Two new highly active esterases were detected by starch electrophoretic studies in Culex pipiens mosquitoes from the area of Montpellier (France) and from Cyprus. We demonstrate here that both the French and the Cyprus esterases B are overproduced due to amplification of the coding gene. The production of the esterase B is approximately 50- and 500-fold higher in mosquitoes from France and Cyprus, respectively, than in susceptible insects, whereas the number of gene copies is about 25 and 250. Differences of about 7- and 95-fold were also found in the degree of chlorpyrifos resistance. RFLP comparison of the amplified region containing the esterase B gene revealed large differences between French and Cyprus mosquitoes. It thus appears that two distinct haplotypes with an esterase B gene coding an enzyme with identical electrophoretic mobility have been amplified. We therefore named the haplotypes in mosquitoes from France and Cyprus B4 and B5, respectively. The estimated genetic distance between these two haplotypes is not smaller than those observed in all pair comparisons of other known esterase B haplotypes. These results are discussed in the context of amplification phenomena.