Location: Vegetable Crops Research
2021 Annual Report
Objectives
Objective 1: Identify pollinator behaviors, pollinator management strategies, and crop production strategies that together mitigate unintended gene flow.
Sub-objective 1.1: Pollinator behavior and plant reproductive strategies affect gene flow risk.
Sub-objective 1.2: Visual and Olfactory cues that attract pollinators can guide the development of pollinator or crop management strategies that reduce gene flow and increase yield.
Objective 2: Determine the impacts of cultivated carrot genes on the genomic landscape of wild carrot.
Approach
Objective 1. This objective is divided into two sub-objectives, each with three hypotheses to be tested.
Sub-Objective 1.1. We will use a combination of field and greenhouse experiments to test the hypotheses within this subobjective. For example, the rules bees use when moving between patches or fields will be tested using patches of distinct sizes and isolation distances and measuring the number of transitions made by bees from a center glyphosate-resistant patch to the different conventional patches. The number of gene flow events in the different conventional patches, identified by the presence of glyphosate-resistant seeds, will also be used to test the decision making process of bumble bees. Greenhouse experiments will examine the pattern of seed deposition on flowers visited in succession by three bee species, honey bees, leafcutting bees and bumble bees. We will use glyphosate-resistant pollen donor and conventional pollen recipients and examine the number and proportion of glyphosate-resistant seeds on flowers visited in succession to determine the seed curve for each bee species.
Sub-Objective 1.2. To determine the preference of each of three bee species to visual and/or olfactory cues, we will perform greenhouse experiments and quantify approaches and landings to different visual and/or olfactory cues. To identify a blend derived from nest cells that attract leafcutting bees, we will capture and identify the chemicals present in the bee cell using Gas Chromatography-Mass Spectrometry (GC-MS); determine whether there is a behavioral response and then use couple gas chromatography – electroantennographic detection (GC-EAD) to identify physiological responses. Finally, the electrophysiologically active constituents will be tested using a behavioral assay.
Objective 2. We will use genotyping by sequencing on both cultivated carrots used in a breeding program and wild carrots in close proximity to the breeding area and far away to detect the presence of cultivated carrot genes in wild carrot populations. The presence of cultivated genes in wild populations represents introgression. We will determine the extent of introgression of cultivar genes in wild carrot populations.
Progress Report
Objective 1, Sub-Objective 1.1. A manuscript on transition rules used by bumble bees was published in 2021 in Scientific reports. These results demonstrate how bumble bees utilize both patch size and distance between patches when selecting the patch (or field) they move to next. Bumble bees can estimate the total amount of resources available in a patch.
A manuscript examining selfing rate in alfalfa seed production was submitted and accepted by Frontiers in Plant Sciences in 2021. This study reported the selfing rate of 32 alfalfa seed production fields and identified a 15% reduction in seed production (seeds per stem) following selfing. Results indicated the prevalence of pollinator-mediated selfing in alfalfa seed production and stressed the fact that increasing pollinator abundance does not reduce selfing. In fact, eliminating selfing in alfalfa seed production would require selection of self-incompatible varieties.
A collaboration to determine the role of factors, besides distance, on gene flow and adventitious presence(AP) of glyphosate resistance in alfalfa seed production fields resulted in a manuscript published in 2021 in PlOs ONE. This study pointed out the roles of the size of the GE pollen pool within the pollinator foraging range, and of the foraging behavior of pollinators on gene flow and AP.
Experiments were performed in the greenhouse to examine the rules of patch transition used by alfalfa leafcutting bees when moving between alfalfa patches and subsequent gene flow.
Experiments were performed in the greenhouse to examine the rules used by bumble bees when moving between patches of alfalfa, most specifically how they use size of fields vs. distance between fields when selecting patches. We compared the results of the experiment ran in the greenhouse to the results of the field experiment.
Objective 1, Sub-objective 1.2. We performed tests in the greenhouse to determine the role of visual and olfactory cues in attracting leafcutting bees to alfalfa plants.
ARS and University of Wisconsin scientists in Madison, Wisconsin, performed behavior experiments to determine the preference of the western tarnished plant bug, Lygus hesperus, to various host plants.
Objective 2. Genotyping by sequencing was performed on 10-20 individuals of each carrot population for the latest populations collected in Iowa, California, Colorado, South Dakota, Minnesota and Massachusetts. Single Nucleotide polymorphisms (SNP) were identified from the GBS data. The SNP data are being analyzed to determine the extent of introgression of cultivar genes into wild carrot populations in different parts of the U.S.
Feral alfalfa populations are being located in Wisconsin, Kansas, California, Washington and Montana and tested for the presence of the glyphosate resistance gene. This project relates to gene flow and introgression from cultivars into feral or wild populations, in alfalfa (Objective 2).
Accomplishments
1. Selfing is prevalent in alfalfa seed production and lead to a 15% reduction in seed yield. Self-pollination is prevalent in alfalfa seed production and leads to a 15% reduction in seed yield. Self pollination or “selfing” can create inbreeding depression and led to a reduction in seed yield. Seed yield is important because it determines how many seeds a farmer produces in a field. ARS researchers in Madison, Wisconsin, estimated the level of selfing using genetic markers in 32 seed-production fields over three alfalfa seed production areas, the Pacific Northwest, the Central Valley, and the Imperial Valley of California. Selfing was detected in each field, with the rate varying between 5.8 and 30%; which means that 5.8 to 30% of the seeds resulted from a self-pollination event. In addition, selfing created a 15% reduction in seeds set per stem. Selfing in alfalfa seed production was pollinator mediated, meaning that it resulted from pollinators (e.g. bees) moving pollen between flowers on the same plant (geitonogamous selfing) and possibly within the same flower (facilitated autogamy). The fact that pollinators mediated selfing implies that selfing will always occur when pollinators are used to produce seeds, whether in a field or greenhouse setting and irrespective of the field dimensions. In addition, contrary to a common misconception, increasing pollinator abundance will not reduce selfing. Stems with many racemes (cluster of flowers) and more open flowers had higher selfing via geitonogamy and higher seed set. Thus, to reduce selfing and maintain seed yield in alfalfa, breeders should select for self-incompatible varieties. Identifying the mating system of a crop and its prevailing mode of selfing can guide the development of effective strategies to reduce selfing and increase yield. This information will be helpful to alfalfa breeders, alfalfa growers and the alfalfa industry interested in increasing alfalfa yield.
2. Netting and pan traps fail to identify pollinators of a crop. Netting and pan traps fail to identify pollinators of alfalfa. Pollinators are in decline and affect the reproduction of many plant species. In agriculture, most fruits, vegetables and forage plants like alfalfa need pollinators for seed production. The decline in pollinators is expected to cause significant reductions in food production and plant reproduction. Netting and pan traps are the two most widely used methods to capture pollinators and measure bee diversity. ARS researchers at Madison, Wisconsin, in collaboration with Oak Ridge Science and Education scientists compared the effectiveness of netting and pan traps at capturing pollinators of alfalfa, Medicago sativa. Netting was more effective at capturing known pollinators of alfalfa, especially bumble bees and honey bees. Pan traps captured a higher bee diversity relative to netting and, like previous studies, each survey method was more efficient at capturing certain types of bees. However, without a prior knowledge of pollinators, neither survey method identified which of the bee species observed by these methods could actually pollinate alfalfa. Therefore, direct observations are recommended when the goal of a study is to identify pollinators or link pollinator decline to a particular crop. These results should be of interest to policy makers, the general public, and the scientific community.
3. Management practices to reduce AP in conventional alfalfa seed fields. Management practices to reduce the unwanted accidental transfer of genetically engineered traits to conventional alfalfa seed fields. In insect-pollinated crops, the transfer of genes from plant to plant is affected by numerous factors including crop characteristics, mating system, life history, pollinators, and planting management practices. Previous studies have concentrated on the impact of distance between genetically engineered (GE) and conventional fields on the accidental, unwanted transfer of genetically engineered traits. The impact of variables other than distance on gene transfer has received little attention. Moreover, some of the parent seed lots used to establish conventional fields already contain unwanted genetically engineered traits. ARS researchers at Pullman, Washington, measured various field and pollinator characteristics in conventional and surrounding GE fields, together with incidence of the genetically engineered trait in the conventional fields. Distance from the GE field explained 66% of the variability in GE trait presence, confirming its importance in affecting unwanted transfer. The area of GE fields within the pollinator foraging range explained an additional 30% of the variation in GE trait in conventional fields. The density of alfalfa leafcutting bee domiciles also influenced GE trait transfer. To minimize GE trait transfer to conventional alfalfa seed fields, management practices should focus on optimizing isolation distances between GE and conventional fields, while also considering the size of the GE pollen pool within the pollinator foraging range, and the foraging behavior of pollinators. Because the GE trait was detected in the conventional seed lot used for planting, screening of parental seed lots for GE trait presence prior to planting is also recommended. This information should benefit growers who want to minimize GE trait transfer to their conventional fields.
Review Publications
Fragoso, F.P., Jiang, Q., Clayton, M.K., Brunet, J. 2021. Patch selection by bumble bees navigating discontinuous landscapes. Scientific Reports. 11. Article 8986. https://doi.org/10.1038/s41598-021-88394-2.
Brunet, J., Flick, A., Bauer, A. 2021. Phenotypic selection on flower color and floral display size by three bee species. Frontiers in Plant Science. 11. Article 587528. https://doi.org/10.3389/fpls.2020.587528.
Boyer, K.J., Fragoso, F.P., Dieterich Mabin, M.E., Brunet, J. 2020. Netting and pan traps fail to identify the pollinator guild of an agricultural crop. Scientific Reports. 10. Article 13819. https://doi.org/10.1038/s41598-020-70518-9.
Kesoju, S., Kramer, M.H., Brunet, J., Greene, S.L., Jordan, A., Martin, R.C. 2021. Gene flow in commercial alfalfa (Medicago sativa subsp. sativa L.) seed production fields: Distance is the primary but not the sole influence on adventitious presence. PLoS ONE. 16(3). Article e0248746. https://doi.org/10.1371/journal.pone.0248746.
Santa-Martinez, E., Castro, C.C., Flick, A.J., Sullivan, M.L., Riday, H., Clayton, M.K., Brunet, J. 2021. Bee species visiting Medicago sativa differ in pollen deposition curves with consequences for gene flow. American Journal of Botany. 108(6):1016-1028. https://doi.org/10.1002/ajb2.1683.
