A new molecular method for the rapid detection of a metamitron-resistant target site in Chenopodium album.

Resistance to photosystem II inhibitors-triazines (atrazine) and triazinones (metamitron, metribuzin)-in Chenopodium album L. is caused by the serine 264 to glycine mutation in the D1 protein. This mutation has been detected in C. album collections from Belgium with unsatisfactory metamitron efficacy in the field and was confirmed in greenhouse resistance bioassays. Incomplete herbicide efficacy in practice can also be caused by reduced uptake due to environmental conditions. Hence, for reliable differentiation and resistance identification, a rapid method for mutation detection in the target gene psbA is required. Dose-response curves obtained in herbicide greenhouse assays with metamitron-resistant and -susceptible reference biotypes showed that a dose of 2 L ha(-1) metamitron was suitable for discrimination. A psbA PCR-RFLP was developed, based on the presence of a FspBI restriction enzyme recognition site, covering D1 codon 264 in susceptible genotypes. A paper-based DNA extraction allowed direct processing of leaf samples already in the field. In order to detect the mutation even in mixed seed samples, a nested PCR-RFLP was also developed. The method allows exhaustive surveys screening C. album leaf or seed samples for the occurrence of the D1 Ser264Gly mutation to confirm or disprove metamitron resistance in the case of unsatisfactory control.