Cold Tolerance Including Rapid Cold-hardening and Inoculative Freezing of Fall Migrant Monarch Butterflies in Ohio

Migrants of the eastern North American population of the monarch butterfly, Danaus plexippus, are often subjected to subzero temperatures, heavy dews and frost in late September and October during the autumn migration to overwintering sites in Mexico. Adults of this generation had the capacity to rapidly increase their cold-hardiness. A chilling period of 1 h at +4OC before exposure to -4OC for 24 h significantly improved the survival ( > 80%) of monarchs over those with no chilling prior to -4OC exposure (~40%). This is the first report of rapid cold-hardening in Lepidoptera, and the capacity to rapidly cold-harden may protect monarchs against cold injury during diurnal changes in temperature. An extended period of progressively lower temperature acclimation enhanced cold-hardiness to a greater extent than did the rapid cold-hardening treatment. When exposed to -4’C, migrants collected in the field from Ohio were less tolerant (LT, = 37.5 h) than migrants gradually acclimated over several weeks from 20 to 25OC in the field to overwintering conditions of +4OC in the laboratory (LT,, = 85.8 h). External moisture on the exoskeleton significantly raised supercooling points (SCPs) of acclimated monarchs from -8.2 to -4.7OC. In addition, external moisture decreased the supercooling capacity and triggered internal ice formation at subzero temperatures above the SCP of dry monarchs. No monarchs survived internal ice formation associated with SCP determination, evidence that the monarch butterfly is a freeze-susceptible insect.