Two novel mechanisms of organophosphate resistance were isolated and characterized from a population of Culex pipiens L. from Cyprus. Two strains, one expressing the novel, highly active esterases A5 and B5 (strain A5B5-R), and one expressing insensitive acetylcholinesterase (strain Ace-R), were developed by single pair crosses and selection with temephos and propoxur, respectively. The A5B5-R strain demonstrated resistance toward organophosphate insecticides that could be suppressed by the esterase inhibitor S,S,S-tributyl phosphorotrithioate (DEF). No cross-resistance to carbamates occurred. The Ace-R strain demonstrated resistance to organophosphate as well as to carbamate insecticides. Propoxur and temephos resistance was not affected by the monooxygenase inhibitor piperonyl butoxide or by DEF. The Ace-R strain possessed a novel toxicologic profile as well as a unique acetylcholinesterase inhibition pattern. Inheritance of temephos or propoxur resistance was codominant in F1 offspring. Backcrosses to a susceptible strain in both cases failed to fit a single gene model, suggesting that multiple loci may be involved. Combining the A5B5-R and the Ace-R strains resulted in high levels of temephos resistance, similar to that of the parents.