Scott J. Nissen

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.

Inheritance of Resistance to The Auxinic Herbicide Dicamba in Kochia (Kochia scoparia)

Abstract The inheritance of resistance to the auxinic herbicide dicamba was examined in a kochia population from Nebraska. An inbred, resistant line was developed by selection and selfing over seven generations to ensure any resistance alleles would be homozygous in the parents. An inbred, susceptible line was similarly developed, but without selection. Dose–response experiments with dicamba determined a glyphosate-resistant concentration required to inhibit dry weight accumulation by 50% (GR50) of 45 and 1,331 g ae ha−1 for the susceptible and resistant populations, respectively. F1 crosses were made between resistant and susceptible inbred individuals by hand-pollination, and the F1 plants were selfed to produce F2 plants. The F2 population was screened with 280 g ha−1 dicamba, a rate that could discriminate between susceptible and resistant plants. A total of eight F2 families were screened twice. In the first screen, seven F2 families segregated in a 3:1 ratio, consistent with a single dominant allele controlling resistance, and in the second screen six F2 families segregated in a 3:1 ratio. F2 individuals were selfed, the F3 progeny were tested with 280 g ha−1 dicamba, and the genotype of each F2 parent was determined based on F3 progeny segregation. F3 family segregation was consistent with the F2 parents having a 1:2:1 homozygous-susceptible:heterozygote:homozygous-resistant pattern, confirming that resistance to dicamba in kochia is likely conferred by a single allele with a high degree of dominance. Nomenclature: Dicamba, kochia, Kochia scoparia (L.) Schrad. KCHSC

Herbicide-resistant weeds: from research and knowledge to future needs.

Synthetic herbicides have been used globally to control weeds in major field crops. This has imposed a strong selection for any trait that enables plant populations to survive and reproduce in the presence of the herbicide. Herbicide resistance in weeds must be minimized because it is a major limiting factor to food security in global agriculture. This represents a huge challenge that will require great research efforts to develop control strategies as alternatives to the dominant and almost exclusive practice of weed control by herbicides. Weed scientists, plant ecologists and evolutionary biologists should join forces and work towards an improved and more integrated understanding of resistance across all scales. This approach will likely facilitate the design of innovative solutions to the global herbicide resistance challenge.

Potato (Solanum tuberosum) Variety and Weed Response to Sulfentrazone and Flumioxazin1

Field and greenhouse studies were conducted to evaluate sulfentrazone and flumioxazin as preemergence (PRE) herbicides for broadleaf weed control in potato. Sulfentrazone and flumioxazin were applied alone and in combination with s-metolachlor to determine the crop response and the weed spectrum controlled. These treatments were compared with metribuzin or rimsulfuron plus s-metolachlor treatments. Potato variety response to sulfentrazone and flumioxazin was evaluated in a separate field study. Sangre, Chipeta, Russet Norkotah, and Russet Nugget were treated with sulfentrazone from 0.14 to 0.28 kg/ha or flumioxazin from 0.035 to 0.07 kg/ha. Sulfentrazone and flumioxazin provided excellent broadleaf weed control at all the rates tested, whereas grass control increased as rate increased. Grass control improved when combined with s-metolachlor. Sulfentrazone and flumioxazin treatments were comparable with metribuzin and rimsulfuron treatments in weed control and total yield. Flumioxazin was safe when applied PRE to four selected varieties, whereas sulfentrazone produced initial phytotoxicity to Sangre and Chipeta at high rates but did not affect yields. Sulfentrazone increased the yield of U.S. No.1 potatoes compared with other treatments in the variety response study. Dose–response curves were used to generate the sulfentrazone, flumioxazin, and metribuzin herbicide rates required to reduce biomass by 50% (I50) for eight common weed species. Herbicides were applied PRE at several rates, and plant response was recorded. Log-logistic analysis was performed on bioassay data generated to estimate species sensitivity to each herbicide. Sulfentrazone reduced the biomass of hairy nightshade, black nightshade, redroot pigweed, kochia, common lambsquarters, and redstem filaree by more than 90% at 0.0175 kg/ha (the lowest rate evaluated), whereas flumioxazin had a similar effect on all broadleaf species except on kochia at 0.004 kg/ha (the lowest rate evaluated). Therefore, it was not possible to calculate I50 or even I80 values for most broadleaf species. Metribuzin I50 values could be calculated for most of the species tested. The metribuzin I50 value for hairy nightshade was 0.28 kg/ha, which was 16 and 70 times higher than the sulfentrazone and flumioxazin rates, respectively, that reduced hairy nightshade biomass by more than 90%. Sulfentrazone and flumioxazin appeared to be sufficiently safe when applied on potato and controlled several weed species common to potato production in the western United States. Nomenclature: Flumioxazin; sulfentrazone; black nightshade, Solanum nigrum L. #3 SOLNI; common lambsquarters, Chenopodium album L. # CHEAL; hairy nightshade, Solanum sarrachoides Sendtner # SOLSA; kochia, Kochia scoparia L. # KOCSC; redstem filaree, Erodium cicutarium L. # EROCI; redroot pigweed, Amaranthus retroflexus L. # AMARE; potato, Solanum tuberosum L. ‘Chipeta’, ‘Russet Norkotah’, ‘Russet Nugget’, ‘Sangre’. Additional index words: Arachis hypogaea, barnyardgrass, crop safety, diquat, ECHCG, Echinochloa crus-galli L., log-logistic analysis, MALNE, Malva neglecta Wallr., Panicum miliaceum L., PANMI, paraquat, Phaseolus lunatus, phytotoxicity, potato weed management, protoporphyrinogen oxidase inhibitors, Setaria viridis L., SETVI, weed dose–response, wild proso millet. Abbreviations: ALS, acetolactate synthase; I50, herbicide rate required to reduce biomass by 50%; OM, organic matter; POST, postemergence; PPO, protoporphyrinogen oxidase; PRE, preemergence; WAT, weeks after treatment.

The importance of analytical techniques in allelopathy studies with the reported allelochemical catechin as an example

Allelopathy can be challenging to demonstrate. Developing rigorous analytical techniques to detect and quantify compound(s) of interest from soil or liquid media lays the foundation for designing ecologically relevant experiments that incorporate candidate allelochemicals. In this paper, fundamental components of analytical techniques, including method development, validation, and appropriate controls are discussed. Research on the candidate allelochemical from spotted knapweed, catechin, is used as an example to demonstrate the importance of including these components both during data collection and in subsequent publications. This example shows how contrasting results between research groups can be difficult to interpret when information on controls and method validation are not included in publications. Recent research suggests that catechin is not likely driving spotted knapweed’s invasion, and thus future research on this system should focus on alternate candidate toxins from spotted knapweed. By employing appropriate analytical techniques, such as those outlined here, a strong foundation can be laid for ecologically oriented experiments that examine the role of allelochemicals in structuring communities.

Effect of Commercial Adjuvants on Vegetable Crop Fungicide Coverage, Absorption, and Efficacy

The addition of an appropriate adjuvant with foliar fungicide can significantly improve coverage, absorption, and efficacy. Laboratory and field studies evaluated coverage, absorption, and efficacy of commercial adjuvants with diverse chemistries on multiple host-pathogen systems. Organosilicone-based adjuvants improved coverage by 26 to 38% compared with a latex spreader-sticker and water. Significant crop by coverage interaction effects were also detected. The organosilicone/methylated seed oil-based adjuvant, Aero Dyne-Amic, significantly improved total [14C]azoxystrobin absorption on onion and potato by 30 and 21%, respectively, compared with water. The spreader-sticker, Bond, improved [14C]azoxystrobin absorption on onion and dry bean by 41 and 39%, respectively, compared with water. In experimental field plots, dry bean rust incidence was reduced by 52% when Kinetic or Latron AG-98 was added to maneb compared with maneb alone. The area under the potato early blight disease progress curve was reduced 29, 24, or 21% when Kinetic, Bond, or Latron AG-98 was added to maneb, respectively, compared with maneb applications alone.

Nonlinear Regression Analysis of Herbicide Absorption Studies

Abstract Although foliar herbicide absorption has been studied intensively, there is currently no standardized method for data analysis when evaluating herbicide absorption over time. Most peer-reviewed journals require the treatment structure of data be incorporated in the analysis; however, many herbicide absorption studies published in the past 5 yr do not account for the time structure of the experiment. Herbicide absorption studies have been presented in a variety of ways, making it difficult to compare results among studies. The objective of this article is to propose possible nonlinear models to analyze herbicide absorption data and to provide a stepwise framework so that researchers may standardize the analysis method in this important research area. Asymptotic regression and rectangular hyperbolic models with similar parameterizations are proposed, so that the maximum herbicide absorption and absorption rate may be adequately modeled and statistically compared among treatments. Adoption of these models for herbicide absorption analysis over time will provide a standardized method making comparison of results within and among studies more practical.

Characterization of glyphosate resistance in Amaranthus tuberculatus populations.

The evolution of glyphosate-resistant weeds has recently increased dramatically. Six suspected glyphosate-resistant Amaranthus tuberculatus populations were studied to confirm resistance and determine the resistance mechanism. Resistance was confirmed in greenhouse for all six populations with glyphosate resistance factors (R/S) between 5.2 and 7.5. No difference in glyphosate absorption or translocation was observed between resistant and susceptible individuals. No mutation at amino acid positions G101, T102, or P106 was detected in the EPSPS gene coding sequence, the target enzyme of glyphosate. Analysis of EPSPS gene copy number revealed that all glyphosate-resistant populations possessed increased EPSPS gene copy number, and this correlated with increased expression at both RNA and protein levels. EPSPS Vmax and Kcat values were more than doubled in resistant plants, indicating higher levels of catalytically active expressed EPSPS protein. EPSPS gene amplification is the main mechanism contributing to glyphosate resistance in the A. tuberculatus populations analyzed.

Genetic diversity of jointed goatgrass (Aegilops cylindrica) determined with RAPD and AFLP markers.

Abstract Two DNA molecular marker techniques were used to evaluate genetic diversity in 58 accessions of jointed goatgrass and 6 accessions of the related wild species barb goatgrass. Random amplified polymorphic DNA (RAPD) assays were performed on 8 U.S. and 50 Eurasian jointed goatgrass accessions using 30 random decamer primers. The frequency of scorable polymorphic bands within jointed goatgrass was 6 out of 90 (6.7%). Cluster analysis of RAPD data showed small genetic distances (values of 0.005 or less) among jointed goatgrass accessions. To validate the effectiveness of RAPD techniques to detect genetic diversity in tetraploid Aegilops species, six accessions of barb goatgrass were assayed using a subset of 20 decamer primers (from the original 30). RAPD data for barb goatgrass were pooled with jointed goatgrass data from the same primers. A total of 63 scorable bands were generated, of which 27 (43%) were polymorphic between two or more accessions. RAPD analysis readily distinguished between the two species and detected much greater levels of genetic diversity within barb goatgrass than between the jointed goatgrass accessions. Amplified fragment length polymorphism (AFLP) assays were performed on a subset of the 58 jointed goatgrass accessions, 3 U.S. and 13 Eurasian. These accessions were selected to represent a range in geographic diversity within our collection. Ten primer combinations generated 560 scorable bands of which 28 (5%) were polymorphic. Cluster analysis of AFLP data showed a slightly smaller range in genetic distance (0.0002 to 0.0022) among accessions compared with RAPD results; however, AFLPs distinguished among all but 2 of the 16 accessions surveyed. Although AFLP produced more scorable bands than RAPD did, both methods revealed limited genetic diversity in jointed goatgrass. Nomenclature: Jointed goatgrass, Aegilops cylindrica L. AEGCY; barb goatgrass, Aegilops triuncialis L. AEGTR.

Above winter wheat

Above, a hard red winter wheat (Triticum aestivum L. em. Thell.), is adapted for dryland production in the west central Great Plains of the United States. It carries a nontransgenic source of tolerance to imidazolinone herbicides derived by mutation induction with sodium azide. Above was developed cooperatively by the Colorado and Texas Agricultural Experiment Stations and released to seed producers in September 2001. Key words: Triticum aestivum, wheat (winter), cultivar description, herbicide tolerance