A sample of Aedes aegypti L. from Santiago de Cuba with a high level of deltamethrin resistance (113.7 x at the 50% lethal concentration [LC50]), was subjected to deltamethrin selection to determine the capacity of this population to evolve higher resistance under intensive laboratory selection pressure, to characterize that resistance, to attempt to identify some of the mechanisms involved, and to use it as a reference strain for future molecular research. High resistance developed after 12 generations of selection (1,425 x). After selection for 12 generations with deltamethrin, the Santiago de Cuba colony (SAN-F12) showed little or no cross-resistance to the organophosphates evaluated, but high cross-resistance was observed for all the pyrethroids in larvae from this strain: lambdacyhalothrin (197.5 x), cypermethrin (45 x), and cyfluthrin (41.2 x). Adult bioassays reveal that a SAN-F12 strain was resistant to the pyrethroid and the organochlorine dichlorodiphenyltrichloroethane (DDT). Synergism tests implicated detoxifying esterase or glutathione S-transferase (GST) and monooxygenase in pyrethroid resistance. Biochemical tests reveal that acetylcholinesterase was not involved in deltamethrin resistance. The frequency of GST enzyme increased from 0.43 in Santiago de Cuba to 0.88 in SAN-F12. Esterase frequency increased from 0.12 in Santiago de Cuba to 0.63 in SAN-F6 and it diminished to 0.38 in SAN-F12. The polyacrylamide gel electrophoresis and inhibition study suggests the presence of elevated esterase activity not associated with pyrethroid resistance. The presence of both DDT and pyrethroid resistance in the SAN-F12 strain suggests the presence of a knockdown (Kdr)-type resistance mechanism, although the frequency of this mechanism was low. Resistance to deltamethrin could be associated with esterase or GST mechanisms, and more investigation is required. This information contributes to the improvement of resistance management strategies in the Cuban Ae. aegypti control program.