Molecular and physiological characterization of six‐way resistance in an <i>Amaranthus tuberculatus</i> var. <i>rudis</i> biotype from Missouri

<jats:title>Abstract</jats:title><jats:sec><jats:title>BACKGROUND</jats:title><jats:p>Previous research reported the first case of six‐way herbicide resistance in a common waterhemp (<jats:italic>Amaranthus tuberculatus</jats:italic> var. <jats:italic>rudis</jats:italic>) biotype from Missouri, USA designated MO‐Ren. This study investigated the mechanisms of multiple‐resistance in the MO‐Ren biotype to herbicides from six site‐of‐action (SOA) groups, i.e. synthetic auxins, 5‐enolypyruvyl‐shikimate‐3‐phosphate synthase (EPSPS)‐, protoporphyrinogen oxidase (PPO)‐, acetolactate synthase (ALS)‐, photosystem II (PSII)‐, and 4‐hydroxyphenyl‐pyruvate‐dioxygenase (HPPD)‐inhibitors.</jats:p></jats:sec><jats:sec><jats:title>RESULTS</jats:title><jats:p>Genomic DNA sequencing confirmed the presence of known mutations associated with ALS‐ or PPO‐inhibiting herbicide resistance: the Trp‐574‐Leu amino acid substitution in the ALS enzyme and the codon deletion corresponding to the ΔG210 in the PPX2 enzyme. No target‐site point mutations associated with resistance to PSII‐ and EPSPS‐inhibitors were detected. Quantitative polymerase chain reaction (qPCR) indicated that MO‐Ren plants contained five‐fold more copies of the <jats:italic>EPSPS</jats:italic> gene than susceptible plants. Malathion in combination with 2,4‐D (2,4<jats:italic>‐</jats:italic>dichlorophenoxyacetic acid), mesotrione, and chlorimuron POST enhanced the activity of these herbicides indicating that metabolism due to cytochrome P450 monooxygenase activity was involved in herbicide resistance. 4‐Chloro‐7‐nitrobenzofurazan (NBD‐Cl), a glutathione‐<jats:italic>S</jats:italic>‐transferase (GST)‐inhibitor, in combination with atrazine did not reduce the biomass accumulation. Reduced absorption or translocation of 2,4‐D did not contribute to resistance. However, the resistant biotype metabolized 2,4‐D, seven‐ to nine‐fold faster than the susceptible.</jats:p></jats:sec><jats:sec><jats:title>CONCLUSION</jats:title><jats:p>Target‐site point mutations, gene amplification, and elevated rates of metabolism contribute to six‐way resistance in the MO‐Ren biotype, suggesting both target site and non‐target site mechanisms contribute to multiple herbicide resistance in this <jats:italic>Amaranthus tuberculatus</jats:italic> biotype. © 2018 Society of Chemical Industry</jats:p></jats:sec>