

Freeze-tolerance and some of the underlying biochemical defence mechanisms in the earthworm Dendrobaena octaedra was investigated. Survival after slow cooling to -2 degrees C, -4 degrees C, or -6 degrees C was analysed in D. octaedra from three geographic regions representing large differences in winter temperature (Denmark, Finland and Greenland). A large variation in freeze-tolerance between the three populations of D. octaedra was found. Earthworms from the northern populations (Finland and Greenland) tolerated lower temperatures (-6 degrees C) than earthworms from the Danish population (poor survival at -4 degrees C and -2 degrees C). In the Finnish population, freezing led to the production of high concentrations of glucose, which reached values much higher than controls (94 mg g(-1) vs. 2 mg g(-1) dry weight). Other potential cryoprotectants were not elevated after freezing. The Danish and Greenlandic populations had substantially lower mean glucose levels after freezing than the Finnish population (about 15 mg g(-1)). Danish earthworms rapidly frozen did not accumulate glucose, and did not survive freezing at -2 degrees C. Danish earthworms exposed to osmotic stress in Ringer's solutions, containing different concentrations of glycerol, showed significantly elevated glucose levels, but did not survive rapid freezing. It was determined if freezing had an influence on the reproduction of the earthworms. After warming to summer temperatures (15 degrees C), survivors of freezing produced viable cocoons. In a field experiment it was tested if natural acclimatization during autumn and winter months had an effect on freeze-tolerance in the Danish population. There was a significant increase of post-freeze survival during this period. The results of the freezing experiments are discussed in relation to the general ecology of D. octaedra.