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Target-site basis for fomesafen resistance in redroot pigweed (Amaranthus retroflexus) from China

Published online by Cambridge University Press:  22 February 2021

Long Du
Research Assistant, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Xiao Li
Research Assistant, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Xiaojing Jiang
Research Assistant, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Qian Ju
Research Assistant, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Wenlei Guo
Research Assistant, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangdong Provincial Key Laboratory of High Technology for Plant Protection, Guangzhou, China
Lingxu Li
Associate Professor, College of Plant Health and Medicine, Qingdao Agricultural University, Qingdao, China
Chunjuan Qu*
Associate Professor, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Mingjing Qu*
Professor, Pest Bio-control Laboratory, Shandong Peanut Research Institute, Qingdao, China
Authors for correspondence: Mingjing Qu, Shandong Peanut Research Institute, No. 126, Wannianquan Road, Qingdao, China. Email:; Chunjuan Qu, Shandong Peanut Research Institute, No. 126, Wannianquan Road, Qingdao, China. Email:
Authors for correspondence: Mingjing Qu, Shandong Peanut Research Institute, No. 126, Wannianquan Road, Qingdao, China. Email:; Chunjuan Qu, Shandong Peanut Research Institute, No. 126, Wannianquan Road, Qingdao, China. Email:


Redroot pigweed (Amaranthus retroflexus L.) is a dominant weed in soybean [Glycine max (L.) Merr.] fields in Heilongjiang Province, China. High selective pressure caused by the extensive application of the protoporphyrinogen oxidase (PPO)-inhibiting herbicide fomesafen has caused A. retroflexus to evolve resistance to this herbicide. Two susceptible and two resistant populations (S1, S2, R1, and R2) were selected in this study to illustrate the target-site resistance mechanism in resistant A. retroflexus. Whole-plant bioassays indicated that R1 and R2 had evolved high-level resistance to fomesafen, with resistance factors of 27.0 to 27.9. Sequence alignment of the PPO gene showed an Arg-128-Gly substitution in PPX2. The basal expression differences of PPX1 and PPX2 between the S1 and R1 plants were essentially nonsignificant, whereas the basal expression of PPX2 in R2 plants was slightly lower than in S1 plants. Compared with the PPX1 gene, the PPX2 gene maintained higher expression in the resistant plants after treatment with fomesafen. An enzyme-linked immunosorbent assay showed a similar basal PPO content between the susceptible and resistant plants without treatment. After fomesafen treatment, the PPO content decreased sharply in the susceptible plants compared with the resistant plants. Furthermore, after 24 h of treatment, the resistant plants showed increased PPO content, whereas the susceptible plants had died. The PPO2 mutation resulted in high extractable PPO activity and low sensitivity to fomesafen along with changes in PPO enzyme kinetics. Although the mutant PPO2 exhibited increased Km values in the resistant plants, the Vmax values in these plants were also increased. Changes in the properties of the PPO enzyme due to an Arg-128-Gly substitution in PPX2, including changes in enzyme sensitivity and enzyme kinetics, are the target-site mechanism of resistance in A. retroflexus.

Research Article
© The Author(s), 2021. Published by Cambridge University Press on behalf of the Weed Science Society of America

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These authors contributed equally to this work.

Associate Editor: Mithila Jugulam, Kansas State University


Bradford, MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248254 CrossRefGoogle ScholarPubMed
Chen, J, Huang, Z, Zhang, C, Huang, H, Wei, S, Chen, J, Wang, X (2015) Molecular basis of resistance to imazethapyr in redroot pigweed (Amaranthus retroflexus L.) populations from China. Pestic Biochem Physiol 124:4347 CrossRefGoogle ScholarPubMed
Collavo, A, Panozzo, S, Lucchesi, G, Scarabel, L, Sattin, M (2011) Characterisation and management of Phalaris paradoxa resistant to ACCase-inhibitors. Crop Prot 30:293299 CrossRefGoogle Scholar
Dayan, FE, Barker, A, Tranel, PJ (2018) Origins and structure of chloroplastic and mitochondrial plant protoporphyrinogen oxidases: implications for the evolution of herbicide resistance. Pest Manag Sci 74:22262234 CrossRefGoogle ScholarPubMed
Dayan, FE, Daga, PR, Duke, SO, Lee, RM, Tranel, PJ, Doerksen, RJ (2010) Biochemical and structural consequences of a glycine deletion in the alpha-8 helix of protoporphyrinogen oxidase. Biochim Biophys Acta 1804:15481556 CrossRefGoogle ScholarPubMed
Dayan, FE, Owens, DK, Tranel, PJ, Preston, C, Duke, SO (2014) Evolution of resistance to phytoene desaturase and protoporphyrinogen oxidase inhibitors—state of knowledge. Pest Manag Sci 70:13581366 CrossRefGoogle ScholarPubMed
Deybach, J-C, Silva, VD, Grandchamp, B, Nordmann, Y (1985) The mitochondrial location of protoporphyrinogen oxidase. Eur J Biochem 149:431435 CrossRefGoogle ScholarPubMed
Emanuelesson, O, Nielsen, H, Brunak, S, Heijne, G von (2000) Predicting subcellular localization of proteins based on their N-terminal amino acid sequence. J Mol Biol 300:10051016 CrossRefGoogle Scholar
Gaines, TA, Zhang, W, Wang, D, Bukun, B, Chisholm, ST, Shaner, DL, Nissen, SJ, Patzoldt, WL, Tranel, PJ, Culpepper, AS, Grey, TL, Webster, TM, Vencill, WK, Sammons, RD, Jiang, J, et al. (2010) Gene amplification confers glyphosate resistance in Amaranthus palmeri . Proc Natl Acad Sci USA 107:10291034 CrossRefGoogle ScholarPubMed
Giacomini, DA, Umphres, AM, Nie, H, Mueller, TC, Steckel, LE, Young, BG, Scott, RC, Tranel, PJ (2017) Two new PPX2 mutations associated with resistance to PPO-inhibiting herbicides in Amaranthus palmeri . Pest Manag Sci 73:15591563 CrossRefGoogle ScholarPubMed
Hao, GF, Tan, Y, Yang, SG, Wang, ZF, Zhan, CG, Xi, Z, Yang, GF (2013) Computational and experimental insights into the mechanism of substrate recognition and feedback inhibition of protoporphyrinogen oxidase. PLoS ONE 8:e69198 CrossRefGoogle ScholarPubMed
Hao, GF, Tan, Y, Xu, WF, Cao, RJ, Xi, Z, Yang, GF (2014) Understanding resistance mechanism of protoporphyrinogen oxidase-inhibiting herbicides: insights from computational mutation scanning and site-directed mutagenesis. J Agric Food Chem 62:72097215 CrossRefGoogle ScholarPubMed
Heap, I (2020) The International Survey of Herbicide Resistant Weeds. Accessed: August 1, 2020Google Scholar
Heinemann, IU, Diekmann, N, Masoumi, A, Koch, M, Messerschmidt, A, Jahn, M, Jahn, D (2007) Functional definition of the tobacco protoporphyrinogen IX oxidase substrate-binding site. Biochem J 402:575580 CrossRefGoogle ScholarPubMed
Huang, Z, Chen, J, Zhang, C, Huang, H, Wei, S, Zhou, X, Chen, J, Wang, X (2016) Target-site basis for resistance to imazethapyr in redroot amaranth (Amaranthus retroflexus L.). Pestic Biochem Physiol 128:1015 CrossRefGoogle ScholarPubMed
Huang, Z, Cui, H, Wang, C, Wu, T, Zhang, C, Huang, H, Wei, S (2020) Investigation of resistance mechanism to fomesafen in Amaranthus retroflexus L. Pestic Biochem Physiol 165, 10.1016/j.pestbp.2020.104560 CrossRefGoogle ScholarPubMed
Ishida, S, Miller-Sulger, R, Kohno, H, Böger, P, Wakabayashi, K (2000) Enzymatic activity of protoporphyrinogen-IX oxidase from various plant species: its sensitivity to peroxidizing herbicides. J Pest Sci 25:1823 CrossRefGoogle Scholar
Jacobs, JM, Jacobs, NJ (1993) Porphyrin accumulation and export by isolated barley (Hordeum vulgare) plastids (effect of diphenyl ether herbicides). Plant Physiol 101:11811187 CrossRefGoogle Scholar
Jacobs, JM, Jacobs, NJ, Sherman, TD, Duke, SO (1991) Effect of diphenyl ether herbicides on oxidation of protoporphyrinogen to protoporphyrin in organellar and plasma membrane enriched fractions of barley. Plant Physiol 97:197203 CrossRefGoogle ScholarPubMed
Koch, M, Breithaupt, C, Kiefersauer, R, Freigang, J, Huber, R, Messerschmidt, A (2004) Crystal structure of protoporphyrinogen IX oxidase: a key enzyme in haem and chlorophyll biosynthesis. EMBO 23:17201728 CrossRefGoogle ScholarPubMed
Lee, HJ, Duke, SO (1994) Protoporphyrinogen IX-oxidizing activities involved in the mode of action of peroxidizing herbicides. J Agric Food Chem 42:26102618 CrossRefGoogle Scholar
Lee, RM, Hager, AG, Tranel, PJ (2008) Prevalence of a novel resistance mechanism to PPO-inhibiting herbicides in waterhemp (Amaranthus tuberculatus). Weed Sci 56:371375 Google Scholar
Legleiter, TR, Bradley, KW, Massey, RE (2009) Glyphosate-resistant waterhemp (Amaranthus rudis) control and economic returns with herbicide programs in soybean. Weed Technol 23:54–61CrossRefGoogle Scholar
Lermontova, I, Kruse, E, Mock, H-P, Grimm, B (1997) Cloning and characterization of a plastidal and a mitochondrial isoform of tobacco protoporphyrinogen IX oxidase. Proc Natl Acad Sci USA 94:88958900 CrossRefGoogle Scholar
Nicolaus, B, Sandmann, G, Böger, P (1993) Molecular aspects of herbicide action on protoporphyrinogen oxidase. Z Naturforsch C J Biosci 48:326333 CrossRefGoogle Scholar
Nie, H, Mansfield, BC, Harre, NT, Young, JM, Steppig, NR, Young, BG (2019) Investigating target-site resistance mechanism to the PPO-inhibiting herbicide fomesafen in waterhemp and interspecific hybridization of Amaranthus species using next generation sequencing. Pest Manag Sci 75:32353244 CrossRefGoogle ScholarPubMed
Obenland, O, Ma, R, O’Brien, S, Lygin, AV, Riechers, DE (2017) Resistance to carfentrazone-ethyl in tall waterhemp. Pages 30–31 in Proceedings of the 72nd Annual Meeting of the North Central Weed Science Society. St Louis, MO: North Central Weed Science Society of America Google Scholar
Odell, JT, Caimi, PG, Yadav, NS, Mauvais, CJ (1990) Comparison of increased expression of wild-type and herbicide-resistant acetolactate synthase genes in transgenic plants, and indication of posttranscriptional limitation on enzyme activity. Plant Physiol 94:16471654 CrossRefGoogle ScholarPubMed
Patzoldt, WL, Hager, AG, McCormick, JS, Tranel, PJ (2006) A codon deletion confers resistance to herbicides inhibiting protoporphyrinogen oxidase. Proc Natl Acad Sci USA 103:1232912334 CrossRefGoogle ScholarPubMed
Poulson, R, Polglase, WJ (1975) The enzymic conversion of protoporphyrinogen IX to protoporphyrin IX. J Biol Chem 250:12691274 CrossRefGoogle ScholarPubMed
Powles, SB (2010) Gene amplification delivers glyphosate-resistant weed evolution. Proc Natl Acad Sci USA 107:955956 CrossRefGoogle ScholarPubMed
Powles, SB, Yu, Q (2010) Evolution in action: plants resistant to herbicides. Annu Rev Plant Biol 61:317347 CrossRefGoogle ScholarPubMed
Rangani, G, Salas-Perez, RA, Aponte, RA, Knapp, M, Craig, IR, Mietzner, T, Langaro, AC, Noguera, MM, Porri, A, Roma-Burgos, N (2019) A novel single-site mutation in the catalytic domain of protoporphyrinogen oxidase IX (PPO) confers resistance to PPO-inhibiting herbicides. Front Plant Sci 10:568 CrossRefGoogle ScholarPubMed
Rousonelos, SL, Lee, RM, Moreira, MS, Vangessel, MJ, Tranel, PJ (2012) Characterization of a common ragweed (Ambrosia artemisiifolia) population resistant to ALS- and PPO-inhibiting herbicides. Weed Sci 60:335344 Google Scholar
Salas, RA, Burgos, NR, Tranel, PJ, Singh, S, Glasgow, L, Scott, RC, Nichols, RL (2016) Resistance to PPO-inhibiting herbicide in Palmer amaranth from Arkansas. Pest Manag Sci 72:864869 CrossRefGoogle ScholarPubMed
Seefeld, SS, Jensen, JE, Fuerst, EP (1995) Log-logistic analysis of herbicide dose-response relationships. Weed Technol 9:218227 CrossRefGoogle Scholar
Shoup, DE, Al-Khatib, K, Peterson, DE (2003) Common waterhemp (Amaranthus rudis) resistance to protoporphyrinogen oxidase-inhibiting herbicides. Weed Sci 51:145150 CrossRefGoogle Scholar
Tranel, PJ (2002) Resistance of weeds to ALS-inhibiting herbicides: what have we learned. Weed Sci 50:700712 CrossRefGoogle Scholar
Trezzi, M, Felippi, C, Mattei, D, Silva, H, Nunes, A, Debastiani, C, Vidal, R, Marques, A (2005) Multiple resistance of acetolactate synthase and protoporphyrinogen oxidase inhibitors in Euphorbia heterophylla biotypes. J Environ Sci Health B 40:101109 CrossRefGoogle ScholarPubMed
Varanasi, VK, Brabham, C, Korres, NE, Norsworthy, JK (2019) Nontarget site resistance in Palmer amaranth [Amaranthus palmeri (S.) Wats.] confers cross-resistance to protoporphyrinogen oxidase-inhibiting herbicides. Weed Technol 33:349354 CrossRefGoogle Scholar
Varanasi, VK, Brabham, C, Norsworthy, JK (2018a) Confirmation and characterization of non–target site resistance to fomesafen in Palmer amaranth (Amaranthus palmeri). Weed Sci 66:702709 Google Scholar
Varanasi, VK, Brabham, C, Norsworthy, JK, Nie, H, Young, BG, Houston, M, Barber, T, Scott, RC (2018b) A Statewide survey of PPO-inhibitor resistance and the prevalent target-site mechanisms in palmer amaranth (Amaranthus palmeri) accessions from Arkansas. Weed Sci 66:149158 Google Scholar
Wang, H, Guo, W, Zhang, L, Zhao, K, Ge, L, Lv, X, Liu, W, Wang, J (2017) Multiple resistance to thifensulfuron-methyl and fomesafen in redroot pigweed (Amaranthus retroflexus L.) from China. Chilean J Agric Res 77:311317 Google Scholar
Wang, H, Wang, H, Zhao, N, Zhu, B, Sun, P, Liu, W, Wang, J (2019) Multiple resistance to PPO and ALS inhibitors in redroot pigweed (Amaranthus retroflexus). Weed Sci 68:18 Google Scholar
Watanabe, N, Che, FS, Iwano, M, Takayama, S, Yoshida, S, Isogai, A (2001) Dual targeting of spinach protoporphyrinogen oxidase II to mitochondria and chloroplasts by alternative use of two in-frame initiation codons. J Biol Chem 276:2047420481 CrossRefGoogle ScholarPubMed
Yang, S, Hao, G, Dayan, FE, Tranel, PJ, Yang, G (2013) Insight into the structural requirements of protoporphyrinogen oxidaseinhibitors: molecular docking and CoMFA of diphenyl ether, isoxazole phenyl, and pyrazole phenyl ether. Chin J Chem 31:11531158 CrossRefGoogle Scholar
Young, BG (2006) Changes in herbicide use patterns and production practices resulting from glyphosate-resistant crops. Weed Technol 20:301307 CrossRefGoogle Scholar
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