Technical Abstract: The occurrence of resistance in Varroa mite populations is a serious threat to the beekeeping industry and crops that rely on the honey bee for pollination. Integrated pest management strategies for control of this pest include the judicious use of insecticides. To monitor field populations of Varroa mite for insecticide resistance, a glass vial bioassay procedure was developed to use in the development of a resistance management strategy. Diagnostic concentrations needed to separate susceptible genotypes from resistant individuals were determined for cypermethrin (0.1 µg per vial), fluvalinate (5 0. µg per vial), malathion (0.01 µg per vial), coumaphos (10.0 µg per vial), diazinon (5.0 µg per vial), methomyl (0.5 µg per vial), propoxur (0.1 µg per vial) and endosulfan (2.5 µg per vial). Resistance to organophosphorus insecticides (malathion, coumaphos) and pyrethroids (cypermetrhrin, fluvalinate) was widespread in both Texas and Florida from 2007 to 2009. There was no resistance to endosulfan, diazinon, methomyl and propoxur in field populations of Varroa mite in the two states. The seasonal patterns of resistance in Florida were different from those of Texas. In Florida, the frequency of resistance to all insecticides tested decreased significantly from 2007 to 2009 while it increased in Texas. Resistance levels were unstable, suggesting that resistance could be successfully managed. The results validate use of the glass vial bioassay to monitor for resistance in Varroa mite and provide the basis for the development of a resistance management strategy designed to extend the efficacy of all classes of insecticides used for control of Varroa mite.