Simultaneous Occurrence of Two Proline-197-Threonine (Pro-197-Thr) and Tryptophan-574-Leucine (Trp-574-Leu) Mutations Associated with Resistance to Iodosulfuron-Methyl-Sodium+Mesosulfuron-Methyl in Annual Ryegrass (Lolium rigidum Gaudin)

Document Type : Research Paper

Authors

1 M.Sc. of Weed Science, Faculty of Agriculture and Natural Resources, University of Mohaghegh Ardabili, Ardabil, Iran.

2 University of Ardebil

3 Iranian Research Institute of Plant Protection

4 Assistant Professor, Iranian Research Iinstitute of Plant Protection

5 Assistant Professor, Department of Genetic Engineering and Biosafety, Agricultural Biotechnology Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

10.22034/ijws.2024.133953

Abstract

The present study aimed to investigate the probability of mutation occurrence at the target site of acetolactate synthase (ALS) inhibitors in annual ryegrass. For this purpose, annual ryegrass seeds were collected from a wheat field in Golestan province, where the occurrence of resistance had been reported. A whole-plant assay was conducted in a greenhouse using the recommended rate (1.5 L ha⁻¹) of the herbicide iodosulfuron-methyl-sodium+mesosulfuron-methyl (Atlantis, OD 1.2%). Four weeks after treatment, samples were collected to evaluate the resistance ratio and study the molecular resistance mechanism at the target site. DNA extraction, amplification, and purification of the ALS gene from the total genomic DNA of the L183 population were performed using PCR and electrophoresis. The excised gel fragment containing the ALS gene was sent for sequencing to facilitate further molecular investigations. The results indicated that the L183 annual ryegrass population, with a survival rate of 89.29%, a dry weight percentage of 86.22%, and a fresh weight reduction of 9.28% compared to the untreated control, was highly resistant (RRR) to iodosulfuron-methyl-sodium+mesosulfuron-methyl. Molecular sequencing of the ALS gene in the L183 ryegrass population, compared to the reference ALS gene of Arabidopsis, revealed the potential simultaneous presence of two mutations. These involved the substitution of proline with threonine at codon 197 (Pro-197-Thr) and the substitution of tryptophan with leucine at codon 574 (Trp-574-Leu). These mutations were identified in the bidirectional sequencing of the ALS gene in the L183 annual ryegrass population from Golestan province, and these alleles are likely responsible for resistance. Based on the screening and sequencing results, the occurrence of known two known point mutations, Pro-197-Thr and Trp-574-Leu, in the ALS gene of the L183 annual ryegrass population led to changes in the amino acids of the target protein. These nucleotide alterations caused reduced effectiveness of the herbicide iodosulfuron-methyl-sodium+mesosulfuron-methyl and the emergence of resistance.

Keywords

Iranian Journal of Weed Science
Volume 20, Issue 1
June 2024
Pages 147-164

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