Multiple Resistance Mechanisms Involved in Glyphosate Resistance in Eleusine indica

Glyphosate is a non-selective herbicide and is widely used for weed control in non-cultivated land in China. One susceptible (S) and five putative glyphosate-resistant (R1, R2, R3, R4, and R5) Eleusine indica biotypes were selected to investigate their resistance levels and the potential resistance mechanisms. Based on the dose-response assays, the R3 and R5 biotypes showed a low-level (2.4 to 3.5-fold) glyphosate resistance, and the R1, R2, and R4 biotypes exhibited a moderate- to high-level (8.6 to 19.2-fold) resistance, compared with the S biotype. The analysis of the target-site resistance (TSR) mechanism revealed that the P106A mutation and the heterozygous double T102I + P106S mutation were found in the R3 and R4 biotypes, respectively. In addition, the similar EPSPS gene overexpression was observed in the R1, R2, and R5 biotypes, suggesting that additional non-target-site resistance (NTSR) mechanisms may contribute to glyphosate resistance in R1 and R2 biotypes. Subsequently, an RNA-Seq analysis was performed to identify candidate genes involved in NTSR. In total, ten differentially expressed contigs between untreated S and R1 or R2 plants, and between glyphosate-treated S and R1 or R2 plants, were identified and further verified with RT-qPCR. One ATP-binding cassette (ABC) transporter gene, one aldo-keto reductases (AKRs) gene and one cytochrome P450 monooxygenase (CytP450) gene were up-regulated in R1 or R2 plants. These results indicated that EPSPS overexpression, single or double mutation was a common TSR mechanisms in E. indica. Additional NTSR mechanisms could play an essential role in glyphosate resistance. Three genes, ABCC4, AKR4C10, and CYP88, could serve as important candidate genes and deserve further functional studies.