**1. Introduction and general overview of resistance**

Since the introduction of 2,4‐D as a first selective herbicide in 1947, herbicides have had a major positive impact on weed management in all over the world. Unfortunately, herbicide resistance developed shortly after the introduction of the herbicides. The phenomenon of resistance can be defined as the decreased response of a species' population to herbicide [1].

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It is also defined as a 'survival of a segment of the population of a weed species following an herbicide dose lethal to the normal population' [2]. In addition, resistance can be defined as 'the inherited ability to survive treatment by a herbicide' [3], or it is a 'phenomen which occurs as a result of heritable changes to biochemical processes that enable weed species sur‐ vival when treated with a herbicide' [4].

Weed resistance to herbicides is a normal and predictable outcome of natural selection. In that context, rare mutations that confer herbicide resistance exist in wild/weed populations before any herbicide introduction. These mutations increase over time after each herbicide applica‐ tion until they become predominant at what time the weed population is called resistant [5]. The first confirmed herbicide‐resistant weed species was *Senecio vulgaris* that had developed resistance to PS II inhibitors (atrazine and simazine) after the herbicides had been applied once or twice annually for 10 years [6]. Therefore, about 30‐resistant weed populations have been confirmed within the first decade, mostly in N. America and W. Europe [7]. Some weed species, such as *Lolium rigidum*, *Echinochloa crus‐galli* var. *crus‐galli*, *Poa annua*, *Alopecurus myo‐ suroides*, *Echinochloa colona*, *Eleusine indica*, *Amaranthus* sp., etc. have a high affinity to develop resistance especially due to their congenital genetic variability. Additionally, herbicides of dif‐ ferent chemical groups and different modes of action (e.g. sulfonylurea and synthetic auxins) can greatly differ in their risk levels for resistance. On the other hand, different chemical groups with the same mode of action such as herbicide inhibitors of acetolactate/acetohydroxyacid synthase (ALS/AHAS) (sulfonylurea, pyrimidinyl(thio)benzoate, sulfonylaminocarbonyl‐tri‐ azolinone, imidazolinone) can also be distinguished in their risk level for resistance.

Currently, herbicide resistance has been reported in 478 weed biotypes (252 weed species) in 67 countries. Many of those biotypes are resistant to ALS inhibitors, B/2 (97 dicots + 62 mono‐ cots), PS II inhibitors (C1/5 = 51 + 23, C2/7 = 10 + 18, C3/6 = 3 + 1), ACC‐ase inhibitors, A/1 (48 monocots) and EPSPS inhibitors, G (19 + 17). The highest number of confirmed resistant weed species belongs to the families: *Poaceae* (80 species), *Asteraceae* (39), *Brassicaceae* (22), *Cyperaceae* (12), *Amaranthaceae* (11), *Scrophulariaceae* (9), *Chenopodiaceae* (8), *Alismataceae* (7), *Polygonaceae* (7) and *Caryophyllaceae* (6). According to the number of active ingredients (a.i.), those four sites of action participate in the next relation: 50 a.i. from ALS inhibitors, 24 a.i. from PS II inhibi‐ tors, 15 a.i. from ACC‐ase inhibitors and 2 a.i. from EPSPS inhibitors. Atrazine (PS II inhibitors) is an active ingredient, which was confirmed by the greatest number of weed resistant spe‐ cies (66), the second is imazethapyr (44), followed by tribenuron‐methyl (43), imazamox (37), chlorsulfuron (36) metsulfuron‐methyl (35), glyphosate (34), iodosulfuron‐methyl‐sodium (33), fenoxaprop‐P‐ethyl (31), simazine (31), bensulfuron‐methyl (29), thifensulfuron‐methyl (27), fluazifop‐P‐bityl (25), pyrazosulfuron‐ethyl (25), etc. In relation to herbicide‐resistant weeds by county and site of action top 10 counties are the United States, Australia, Canada, France, Brazil, China, Spain, Israel, Japan and Germany [7] (**Table 1**).

In Serbia, study of weed resistance to herbicides started in the 1990s with resistance of *Amaranthus retroflexus* and *Chenopodium hybridum* to PS II inhibitors (atrazine) [8–10]. Until today, in Serbia, as a small county with less than 3 million ha arable lands, in the last 15 years, eight herbicide‐ resistant weed species were confirmed: *A. retroflexus*, *Setaria viridis*, *C. hybridum* and *Abutilon theo‐ phrasti* to PS II inhibitors, as well as *A. retroflexus*, *E. crus‐galli*, *Datura stramonium*, *Chenopodium* 


**Table 1.** Top 10 countries with the most number of confirmed resistant weed species.

It is also defined as a 'survival of a segment of the population of a weed species following an herbicide dose lethal to the normal population' [2]. In addition, resistance can be defined as 'the inherited ability to survive treatment by a herbicide' [3], or it is a 'phenomen which occurs as a result of heritable changes to biochemical processes that enable weed species sur‐

Weed resistance to herbicides is a normal and predictable outcome of natural selection. In that context, rare mutations that confer herbicide resistance exist in wild/weed populations before any herbicide introduction. These mutations increase over time after each herbicide applica‐ tion until they become predominant at what time the weed population is called resistant [5]. The first confirmed herbicide‐resistant weed species was *Senecio vulgaris* that had developed resistance to PS II inhibitors (atrazine and simazine) after the herbicides had been applied once or twice annually for 10 years [6]. Therefore, about 30‐resistant weed populations have been confirmed within the first decade, mostly in N. America and W. Europe [7]. Some weed species, such as *Lolium rigidum*, *Echinochloa crus‐galli* var. *crus‐galli*, *Poa annua*, *Alopecurus myo‐ suroides*, *Echinochloa colona*, *Eleusine indica*, *Amaranthus* sp., etc. have a high affinity to develop resistance especially due to their congenital genetic variability. Additionally, herbicides of dif‐ ferent chemical groups and different modes of action (e.g. sulfonylurea and synthetic auxins) can greatly differ in their risk levels for resistance. On the other hand, different chemical groups with the same mode of action such as herbicide inhibitors of acetolactate/acetohydroxyacid synthase (ALS/AHAS) (sulfonylurea, pyrimidinyl(thio)benzoate, sulfonylaminocarbonyl‐tri‐

azolinone, imidazolinone) can also be distinguished in their risk level for resistance.

France, Brazil, China, Spain, Israel, Japan and Germany [7] (**Table 1**).

Currently, herbicide resistance has been reported in 478 weed biotypes (252 weed species) in 67 countries. Many of those biotypes are resistant to ALS inhibitors, B/2 (97 dicots + 62 mono‐ cots), PS II inhibitors (C1/5 = 51 + 23, C2/7 = 10 + 18, C3/6 = 3 + 1), ACC‐ase inhibitors, A/1 (48 monocots) and EPSPS inhibitors, G (19 + 17). The highest number of confirmed resistant weed species belongs to the families: *Poaceae* (80 species), *Asteraceae* (39), *Brassicaceae* (22), *Cyperaceae* (12), *Amaranthaceae* (11), *Scrophulariaceae* (9), *Chenopodiaceae* (8), *Alismataceae* (7), *Polygonaceae* (7) and *Caryophyllaceae* (6). According to the number of active ingredients (a.i.), those four sites of action participate in the next relation: 50 a.i. from ALS inhibitors, 24 a.i. from PS II inhibi‐ tors, 15 a.i. from ACC‐ase inhibitors and 2 a.i. from EPSPS inhibitors. Atrazine (PS II inhibitors) is an active ingredient, which was confirmed by the greatest number of weed resistant spe‐ cies (66), the second is imazethapyr (44), followed by tribenuron‐methyl (43), imazamox (37), chlorsulfuron (36) metsulfuron‐methyl (35), glyphosate (34), iodosulfuron‐methyl‐sodium (33), fenoxaprop‐P‐ethyl (31), simazine (31), bensulfuron‐methyl (29), thifensulfuron‐methyl (27), fluazifop‐P‐bityl (25), pyrazosulfuron‐ethyl (25), etc. In relation to herbicide‐resistant weeds by county and site of action top 10 counties are the United States, Australia, Canada,

In Serbia, study of weed resistance to herbicides started in the 1990s with resistance of *Amaranthus retroflexus* and *Chenopodium hybridum* to PS II inhibitors (atrazine) [8–10]. Until today, in Serbia, as a small county with less than 3 million ha arable lands, in the last 15 years, eight herbicide‐ resistant weed species were confirmed: *A. retroflexus*, *Setaria viridis*, *C. hybridum* and *Abutilon theo‐ phrasti* to PS II inhibitors, as well as *A. retroflexus*, *E. crus‐galli*, *Datura stramonium*, *Chenopodium* 

vival when treated with a herbicide' [4].

8 Herbicide Resistance in Weeds and Crops

*album* and *Sorghum halepense* to ALS inhibitors [11–19]. According to the herbicide resistance mechanisms, all processes can be grouped as follows: target‐site resistance, non‐target‐site resis‐ tance, cross‐resistance and multiple‐resistance [20–22].

*Target‐site resistance* (TSR) is generally due to a single or several mutations in the gene encod‐ ing the herbicide‐target enzyme, which, in turn, decreases the affinity for herbicide binding to that enzyme. Most, but not all cases of resistance to herbicide ALS inhibitors, ACC‐ase, triazine, dinitroaniline etc. are due to modifications of the site of action of the herbicide. In addition, gene overproduction (amplification) is the most recently identified herbicide resis‐ tance mechanism, for example, EPSPS gene amplification correlates with glyphosate resis‐ tance in *Amaranthus palmeri* and *Kochia scoparia* [23–25], and causes resistance by increasing the production of the target enzyme, effectively diluting the herbicide in relation to the target site (**Figure 1**).

*Non‐target‐site resistance* **(NTSR)** is caused by mechanisms that reduce the amount of herbi‐ cidal active compound before it can attack the plant. Reduced absorption (penetration) or altered translocation, increased herbicide sequestration or enhanced herbicide metabolism (detoxification) can cause resistance due to the restriction of herbicide movement where the herbicide does not reach its site of action in sufficient concentration to cause plant mortality. Active vacuolar or cell walls sequestration can keep the herbicide from the site of action lead‐ ing to resistance. For example, vacuolar herbicide sequestration correlates with glyphosate resistance in *Conyza canadensis*, *Lolium* sp. etc. [26, 27] (**Figure 1**). Finally, the biochemical reac‐ tions that detoxify herbicides can be grouped into four major categories: oxidation, reduction, hydrolysis and conjugation [28].

**Figure 1.** The route of the herbicide after the application, and the possible mechanisms of resistance in plant. After application: (1) herbicide absorption/penetration, (2) translocation, (3) accumulate at the target protein location, and (4) binding to the target protein, (5) disruption of the biosynthesis pathways or cell structures, and/or generation of cytotoxic molecules. NTSR mechanisms: (A) reduction in herbicide penetration, (B) altered translocation of the herbicide away from the target protein, (C) enhanced detoxification of the herbicide, or (D) enhanced neutralization of cytotoxic molecules generated by herbicide action. TSR mechanisms: (E) target protein overproduction, and/or (F) structural mutations that modify the 3D structure and electrochemical properties of the target protein. Structural mutations can have no, moderate or strong negative effects on the stability of herbicide binding to the target protein, which results in (F‐a) no, (F‐b) moderate or (F‐c) marked reduction in herbicide sensitivity at the protein level, respectively; or can (F‐d) increase the stability of herbicide binding to the target protein, which results in an increase in herbicide sensitivity at the protein level (downloaded from Ref. [32]).

*Cross‐resistance* **(CR)** means that a single‐resistance mechanism causes resistance to several herbicides. CR can be conferred by a single gene or by two or more genes influencing a single mechanism. There are two types of CR: target‐site cross‐resistance (TS‐CR) and non‐target‐site cross‐resistance (NTS‐CR). The most common type of CR is TS‐CR where an altered target site confers resistance to many or all of the herbicides that inhibit the same enzyme, for example, Trp‐574‐Leu amino acid substitution within the ALS gene was found in two populations of *Cyperus iria* after exposition to bispyribac‐sodium, halosulfuron, imazamox and penoxsulam [29]. On the other hand, NTS‐CR is type of herbicide resistance in which a mechanism other than resistant enzyme target sites is involved (e.g. reduced absorption, translocation, or enhanced herbicide detoxification) [30].

*Multiple‐resistance* is a situation where two or more resistance mechanisms are present within the same plant, often due to sequential selection by herbicides with different modes of action (e.g. resistance of *Lolium* sp. populations to glyphosate and ACC‐ase inhibitors, as well as resistance to glyphosate and ALS inhibitors were confirmed by multiple‐resistance [31]).
