Progress 01/01/24 to 12/31/24
Outputs
Target Audience:The target audience for this project includes scientists, students, farmers, industry, and members of the broader public. Our efforts to reach these groups during the previous reporting period included the following: To communicate our research with the scientific community, we presented findings from this research at conferences and seminars. In 2024, PI Helms presented this research in seminars at the University of Florida and Louisiana State University. PI Helms and two graduate students presented findings related to this research at the Entomological Society of America Southwestern Branch and Annual meetings, and International Congress of Entomology. To reach students, PI Helms taught an undergraduate course in insect ecology where she discussed this research. PI Helms also mentored research students, including 2 graduate students and 3 undergraduate students working on this project. PI Helms and two undergraduate students participated in a USDA-funded summer REEU to increase participation in entomology research. In spring 2024, PI Helms and her students presented research from this project at the Texas A&M University Darwin Day, which is a community-wide outreach event to increase science literacy for the public with approximately 1,500 attendees. Changes/Problems:During the previous reporting period, we encountered difficulties with technical aspects of the project and project personnel. First, we initially struggled with infecting rootworms and coordinating sample shipments of nematode-infected rootworm cadavers for pheromone extractions. We have worked through these issues, refined our methodologies, and should be able to complete the extractions with the additional time offered by an extension. This extension will also give us time to complete the pheromone-plant bioassays, which were delayed because of difficulties with the pheromone extractions. Second, we lost key personnel on the project when a graduate student unexpectedly left our program this spring. We had to recruit a new student to work on the project and should now be able to complete the unfinished assays with an extension. What opportunities for training and professional development has the project provided?PI Helms mentored 3 undergraduate and 2 graduate research students through this project. Students are gaining technical training in designing and conducting experiments and analyzing and interpreting data. They are also gaining professional development opportunities through participation in seminars and workshops in teaching/mentoring and statistical methods. One undergraduate student will be enrolling in a master's program this fall. Co-PI Kaplan is mentoring a research scientist to work on this project, and he is being trained in a variety of scientific techniques, including pheromone extractions and metabolomics, and plans to start a graduate program this coming year. How have the results been disseminated to communities of interest?PI Helms presented research findings from this project in a symposium on advances in entomopathogenic nematode research at the International Congress of Entomology. Two graduate students also presented findings from this project at the Entomological Society of America regional and national conferences. Results were presented in a community outreach event. What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period, we will continue working toward completing our research objectives. We will communicate our research findings to the scientific community, farmers, and the broader public through conference presentations, drafting manuscripts, and participating in Extension and outreach events.
Impacts
What was accomplished under these goals?
Impact The major goal of this project is to expand our knowledge about the ecological interactions underlying biological control with beneficial entomopathogenic nematodes (EPNs) to 1) maximize its effectiveness for controlling pest insects and 2) improve public perception and adoption rates. During the second year of this project, we recruited and trained additional personnel and conducted experiments for the proposed objectives. The findings from this research are increasing our knowledge about EPN ecology and biological control, which will greatly advance the fields of nematology, chemical ecology, and plant biology. Our findings have already led to greater adoption of beneficial nematodes for biological control of pest insects, including vegetable growers in Texas and New York and many home gardeners, and we expect these rates to further increase in the future. Overall, this research is contributing to more sustainable and profitable agriculture through reduced pesticide inputs, reduced pest damage, and increased yields for vegetable growers and home gardeners. Objective 1. Characterize the chemical cues produced by different EPN species with different species of insect hosts. Major activities completed/experiments conducted: During this reporting period, we made substantial progress characterizing the pheromones produced by three different EPN species (Heterorhabditis bacteriophora, Steinernema riobrave, and Steinernema carpocapsae) infecting two insect host species, the striped cucumber beetle (Acalymma vittatum) and the spotted cucumber beetle (Diabrotica undecimpunctata howardi). Data collected: We are making progress on identifying and quantifying pheromone blend components for these three EPN species in two insect host species (A. vittatum and D. undecimpunctata howardi). Summary statistics and discussion of results: EPN-infected cadavers contain pheromone compounds and blends are characteristic of each species. Key outcomes or other accomplishments realized: We confirmed that we can extract and analyze the non-volatile EPN pheromone compounds from different species of infected insect cadavers. We are now working to finalize these analyses and will use the pheromone extracts for future experiments. Objective 2. Quantify changes in plant defenses following exposure to EPN-produced cues. Major activities completed/experiments conducted: During this reporting period, we conducted experiments evaluating how plant treatment with EPNs influences defense-related volatile compounds and phytohormones in cucumber plants. Cucumber plants were exposed to different EPN species (Heterorhabditis bacteriophora, Steinernema riobrave, and Steinernema carpocapsae) or controls and their volatile emissions were collected and analyzed before and during herbivory by adult striped cucumber beetles (A. vittatum). Levels of the defense hormones jasmonic acid and salicylic acid were also quantified from foliar tissue before and after beetle herbivory. Experiments were conducted to determine the influence of EPN exposure on squash plant defenses. We exposed squash plants to (Heterorhabditis bacteriophora) either alone or in combination with arbuscular mycorrhizal fungi (AMF) and evaluated plant defense volatile emissions before and during beetle herbivory. Data collected: We identified and quantified volatile compounds and defense hormones produced by cucumber and squash plants treated with EPNs both with and without cucumber beetle herbivory or with arbuscular mycorrhizal fungi (AMF). Summary statistics and discussion of results: Treating squash plants with EPNs (H. bacteriophora) did not directly alter their volatile emissions. Plant colonization with AMF had a small effect on volatile emissions. Herbivory by adult cucumber beetles on squash plants induced higher production of volatiles, although prior plant exposure to EPNs or AMF did not affect the herbivore-induced plant volatiles. For cucumber plants, we found no evidence that EPN treatment alone alters plant volatile emissions. Cucumber plants exposed to EPNs emitted similar volatile profiles and abundances compared to unexposed control plants. Adult cucumber beetle herbivory induced higher production of volatiles, regardless of EPN exposure. EPN exposure increased levels of salicylic acid and decreased jasmonic acid in cucumber plants. Key outcomes or other accomplishments realized: We made progress characterizing the influence of different EPN species on plant defenses expressed systemically in leaves. We are now examining the effects of different EPN species on plant volatile emissions from leaves and roots and quantifying levels of additional plant defense compounds. Objective 3. Quantify effects of plant exposure to EPN cues on plant resistance to pests and plant recruitment of beneficial organisms. Major activities completed/experiments conducted: We conducted bioassays with squash plants exposed to cues from EPNs (H. bacteriophora) alone or in combination with AMF and quantified plant resistance adult cucumber beetles and melon aphids. We also conducted feeding bioassays with cucumber plants exposed to EPNs (H. bacteriophora, S. carpocapsae, S. riobrave) and quantified plant resistance to adult cucumber beetles. We additionally evaluated adult cucumber beetle preference for cucumber plants exposed to EPNs or controls. Data collected: We quantified leaf area removed and beetle settling behavior on plants exposed to EPN cues or controls. We quantified numbers of aphids on treated and control plants. Summary statistics and discussion of results: We found that AMF colonization increases aphid performance, regardless of EPN treatment, while EPN exposure alone, reduced aphid performance. EPN or AMF exposures did not affect cucumber beetle feeding on squash. We found that adult cucumber beetles consumed more leaf tissue on cucumber plants exposed to S. riobrave IJs than control plants, suggesting that plant exposure to this EPN species increased their susceptibility to herbivore feeding damage. We also found that adult beetles preferred to settle on the S. riobrave-exposed cucumber plants compared to controls. In contrast, we found that adult beetles consumed less leaf tissue on cucumber plants exposed to H. bacteriophora IJs than control plants, suggesting that plant exposure to this EPN species increased their resistance to herbivore feeding damage. However, beetles preferred to settle on H. bacteriophora-exposed cucumber plants compared to controls. We found that adult cucumber beetles consumed similar amounts of leaf tissue on squash plants exposed to S. carpocapsae EPNs and controls. Overall, these findings highlight that EPNs can have species-specific effects on plant resistance to herbivores. Key outcomes or other accomplishments realized: We identified species-specific effects of EPNs on plant resistance to herbivores. Our results suggest that different plant species may have different responses to EPNs and different species of EPNs can have contrasting effects on plant resistance to a foliar-feeding herbivore. Future experiments will build on these findings to further examine the mechanisms underlying these differences in plant responses to EPN cues.
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Fifield, Adriane. Suh, Charles., Helms, Anjel M. Entomopathogenic Nematodes Influence on Cucumis sativus Volatile Organic Compounds and Adult Acalymma vittatum Feeding. Entomological Society of America Southwestern Branch Meeting. March 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Peterson, Skai and Helms, Anjel M. The Effects of Arbuscular Mycorrhizal Fungal Colonization and Entomopathogenic Nematodes on Volatile Organic Compound-Mediated Systemic Defenses of Squash Plants (Cucurbita pepo). Texas A&M University College of Agriculture and Life Sciences Graduate Research Symposium. May 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Helms, Anjel M. Chemical cues from entomopathogenic nematodes influence plant-insect interactions and enhance biological control. International Congress of Entomology. August 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Fifield, Adriane and Helms, Anjel M. Rooted defenders: The role of entomopathogenic nematodes in enhancing systemic cucumber plant resistance to adult striped cucumber beetles. Entomological Society of America Annual Meeting. November 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Peterson, Skai and Helms, Anjel M. Effects of Arbuscular Mycorrhizal Fungi Colonization and Entomopathogenic Nematode Exposure on Plant Systemic Defenses in Cucurbita pepo. Entomological Society of America Annual Meeting. November 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Helms, Anjel M. 2024. Chemical Cues Linked to Risk: How Organisms Navigate Conflict in a Chemical World. Louisiana State University Department of Entomology Seminar. December 2024
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Fifield, Adriane. Suh, Charles., Helms, Anjel M. Investigating How Entomopathogenic Nematode Chemical Cues Mediate Cotton Plant Defense. Cotton Beltwide Conference. January 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Helms, Anjel M. 2024. Chemical Cues Linked to Risk: How Organisms Navigate Conflict in a Chemical World. University of Florida Department of Entomology and Nematology Seminar. January 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Monahan, Scott, Grunseich, John M., Helms, Anjel M. Herbivore-Induced Plant Volatiles Mediate Belowground Tritrophic Interactions. San Diego State University Student Symposium. March 2024.
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Progress 01/01/23 to 12/31/23
Outputs
Target Audience:The target audience for this project includes scientists, students, farmers, industry, and members of the broader public. Our efforts to reach these groups during the previous reporting period included the following: To communicate our research with the scientific community, we presented findings from this research at conferences and seminars. In 2023, PI Helms presented this research in seminars at the University of Idaho and Penn State University. PI Helms and two graduate students presented findings related to this research at the Entomological Society of America annual meeting. To reach students, PI Helms taught an undergraduate course in insect ecology and a graduate course in chemical ecology where she discussed this research. PI Helms also mentored graduate and undergraduate research students, including 2 graduate students and 1 undergraduate student working on this project. During the summer of 2023, Co-PI Shapiro-Ilan hosted two student interns from Fort Valley State University in the Louis Stokes Alliances for Minority Participation (LSAMP). Changes/Problems:During the previous reporting period, we encountered some difficulties preparing infected larvae for EPN pheromone extractions that resulted in some delays for the project. We eventually identified a solution to this challenge and have made good progress with the analyses for objective 1. What opportunities for training and professional development has the project provided?PI Helms mentored 1 undergraduate and 2 graduate research students through this project. Students are gaining technical training in designing and conducting experiments and analyzing and interpreting data. They are also gaining professional development opportunities through participation in seminars and workshops in teaching/mentoring and statistical methods. Co-PI Kaplan is mentoring a research scientist to work on this project and he is being trained in a variety of scientific techniques, including pheromone extractions and metabolomics. How have the results been disseminated to communities of interest?PI Helms presented information on biocontrol and research findings from this project in a symposium on advances in biological control applications at the Entomological Society of America meeting. Two graduate students also presented findings from this project at this ESA conference. What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period, we will continue working toward completing our research objectives. We will communicate our research findings to the scientific community, farmers, and the broader public through conference presentations, drafting manuscripts, and participating in Extension and outreach events.
Impacts
What was accomplished under these goals?
Impact: The major goal of this project is to expand our knowledge about the ecological interactions underlying biological control with beneficial entomopathogenic nematodes (EPNs) to 1) maximize its effectiveness for controlling pest insects and 2) improve public perception and adoption rates. During the second year of this project, we recruited and trained additional personnel and conducted experiments for the proposed objectives. The findings from this research are increasing our knowledge about EPN ecology and biological control, which will greatly advance the fields of nematology, chemical ecology, and plant biology. Our findings have already led to greater adoption of beneficial nematodes for biological control of pest insects, including vegetable growers in Texas and New York and many home gardeners, and we expect these rates to further increase in the future. Overall, this research is contributing to more sustainable and profitable agriculture through reduced pesticide inputs, reduced pest damage, and increased yields for vegetable growers and home gardeners. Objective 1. Characterize the chemical cues produced by different EPN species with different species of insect hosts. Major activities completed/experiments conducted: During this reporting period, we characterized the pheromones produced by three different EPN species (Heterorhabditis bacteriophora, Steinernema riobrave, and Steinernema carpocapsae) infecting the insect host Galleria mellonella. We have also made substantial progress characterizing the pheromones produced by these three EPN species infecting two additional insect host species, the striped cucumber beetle (Acalymma vittatum) and the spotted cucumber beetle (Diabrotica undecimpunctata howardi). Data collected: We identified and quantified pheromone blend components for three different EPN species (H. bacteriophora, S. riobrave, and S. carpocapsae) infecting the insect host G. mellonella. We have begun identifying and quantifying pheromone blend components for these three EPN species in two additional insect host species (A. vittatum and D. undecimpunctata howardi). Summary statistics and discussion of results: EPN-infected cadavers contain pheromone compounds. Key outcomes or other accomplishments realized: We confirmed that we can extract and analyze the non-volatile EPN pheromone compounds from different species of infected insect cadavers. We are now working to finalize these analyses and will use the pheromone extracts for future experiments. Objective 2. Quantify changes in plant defenses following exposure to EPN-produced cues. Major activities completed/experiments conducted: During this reporting period, we conducted experiments evaluating how plant treatment with EPNs influences the production of volatile plant defense compounds from cucumber and squash plants. For these experiments, we exposed squash plants to EPNs (H. bacteriophora) or controls and analyzed emissions of plant volatile compounds. We exposed cucumber plants to EPNs (S. riobrave) or controls and characterized their volatile emissions. We also challenged squash and cucumber plants that had been exposed to EPNs with herbivory by striped cucumber beetle (A. vittatum) adults to determine how prior EPN exposure affects the herbivore-induced plant volatile emissions. Data collected: We identified and quantified volatile compounds emitted by cucumber and squash plants treated with EPNs both with and without cucumber beetle herbivory. Summary statistics and discussion of results: We found that treating squash plants with EPNs (H. bacteriophora) did not directly alter their volatile emissions. Herbivory by adult cucumber beetles on squash plants induced higher production of volatiles, including the compound (E)-beta-Ocimene. However, prior plant exposure to EPNs did not affect the herbivore-induced plant volatiles. For cucumber plants, we also found no evidence that EPN treatment alone alters plant volatile emissions. Cucumber plants exposed to EPNs (S. riobrave) emitted similar volatile profiles and abundances compared to unexposed control plants. Adult cucumber beetle herbivory induced higher production of volatiles, regardless of EPN exposure. These findings suggest that the production of volatile plant defense compounds is not impaired by plant exposure to EPNs. Key outcomes or other accomplishments realized: We made progress characterizing the influence of different EPN species on plant defenses expressed systemically in leaves. We are now examining the effects of different EPN species on plant volatile emissions from leaves and roots and quantifying levels of additional plant defense compounds. Objective 3. Quantify effects of plant exposure to EPN cues on plant resistance to pests and plant recruitment of beneficial organisms. Major activities completed/experiments conducted: We conducted bioassays with squash plants exposed to cues from EPNs (H. bacteriophora) or controls and quantified plant resistance adult cucumber beetles. We also conducted feeding bioassays with cucumber plants exposed to EPNs (H. bacteriophora and S. riobrave) or controls and quantified plant resistance to adult cucumber beetles. We additionally evaluated adult cucumber beetle preference for cucumber plants exposed to S. riobrave or controls. Data collected: We quantified leaf area removed and beetle settling behavior on plants exposed to EPN cues. Summary statistics and discussion of results: We found that adult cucumber beetles consumed more leaf tissue on cucumber plants exposed to S. riobrave IJs than control plants, suggesting that plant exposure to this EPN species increased their susceptibility to herbivore feeding damage. We also found that adult beetles preferred to settle on the EPN-exposed cucumber plants compared to controls. In contrast, we found that adult beetles consumed less leaf tissue on cucumber plants exposed to H. bacteriophora IJs than control plants, suggesting that plant exposure to this EPN species increased their resistance to herbivore feeding damage. We found that adult cucumber beetles consumed similar amounts of leaf tissue on squash plants exposed to H. bacteriophora EPNs. Overall, these findings highlight that EPNs can have species-specific effects on plant resistance to herbivores. Key outcomes or other accomplishments realized: We identified species-specific effects of EPNs on plant resistance to herbivores. Our results suggest that different plant species may have different responses to EPNs and different species of EPNs can have contrasting effects on plant resistance to a foliar-feeding herbivore. Future experiments will build on these findings to further examine the mechanisms underlying these differences in plant responses to EPN cues.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Peterson, Skai, and Anjel M. Helms. 2023. Effects of Entomopathogenic Nematode Inoculations on Herbivory-Induced Plant Volatile Production. Entomological Society of America Annual Meeting. November 2023
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Fifield, Adriane, Charles Suh, and Anjel M. Helms. 2023. Investigating How Chemical Cues from
Entomopathogenic Nematodes Modulate Cotton Plant Defenses. Entomological Society of America Annual Meeting. November 2023
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Helms, Anjel M. 2023. Out of the Lab and into the Field: Can Indirect Plant Defense Play a Meaningful Role in Insect Pest Management?. Entomological Society of America Annual Meeting. November 2023
- Type:
Other
Status:
Published
Year Published:
2023
Citation:
Helms, Anjel M. 2023. Chemical Cues Linked to Risk: How Organisms Navigate Conflict in a Chemical World. University of Idaho Department of Entomology, Plant Pathology and Nematology Seminar. February 2023
- Type:
Other
Status:
Published
Year Published:
2023
Citation:
Helms, Anjel M. 2023. Chemical Cues Linked to Risk: How Organisms Navigate Conflict in a Chemical World. The Pennsylvania State University Department of Entomology Seminar. March 2023
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Progress 01/01/22 to 12/31/22
Outputs
Target Audience:The target audience for this project includes scientists, students, farmers, industry, and members of the broader public. Our efforts to reach these groups during the previous reporting period included the following: To communicate our research with the scientific community, we published findings from this project in a peer-reviewed scientific journal article (Grunseich et al.). In 2022, PI Helms also communicated findings from this research through invited seminars at Kansas State University, University of Nebraska Lincoln, and University of Houston. To reach students, PI Helms taught an undergraduate course in insect ecology at Texas A&M University where she discussed this research. PI Helms also mentored graduate and undergraduate research students, including 3 graduate students working on this project and a student from an underrepresented group in STEM recruited through an REU at Texas A&M. PI Helms presented information on biocontrol and relevant research findings to Extension faculty in Texas and New York State, including presenting on a local Extension radio program "Garden Success". She also communicated these findings to the broader public during the 2022 Ronin Spring Farm Day event through an exhibit on entomopathogenic nematode biocontrol. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?PI Helms mentored 3 graduate and 2 undergraduate research students through this project. Two graduate students (1 Ph.D. and 1 M.S.) were recruited to work on this project starting in fall and summer 2022, respectively. Students are gaining technical training in designing and conducting experiments and analyzing and interpreting data. They are also gaining professional development opportunities through participation in seminars and workshops in teaching/mentoring and statistical methods. Co-PI Kaplan recruited a research scientist to work on this project and he is being trained in a variety of scientific techniques, including pheromone extractions and metabolomics. How have the results been disseminated to communities of interest?PI Helms presented information on biocontrol and relevant research findings to Extension faculty in Texas and New York State and members of the public, including presenting on a local Extension radio program "Garden Success". This program aired on the local public radio station, KAMU, and is available online and through various podcast platforms. PI Helms and her students also communicated findings related to this project to the broader public during the 2022 Ronin Spring Farm Day event through an exhibit on entomopathogenic nematode biocontrol. This was done in conjunction with an on-farm demonstration about beneficial nematode applications. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Impact The major goal of this project is to expand our knowledge about the ecological interactions underlying biological control with beneficial entomopathogenic nematodes (EPNs) to 1) maximize its effectiveness for controlling pest insects and 2) improve public perception and adoption rates. During the first year of this project, we recruited and trained key personnel and began conducting experiments for the proposed objectives. We also engaged in scientific outreach to communicate our findings with growers, Extension agents, and the broader public. The findings from this research will increase our knowledge about EPN ecology and biological control, which will greatly advance the fields of nematology, chemical ecology, and plant biology. This research will also contribute to improving sustainable pest management by identifying new approaches for harnessing ecological interactions in agricultural systems. Objective 1. Characterize the chemical cues produced by different EPN species with different species of insect hosts. Major activities completed/experiments conducted: We have characterized the volatile compounds produced by three different EPN species (Heterorhabditis bacteriophora, Steinernema riobrave, and Steinernema carpocapsae) with two different species of insect hosts (Galleria mellonella and Acalymma vittatum). We also began extracting the non-volatile pheromones produced by these three EPN species with the two different species of insect hosts. Data collected: We identified volatile and pheromone compounds and measured their relative abundances. Summary statistics and discussion of results: We found that the volatile blends produced by insect cadavers infected with each of the three different EPN species were unique. Cadavers infected with either of the two Steinernema species had more similar blends to each other than those of dead insect cadavers without nematodes, or those infected with Heterorhabditis. Our findings revealed that volatile blends from nematode-infected insect cadavers are indeed characteristic and are species specific. We also confirmed that we are able to extract and analyze the non-volatile nematode pheromones from infected insect cadavers. We are now working to finalize these analyses and will use the extracts for future experiments. Objective 2. Quantify changes in plant defenses following exposure to EPN-produced cues. Nothing to report Objective 3. Quantify effects of plant exposure to EPN cues on plant resistance to pests and plant recruitment of beneficial organisms. Major activities completed/experiments conducted: We have conducted bioassays with squash plants exposed to cues from EPNs (live or heat-killed IJs or water controls) and quantified plant resistance and tolerance to melon aphids. Data collected: We quantified aphid population growth on plants exposed to EPN cues and measured plant growth parameters. Summary statistics and discussion of results: We found that plants exposed to Heterorhabditis bacteriophora were more resistant to melon aphids, while plant exposure to Steinernema riobrave had no effect on plant resistance. No differences in plant growth or reproductive traits were observed.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Grunseich, J.M., Aguirre, N.M., Thompson, M.N., Ali, J.G., Helms A.M. (2021) Olfactory cues from entomopathogenic nematodes vary across species with different hunting strategies, triggering different behavioral responses in prey and competitors. Journal of Chemical Ecology. 47, 822833 https://doi.org/10.1007/s10886-021-01304-8
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