Source: TEXAS A&M UNIVERSITY submitted to
THE CAUSES AND CONSEQUENCES OF SECONDARY PEST OUTBREAKS: DIRECT EFFECTS OF PESTICIDES ON PLANT DEFENSE AGAINST HERBIVORES
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
0228232
Grant No.
2012-67013-19346
Cumulative Award Amt.
$499,977.00
Proposal No.
2011-04659
Multistate No.
(N/A)
Project Start Date
Mar 1, 2012
Project End Date
Feb 29, 2016
Grant Year
2012
Program Code
[A1111]- Plant Health and Production and Plant Products: Insects and Nematodes
Project Director
Eubanks, M. D.
Recipient Organization
TEXAS A&M UNIVERSITY
750 AGRONOMY RD STE 2701
COLLEGE STATION,TX 77843-0001
Performing Department
Entomology
Non Technical Summary
The long-term goal of this research is to improve our understanding of the effects of agricultural chemicals on plants and plant-pest interactions to optimize sustainable pest management. Specifically, we propose to study signaling mechanisms involved in induced defenses in cotton (Gossypium hirsutum L.) and corn (Zea mays L.) to agriculturally important pests, spider mites, and the effects of neonicotinoid insecticides on these induced defenses. Our novel approach will improve our understanding of genetic interactions between plants and mites and the effect of neonicotinoid pesticides on these interactions, particularly on expression of defense genes. By quantifying the ecological impact of systemic pesticides on the abundance of secondary pests and their natural predators, we will improve our ability to predict secondary pest outbreaks following the use of neonicotinoid pesticides. The expected outcomes are: (1) increased awareness of secondary pest outbreaks following neonicotinoid applications, (2) more judicious use of neonicotinoid pesticides, currently the standard arthropod pest control in agro-ecosystems, and (3) implementation of sustainable management strategies for predicted pest outbreaks by growers using neonicotinoid pesticides. The adoption of these sustainable practices will have significant implications for pesticide resistance management and will reduce pesticide exposure to humans and the environment. Furthermore, this research will increase our fundamental understanding of signaling mechanisms involved in induced plant defenses against pests in agro-ecosystems.
Animal Health Component
50%
Research Effort Categories
Basic
50%
Applied
50%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2011510107034%
2031710113033%
2111510104033%
Knowledge Area
211 - Insects, Mites, and Other Arthropods Affecting Plants; 203 - Plant Biological Efficiency and Abiotic Stresses Affecting Plants; 201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1510 - Corn; 1710 - Upland cotton;

Field Of Science
1040 - Molecular biology; 1130 - Entomology and acarology; 1070 - Ecology;
Goals / Objectives
Neonicotinoid pesticides are one of the most widely used pesticides in the world. Applications of neonicotinoids, however, have been linked to outbreaks of non-susceptible, secondary pests, especially spider mites. Our preliminary data suggests that neonicotinoid pesticides suppress the expression of plant genes involved in induced defense against pests and that corresponding reductions in plant defense drives outbreaks of spider mites. Our goal is to quantify the effect of neonicotinoid pesticides on induced defense in cotton and corn and determine the consequences for spider mite populations and pest management. Our specific objectives are to 1) Determine changes in the transcriptomes of cotton and corn plants exposed to spider mites and treated with neonicotinoid pesticides; 2) Quantify the effect of neonicotinoid pesticides on the production of plant defense proteins and phytohormone levels in cotton and corn; 3) Quantify the direct and plant-mediated effect of neonicotinoid pesticides on spider mite fecundity; 4) Quantify the effect of neonicotinoid pesticides on the incidence and severity of spider mite outbreaks and abundance of natural enemies of spider mites on cotton and corn and 5) Evaluate the effect of neonicotinoid pesticides on miticide use and the economic cost of spider mite outbreaks on cotton and corn.
Project Methods
Our preliminary data show that spider mites on neonicotinoid pesticide-treated plants elicit weaker expression of a few selected genes involved in defense compared to untreated plants. Clearly, neonicotinoid pesticides change plant quality. These changes likely involve defense mechanisms or primary metabolism. To understand spider mite outbreaks after neonicotinoid pesticide applications, we will survey global transcriptome changes in cotton and corn using the Illumina sequencing technology, which is preferred to microarrays because it has the potential of surveying all transcripts from a tissue and it is not limited to the probes present in a microarray. Moreover, next generation sequencing technology is not subject to dynamic range limitation of microarrays. The combination of non-normalized samples and deep sequencing will help characterize the complete transcript profiles of plants in each treatment while surveying low-expression transcripts. Using full-length cDNAs will allow for monitoring differences of expression for splicing variants or very similar members of gene families. Results from the sequencing experiments will be compared with results from plant protein and phytohormone analysis to provide insight into what genes are associated with defensive proteins and phytohormones. Moreover, the information gained through transcriptome profiling will extend beyond the effects of neonicotinoid pesticides on plant defense. It will contribute to our general knowledge of the influence of systemic neonicotinoid pesticides on expression of genes involved in other plant functions such as metabolism. For example, if neonicotinoid pesticides affect photosynthetic rates and plant growth, this study will detect these changes. This will be the first study to quantify the effect of systemic pesticides on transcriptome profiles of plants. In addition, we will examine interactions between spider mite herbivory and neonicotinoid pesticides on levels of defense proteins and phytohormones that regulate plant defense pathways. We hypothesize that concentrations of defense proteins and phytohormones will decrease following neonicotinoid pesticide applications making plants more susceptible to spider mite herbivory. Testing this hypothesis would elucidate the mechanism underlying outbreaks of Tetranychidae on cotton and corn plants. In addition, field experiments and surveys of commercial cotton and corn production will determine the effect of neonicotinoid pesticides on spider mite abundance. Together, field experiments and surveys will establish a correlation between neonicotinoid pesticide use and spider mite abundance under field conditions and production management.

Progress 03/01/12 to 02/29/16

Outputs
Target Audience:Academic, government, and industry scientists, graduate and undergraduate students, and extension agents and agricultural consultants. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The postdoctoral research fellow and students were provided with training in RNA extraction, next generation sequencing, bioinformatics analysis, and qPCR. The also received mentoring on scientific presentations and writing. How have the results been disseminated to communities of interest?Publications in scientific journals, presentations at scientific meetings, and in presentations to extension agents and crop consultants. What do you plan to do during the next reporting period to accomplish the goals?Finish transcriptomic analysis and publish scientific manuscripts.

Impacts
What was accomplished under these goals? Insecticides can have a range of unintended effects on non-target organisms. In particular, neonicotinoids, a widely used class of insecticides, modify plant defense gene regulation. This likely increases host plant vulnerability to outbreaks of the non-susceptible two spotted spider mite, Tetranychus schoenei (Acari: Tetranychidae). Neonicotinoid treatments also potentially benefit plant development, which may further provision spider mite outbreaks. To determine the extent of these neonicotinoid mediated endogenous changes, we used RNA-Seq to develop a broader picture of the differential gene expression cascades that may increase plant quality and susceptibility for spider mite infestations. We exposed a commercial variety of corn, Zea mays L., to three treatments, neonicotinoid seed treatment (Thiamethoxam), a foliar treatment (Imidacloprid) or untreated with neonicotinoids. We extract total RNA from leaf tissue, four weeks after germination, for three replicates from each of the three treatments. These were sequenced on one lane of an Illumina HiSeq 2500 system, which produced 250 million, 100 bp SE, transcripts. We mapped the reads to the maize genome sequence v2 using the Tophat v2.0 program, assembled the transcripts (Cufflinks and Cuffmerge), and compared gene expression amongst our treatments using EdgeR. We identified several differentially expressed genes involved in plant response to biotic stress. We selected a subset of these, jasmonic acid synthesis, starch synthesis, and pathogen response genes, to verify through qRT-PCR. Overall, we found that both foliar and seed neonicotinoid treatments modify the expression of several genes involved in defensive and metabolic pathways.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Raupp, M.J, A. Szczepaniec, and S. Creary. 2015. Unravelling direct and indirect effects of�� insecticides on pollinators and natural enemies in managed landscapes. Proceedings of the Conference on Protecting Pollinators in Ornamental Landscapes, North Carolina, USA.��
  • Type: Other Status: Published Year Published: 2015 Citation: Szczepaniec, A., B. Fuller, and K.J. Tilmon. 2014. Miticide and insecticide control of spider mites in��soybeans. South Dakota State University. Extension Publication 03-7000-2015


Progress 03/01/13 to 02/28/14

Outputs
Target Audience: Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? The postdoctoral research fellow and students were provided with training in RNA extraction, next generation sequencing, bioinformatics analysis, and qPCR. The also received mentoring on scientific presentations and writing. How have the results been disseminated to communities of interest? Publications in scientific journals, scientific meetings, and in presentations to extension agents and crop consultants. What do you plan to do during the next reporting period to accomplish the goals? Finish transcriptomic and related studies and pubication of the final results of the project.

Impacts
What was accomplished under these goals? Insecticides can have a range of unintended effects on non-target organisms. In particular, neonicotinoids, a widely used class of insecticides, modify plant defense gene regulation. This likely increases host plant vulnerability to outbreaks of the non-susceptible two spotted spider mite, Tetranychus schoenei (Acari: Tetranychidae). Neonicotinoid treatments also potentially benefit plant development, which may further provision spider mite outbreaks. To determine the extent of these neonicotinoid mediated endogenous changes, we used RNA-Seq to develop a broader picture of the differential gene expression cascades that may increase plant quality and susceptibility for spider mite infestations. We exposed a commercial variety of corn, Zea mays L., to three treatments, neonicotinoid seed treatment (Thiamethoxam), a foliar treatment (Imidacloprid) or untreated with neonicotinoids. We extract total RNA from leaf tissue, four weeks after germination, for three replicates from each of the three treatments. These were sequenced on one lane of an Illumina HiSeq 2500 system, which produced 250 million, 100 bp SE, transcripts. We mapped the reads to the maize genome sequence v2 using the Tophat v2.0 program, assembled the transcripts (Cufflinks and Cuffmerge), and compared gene expression amongst our treatments using EdgeR. We identified several differentially expressed genes involved in plant response to biotic stress. We selected a subset of these, jasmonic acid synthesis, starch synthesis, and pathogen response genes, to verify through qRT-PCR. Overall, we found that both foliar and seed neonicotinoid treatments modify the expression of several genes involved in defensive and metabolic pathways.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Rice, K.B. and M.D. Eubanks. 2013. No enemies needed: Cotton aphids (Hemiptera: Aphididae) directly benefit from red imported fire ant (Hymenoptera: Formicidae) tending. Florida Entomologist 96(3):929-932.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Eubanks, M.D. and D.L. Finke. 2014. Interaction webs in agroecosytems: Beyond who eats whom. Current Opinion in Insect Science 2:1-6
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Kulmatiski, A., A. Anderson-Smith, K.H. Beard, S. Doucette-Riise, M. Mazzacavallo, N.E. Nolan, R.A Ramirez, and J.R. Stevens. Most soil trophic guilds increase plant growth: a meta-analytical review. Oikos 123:1409-1419.


Progress 03/01/12 to 02/28/13

Outputs
Target Audience: Agricultural scientists, extension personnel, and growers are the target audience of this project. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? This project provided training in experimental design, the execution of experiments, and data analysis for two undergraduates and two graduate students as well as the development of a research technician. A postdoctoral fellow will be added to the project soon. How have the results been disseminated to communities of interest? Publication in a scientific journal, presentations at national and regional scientific meetings, and presentations at extension meetings that include extension specialists and agents and growers in Texas, Utah, and South Dakota. What do you plan to do during the next reporting period to accomplish the goals? We are now starting the transcriptomic portion of the project and will begin to identify the changes in gene expression that are induced in plants by neonicotinoid insecticides.

Impacts
What was accomplished under these goals? We demonstrated that applications of neonicotinoid insecticides, one of the most important insecticide classes worldwide, suppress expression of important plant defense genes, alter levels of phytohormones involved in plant defense, and decrease plant resistance to unsusceptible herbivores, spider mites (Tetranychus urticae), in multiple, distantly related crop plants. Using cotton (Gossypium hirsutum), corn (Zea mays) and tomato (Solanum lycopersicum) plants, we show that transcription of phenylalanine amonia lyase, coenzyme A ligase, trypsin protease inhibitor and chitinase are suppressed and concentrations of the phytohormone OPDA and salicylic acid were altered by neonicotinoid insecticides. Consequently, the population growth of spider mites increased from 30% to over 100% on neonicotinoid-treated plants in the greenhouse and by nearly 200% in the field experiment. These results are important because applications of neonicotinoid insecticides have been associated with outbreaks of spider mites in several unrelated plant species and this is the first study to document insecticide-mediated disruption of plant defenses and link it to increased population growth of an unsusceptible herbivore. This study adds to growing evidence that agrochemicals can have unanticipated ecological effects and suggests that the direct effects of insecticides on plant defenses should be considered when the ecological costs of insecticides are evaluated.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Szczepaniec, A., M.J. Raupp, R.D. Parker, D. Kerns, and M.D. Eubanks. 2013. Neonicotinoid insecticides alter induced defenses and increase susceptibility to spider mites in distantly related crop plants. PLoS ONE 8(5): e62620.