Christopher Preston

Gene amplification confers glyphosate resistance in Amaranthus palmeri

The herbicide glyphosate became widely used in the United States and other parts of the world after the commercialization of glyphosate-resistant crops. These crops have constitutive overexpression of a glyphosate-insensitive form of the herbicide target site gene, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). Increased use of glyphosate over multiple years imposes selective genetic pressure on weed populations. We investigated recently discovered glyphosate-resistant Amaranthus palmeri populations from Georgia, in comparison with normally sensitive populations. EPSPS enzyme activity from resistant and susceptible plants was equally inhibited by glyphosate, which led us to use quantitative PCR to measure relative copy numbers of the EPSPS gene. Genomes of resistant plants contained from 5-fold to more than 160-fold more copies of the EPSPS gene than did genomes of susceptible plants. Quantitative RT-PCR on cDNA revealed that EPSPS expression was positively correlated with genomic EPSPS relative copy number. Immunoblot analyses showed that increased EPSPS protein level also correlated with EPSPS genomic copy number. EPSPS gene amplification was heritable, correlated with resistance in pseudo-F2 populations, and is proposed to be the molecular basis of glyphosate resistance. FISH revealed that EPSPS genes were present on every chromosome and, therefore, gene amplification was likely not caused by unequal chromosome crossing over. This occurrence of gene amplification as an herbicide resistance mechanism in a naturally occurring weed population is particularly significant because it could threaten the sustainable use of glyphosate-resistant crop technology.

Evolved Glyphosate Resistance in Plants: Biochemical and Genetic Basis of Resistance'

Resistance to the herbicide glyphosate is currently known in at least eight weed species from many countries. Some populations of goosegrass from Malaysia, rigid ryegrass from Australia, and Italian ryegrass from Chile exhibit target site–based resistance to glyphosate through changes at amino acid 106 of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene. Mutations change amino acid 106 from proline to either serine or threonine, conferring an EPSPS weakly resistant to glyphosate. The moderate level of resistance is sufficient for commercial failure of the herbicide to control these plants in the field. Conversely, a nontarget site resistance mechanism has been documented in glyphosate-resistant populations of horseweed and rigid ryegrass from the United States and Australia, respectively. In these resistant plants, there is reduced translocation of glyphosate to meristematic tissues. Both of these mechanisms are inherited as a single, nuclear gene trait. Although at present only two glyphosate-resistance mechanisms are known, it is likely that other mechanisms will become evident. The already very large and still increasing reliance on glyphosate in many parts of the world will inevitably result in more glyphosate-resistant weeds, placing the sustainability of this precious herbicide resource at risk. Nomenclature: Glyphosate; goosegrass, Eleusine indica (L.) Gaertn. #3 ELEIN; horseweed, Conyza canadensis (L.) Cronq. # ERICA; rigid ryegrass, Lolium rigidum Gaud. # LOLRI; Italian ryegrass, Lolium multiflorum Lam. # LOLMU. Additional index words: EPSPS, herbicide resistance, herbicide translocation. Abbreviations: ACCase, acetyl-coenzyme A carboxylase; ALS, acetolactate synthase; ESPS, 5-enolpyruvylshikimate-3-phosphate synthase.

Herbicide Resistance: Impact and Management

Publisher Summary This chapter focuses on the threat to the productivity of world agriculture imposed by the evolution of herbicide-resistant weed populations. Implicit in this chapter is that herbicides should and will continue to be a major tool for weed control. It is believed that modern herbicides are a cost-effective, efficient, and environmentally benign means for obtaining weed control. Proponents rightly identify the benefits of herbicides in achieving weed control as well as positive environmental influences in substituting for soil cultivation in weed management. Reliance on herbicides for weed control is expected to continue because there is no attractive superior technology available. However, for sustainable weed management to be achieved, changes to current herbicide use patterns are required. Multiple-resistant populations of weeds, such as L. rigidum and A . myosuroides, are current indicators of potential worst-case weed problems of the future. Resistance will continue to increase if present herbicide use patterns are not altered.

Investigations into the mechanism of glyphosate resistance in Lolium rigidum

Glyphosate is a widely used non-selective herbicide to which so far only three weed species have evolved resistance. Here, we report on the mechanism of glyphosate resistance in one resistant population of Lolium rigidum (Gaud.). Experiments demonstrate that glyphosate resistance in this population is directly correlated with increased transport of the herbicide to leaf tips. No significant differences in the level of expression of the herbicide target site, EPSP synthase, between resistant and susceptible plants were found and the enzyme is equally sensitive to inhibition by glyphosate in both populations. Similarly, plant metabolism of glyphosate does not contribute to resistance. Resistant and susceptible plants are equally capable of absorbing the applied herbicide. The most notable difference between resistant and susceptible populations is found in the translocation of glyphosate. Following treatment, an accumulation of glyphosate in the roots of susceptible plants is observed, whereas glyphosate accumulates in the leaf tips of resistant plants. Taken together, it seems likely that an alteration to the cellular transport of glyphosate confers resistance.

Resistance to glyphosate in Lolium rigidum

Annual ryegrass (Lolium rigidum) is a widespread and important weed of Australia and populations of this weed have developed resistance to most major herbicides, including glyphosate. The possible mechanisms of resistance have been examined in one glyphosate-resistant Lolium population. No major differences were observed between resistant and susceptible biotypes in respect of (i) the target enzyme (EPSP synthase), (ii) DAHP synthase, the first enzyme of the target (shikimate) pathway, (iii) absorption of glyphosate, or (iv) translocation. Following treatment with glyphosate, there was greater accumulation of shikimate (derived from shikimate-3-Pi) in susceptible than in resistant plants. In addition, the resistant population exhibited cross-resistance to 2-hydroxy-3-(1,2,4-triazol-1-yl)propyl phosphonate, a herbicide which, although structurally similar to glyphosate, acts at an unrelated target site. On the basis of these observations we speculate that movement of glyphosate to its site of action in the plastid is involved in the resistance mechanism. © 1999 Society of Chemical Industry

Tillage system effects on weed ecology, herbicide activity and persistence: a review

In the past few years, there has been a growing trend towards reducing tillage in cropping systems to allow stubble retention, earlier planting and improved soil structure. However, the adoption of conservation tillage systems will change weed control practices. Different tillage systems interact with the micro-environment of weed seeds and can influence the pattern of recruitment from the weed seed bank. Here, we present a review of the effect of different tillage systems on weed ecology, herbicide activity and herbicide persistence. Tillage systems can have a major influence on the vertical distribution of weed seeds in the soil seed bank. However, the impact of the changes in the vertical seed distribution on weed seedling recruitment is not well understood. Usually weed seedling recruitment increases if tillage equipment brings buried seed to, or close to, the soil surface, and seedling recruitment decreases if surface seed is buried deeper in the soil. However, tillage responses have a tendency to be species specific and can also be influenced by the intensity of tillage. Any weed species in which germination is stimulated by exposure to light is likely to become more prevalent under reduced tillage systems. Similarly, species that require burial for germination may become less prevalent. Crop residue present on the soil surface can also influence weed seedling recruitment by modifying the physical environment (mainly temperature) of weed seeds. Weed responses to plant residue could also be influenced by the allelopathic activity of the residue and the sensitivity of the weed species present. Few studies have investigated the fate of weed seeds that fail to germinate under any tillage system. Further research is needed to determine whether the weed seeds that fail to germinate decay before the start of the next growing season or become part of a persistent seed bank. Crop residues present on the soil surface can intercept a considerable amount of the applied herbicide and, depending on the herbicide, this intercepted component is susceptible to losses. Therefore, conservation tillage systems are expected to have lower efficacy of soil active herbicides. However, there has been little investigation of rate of loss of soil active herbicides under reduced tillage systems and the results reported have been inconsistent. Much of the research on these effects is from overseas and may not be true in Australian conditions. Therefore, further work is needed to clearly understand the impact of changing tillage systems on weed ecology, herbicide performance and persistence.

Evolution of herbicide resistance in weeds: initial frequency of target site-based resistance to acetolactate synthase-inhibiting herbicides in Lolium rigidum

The frequency of individuals resistant to two acetolactate synthase (ALS)-inhibiting herbicides in three previously untreated populations of Lolium rigidum was determined. The frequency of individuals resistant to the sulfonylurea herbicide sulfometuron-methyl varied from 2.2 × 10−5 to 1.2 × 10−4 and the frequency of individuals resistant to the imidazolinone herbicide imazapyr varied from 1 × 10−5 to 5.8 × 10−5 depending on the population. Application of sulfometuron-methyl selected individuals with a herbicide-insensitive ALS, which was also cross-resistant to imazapyr. The high initial frequency of individuals resistant to ALS-inhibiting herbicides in L. rigidumpopulations never previously exposed to these herbicides helps explain the rapid evolution of herbicide resistance in this species once ALS-inhibiting herbicides were used.

A Decade of Glyphosate-Resistant Lolium around the World: Mechanisms, Genes, Fitness, and Agronomic Management

Abstract Glyphosate resistance was first discovered in populations of rigid ryegrass in Australia in 1996. Since then, glyphosate resistance has been detected in additional populations of rigid ryegrass and Italian ryegrass in several other countries. Glyphosate-resistant rigid ryegrass and Italian ryegrass have been selected in situations where there is an overreliance on glyphosate to the exclusion of other weed control tactics. Two major mechanisms of glyphosate resistance have been discovered in these two species: a change in the pattern of glyphosate translocation such that glyphosate accumulates in the leaf tips of resistant plants instead of in the shoot meristem; and amino acid substitutions at Pro 106 within the target site, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). There are also populations with both mechanisms. In the case of glyphosate resistance, the target site mutations tend to provide a lower level of resistance than does the altered translocation mechanism. Each of these resistance mechanisms is inherited as a single gene trait that is largely dominant. As these ryegrass species are obligate outcrossers, this ensures resistance alleles can move in both pollen and seed. Some glyphosate-resistant rigid ryegrass populations appear to have a significant fitness penalty associated with the resistance allele. Field surveys show that strategies vary in their ability to reduce the frequency of glyphosate resistance in populations and weed population size, with integrated strategies—including alternative weed management and controlling seed set of surviving plants—the most effective. Nomenclature: Glyphosate; rigid ryegrass, Lolium rigidum Gaud. LOLRI; Italian ryegrass, Lolium multiflorum Lam. LOLMU.

Influence of tillage systems on vertical distribution, seedling recruitment and persistence of rigid ryegrass (Lolium rigidum) seed bank

Abstract Several studies were conducted to evaluate the effects of different tillage systems on the vertical seed distribution, seedling recruitment pattern, and persistence of the rigid ryegrass seed bank. Experiments were conducted in South Australia at two locations (Roseworthy Campus and Minlaton, a site on the Yorke Peninsula) in 2003 and 2005. The distribution of surface seeds through the soil profile was associated with the level of soil disturbance. The low–soil-disturbance tillage systems left more seed on the soil surface, whereas the high–soil-disturbance systems buried most of the seeds. The seedling recruitment of rigid ryegrass was lower under the low–soil-disturbance tillage systems than under the high–soil-disturbance tillage systems at both locations. The seedling recruitment was two- to fourfold greater under minimum tillage than under no-till. Not only was the seedling recruitment lower under the low–soil-disturbance tillage systems, biomass accumulation by rigid ryegrass seedlings was also lower under these systems. The carryover of residual viable seeds from one season to the next was similar between the tillage systems. However, seed decay under no-till (48 to 60%) was much greater than under minimum tillage (12 to 39%). Nomenclature: Rigid ryegrass, Lolium rigidum Gaudin, LOLRI.

Herbicide resistance in weeds endowed by enhanced detoxification: complications for management

Abstract Herbicide-resistant weeds are a constraint to weed management in many cropping regions around the world. Of the numerous populations of weeds with resistance to herbicides, it appears that most have resistance due to an alteration to the target enzyme. Use of herbicides with alternative modes of action has relatively easily controlled these populations. In stark contrast are a much smaller number of populations with resistance due to increased rates of herbicide detoxification. These populations may be cross-resistant to herbicides with other modes of action. Such cross-resistance can severely compromise weed control because alternative herbicides may fail on their first use. It has proved extremely difficult to predict cross-resistance due to increased herbicide detoxification in weed populations and hence, difficult to provide adequate advice to growers on how to avoid or manage the problem. Most commonly, such cross-resistance has been selected by certain aryloxyphenoxypropanoate herbicides such as diclofop-methyl and phenylurea herbicides such as chlorotoluron and isoproturon; however, other herbicides can also act as selecting agents for this type of resistance. Illustrative examples from rigid ryegrass in Australia and blackgrass in Europe demonstrate the breadth of the problem and the magnitude of the effort required to understand increased herbicide detoxification as a resistance mechanism. Recent work has elucidated the genetic basis of cross-resistance in some populations, but this has so far not provided new predictive tools useful to growers. Despite more than a decade of research aimed at unraveling the complexities of cross-resistance due to increased herbicide detoxification, management of these cross-resistant populations remains a significant challenge. Nomenclature: Chlorotoluron; diclofop-methyl; isoproturon; blackgrass, Alopecurus myosuroides Huds. ALOMY; rigid ryegrass, Lolium rigidum Gaud. LOLRI.