Recipient Organization
TEXAS A&M UNIVERSITY
750 AGRONOMY RD STE 2701
COLLEGE STATION,TX 77843-0001
Performing Department
(N/A)
Non Technical Summary
Increased agricultural pest pressure and heightened demands for reducing reliance on chemical controls necessitates finding alternative sustainable pest management strategies. Naturally occurring soil-dwelling microorganisms can boost agricultural crop resistance to environmental stress, including insect pests, but harnessing microorganisms for improved crop production remains challenging. This project aims to characterize crop-protective microorganisms from different soil types in order to strengthen crop resistance. Specifically, the project will evaluate the role of microorganisms in corn and cotton resistance to a challenging insect pest, fall armyworm (Spodoptera frugiperda). Fall armyworm is currently devastating corn and cotton production across the globe. In the United States, fall armyworm persists as a significant agricultural pest with few tools to combat its spread, calling for innovative control approaches. Utilizing naturally occurring microorganisms to bolster corn and cotton resistance against fall armyworm infestations provides a novel and environmentally friendly strategy for management of this insect pest. To isolate and identify crop-protective microorganisms, soil from fields with different histories of plantings will be collected and DNA from microorganisms will be extracted in the laboratory. Extracted DNA will allow for the identification of microorganisms present in soil samples. The effect of different microorganisms on corn and cotton plants will be evaluated in a greenhouse experiment. Corn and cotton plants will be grown with different microorganisms and then challenged by fall armyworm feeding, assessing changes in crop resistance to insect pests that are dependent on different microorganisms. How fall armyworm feeding alters microorganisms will also be evaluated, determining if corn or cotton plants recruit specific microorganisms to enhance resistance. Ultimately, the goal of this project is to advance insect pest control through the use of naturally occurring microorganisms. Improving sustainable pest management strategies will increase agricultural crop production and reduce reliance on chemical insecticides to control insect pests. Long-term, this project will enhance safety of food and fiber supply across the United States and inspire future generations of agricultural scientists.
Animal Health Component
0%
Research Effort Categories
Basic
100%
Applied
0%
Developmental
0%
Goals / Objectives
The over-arching goal for this project is to determine how crop rotation influences corn and cotton microbiota and crop-pest interactions, ultimately leading to the identification of crop-protective microbes and enhancing sustainable insect pest management strategies. To accomplish this over-arching goal, I will pursue three specific research objectives:1) Characterize soil-dwelling microbiota from fields with distinct crop rotation legacies2) Quantify microbiota-mediated crop resistance to insect pests across different soil legacies3) Identify insect-driven changes in microbiota of plants grown in varied soil legacies
Project Methods
Objective 1 Methods: Characterize soil-dwelling microbiota from fields with distinct crop rotation legacies I will collect soil at the USDA-ARS Southern Plains Agricultural Research Center in College Station, TX, from fields with a legacy of consecutive corn or cotton plantings. Microbial DNA will be extracted from soils and sequenced to determine microbial community diversity. Soil collected from different fields will be used for experiments in Objectives 2 & 3. A) Soil collection: Soil will be collected from three distinct USDA fields: 1) consecutive corn field, 2) consecutive cotton field, and 3) crop rotation field. Each field will have a legacy of consecutive planting or crop rotation for at least 5 years prior to soil collection. From each field, the top 10-20 cm of soil will be removed at 10 different locations and sieved to remove rocks and other debris. From each of the 10 locations sampled per field, 250 mg of soil will be set aside for DNA extraction. The remaining soil from the 10 locations will be homogenized per field and stored in sterilized plastic buckets placed on ice during transportation to the greenhouse. In the greenhouse, I will generate two separate soil inoculants for use in Objectives 2 & 3. B) DNA extraction and sequencing: To characterize microbiota, microbial DNA will be isolated from soil samples. DNeasy PowerSoil Pro Kit (Qiagen) will be used to extract microbial DNA. Extracted DNA will be sequenced using MiSeq amplicon sequencing following previously described methods. Samples will be sequenced to characterize the bacterial V4-V5 region of the 16S ribosomal RNA region and the fungal internal transcribed spacer (ITS) region of the rRNA gene. After obtaining the sequencing data, the sequences will be processed using QIIME and further cleaned using MOTHUR following previously described methods to assign operational taxonomic units (OTUs) to bacterial and fungal sequences. Sequences will be normalized using rarefication (to 1,000 sequences) to estimate relative abundance. C) Statistical analyses: Microbial community richness, evenness, and diversity will be analyzed across fields. Dissimilarity across microbial communities will be assessed using non-metric multidimensional scaling (NMDS) ordinations of Bray-Curtis distances. Differences across fields in microbial community diversity will be analyzed using PERMANOVA.Objective 2 Methods: Quantify microbiota-mediated crop resistance to insect pests across different soil legacies A greenhouse experiment will be conducted with corn and cotton plants grown in different microbiota treatments generated by field-collected soil (Objective 1). I will assess plant resistance by measuring fall armyworm (S. frugiperda) mass gain and characterizing plant defensive chemicals. A) Microbiota inoculation: Corn (Z. mays cv. B73) and cotton (G. hirsutum cv. FM966) plants will be grown in the USDA-ARS greenhouse in a mixture of 95% (v/v) sterilized potting soil and 5% (v/v) microbiota inoculant from the field. Potting soil will be sterilized by autoclaving soil three times for 25 min, with 24 hours between sterilizations. Sterilized soil and microbiota inoculant will be mixed by hand in sterile bins for 5 mins prior to planting. Data loggers will also be placed in the soil of each plant to monitor soil temperatures and moisture. B) Insect pest bioassays: After 3 weeks of growth (3-4 true leaves), each plant in the greenhouse will be challenged by a leaf-feeing insect pest, second-instar fall armyworm larvae (purchased from Benzon). Each larva will be pre-weighed and 2 larvae will be caged on the two youngest, fully expanded true leaves of each plant. Larvae will be allowed to feed for 72 hr, then larvae will be removed from each plant and re-weighed to quantify mass gain. C) Crop defensive chemical extraction and quantification: Leaf tissue from each plant challenged by larvae will be flash frozen in liquid nitrogen. One leaf will be used to measure phytohormone concentrations using previously described methods. Briefly, phytohormones will be methylated, collected using vapor-phase extraction, and analyzed using a GC/CI-MS. The other leaf from each plant will be lyophilized and used for defensive chemistry analysis (e.g., cotton--terpenoid aldehydes; corn--benzoxazinoids) using previously described extraction methods and quantification on an HPLC. D) Statistical analyses: Larvae mass gain will be analyzed using a nested-ANOVA to account for non-independence of two larva feeding on the same plant. Defensive chemicals will be analyzed using one-way ANOVAs.Objective 3 Methods: Identify insect-driven changes in microbiota of plants grown in varied soil legacies Soil from all plants used in Objective 2 will be collected to determine how aboveground fall armyworm herbivory alters crop-associated microbiota. A) DNA extraction, sequencing, & statistical analyses: The same methods described above in Objective 1 will be used to extract, sequence, and analyze microbial DNA from each sample. Analyzes will reveal differences in microbial community richness, evenness, and diversity following aboveground fall armyworm herbivory.Efforts used to cause a change in knowledge of target audiencesTarget audiences that are the focus of effort for the duration of the project include 1) undergraduate students mentored in scientific research, and 2) corn and cotton growers. When mentoring undergraduate students, PD Morgan Thompson will strive to teach students new technical skills, advance their professional development, and improve their understanding of core scientific concepts related to the project. PD Thompson approaches work with students as a research collaboration. To empower students to collaborate, PD Thompson will continue to lead the weekly journal club she began with undergraduate mentees this past year. PD Thompson found that when students gained the confidence to express scientific ideas during journal club, students were more likely to speak up and provide unique insights on research in the laboratory. To reach corn and cotton growers, PD Thompson will organize and host 1-2 farms days throughout the duration of the project. Farm days will inform growers about the research findings, offering novel insect pest management strategies. Growers can also provide feedback and insights on the research project at farm days, which may shape the trajectory of the research. As growers are the ultimate practitioners of pest management in agricultural fields, their perspective is highly valuable.How outputs will be evaluated for impact on target audiencesTo evaluate research progress throughout the project, PD Thompson will meet with her mentors, Drs. Helms and Suh, on a weekly basis. Weekly meetings will ensure consistent progress is made on the proposed research, confirming the project is on track for successful completion. Meetings will facilitate experimental troubleshooting, manuscript preparation, and presentation of findings. To monitor undergraduate student mentorship, PD Thompson will share updates on undergraduate student progress in weekly meetings with Dr. Helms. Undergraduate students will be encouraged to publish research findings and present at conferences. To assess extension outputs, Dr. Suh will oversee organization of farm days and connect PD Thompson with local growers. Additionally, findings will be communicated through ResearchGate, LinkedIn, and Twitter.