**5. Weed resistance to herbicide EPSPS inhibitors, glyphosate**

Glyphosate was discovered and developed as a non‐selective herbicide by Chemical Company Monsanto in 1974. N‐(phosphonometil) glycine, the active ingredient in glyphosate, is a deri‐ vate of the amino acid glycine and phosphonic acid. It's mode of action in relation to the enzyme EPSPS (5‐enolpyruvylshikimate‐3‐phosphate synthase) preventing the biosynthesis of the aromatic amino acids required for the production of growth regulators, anthocyanins, phenolics and proteins [146]. The site of action is located in the chloroplast and it was con‐ firmed 8 years after glyphosate introduction [146]. Broad weed spectrum (annual and peren‐ nial, monocots and dicots), high efficacy, lack of soil activity and low mammalian toxicity are key characteristics that make glyphosate the world's most widely used herbicide [147]. Because glyphosate is inherently non‐selective, selectivity has often been achieved by place‐ ment and timing, for example, as a pre‐plant or pre‐emergence herbicide for the control of weeds in no‐till systems and for turf‐grass renovation [104]. The introduction of genetically modified glyphosate resistant crops in the United States and other parts of the world [148] has led to enormous increase of glyphosate use on arable land (cotton, canola, corn, wheat, sugar beets, potatoes, etc.) as a post‐emergence herbicide.

Glyphosate‐resistant weeds were not found during the first 15 years of glyphosate use (1972–1997). Based on the resistance risk criteria for assessing the risk of developing weed resistance to glyphosate, it was estimated that the glyphosate has low risk for the evolution of weed resistance [149]. However, in the last 19 years (1998–2016), glyphosate resistance in 36 weed species was confirmed and according to the decades, it looks like this: 2 (first decade), 18 (second) and 16 species (the last, third) (**Table 2**) [7]. Mechanism of glyphosate resistance to weed species includes target‐site mutation, target‐site gene amplification/expression, active vacuole sequestration, limited cellular uptake and a rapid necrosis response [21].

In a number of cases of confirmed weed resistance to glyphosate, the resistance was based on some different mechanisms which include non‐target‐site (limited absorption and transloca‐ tion, vacuolar sequestration) and target‐site resistance (amino acid substitution, ESPSP gene expression/amplification, altered enzyme activity) (**Table 6**). Generally, usually confirmed


**5. Weed resistance to herbicide EPSPS inhibitors, glyphosate**

**Table 5.** Confirmed mechanisms of resistance to herbicide ACC‐ase inhibitors in some weed species.

**Weed species Mechanism of resistance**

*Alopecurus myosuroides* Huds. Ile‐1781‐Leu, Trp‐2027‐Cys, Ile‐2041‐

*Avena fatua* L. Ile‐1781‐Leu, Trp‐1999‐Cys,Trp‐2027‐

*Avena sterilis* L. Ile‐1781‐Leu, Trp‐1999‐Cys, Trp‐2027‐

*Eleusine indica* (L.) Gaertn Asp‐2078‐Gly, Thr‐1805‐Ser [131] *Hordeum glaucum* (Steud.) Tzvelev Ile‐1781‐Leu, Gly‐2096‐Ala [132]

*Lolium multiflorum* Lam. Ile‐1781‐Leu (Ile‐418‐Leu), Cys‐2088‐

*Lolium rigidum* Gaud. Ile‐1781‐Leu, Ile‐2041‐Asn, Asp‐2078‐

*Lolium* sp. Ile‐1781‐Leu, Trp‐1999‐Cys, Ile‐2041‐

*Pseudosclerochloa kengiana* Trp‐1999‐Ser [141]

[110, 112]

Asn, Asp‐2078‐Gly, Gly‐2096‐Ala

Cys, Ile‐2041‐Asn, Asp‐2078‐Gly, Cys‐2088‐Arg, Gly‐2096‐Ser [109, 114]

Cys, Ile‐2041‐Asn, Asp‐2078‐Gly,

*Echinochloa crus‐galli* (L.) Beauv. Ile‐1781‐Leu [128] Altered enzyme activity, gene

*Echinochloa colona* (L.) Link. altered enzyme activity [130]

*Hordeum leporinum* (Link) Arcang. Ile‐1781‐Leu, Gly‐2096‐Ala [132] Detoxification, altered enzyme

Ile‐1781‐Leu, Trp‐2027‐Cys, Ile‐2041‐ Asn, Asp‐2078‐Gly [137, 138]

Gly, Cys‐2088‐Arg, Gly‐2096‐Ala, Trp‐2027‐Cys [111, 116]

Asn/Val, Asp‐2078‐Gly, Cys‐2088‐ Arg, Gly‐2096‐Ala [113]

*Setaria viridis* L. Beauv. Ile‐1780‐Leu [142] Altered enzyme activity [143] *Sorghum halepense* (L.) Pers. Ile‐2041‐Asn [144] Altered enzyme activity [145]

Ile‐1781‐Leu, Ile‐2041‐Asn, Asp‐2078‐

Cys‐2088‐Arg [115]

Gly [126, 127]

Arg [134, 135]

*Alopecurus aequalis* Sobol. Ile‐1781‐Leu [119]

*Bechmannia syzigachne* (Steud.)

18 Herbicide Resistance in Weeds and Crops

*Lolium perenne* L. ssp. *multiflorum*

Fernald

Lam.

**Amino acid substitutions Other mechanisms of resistance**

[120–122]

Detoxification, gene expression

Detoxification [123, 124]

Detoxification [123, 125]

expression [129]

activity [133]

Detoxification [136]

Detoxification [139, 140]

Glyphosate was discovered and developed as a non‐selective herbicide by Chemical Company Monsanto in 1974. N‐(phosphonometil) glycine, the active ingredient in glyphosate, is a deri‐ vate of the amino acid glycine and phosphonic acid. It's mode of action in relation to the enzyme EPSPS (5‐enolpyruvylshikimate‐3‐phosphate synthase) preventing the biosynthesis


**Table 6.** Confirmed mechanisms of resistance to EPSPS inhibitor in some weed species.

cases of weed resistance to glyphosate were due to reduced absorption and translocation of the herbicide. Further, cDNA sequence analysis of the EPSPS gene indicated that resis‐ tance to glyphosate was based on substitution of proline with serine (Pro‐106‐Ser), alanine (Pro‐106‐Ala), threonine (Pro‐106‐Thr), or leucine (Pro‐106‐Leu)) at the position 106 of the EPSPS protein in many weed species (*Amaranthus tuberculatus*, *E. indica*, *E. colona*, *L. rigidum*, *Lolium multiflorum*, *P. annua*).

## **6. Management strategies for herbicide‐resistant weeds**

Strategy for herbicide‐resistance weed management must involve all the available preven‐ tive, cultural, mechanical and chemical measures for effective, safe and cost‐effective weed control [183]: (a) survey of present weed flora; (b) preventing weed seed production and reduction of weed seed in the soil seed‐bank; (c) prevention of the movement of seeds and vegetative propagules from field to field or from field margins (or lost field) to field; (d) keep arable and non‐arable land as weed free as possible; (e) sowing pure crop seeds; (f) growing competitive crops that can suppress weeds; (g) destruction of weed seeds in post‐harvest materials (e.g. Integrated Harrington Seed Destructor); (h) use mechanical and physical measures where appropriate; (i) using herbicides with different modes of action, tank mixtures and sequential applications; (j) use of recommended herbicide rate for certain number of weed populations; (k) adopting crop rotations that allow use of herbicides of alternative mode of action; (l) intensify research and professional commu‐ nication and grower education programs and (m) publish guidelines for managing anti‐ resistant strategy.

The state government sectors, universities and research institutes, technology development centres, farmers and other relevant stakeholders were called to proactively address emerging weed resistance problems and to develop cost‐effective resistance‐management strategy and practices that support effective weed control.
