Anticoagulant rodenticides are commonly used to control rodent pests worldwide. They specifically inhibit the vitamin K epoxide reductase (VKORC1), which is an enzyme encoded by the Vkorc1 gene, involved in the recycling of vitamin K. Therefore, they prevent blood clotting. Numerous mutations of Vkorc1 gene were reported in rodents, and some are involved in the resistant to rodenticides phenotype. Two hundred and sixty‐six mice tails were received from 65 different locations in France. Coding sequences of Vkorc1 gene were sequenced in order to detect mutations. Consequences of the observed mutations were evaluated by the use of recombinant VKORC1. More than 70% of mice presented Vkorc1 mutations. Among these mice, 80% were homozygous. Contrary to brown rats for which only one predominant Vkorc1 genotype was found in France, nine missense single mutations and four double mutations were observed in house mice. The single mutations lead to resistance to first‐generation antivitamin K (AVKs) only and are certainly associated with the use of these first‐generation molecules by nonprofessionals for the control of mice populations. The double mutations, probably obtained by genetic recombination, lead to in vitro resistance to all AVKs. They must be regarded as an adaptive evolution to the current use of second‐generation AVKs. The intensive use of first‐generation anticoagulants probably allowed the selection of a high diversity of mutations, which makes possible the genetic recombination and consequently provokes the emergence of the more resistant mutated Vkorc1 described to date.

The four introgressed polymorphisms were clearly the cause of the strong resistant phenotype observed in the field. In the context of strong selection pressure due to the extensive use of AVKs, this resistant phenotype may explain the widespread distribution of this genotype from Spain to Germany. © 2016 Society of Chemical Industry.

The p.Y25F mutation detected in the Vkorc1 gene in R. rattus trapped on the Spanish farm is associated with the resistance phenotype to bromadiolone that has been observed. It is the first evidence of target resistance to antivitamin K anticoagulants in R. rattus.