Identificação e caracterização de biótipos de leiteira resistentes aos herbicidas inibidores da enzima HPPD

Abstract

Wild poinsettia (Euphorbia heterophylla) is one of the main troublesome weed of summer crops, due to the intrinsic aspects of competitiveness, combined with the occurrence of resistant biotypes. Dose-response curve studies demonstrated unsatisfactory control of some biotypes by hydroxyphenyl pyruvate dioxygenase (HPPD) inhibitors, suggesting the occurrence of resistant biotypes. Thus, the objectives of this work were to identify and characterize wild poinsettia resistance, elucidate resistance mechanisms, verify changes in plant metabolism and characterize the inheritance of resistance of Euphorbia heterophylla to HPPD-inhibiting herbicides. To this end, a series of studies were carried out that were framed in four chapters, in which they were carried out from March 2018 to April 2022, in a greenhouse and laboratories at the Federal University of Pelotas. Initially, seeds collected in different regions of Rio Grande do Sul state were evaluated for resistance to the herbicide tembotrione, and control maps, dose-response curves, cross-resistance investigation and alternative control were carried out (chapter 1). To elucidate the mechanisms of resistance involved, studies of absorption, translocation, metabolism and identification of known metabolites of tembotrione were evaluated, as well as the effect of inhibition of P450 enzymes under exposure to the herbicide tembotrione and the quantification of the expression of the HPPD gene (chapter two). To verify the changes in plant metabolism between the biotypes, aspects related to photosynthesis, chlorophyll fluorescence, pigments, cellular integrity and activity of antioxidant enzymes were quantified (chapter 3). The mode and type of inheritance of resistance was determined for the wild poinsettia biotypes resistant to the HPPD-inhibiting herbicides (chapter 4). The results showed that biotype 32.2 presents multiple resistance to HPPD and PPO inhibitors and biotype 32.1 presents multiple resistance to HPPD, PPO and ALS. The pre-emergent atrazine, flumioxazin and sulfentrazone and the post-emergent atrazine, paraquat, paraquat+diuron, saflufenacil, glyphosate, ammonium glufosinate demonstrated satisfactory control of all wild poinsettia biotypes. Wild poinsettia biotypes resistant to HPPD inhibitors present as a mechanism of resistance the rapid metabolism of the herbicide tembotrione, to a non-toxic metabolite called TCMBA, and no differential responses were observed between biotypes regarding absorption, P450 inhibition and HPPD gene expression. The increase in exposure of the susceptible biotype to tembotrione results in a rapid increase in F0 and a marked reduction in Fm and Fv/Fm, compared to resistant biotypes. Biotypes resistant to HPPD inhibitors show a smaller decline in photosynthetic variables (A, Gs, E, CE and iWUE), in addition to a smaller increase in Ci, in relation to the susceptible biotype, during the period of exposure to the herbicide tembotrione. The increase in the period of exposure to the herbicide tembotrione resulted in a significant increase in oxidative stress in the susceptible biotype compared to the resistant ones, as evidenced by the increase in TBARs, H2O2, cellular extravasation and lower carotenoid content. The higher activity of APX 96 hours after application of tembotrione, in resistant biotypes resulted in lower levels of H2O2 and less damage to cellular components. The inheritance of tembotrione herbicide resistance is complex, being encoded by incompletely dominant nuclear genes. Heterozygotes have intermediate sensitivity to the herbicide tembotrione, where F1 R♂ x S♀ has a behavior closer to the susceptible parent, and F1 S♂ x R♀ is closer to the resistant parent.