W. Brien Henry

A rapid in vivo shikimate accumulation assay with excised leaf discs

Abstract An in vivo shikimate accumulation assay with excised leaf tissue was developed to provide a fast and reliable method for identifying glyphosate-resistant plants. The assay is based on glyphosate-induced accumulation of shikimate. There was a linear accumulation of shikimate in excised leaf discs of soybean and canola treated with 250 μM glyphosate for 48 h. The IC50 for the accumulation of shikimate in soybean and corn leaf discs was 34 and 87 μM, respectively. Leaf discs excised from glyphosate-resistant corn or soybean did not accumulate shikimate when treated with 500 μM glyphosate. Leaf discs taken from a number of field-grown plants accumulated shikimate in a glyphosate dose–dependent manner. The accumulation of shikimate was dependent on light and the age of the leaf from which the disc was taken. The assay worked either in 96-well microtiter plates or in vials, and it clearly differentiated between glyphosate-resistant and -susceptible crops in which the resistance is due to an alteration of the target site for glyphosate. The assay was simple and robust and has the potential to be used as a high throughput assay to detect glyphosate resistance in weeds. Nomenclature: Glyphosate; canola, Brassica napus L. ‘Hayola 420’; corn, Zea mays L. ‘Pioneer 37M34’, ‘Dekalb DK493RR/BTY’; soybean, Glycine max (L.) Merr., ‘Asgrow A2869’, ‘Asgrow AG3003’.

Assessment of two nondestructive assays for detecting glyphosate resistance in horseweed (Conyza canadensis)

Abstract Two rapid, nondestructive assays were developed and tested for their potential in differentiating glyphosate-resistant from glyphosate-susceptible biotypes of horseweed. In one assay, leaves of glyphosate-resistant and -susceptible corn, cotton, and soybean plants as well as glyphosate-resistant and -susceptible horseweed plants were dipped in solutions of 0, 300, 600, and 1200 mg ae L−1 glyphosate for 3 d and subsequent injury was evaluated. In the second assay, plant sensitivity to glyphosate was evaluated in vivo by incubating excised leaf disc tissue from the same plants used in the first assay in 0.7, 1.3, 2.6, 5.3, 10.6, 21.1, 42.3, and 84.5 mg ae L−1 glyphosate solutions for 16 h and measuring shikimate levels with a spectrophotometer. The leaf-dip assay differentiated between glyphosate-resistant and -susceptible crops and horseweed biotypes. The 600 mg L−1 rate of glyphosate was more consistent in differentiating resistant and susceptible plants compared with the 300 and 1,200 mg L−1 rates. The in vivo assay detected significant differences between susceptible and glyphosate-resistant plants of all species. Shikimate accumulated in a glyphosate dose-dependent manner in leaf discs from susceptible crops, but shikimate did not accumulate in leaf discs from resistant crops and levels were similar to nontreated leaf discs. Shikimate accumulated at high (≥ 21.1 mg ae L−1) concentrations of glyphosate in leaf discs from all horseweed biotypes. Shikimate accumulated at low glyphosate concentrations (≤ 10.6 mg L−1) in leaf discs from susceptible horseweed biotypes but not in resistant biotypes. Both assays were able to differentiate resistant from susceptible biotypes of horseweed and might have utility for screening other weed populations for resistance to glyphosate. Nomenclature: Glyphosate; horseweed, Conyza canadensis (L.) Cronq. ERICA; corn, Zea mays L. ‘Dekalb 687RR’, ‘Pioneer 31B13’; cotton, Gossypium hirsutum L. ‘Delta and Pine Land 444RR’, ‘Suregrow 747’; soybean, Glycine max (L.) Merr. ‘Delta and Pine Land 4748’, ‘Asgrow 4702RR’.

Shikimate Accumulation in Sunflower, Wheat, and Proso Millet after Glyphosate Application

Abstract Experiments were conducted to examine the utility of a spectrophometric leaf disc assay for detecting shikimate accumulation after glyphosate application in sunflower, proso millet, and wheat. The assay was conducted on both greenhouse- and field-grown plants. Glyphosate was applied at five rates ranging from 840 to 53 g ae ha−1. Shikimate accumulation data were generated at 1, 4, 7, and 14 d after application (DAA). Sunflower accumulated shikimate more rapidly and at lower glyphosate rates than the other two species. At 14 DAA, glyphosate at the two highest rates remained detectable in all three species. Plants receiving lower glyphosate doses (210, 105, and 53 g ae ha−1) had begun to grow out of the injury, or at least the shikimate levels in the plants were no longer significantly different than that present in the untreated controls. This spectrophotometric assay is both rapid and simple, with respect to other means of detecting shikimate, and it can be used to detect glyphosate drift. For it to be used by crop managers, samples from potentially drift-affected crops should be taken as soon as possible after the suspected drift event or immediately after the appearance of glyphosate injury. Nomenclature: Glyphosate; proso millet, Panicum miliaceum L. ‘Sunup’; sunflower, Helianthus annus L. ‘Triumph 765C’; wheat, Triticum aestivum, L. ‘Stak-Tite N25550’.

Rapid Assay for Detecting Enhanced Atrazine Degradation in Soil

Abstract Atrazine is widely used to control broadleaf weeds and grasses in corn, sorghum, and sugarcane. Field persistence data published before 1995 showed that the average half-life of atrazine in soil was 66 d, and farmers expect to achieve weed control with a single application for the full season. However, reports of enhanced atrazine degradation in soil from fields that have a history of atrazine applications are increasing. A rapid laboratory assay was developed to screen soils for enhanced atrazine degradation. Soil (50 g) was placed in a 250 ml glass jar and treated with 7.5 ml of water containing atrazine (5 µg ai ml−1) and capped with a Teflon-lined lid. The assay was conducted at room temperature (25 C). Soil subsamples (1.5 to 3 g) were removed at 0, 1, 2, 4, 8, and 16 d after treatment and extracted with an equal weight of water (wt/vol). The atrazine in the water extract was assayed with high-pressure liquid chromatography (HPLC). The half-life of atrazine in soils with a history of use was ≤ 1.5 d, whereas the half-life in soils with no history of atrazine use was > 8 d. The advantages of this assay are (1) the ease of set up; (2) the rapidity of extraction, and (3) the simplicity of the quantification of the atrazine. Nomenclature: Atrazine; corn, Zea mays L; sorghum, Sorghum bicolor (L.) Moench; sugarcane, Saccharum officinarum L.

Jointed Goatgrass (Aegilops Cylindrica) by Imidazolinone-Resistant Wheat Hybridization under Field Conditions

Abstract Gene flow between jointed goatgrass and winter wheat is a concern because transfer of herbicide-resistance genes from imidazolinone-resistant (IR) winter wheat cultivars to jointed goatgrass could restrict weed-management options for this serious weed of winter wheat cropping systems. The objectives of this study were (1) to investigate the frequency of interspecific hybridization between IR wheat and jointed goatgrass in eastern Colorado, and (2) to determine the gene action of the IR acetolactate synthase (ALS) allele in IR wheat by jointed goatgrass and in IR wheat by imidazolinone-susceptible (IS) wheat backgrounds. Jointed goatgrass was sampled side-by-side with IR wheat and at distances up to 53 m away in both experimental plots and at commercial field study sites in 2003, 2004, and 2005. A greenhouse-screening method was used to identify IR hybrids in collected jointed goatgrass seed. The average percentage of hybridization across sites and years when IR wheat and jointed goatgrass were grown side-by-side was 0.1%, and the maximum was 1.6%. The greatest distance over which hybridization was documented was 16 m. The IR ALS allele contributed 25% of untreated ALS activity in jointed goatgrass by IR wheat F1 plants, as measured by an in vitro ALS assay. The hybridization rate between wheat and jointed goatgrass and the expression of the IR wheat ALS allele in hybrid plants will both influence trait introgression into jointed goatgrass. Nomenclature: Jointed goatgrass, Aegilops cylindrica Host AEGCY; hard red winter wheat, Triticum aestivum L. ‘Above’, ‘Bond’, ‘Prairie Red’, ‘Halt’.

Remote Sensing to Detect Herbicide Drift on Crops

Glyphosate and paraquat herbicide drift injury to crops may substantially reduce growth or yield. Determining the type and degree of injury is of importance to a producer. This research was conducted to determine whether remote sensing could be used to identify and quantify herbicide injury to crops. Soybean and corn plants were grown in 3.8-L pots to the five- to seven-leaf stage, at which time, applications of nonselective herbicides were made. Visual injury estimates were made, and hyperspectral reflectance data were recorded 1, 4, and 7 d after application (DAA). Several analysis techniques including multiple indices, signature amplitude (SA) with spectral bands as features, and wavelet analysis were used to distinguish between herbicide-treated and nontreated plants. Classification accuracy using SA analysis of paraquat injury on soybean was better than 75% for both 1/2- and 1/8× rates at 1, 4, and 7 DAA. Classification accuracy of paraquat injury on corn was better than 72% for the 1/2× rate at 1, 4, and 7 DAA. These data suggest that hyperspectral reflectance may be used to distinguish between healthy plants and injured plants to which herbicides have been applied; however, the classification accuracies remained at 75% or higher only when the higher rates of herbicide were applied. Applications of a 1/2× rate of glyphosate produced 55 to 81% soybean injury and 20 to 50% corn injury 4 and 7 DAA, respectively. However, using SA analysis, the moderately injured plants were indistinguishable from the uninjured controls, as represented by the low classification accuracies at the 1/8-, 1/32-, and 1/64× rates. The most promising technique for identifying drift injury was wavelet analysis, which successfully distinguished between corn plants treated with either the 1/8- or the 1/2× rates of paraquat compared with the nontreated corn plants better than 92% 1, 4, and 7 DAA. These analysis techniques, once tested and validated on field scale data, may help determine the extent and the degree of herbicide drift for making appropriate and, more importantly, timely management decisions. Nomenclature: Corn, Zea mays L.; soybean, Glycine max (L.) Merr. Additional index words: Glyphosate, hyperspectral imagery, indices, paraquat, ROC curve, wavelet analysis. Abbreviations: DAA, days after application; DINO, differential index of normalized observations; DWT, discrete wavelet transform; EPSP, 5-enolpyruvylshikimate-3-phosphate; LDA, linear discriminant analysis; NDVI, normalized difference vegetation index; NIR, near-infrared; POST, postemergence; ROC, receiver operator characteristics; SA, signature amplitudes.

Interactive effects on CO2, drought, and ultraviolet-B radiation on maize growth and development

Crop growth and development are highly responsive to global climate change components such as elevated carbon dioxide (CO2), drought, and ultraviolet-B (UV-B) radiation. Plant tolerance to these environmental stresses comprises its genetic potential, physiological changes, metabolism, and signaling pathways. An inclusive understanding of morphological, physiological, and biochemical responses to these abiotic stresses is imperative for the development of stress tolerant varieties for future environments. The objectives of this study were to characterize the changes in vegetative and physiological traits in maize hybrids in their response to multiple environmental factors of (CO2) [400 and 750μmolmol(-1) (+(CO2)], irrigation treatments based evapotranspiration (ET) [100 and 50% (-ET)], and UV-B radiation [0 and 10kJm(-2)d(-1) (+UV-B)] and to identify the multiple stress tolerant hybrids aid in mitigating projected climate change for shaping future agriculture. Six maize hybrids (P1498, DKC 65-81, N75H-GTA, P1319, DKC 66-97, and N77P-3111) with known drought tolerance variability were grown in eight sunlit, controlled environment chambers in which control treatment consisted of 400μmolmol(-1) [CO2], 100% ET-based irrigation, and 0kJ UV-B. Plants grown at +UV-B alone or combination with 50% ET produced shorter plants and smaller leaf area while elevated CO2 treatments ameliorated the damaging effects of drought and higher UV-B levels on maize hybrids. Plant height, leaf area, total dry matter chlorophyll, carotenoids, and net photosynthesis measured were increased in response to CO2 enrichment. Total stress response index (TSRI) for each hybrid, developed from the cumulative sum of response indices of vegetative and physiological parameters, varied among the maize hybrids. The hybrids were classified as tolerant (P1498), intermediate (DKC 65-81, N75H-GTA, N77P-3111) and sensitive (P1319 and DKC 66-97) to multiple environmental stresses. The positive correlation between TSRI and vegetative and physiological index developed in this study demonstrates that a combination of vegetative and physiological traits is an effective screening tool to identify germplasm best suited to cope with future changing climates. Furthermore, the tolerant hybrids identified in this study indicate that the possibility of cultivar selection for enhanced agronomic performance and stability in a water limited environment with higher UV-B, anticipated to occur in future climates.

Growth and physiological trait variation among corn hybrids for cold tolerance

Abstract: Global food demand has risen continuously because of increasing population with greater food and energy needs. Corn production, however, is constrained by current and possible increased future variability in the weather. Earlier planting is a strategy for U.S. Mid-South corn producers to avoid typical summer droughts. However, planting early will increase the likelihood of seedlings exposure to cold temperatures. The objectives of this study were to evaluate corn hybrids planted when the conditions are desirable followed by low and moderately low temperatures to assess the variability among the vegetative and physiological parameters and to classify hybrids into different cold tolerant groups. Twenty one commercially-grown hybrids were subjected to three day/night temperature treatments; 29/21 °C (optimum), 25/17 °C (moderately low), and 21/13 °C (low) from 15 d after planting (DAP) for plants grown at optimum temperature. Shoot, root, and physiological parameters were measured, 32-34 DAP. Significant differences and interactions were observed among the temperature treatments and hybrids for most of the traits measured. Based on relative scores, developed in this study, AR1262 and P1636YHR were classified as cold tolerant and H68B and ST11504VT3 as cold sensitive. Cold tolerant hybrids and their associated morpho-physiological characteristics may be useful for breeders to develop new hybrids that could withstand low and variable temperatures during seedling growth and developmental period.

Spatial variability of atrazine and metolachlor dissipation on dryland no-tillage crop fields in Colorado.

An area of interest in precision farming is variable-rate application of herbicides to optimize herbicide use efficiency and minimize negative off-site and non-target effects. Site-specific weed management based on field scale management zones derived from soil characteristics known to affect soil-applied herbicide efficacy could alleviate challenges posed by post-emergence precision weed management. Two commonly used soil-applied herbicides in dryland corn (Zea mays L.) production are atrazine and metolachlor. Accelerated dissipation of atrazine has been discovered recently in irrigated corn fields in eastern Colorado. The objectives of this study were (i) to compare the rates of dissipation of atrazine and metolachlor across different soil zones from three dryland no-tillage fields under laboratory incubation conditions and (ii) to determine if rapid dissipation of atrazine and/or metolachlor occurred in dryland soils. Herbicide dissipation was evaluated at time points between 0 and 35 d after soil treatment using a toluene extraction procedure with GC/MS analysis. Differential rates of atrazine and metolachlor dissipation occurred between two soil zones on two of three fields evaluated. Accelerated atrazine dissipation occurred in soil from all fields of this study, with half-lives ranging from 1.8 to 3.2 d in the laboratory. The rapid atrazine dissipation rates were likely attributed to the history of atrazine use on all fields investigated in this study. Metolachlor dissipation was not considered accelerated and exhibited half-lives ranging from 9.0 to 10.7 d in the laboratory.

Comparison of the side-needle and knife techniques for inducing Aspergillus flavus infection and aflatoxin accumulation in corn hybrids.

Evaluation of corn genotypes for resistance to aflatoxin accumulation has evolved over the past 30 years. Inoculation techniques have been developed to ensure that plants are exposed to Aspergillus flavus and resistant genotypes can be identified. We compared two inoculation techniques (side needle and knife) and different inoculation rates in Georgia and Mississippi. The relative performance of side-needle and knife inoculation techniques did not change or interact significantly across site years, suggesting that both the needle and knife are consistent inoculation techniques for discriminating between resistant and susceptible corn hybrids at the Mississippi State and Tifton locations.