Multiple herbicide resistance in littleseed canarygrass (Phalaris minor): A threat to wheat production in India

Seed collection of Phalaris minor populations

The seeds of P. minor populations were collected during two consecutive rabi seasons (the time period from October to April, during which the wheat crop is grown in India), 2004 to 2005 and 2005 to 2006, from wheat fields with persistent herbicide selection pressure under the rice–wheat system of the Indo-Gangetic Plains, India (Table 1). These fields had a history of poor or no control of P. minor with clodinafop and/or sulfosulfuron. The populations that were earlier confirmed as sensitive were used as susceptible (S) stock for comparison (DWR, PATBIH) and these were from an area having no herbicide use history (PATBIH) or having different herbicide and crop rotations (DWR). The details of the herbicide use history and crop rotations are mentioned in Table 1. The seeds were collected in the first week of April and then stored in the laboratory till November for bioassay studies.

Quantification of the herbicide resistance profile for clodinafop, fenoxaprop, pinoxaden, and sulfosulfuron

Quantification of the herbicide resistance profile in pots was done for two consecutive years (2005 to 2006 and 2006 to 2007). Each year, in the month of November, P. minor populations were grown in pots of 15 cm diameter. The pots were filled with soil and well-rotted farmyard manure in a 6:1 ratio by volume after being passed through a 2 mm sieve. The soil was collected from a field that had no previous infestation of P. minor. It was a sandy clay loam with a pH of 8.1 and an organic matter content of 0.4%. The pots were watered to deplete the soil seed bank before sowing.

The P. minor seedlings were established by seeding 50–60 seeds at a depth of 2.5 cm. Three pots were used for each herbicide treatment (Table 2). The pots were labeled and arranged in a completely randomized design and were watered as required. Two weeks after emergence, thinning was done to maintain 10 plants per pot. The herbicides were sprayed 30–40 days after sowing (DAS) with a knapsack sprayer fitted with flat fan nozzles (Aspee, Navyug Krishi Sadhan, Mumbai, India), delivering 350 L ha⁻¹ of water, when P. minor was at the three-to-four-leaf stage.

The various rates of isoproturon (Isoguard 75 WP; Gharda Chemicals, Mumbai, India), clodinafop (Topik 15 WP; Syngenta India, Mumbai, India), fenoxaprop (Puma super 10 EC; Bayer Crop Science, Mumbai, India), sulfosulfuron (Leader 75 WG; Monsanto India, Mumbai, India), and pinoxaden (Axial 5 EC; Syngenta India, Mumbai, India) used for the resistance profile study are given in Table 2. The surfactant, Leader Mix (polyethylene amine; Monsanto India, Mumbai, India), was used at a concentration of 0.35% (v/v) in the sulfosulfuron treatment. Thirty days after the herbicide spray, the dry weight of the seedlings was recorded and, based on the biomass reduction, the 50% growth reduction (GR₅₀) values were determined (Finney 1971). The resistance index for the different populations was calculated by dividing the GR₅₀ values of the populations with the GR₅₀ value of the S population (Tables 3,4). The values >2 were considered to be resistant.

Response of triazine and dinitroaniline against multiple herbicide-resistant and herbicide-susceptible Phalaris minor populations

The filling of the pots and the seeding of P. minor was done as mentioned above. The population, BHKAI-1, confirmed to have multiple herbicide resistance (isoproturon, clodinafop, fenoxaprop, pinoxaden, and sulfosulfuron), along with a sensitive biotype, DWR, was used for the response study against triazine (metribuzin [Sencor 70 WP; Bayer Crop Science, Mumbai, India], terbutryn [Igran 500 SC; Syngenta India, Mumbai, India]), and dinitroaniline (pendimethalin [Stomp 30 EC; BASF, Mumbai, India]) herbicides in two sets of experiments.

In the first set, metribuzin at 0, 25, 50, 100, 200, and 400 g ha⁻¹ and terbutryn at 0, 200, 400, 800, and 1600 g ha⁻¹ were sprayed at the three-to-four-leaf stage of P. minor. For each treatment, three pots were maintained and, 4 weeks after the herbicide application, the dry weight was taken for calculating the GR₅₀ values.

In the second set, pre-emergence pendimethalin was evaluated against the R BHKAI-1 and S DWR populations. The BHKAI-1 population was resistant to four groups: phenylurea, aryloxyphenoxypropionate, phenylpyrazolin, and sulfonylurea. Fifty seeds per pot of the P. minor population were sown at a depth of 2.5 cm. Next day, pendimethalin was applied at 100, 200, 400, 800, and 1600 g ha⁻¹ with the knapsack sprayer fitted with flat fan nozzles using 350 L ha⁻¹ of water. Three pots were maintained for each treatment. One month after seeding, the seedling biomass was recorded and, based on the biomass reduction, the GR₅₀ values were worked out.

Control of isoproturon-resistant and clodinafop-resistant Phalaris minor with sulfosulfuron under field conditions

At three locations with clodinafop-R P. minor, sulfosulfuron was evaluated in wheat from 2006 to 2007. These three populations (CHKUR, KAKAR, and GHKAR) also were assessed for their resistance profiles under the pot bioassay studies (Table 4). In order to compare their performance, sulfosulfuron and clodinafop also were evaluated against the S DWR population of P. minor in wheat at the Directorate of Wheat Research Farm, Karnal, from 2005 to 2006 and 2006 to 2007. Phalaris minor was the dominant weed at all the locations. The herbicides, sulfosulfuron (25 g ha⁻¹) and clodinafop (60 g ha⁻¹), were applied at 35–40 DAS (two-to-four-leaf stage of P. minor) using the knapsack sprayer equipped with flat fan nozzles at 350 L ha⁻¹ of water. For the control of broad-leaved weeds, metsulfuron-methyl (Algrip 20 WP EI; DuPont India, Gujarat, India) was applied at 4 g ha⁻¹ at 30–35 DAS, that is, 5 days before sulfosulfuron or clodinafop application. The cationic surfactant, Leader Mix (polyethylene amine), was used with the spray mixture at a concentration of 0.35% (v/v) in the sulfosulfuron treatment. In all these field evaluations, wheat (cv. PBW-343; Punjab Agricultural University, Ludhiana, India) was sown at a spacing of 20 cm (row-to-row), with a 100 kg ha⁻¹ seed rate during the second-to-third week of November. Fertilization (150 kg N, 60 kg P₂O₅, and 40 kg K₂ ha⁻¹) and irrigation were performed according to the recommended practises for wheat. The studies were conducted in a randomized block design, with each treatment replicated thrice. Observations on the weed dry weight (g m⁻²) were recorded at 120 DAS using a 0.5 m × 0.5 m quadrat. The weeds were cut at ground level by a sickle, sun-dried for 7 days, and then oven-dried at 70 ± 5°C in order to record the dry weight. The crop was harvested in the first fortnight of April in both years. The wheat grain yield was expressed at 12% moisture. The significance of the treatment means was compared using the paired t-test (Panse & Sukhatme 1995).

Quantification of the herbicide resistance profile

The P. minor populations collected from farmers' fields having a history of poor or no control with the ACCase inhibitor (clodinafop) or the ALS inhibitor (sulfosulfuron) or both were evaluated under pot conditions against different herbicides (Tables 3,4). Most of the populations were resistant to isoproturon, indicating its widespread presence in the Indo-Gangetic Plains. The level of isoproturon resistance varied among the different populations (2.6–13.7-fold) depending on the selection pressure imposed by isoproturon. The varying level of isoproturon resistance in P. minor also has been reported earlier (Malik & Singh 1995; Chhokar & Malik 2002).

For clodinafop, the most S population had GR₅₀ values of 10.3 and 20.6 g ha⁻¹ during the rabi seasons of 2005 to 2006 and 2006 to 2007, respectively. The slightly greater GR₅₀ value during 2006 and 2007, compared with 2005 and 2006, was related to P. minor being at the three-to-four-leaf stage, whereas it was at the two-to-three-leaf stage from 2005 to 2006, at the time of herbicide application. Some of the populations (SAKAR-1, HALUD, BHKAI-1, CHKUR, GHKAR, and KAKAR) needed 11.65-fold more clodinafop for the same level of effect as observed with the most susceptible biotype (Tables 3,4). Some of the clodinafop-R populations had GR₅₀ values greater than four-fold (240 g ha⁻¹) the recommended field application rate (60 g ha⁻¹) from 2006 to 2007. The fields that were detected with clodinafop resistance had almost a similar frequency of clodinafop use (Table 1), as the rate of resistance development relates to the intensity of the selection for resistance (Martinez-Ghersa et al. 1997). The populations that were resistant to clodinafop were cross-resistant to fenoxaprop and pinoxaden. The cross-resistance level was higher for fenoxaprop and lower for pinoxaden. Five cross-resistant populations exhibited GR₅₀ values for fenoxaprop >2.4-fold (240 g ha⁻¹) the field application rate (100 g ha⁻¹), whereas for pinoxaden, it was around the field application rate (35–40 g ha⁻¹). However, pinoxaden can be targeted for the control of R populations of isoproturon or sulfosulfuron or both.

Generally, grass weeds have been reported to show cross-resistance to both aryloxyphenoxypropionate (fops group) and cyclohexanedione (dims group) herbicides (Mansooji et al. 1992; Heap et al. 1993; Tardiff et al. 1993; Seefeldt et al. 1994; Cocker et al. 2001). However, some grass weed populations show a high level of resistance to aryloxyphenoxypropionate, but no cross-resistance to cyclohexanediones (Gronwald et al. 1992; Mansooji et al. 1992; Maneechote et al. 1994; Seefeldt et al. 1994; Cocker et al. 2001). Triazine-R biotypes of Phalaris paradoxa and Alopecurus myosuroides and triallate-R Avena fatua showed cross-resistance to diclofop methyl (Rubin et al. 1985; Thai et al. 1985; Yaacoby et al. 1986). However, the atrazine-R Echinochloa crus-galli biotype (53-fold more resistant than susceptible) is 33-fold and two-fold more susceptible to fluazifop and sethoxydim, respectively (Gadamski et al. 2000). The diclofop-R biotype of annual ryegrass (L. rigidum) was cross-resistant to fluazifop and chlorotoluron, but not to oxyfluorfen (Heap & Knight 1986). Therefore, the various grassy weeds show a different cross-resistance spectrum.

Compared to the fop herbicides, the level of sulfosulfuron resistance was low. The GR₅₀ values for sulfosulfuron were less than the recommended field dose (25 g ha⁻¹). This partial resistance is responsible for the poor control with sulfosulfuron, particularly with delayed herbicide application. Like the results with sulfosulfuron resistance in the present study, Volenberg et al. (2000) reported a 5.9-fold resistance to primisulfuron-methyl, but the plants were controlled by the field-use rate of primisulfuron. The populations, HALUD, SAKAR-2, BHKAI-1, BHKAI-2, UCKAR, DHKUR, and SAKAR, besides exhibiting resistance across herbicides from two different ACCase-inhibiting herbicidal chemistries (fop and den), also exhibited resistance to the photosystem II (PS II)-inhibiting herbicide (isoproturon) and ALS-inhibiting herbicide (sulfosulfuron). Thus, the P. minor populations exhibited multiple herbicide resistance across a range of dissimilar herbicidal chemistries and sites of action. Multiple herbicide resistance evolved because of the imposition of sequential selection pressure. Firstly, the isoproturon was applied continuously for 10–15 years. After the evolution of isoproturon resistance, sulfosulfuron or clodinafop were used continuously. This led to multiple herbicide resistance against four groups of herbicides: phenylureas (isoproturon), sulfonylurea (sulfosulfuron), aryloxyphenoxypropionate (clodinafop, fenoxaprop), and phenylpyrazolin (pinoxaden).

Multiple herbicide resistance is more serious, as has happened in Australia with the Lolium spp., which is highly resistant to ACCase- and ALS-inhibitor herbicides (Gill 1995; Llewellyn & Powles 2001; Heap 2007). Phalaris minor in Mexico and Israel (Tal et al. 1996) has evolved resistance to fenoxaprop. In Israel, fenoxaprop resistance is target site-based, with low levels of cross-resistance. In India, clodinafop-R populations of P. minor are highly cross-resistant to fenoxaprop and moderately cross-resistant to pinoxaden. The resistance mechanism in P. minor for ALS and ACCase inhibitors is not known. However, isoproturon resistance in P. minor in India is metabolic (Singh et al. 1997), which might have evolved because of suboptimal dosages (Wrubel & Gressel 1994; Chhokar & Malik 2002). Like isoproturon, initially sulfosulfuron and clodinafop also were used at less than the recommended rates and farmers increased the dosage over time with the decline in P. minor control. Some farmers failed to control it with 2–2.5-fold the recommended dose of clodinafop, which is well-supported by the GR₅₀ values for clodinafop, as they were >240 g ha⁻¹ for some of the populations (Tables 3,4).

Earlier, Lolium multiflorum populations with both enhanced metabolism and insensitive ACCase (target site) resistance against a range of aryloxyphenoxypropionate and cyclohexanedione herbicides have been reported (Cocker et al. 2001). Metabolism-based ACCase-inhibiting herbicide resistance is complex and can confer resistance to herbicides with different target sites (Delye 2005). De Prado & Franco (2004) also reported that intergroup herbicide resistance can be conferred by a non-target mechanism.

The differential herbicide effects against the P. minor population were dependent on the history of herbicide use. The S populations were from areas where either the particular herbicide was not applied earlier (PATBIH) or was used in rotation with other herbicides (DWR), having different sites of action (Table 1). The MHR populations were from areas where the farmers imposed continuous selection pressure by the sequential application of herbicide in an unbroken rice–wheat sequence. The rice–wheat cropping sequence provides favorable conditions for P. minor, resulting in heavy infestation. Heavy population pressure increases the chances of selection for R populations. High-risk herbicides should be used with caution in fields with high weed densities because the number of herbicide-resistant mutants is proportional to the population size (Jasieniuk et al. 1996). Crop and herbicide rotations are attributed to lowering the selection pressure (Gressel & Segel 1990). Crop rotations do not merely delay resistance by allowing the use of different management options, but also restore diversity in weed flora. Some crop rotations (growing Egyptian clover, Trifolium alexandrinum, for 2 years) might even exhaust the soil seed bank of P. minor, thus providing an effective solution. In a survey, isoproturon resistance in P. minor was observed in 67% of the fields under rice–wheat rotation compared to 8, 9, and 16% when rice–berseem–sunflower–wheat, sugar cane–vegetables–wheat, and cotton–pigeonpea–wheat rotations, respectively, were followed (Malik & Singh 1995). In the present study also, multiple herbicide resistance was not observed in the biotypes, DWR, NAKAR, STKAR, SIKAI, and RAKAR, where vegetables and sugar cane were grown in rotation with rice and wheat crops.

As a consequence of P. minor being a major weed and the preference for its chemical control, the emergence of multiple herbicide resistance is a major threat to the sustainability of wheat production. It is not only a threat to the farmers but also to industries. Presently, multiple herbicide resistance is sporadic and its spread to new areas needs to be checked immediately; if the problem is not curbed, it might become an epidemic within a few years and will threaten food security. One of the most promising tactics that can reduce the R population is the use of effective alternative herbicides and the quick test (Boutsalis 2001) can help to identify which herbicide to use in problematic fields.

Response of triazine and dinitroaniline against multiple herbicide-resistant Phalaris minor

Triazine herbicides (metribuzin and terbutryn) were found to be equally effective in controlling both the MHR (BHKAI-1) and multiple herbicide-susceptible (DWR) populations. The BHKAI-1 population was resistant to herbicides from four groups: isoproturon, clodinafop, fenoxaprop, pinoxaden, and sulfosulfuron (Tables 3,4). The GR₅₀ values for the R and S populations were 145.6 and 153.1 g ha⁻¹ for terbutryn and 35.4 and 37.3 g ha⁻¹ for metribuzin. However, their adoption at farmers' fields will depend on their selectivity range. The differential tolerance of wheat genotypes to metribuzin has been reported (Runyan et al. 1982). Metribuzin was found to be effective for the control of isoproturon-R P. minor in wheat but its application timing is narrow, that is, early application can be toxic and delayed application, particularly under untilled conditions without preseeding and an unselective herbicide (glyphosate/paraquat) being less effective (Chhokar et al. 2006). To increase the safety margin, it will be better if some safeners are developed. The effectiveness of triazine herbicides (atrazine) against Eastern black nightshade (Solanum ptycanthum) populations that were resistant to ALS inhibitors (imazethapyr, flumetsulam, cloransulam, nicosulfuron, prosulfuron, and rimsulfuron) was reported by Ashigh and Tardif (2006). Solanum ptycanthum populations resistant to ALS herbicides also were susceptible to chloroacetamide (metolachlor, dimethenamid, and flufenacet). Flufenacet is also effective for the control of isoproturon-R P. minor, but it had some toxic effects on wheat crops (Chhokar et al. 2006).

The most triazine-R biotypes show no cross-resistance to phenylurea herbicides (Devine & Shukla 2000) and even negative cross-resistance has been reported for triazine-R weed populations for phenylureas and other PS II-inhibiting herbicides (DePrado et al. 1992). The present study also demonstrates the susceptibility of isoproturon-R P. minor to triazine herbicides. However, Burnet et al. (1991) reported a biotype of L. rigidum that was resistant to atrazine as being cross-resistant to chlorotoluron. Triazine and phenylurea have similar chemical and physical characteristics, which are manifested in their ability to inhibit the same protein with overlapping binding domains (Arntzen et al. 1982). Similarly, Milliman et al. (2003) reported that the population of black nightshade resistant to imidazolinone (imazethapyr and imazamox) was found to be susceptible to the sulfonylurea herbicide, primisulfuron-methyl, although both have the same site of action, reflecting different resistance behavior for different weeds.

Pre-emergence pendimethalin was also effective in controlling the MHR P. minor population. The GR₅₀ values for the R and S populations were 467.4 and 493.7 g ha⁻¹, respectively. Therefore, until new herbicides from other groups are made available, pendimethalin can be used pre-emergence in the conventional-tillage system. In the zero tillage (ZT) system, this can be used in combination with an unselective herbicide, like glyphosate, to further improve weed control. Although dinitroaniline resistance has been reported in goosegrass (Elucine indica L. Gaertn.), green foxtail, rigid ryegrass, and black grass (Mudge et al. 1984; Heap & Knight 1986; Vaughn et al. 1990; Morrison et al. 1991; Moss & Cussans 1991), the resistance level is still low, mainly related to the paucity of a detoxification mechanism in weeds. The MHR (resistance to triazines, and the fop and dims groups) population of downy brome (Bromus tectorum) showed sensitivity to trifluralin and glyphosate (Park & Smith 2005). Uludag et al. (2007) also reported the effectiveness of a dinitroaniline herbicide (trifluralin) in controlling wild oat populations resistant to ACCase inhibitor herbicides.

Controlling isoproturon-resistant and clodinafop-resistant populations with sulfosulfuron under field conditions

Sulfosulfuron and clodinafop were evaluated against the S DWR and MHR (clodinafop and isoproturon) populations (CHKUR, KAKAR, and GHKAR) in wheat fields (Table 5). Both the herbicides were effective against the S population and the P. minor dry weight was only 1.3 and 0.2 g m⁻², with sulfosulfuron and clodinafop application, respectively. The mean P. minor dry weight and wheat yield did not differ significantly between the two herbicide treatments with the S population. Whereas, clodinafop failed to control the R populations and the mean dry weight (342.2 g m⁻²) was significantly higher than with sulfosulfuron (3.4 g m⁻²). Uludag et al. (2007) also observed the effectiveness of a sulfonylurea herbicide (mesosulfuron) in controlling wild oat populations with resistance to ACCase-inhibitor herbicides.

The ineffectiveness of clodinafop against the R populations resulted in a significant yield reduction (49.8%) over the sulfosulfuron treatment because of strong competition combined with the lodging induced by this weed. Severe yield reductions by P. minor competition also have been reported earlier (Afentouli & Eleftherohorinos 1996; Chhokar & Malik 2002; Chhokar et al. 2006). Clodinafop resistance is the result of exclusive dependence on clodinafop. Farmers resorted to extensive clodinafop use because of a lack of carry-over effect on succeeding crops and also because it is safe to a mustard crop intercropped with wheat. Presently, the yield reduction experienced in areas having clodinafop-R populations can be resolved through using sulfonylurea herbicides (sulfosulfuron/mesosulfuron). The life expectancy of sulfosulfuron in such areas can be improved by rotating it with pendimethalin, along with other control measures.

With the evolution of multiple resistance, the situation of P. minor interference in some wheat fields could become similar to that of the late 1970s, before isoproturon introduction, and the late 1990s, after the evolution of isoproturon resistance, when this weed caused severe yield reductions, to the extent of complete crop failure.

The cases of herbicide resistance evolution are more frequent with the continuous usage of a herbicide or herbicides of the same group (Burnet et al. 1991; Jasieniuk et al. 1996; Beckie 2006). The reliance on in-crop selective herbicides has led to the widespread evolution of the rigid ryegrass population with multiple herbicide resistance (Llewellyn & Powles 2001). A similar situation might have happened in the P. minor case because exclusive herbicide pressure was imposed in a sequence (isoproturon followed by clodinafop or sulfosulfuron or both), which resulted in multiple herbicide resistance.

Multiple and cross-resistance cases have been reported widely (Heap & Knight 1986; Powles et al. 1989; Holt & LeBaron 1990; Burnet et al. 1991; Heap 1991; Tucker & Powles 1991; Gossett et al. 1992; Holt 1992) but such types of resistance are lacking or rare with glyphosate, even though it has been intensively used worldwide for a long time. This is related to the unique properties of glyphosate with regard to its mode of action, metabolism, chemical structure, and lack of residual activity in the soil (Bradshaw et al. 1997). As a preseeding option, glyphosate can be used for multiple herbicide resistance management.

As metabolic resistance evolves because of under-dosing (Wrubel & Gressel 1994; Gressel 1995), attention will have to be paid to ensure the application of herbicides in rotation at the proper dose and time. The effectiveness of grass herbicides is generally reduced when mixed with broad-leaved herbicides (Zhang et al. 1995; Damalas & Eleftherohorinos 2001; Damalas 2004). Mathiassen and Kudsk (1998) also reported the reduced efficacy of clodinafop or fenoxaprop when applied as a tank mix with either 2,4-D or metsulfuron. Most of the farmers adhered to a tank-mix application of clodinafop with either 2,4-D or metsulfuron. Such antagonistic performances allow the survival, seed production, and build-up of the population in subsequent seasons, thereby increasing the selection pressure for resistance evolution. The sequential application of grass and broad-leaved herbicides needs to be used to overcome the antagonism. Also, a strategy consisting of revolving herbicide doses (Gardner et al. 1998) can help in delaying the evolution of target site and quantitative resistance.

Mostly, if resistance develops in a herbicide, then other herbicides of that group generally become ineffective. Similarly, in the present study, the clodinafop-R populations showed cross-resistance to fenoxaprop and pinoxaden. In earlier studies, L. rigidum (Burnet et al. 1993; Preston & Powles 1997) and A. myosuroides (Kemp et al. 1990) populations that were resistant to chlorotoluron were also resistant to isoproturon. However, with P. minor, the isoproturon-R populations were resistant to diclofop, metoxuron, and methabenzthiazuron, but they were susceptible to chlorotoluron (Singh 2007); all except diclofop belong to the urea group. This shows the diversity of genes encoding cytochrome P 450 mono-oxygenases and means that, in different weed species, different enzymes might be employed to metabolize a single herbicide (Preston & Smith 2001).

Furthermore, the rotation of herbicide groups with different mechanisms of action should be followed strictly as it has a strong effect in delaying the appearance of resistance, as shown by a simulation model (Jasieniuk et al. 1996; Cavan et al. 2000).

With the evolution of multiple herbicide resistance, more focus has to be given to non-chemical weed control measures. Wheat varieties with early vigour and smothering and allelopathic effects on weeds should be developed. Adjusting the sowing time, either early or late, also can help in reducing the impact of P. minor. Early wheat sowing avoids P. minor emergence with the crop as a result of a higher temperature (Chhokar & Malik 1999), thereby reducing competition. Whereas, late sowing helps to deplete the soil seed bank by germination and killing. These two practises can be implemented through the adoption of the ZT system and crop rotation. The ZT system in wheat under rice–wheat rotation can advance the sowing time and also discourage P. minor emergence related to a higher soil strength (Chhokar et al. 2007). Additionally, the usage of unselective herbicides, like glyphosate/paraquat, in the untilled system will help in the management of MHR P. minor. Moreover, unselective herbicides can be combined with pre-emergence pendimethalin or trifluralin to check the later emerging flush.

Adopting crop rotation consisting of competitive crops (barley/mustard) or crops that do not allow weeds to form seeds due to the difference in the maturity period (pea, potato or other vegetable crops after rice) or repeated cutting (green fodder crops like berseem, lucerne, and oat) will help to deplete the seed bank of R populations. Also, crop rotation involving broad-leaved crops, like mustard, will allow the use of graminicides against P. minor that are not selective to wheat. This will help in reducing its seed bank and, thereby, the problem in the next season. Thus, crop rotation, through its effect on weed seed germination and mortality, affects the weed seed bank in the soil, which can conserve the R or S gene within a plant population (Maxwell et al. 1990).

Generally, most of the farmers in north-western India sow wheat seeds contaminated with P. minor seeds. Also, the seeds of new varieties move very fast from farmer to farmer. If contaminated seeds move from a multiple herbicide resistance-prone area to a new area, then resistance will spread. Other factors that also can spread resistance are the use of unrotted farmyard manure and uncleaned farm machinery. Measures should be taken to check the spread of R populations to new areas by encouraging the use of certified seeds and by restricting the movement of contaminated seeds and farmyard manure.

Considering the causes of evolution and spread of herbicide resistance, the strategies comprising crop rotation, herbicide rotation, herbicide mixture and sanitation practises (weed-free crop seeds, well-rotted manure, and clean machinery), along with various other agronomic tactics (competitive variety, early sowing, higher seed rate, ZT system, stale seed bed), should be integrated for the effective management of herbicide-R P. minor in wheat. A detailed study of P. minor biology is also required, which will further strengthen herbicide resistance management. Whenever and wherever possible, consideration also should be given to the use of mechanical weed control to remove the weeds that survive the herbicide application before seed setting. The integration of all these approaches will lower the impact of herbicide resistance on wheat production and farm income.