Recipient Organization
PURDUE UNIVERSITY
(N/A)
WEST LAFAYETTE,IN 47907
Performing Department
Entomology
Non Technical Summary
Proteins from the soil bacterium Bacillus thuringiensis (B.t.) offer high specificity in insect toxicity. Genes that induce crop plants to express these insecticidal proteins have been successfully inserted into corn and cotton. The vast majority of corn (over 80%) planted in the US expresses at least one Bt toxin for insect management. While this approach has largely been successful on a number of fronts, problems with resistance development by target pests are a constant threat, as they are with constant exposure to any pest management approach.Insect resistance management plans have been implemented to delay the onset of resistance targeting the corn rootworm complex. A mandatory refuge that consists of corn plants that do not express the Bt toxin must be part of every Bt corn planting. This ensures that susceptible insects will always be present to mate with any that survive Bt exposure - effectively "diluting" the population of resistant insects. In other words, this approach seeks to have abundant, susceptible, insects from refuge plants without the pest-specific toxin mating with relatively rare, resistant, insects from toxin-producing plants ("Bt" plants). However, prior research from our group on the key pest of corn, the western corn rootworm (WCR), showed that the numbers of insects generated in currently implemented refuges are insufficient to mate with those emerging from Bt corn in the same field. One possible factor limiting adult emergence from refuge plants are the neonicotinoid insecticides applied to seed. Currently, all Bt corn is sold with neonicotinoid seed treatments (NSTs).Despite this widespread use, the contribution of NSTs to corn rootworm control (including resistance management) in Bt corn is completely unknown. For example, the insecticide may be detrimental to resistance management, by causing mortality in the refuge insects that we are trying to preserve. On the other hand, there are other, non-rootworm soil pests that can be effectively managed with NSTs. We will examine the economic need for NSTs to prevent root injury and yield loss from soil insects in two ecologically distinct states where root feeding insect pests routinely occur, Indiana and Virginia.Aside from hybrid differences, most corn in the US is grown using very similar approaches - both expressing Bt toxins and using NSTs. However, the below-ground pest complexes are very different: corn rootworms dictate pest management in Indiana, whereas wireworms, white grubs, and rootworms are concerns in Virginia. While abundant data on environmental impacts exist for Bt products, the costs/benefits of persistent NST residues on soil health have not been measured - Objective 1 of our proposed work will fill this information gap. Using novel methods developed by our group, we will measure rootworm emergence from refuges with and without NSTs, then characterize the value of the control NSTs provide by complementing insect-resistant Bt maize hybrids. During Objective 2, we will collect data on insecticide residues in soil and water in these experiments. We hypothesize that NSTs can be removed from GM corn seeds grown in some regions without increase in pest damage or diminished refuge function (i.e., number of insects produced from refuge plants). Further, we hypothesize that removing NSTs will have the potential for a beneficial effect upon soil and watershed health via reduced residues. The overall goal of this research is to provide information to guide the use of NSTs in a way that does not compromise refuge function, while providing growers protection from root-feeding pests where appropriate. Reductions of levels of pesticide in soil is a stated goal of the 2017 BRAG program as well. The proposed work is directly applicable to the following specified areas from the BRAG 2017 RFA: Environmental impacts of GE relative to non-GE organisms in the context of production systemsc) Comparative assessment of environmental impacts of agricultural production systems using organic and/or conventional methods with those involving plant, animal, or biotechnology.
Animal Health Component
75%
Research Effort Categories
Basic
25%
Applied
75%
Developmental
(N/A)
Goals / Objectives
Objective 1: Quantify effects NSTs on refuge performance, Years 1-3We will measure the production of refuge insects and mating between refuge and Bt insects in corn fields. This will accomplished by collecting data on emergence and mating rates of refuge/Bt insects in 5% seed blends. We will determine if removing NSTs 1) disproportionally affects the respective population sizes of Bt/refuge insects; and 2) affects how often insects from different hosts mate.Objective 2: Quantify the relative benefit of NSTs in combination with Bt toxins for management of WCR and other soil pests, Years 1-3In this objective we will determine the contribution of each approach (Bt vs. NSTs vs. both) to pest management in the field in both VA and IN. This will allow us to test different pest complexes in different regions and growing conditions and document whether effects of both approaches are additive, synergistic or neutral.Objective 3: Quantify the levels of insecticide residues found in soil and water collected from fields planted with and without NSTs, Years 2-3.Using the same fields as experiments outlined above, we will determine whether insecticide residues change significantly as a function of pest management approach. Because we will already have data on pest management efficacy from Objective 1 and 2, this work will allow us to include a previously uninvestigated parameter of NST use, namely the accumulation (or lack thereof) of insecticide residues in soil and water in and near production fields).
Project Methods
Objective 1: Quantify effects NSTs on refuge performance Years 1-3All experiments will be conducted in corn fields in both VA and IN (four locations/state). Treatments include: 1) untreated seed, 5% refuge; 2) NST 1.25 mg/kernel clothianidin, 5% refuge. The manufacturer premixes seeds for seed blend refuges. In order to create pure stands of Bt plants, only seeds containing rootworm-specific Bt traits will be initially planted. Seeds will be separated visually by color; seeds that contain the rootworm-specific Bt trait are a different color (typically green) than refuge seeds (typically purple). Immediately following planting, 345 randomly chosen seeds (representing 5% of the 6,900 seeds used per plot, based on planting rate of 27,700 seeds per acre) will be removed and replaced with two refuge seeds. Refuge plants will be labeled with 15N, as follows: an aqueous solution of ammonium nitrate 15N (~98% 15N) (Cambridge Isotope Laboratories, Inc. Andover, MA) and distilled water will be applied as a soil drench to a 1 cm deep hole at the base of refuge plants in the V2 stage using a CO2-pressurized backpack sprayer. A rate of 0.6125 g of ammonium nitrate (~98% 15N; Cambridge Isotopes) per liter of dH20 will used; 10 mL of solution will added to each refuge plant.We will sample eight rows per plot three times per week beginning at appearance of the first adult rootworm beetle in each location. Plots and rows will be sampled in random order. Beetles will be collected into plastic bags and labeled with the date and location; mating pairs will be stored together until processing. Samples will be stored at -20°C.Stable isotope testingWe will use only elytra and head capsules from beetles for isotope analysis to avoid nitrogen from plant matter in the digestive tract and, in females, nitrogen received from male spermatophores during mating (Murphy and Krupke, 2011). Dried elytra and head capsules will be crushed between layers of wax paper, weighed to the nearest 0.001 g and placed into 4 x 6 mm mass spectrometry tin capsules (Costech Analytical Technologies, Inc., Valencia, CA). We will use fresh wax paper for each sample and clean all instruments and workspace with >70% ethanol between samples. The Purdue Stable Isotope Laboratory (Isotope Ratio Mass Spectrometer-IRMS) will perform mass spectrometry testing.Objective 2: Quantify the relative benefit of NSTs in combination with Bt toxins for management of WCR and other soil pests, Years 1-3Treatments include: 1) untreated, Bt seed; 2) NST, Bt seed; 3) untreated, non-Bt seed; 4) NST, non-Bt seed. We will measure root damage in each plot twice, ca. July 20 and Aug 1. At each interval, four consecutive plants/row (two Bt and two refuge) will be removed from 10 rows and root injury assessed following the 0-3 node injury scale, the standard approach for measuring insect feeding. Plants will be harvested, and yield and grain moisture measured, between mid and late September. Secondary soil pests (wireworms, white grubs) and their damage will be assessed by counting the number of plants in 12.2 m of row from one of the center two rows in each plot and converting this value to plants per hectare.At the same time, we will assess the presence and abundance of non-WCR soil pests, by taking soil samples from four locations in each plot prior to planting. We will sample for soil insects by excavating a 20 cm by 15 cm deep volume of soil and sifting it for insect larvae. We will identify insect larvae to species. Grain moisture and yield estimates will be obtained from the two center rows using a Kincaid® 8-XP plot combine.Objective 3: Quantify the levels of insecticide residues found in soil and water collected from fields planted with and without NSTs, Years 2-3.We will conduct these experiments in plots described above. A "reference site," as a negative control, will be located at a forested area near (<10 kilometers) field plots. Plots will be assessed for residues of clothianidin, the NST used in our field plots and the most commonly NST used on corn. ZIn addition, we will measure residue levels in soils and surface water from a wide range of sites (wetlands, streams, ditches) within Indiana and Virginia that are located near corn, soybean and/or fresh market crops to capture a range of potential neonicotinoid residue levels. A critical question regarding neonicotinoid residues is whether residues in agricultural ditches increase or decrease during storm events and how those levels ebb and flow throughout the growing season. Soil samples will be collected from 4 cm below the soil surface to avoid sampling topsoil. Using 10 cm diameter x 15 cm soil cores, we will collect samples at five random locations in the field. These samples will be combined in a 3.78 liter bucket, mixed, and a subsample of the pooled soil transported to the lab in paper bags stored in coolers to minimize the possibility of photolysis. Samples will be stored at -20°C until analysis. Water samples will be collected from ditches and waterways adjacent to fields, using 1-L amber glass bottles and transported to the laboratory on ice. Water samples will be processed within 96 h after collection. Where necessary, the pesticides will be extracted from the matrix (i.e., soil) by agitation with water, acetonitrile and the salts magnesium sulfate and sodium acetate. After centrifuging, a portion of the supernatant acetonitrile layer containing the pesticides can be further cleaned by the addition of solid phase dispersants (primary secondary amine, magnesium sulfate, C-18 silica).