MINISTRY OF EDUCATION AND TRAINING CAN THO UNIVERSITY SUMMARY OF DOCTORAL DISSERTATION Specialization: Plant Protection Code: 9620112 LE DUY STUDY ON THE RESISTANCE MECHANISM OF BARNYARDGRASS (Echinochloa crus-galli (L.) Beauv.) TO QUINCLORAC IN THE MEKONG DELTA OF VIETNAM Can Tho, 2018 The dissertation was completed at College of Agriculture and Applied biology, Can Tho University Scientific supervisor: Associate Professor PhD Nguyen Minh Chon Reviewer 1: Reviewer 2: Reviewer 3: .…………… The thesis is defended in front of the University Examination Council in Can Tho University Time:………… …………… Date:…………….…………… Further information of the thesis could be found at: Learning Resource Center of Can Tho University National library of Vietnam PUBLISHED RESEARCH Duy Le, C M Nguyen, R K Mann, C N Yerkes and B V N Kumar 2017 Genetic diversity and herbicide resistance of 15 Echinochloa crus-galli populations to quinclorac in Mekong Delta of Vietnam and Arkansas of United States Journal of Plant Biotechnology, 44: 472-477 Duy Le, C M Nguyen, R K Mann, B V N Kumar and M Morell 2018 Efficacy of Rinskor (florpyrauxifen-benzyl ester) on Herbicide Resistant Barnyardgrass (Echinochloa crus-galli) in Rice Fields of Mekong Delta, Vietnam Journal of Crop Science and Biotechnology, 21: 75-81 Duy Le, C M Nguyen, B V N Kumar and R K Mann 2018 Weed management practices to control herbicideresistant Echinochloa crus galli in rice in Mekong Delta, Vietnam Research On Crops , 19: 20-27 Chapter 1: INTRODUCTION 1.1 Problem statement The barnyardgrass (Echinochloa crus-galli) had been considered as the most notorious for a long time, as a C4 plant, the growth rate of barnyardgrass is far dominant to rice In addition, Echinochloa crus-galli and Echinochloa colona can mimic the rice appearance at the early stage of weed seedlings and make it hard to control by hand weeding Herbicide is an effective solution for controlling the barnyardgrass, however, the herbicide resistant barnyardgrass is a critical issue to the weed management in the rice field The barnyardgrass could evolve resistance to several current herbicide active ingredients Further research about herbicide resistance in barnyardgrass is critical As an outcome, the dissertation of “Study on the resistance mechanism of barnyardgrass (Echinochloa crus-galli) to quinclorac in the Mekong Delta of Vietnam” was proposed 1.2 Objective of dissertation (1) Evaluate the morphological variability and genetic diversity of Echinochloa spp populations in Mekong Delta of Vietnam (2) Evaluate the herbicide-resistance level of Echinochloa spp to bispyribac, penoxsulam and quinclorac in rice fields at the Mekong Delta of Vietnam (3) Explain the biochemical mechanism and the molecular mechanism of quinclorac-resistance in Echinochloa crus-galli 1.3 Major research topics of the dissertation (1) Survey the rice cultivation and weed management practice in rice fields at the Mekong Delta of Vietnam (2) Evaluate the diversity of Echinochloa spp population in rice fields at the Mekong Delta of Vietnam (3) Evaluate the herbicide-resistance level of the collected Echinochloa spp samples to bispyribac-sodium, penoxsulam and quinclorac by dose-response screening method (4) Evaluate the efficacy of rinskor, a new herbicide against the herbicide-resistant Echinochloa crus-galli to find new effective herbicide for current herbicide-resistant barnyardgrass (5) Use RAPD analysis to evaluate the genetic diversity of Echinochloa crus-galli populations, and the correspondence of quinclorac-resistance and genetic distance of Echinochloa crus-galli populations in the Mekong delta (6) Measure the activity of β-cyanoalanine synthase in the leaf tissue of Echinochloa crus-galli to study the biochemical mechanism of quinclorac-resistance in barnyardgrass 1.4 Time and location Time of research was from 2014 to 2018 Experiment 1; and was carried out at households in College of Agriculture and Applied Biology, Can Tho University; Experiment 4; and in Discovery Center of Dow AgroSciences in Indianapolis, Indiana State, United States of America 1.5 Novel aspects The research has confirmed the existence of herbicideresistant Echinochloa spp populations in Mekong Delta of Vietnam This research also evaluates the relationship between farmers’ weed management practice and the herbicide-resistance, therefore, the practical solutions for herbicide-resistant weed were also determined and suggested in the dissertation The mechanisms of quinclorac-resistance in barnyardgrass were confirmed and elucidated at enzyme and molecular level, through the measurement of quinclorac detoxifying enzyme activity and its gene expression level in barnyardgrass The results establish important information for the further study about the mechanism of the herbicide-resistance in weeds 1.6 Layout The dissertation consists of 102 pages (21 tables, 38 pictures and 202 references) In that, introduction: pages, background: 33 pages; materials and methods: 18 pages; results and discussion 43 pages; conclusions and suggestions: page; reference: 18 pages; appendix 42 pages for both mumber and pictures Chapter 2: BACKGROUND OF DISSERTATION The genus Echinochloa includes over 250 species, and most of these are considered as weeds The barnyardgrass (Echinochloa spp.) is C4 monocots and originated from Europe and has been now documented worldwide The morphology of Echinochloa spp is highly diverse, in many cases, there are several species confused with one another In Asia, rice fields have three main barnyardgrass species, Echinochloa crus-galli, Echinocloa colona, and Echinochloa glabrescens, which found in Vietnam The bispyribac and penoxsulam are two popular ALS inhibitor herbicide on rice, those two herbicide is mainly used for barnyardgrass control in Vietnam Quinclorac is a synthetic auxin herbicide, and the product contains quinclorac is wide used for controlling barnyardgrass in the rice field Rinskor is a new synthetic auxin herbicide, and this herbicide is not used for weed management in Vietnam when the dissertation started in 2014 The weed seed of 78 populations of Echinochloa spp collected from rice field in provinces of the Mekong Delta of Vietnam, the seedling from collected seed was tested with bispyribac, penoxsulam, quinclorac and rinskor to evaluating the LD90 and herbicide resistance level To continue the study on the mechanism of resistance, 15 selected weed populations were selected for genetic distance analysis by using the RAPD method The activity of cyanoalanide synthase was analyzed to evaluate the enzyme level in leaf of different weed populations under the treatment of quinclorac Finally, the expression level of mRNA of cyanoalanide synthase in populations was measured by the comparative methods of 2−ΔCt, the activity of enzyme and the expression level of mRNA in the populations was analyzed to find the association of two value, the result of this research was used to supporting the study of herbicide mechanism of quinclorac in barnyardgrass CHAPTER 3: MATERIALS AND METHODS 3.1 Experiment Survey on farmer practice in rice cultivation and weed management in the Mekong Delta Target of research: Study on the farmer practice of weed management on the rice field in Mekong Delta of Vietnam Barnyardgrass seed sample was collected from total 90 rice field, after germination test in greenhouse, 78 quality samples were selected for further experiments During the weed seed sample collecting on the field, the researcher had interviewed the field owner about the cultivation practice in the field, the questions were included questions about: - Number of rice seasons per year - Field size that he/she is growing rice (hectare) - Number of herbicide application per season - Condition of water management - Perception about the important weed species in rice field - Most popular herbicides used for controlling barnyardgrass - Cost for weed management per hectare per season - Cost for herbicide per hectare per season - Cost for hand-weeding per hectare per season The questions were designed to get essential information about the field condition, and the weed management practice of the field owner, the questions were asked after seed sample collected, there were 17 seed samples collected without input from farmer Statistical analysis After cross-check, the inputs with unreliable information or unclear data were excluded from the final analysis, final answer from 71 farmers were processed for analysis Collected data were analyzed using ANOVA with mean comparison by Tukey-Kramer test at α=0.05 One way ANOVA ttest with p