Source: MICHIGAN STATE UNIV submitted to
DEVELOPING A SUSTAINABLE STEVIA INDUSTRY IN THE UNITED STATES.
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
OTHER GRANTS
Reporting Frequency
Annual
Accession No.
1013033
Grant No.
2017-51181-26828
Cumulative Award Amt.
$3,208,657.00
Proposal No.
2017-03233
Multistate No.
(N/A)
Project Start Date
Sep 1, 2017
Project End Date
Aug 31, 2023
Grant Year
2017
Program Code
[SCRI]- Specialty Crop Research Initiative
Project Director
Warner, R.
Recipient Organization
MICHIGAN STATE UNIV
(N/A)
EAST LANSING,MI 48824
Performing Department
HORTICULTURE
Non Technical Summary
Stevia rebaudiana (stevia) is a rapidly emerging new crop in the U.S., grown for the extraction of sweet-tasting steviol glycosides used as non-caloric sweeteners. Most stevia is currently produced in China, though production is increasing rapidly in the U.S. The combination of a favorable climate and existing infrastructure necessary for stevia production in the southeastern U.S. make this an ideal region for production of high quality stevia needed to meet the increasing demand for steviol glycosides. However, U.S. stevia growers confront multiple challenges, including a lack of varieties developed for U.S. conditions, a paucity of information on optimizing production practices, such as planting density, fertilization, weed and disease control, and a lack of information on costs of production, consumer preferences, and potential profitability. The specific objectives of this proposal, which were developed with stakeholders through an SCRI Planning Grant, are to: 1. Develop Best Management Practices to optimize stevia biomass and glycoside production in the southeastern U.S.; 2. Develop germplasm and genetic/genomic resources to enable selection and breeding of stevia cultivars adapted to U.S. growing conditions and producing consumer-desired steviol glycosides; 3. Develop consumer profile and market potential analyses for U.S. stevia products and determine consumer-preferred combinations of steviol glycosides.; 4. Develop cost of production estimates for stevia in different cropping systems; and 5. Conduct outreach programs for producers, extension service providers, and industry professionals. Completing these objectives will contribute substantially to filling the knowledge gaps, andfostering the development of new varieties to meet industry demand for "next-generation" sweeteners.
Animal Health Component
60%
Research Effort Categories
Basic
30%
Applied
60%
Developmental
10%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
1022239106015%
2012239108020%
2052239106015%
2122239116015%
6012239301015%
6032239301010%
5022239309010%
Knowledge Area
212 - Pathogens and Nematodes Affecting Plants; 601 - Economics of Agricultural Production and Farm Management; 102 - Soil, Plant, Water, Nutrient Relationships; 603 - Market Economics; 205 - Plant Management Systems; 201 - Plant Genome, Genetics, and Genetic Mechanisms; 502 - New and Improved Food Products;

Subject Of Investigation
2239 - Flavoring and beverage plants, general/other;

Field Of Science
1060 - Biology (whole systems); 3090 - Sensory science (human senses); 1160 - Pathology; 1080 - Genetics; 3010 - Economics;
Goals / Objectives
The long-term goals of this project are to promote development of a sustainable stevia industry in the United States by: 1) developing information and resources to help farmers successfully produce this emerging crop by optimizing agronomic practices for currently available varieties; 2) developing the genomic resources and understanding of consumer preferences necessary to breed high-yielding, disease-resistant stevia varieties producing consumer-desired steviol glycosides to help food and beverage manufacturers create the next generation of stevia-containing products; and 3) developing cost of production models for stevia in different production systems.In support of our long-term goals, the specific objectives for this proposal are: 1. Develop Best Management Practices to optimize stevia biomass and glycoside production in the southeastern U.S. 2. Develop germplasm and genetic/genomic resources to enable selection and breeding of stevia cultivars adapted to U.S. growing conditions and producing consumer-desired steviol glycosides. 3. Develop consumer profile and market potential analyses for US stevia products and determine consumer-preferred combinations of steviol glycosides. 4. Develop cost of production estimates for stevia in different cropping systems. 5. Conduct outreach programs for producers, extension service providers, and industry professionals.
Project Methods
In order to develop Best Management Practices, we will: 1)Conduct multi-state field trials to determine optimal transplanting date, plant density and overwintering survival strategies for stevia; 2)Optimize nutrient management strategies for organic and conventional cropping systems; 3)Identify herbicides that are both safe and effective for stevia production and assist in obtaining labels for these products for growers; and 4)Elucidate disease cycles of major diseases that occur on stevia in the southeastern U.S. and develop best management practices (conventional and organic) for these diseases. Toenable selection and breeding of stevia cultivars adapted to U.S. growing conditions and producing consumer-desired steviol glycosides, we will: 1)Collect and characterize a broad panel of stevia germplasm to assess variation for adaptation, growth and development, overwintering, disease resistance, biomass and steviol glycoside production in the southeastern U.S.; 2)Generate a genome sequence for stevia to elucidate control of steviol glycoside biosynthesis and facilitate molecular marker development; 3)Generate molecular markers and quantitative trait locifor traits of interest to facilitate breeding of superior cultivars for U.S. conditions and consumer-preferred steviol glycosides. To develop consumer profile and market potential analyses for US stevia products and determine consumer-preferred combinations of steviol glycosides we will: 1)Conduct sensory evaluation to determine consumer-preferred combinations of the steviol glycosides rebaudiosides A, D and M in carbonated beverage and ice cream applications; and 2)Conduct online and in-person surveys to understand consumer perceptions, attitudes and purchasing decisions for products containing steviol glycosides. In order to develop cost of production estimates for stevia in different cropping systems, we will: 1) Develop a capital budget and conduct financial analysis of farmer investment costs associated with stevia production; 2)Develop and analyze farm level cost of production numbers and conduct breakeven analysis associated with stevia production; and 3)Conduct financial analysis of selected crop rotations associated with stevia production and determine cost/benefit ratios for organic and conventional production systems. Additionally, we will conduct outreach programs for producers, extension service providers, and other industry professionals.

Progress 09/01/17 to 08/31/23

Outputs
Target Audience:Stevia growers, breeders, processors and consumers. The ag-chemical industry, as well as state and local extension agents Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Five graduate students were mentored in crop production, plant breeding and genetics, and food science. How have the results been disseminated to communities of interest?Results have been disseminated thorough scientific presentations and publications, outreach presentations, and through electronic communications with our stakeholder advisory board. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Using a biparental genetic mapping population evaluated at three sites over each of two years and a SNP-based genetic linkage map developed through this project, we identified quantitative trait loci (QTL) for production of numerous steviol glycosides (stevioside, Reb A, Reb B, Reb C, Reb D, Reb E, Reb N and total steviol glycosides), and several traits related to stevia leaf biomass production (branch number, secondary branch number, leaf length and width, plant height and width, stem caliper). Identified stevia germplasm with desirable steviol glycoside yied and profile. Employed stevia genotypes exhibiting contrasting node development rates to identified differentially expressed genes between fast- and slow-developing lines. Evaluated consumer acceptability of no added sugar pound cakes under blind and informed testing conditions. Utilized various blends of Reb A, D, and M to correlate descriptive human panel data with the electronic tongue data. We found significant correlations between the human panel and the electronic tongue responses of "bitter taste at 90 seconds," confirming that e-tongue can be used to estimate the bitterness of the stevia solutions. Applied different Reb A, D, and M blends in food matrices and characterized sensory attributes using a time-intensity study. Ice cream and carbonated beverage were formulated using the six blends of Rebs. The time-intensity study demonstrated that a blend of Reb A (major glycoside) and Reb D or Reb M (minor glycosides) could produce an optimized taste profile of stevia-sweetened product, but an optimized ratio depends on a food products matrix.

Publications

  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Williams, R., Moazzem M.D., Dees, J., Hyden, M., Traynor, M., Cho, S. 2023. Consumer acceptability of no added sugar pound cakes under blind and informed testing conditions. J. Food Sci. 88:4677-4692
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Warner RM, Abeli PJ, and Beaudry RM. 2023. Development of synthetic cultivars to improve production of desired steviol glycosides in stevia. Acta Horticulturae 1362:85
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 1. Biswas, B. 2023. Field study of Stevia (Stevia rebaudiana Bertoni) in Middle Georgia and Its Leaf Analysis for Various Glycosides Production. Poster presented at the American Society for Horticultural Science 2023 Annual Conference, in Orlando, Florida, July 31-August 4.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: 2. SAHA, A., A. Howlader, N. Mullen, A. Roberson, and B. Biswas 2023. In vitro Study of Selected Stevia Plant (Stevia rebaudiana Bertoni) Grown and Selected from the USDA Hardiness Zone 8A, Located at the CAFST, FVSU Research Plots. Presented at 2023 In Vitro Biology meeting, Norfolk, Virginia, June 10-14.


Progress 09/01/21 to 08/31/22

Outputs
Target Audience:Stevia growers, breeders, processors and consumers. The ag-chemical industry, as well as state and local extension agents. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?Results were disseminated primarily through conference presentations, journal articles, popular press, and online bulletins. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Conducted a consumer test to compare the physicochemical and sensory properties of pound cakes sweetened with Reb A, D, and M and sucrose as a control. The consumer panel evaluated the products twice. The panel evaluated the samples at the first visit with no sample information (blind condition). At the second visit (2 weeks after the first visit), the panel was instructed to read a sample information sheet (e.g., stevia, calories, etc.) and then evaluate the same samples (informed session). The objective was to investigate the effect of stevia knowledge on the acceptability of stevia-sweetened samples. Developed cola syrup for the time-intensity study of stevia-sweetened cola beverages (Reb A, D, and M). Worked with an international ingredient company to find a relationship between electronic tongue (e-tongue, Alpha MOS) and descriptive sensory panel data. Developed 7 different ratios of Reb A, D, and M and conducted e-tongue analysis in Cho's lab. The company used a highly trained descriptive panel (n=8) to analyze the same ratios of the blends. We will correlate these data in Spring 2023. Stevia genotypes varying in vegetative development rate were employed to identify differentially expressed genes between fast- and slow-developing lines. Completed yield trials for plants grown under various disease control application strategy plots.

Publications

  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Muenprasitivej N, Tao R, Nardone SJ, Cho S. The Effect of Steviol Glycosides on Sensory Properties and Acceptability of Ice Cream. Foods. 2022 11(12):1745. doi: 10.3390/foods11121745.
  • Type: Journal Articles Status: Accepted Year Published: 2022 Citation: Warner RM, Abeli PJ, and Beaudry RM. Development of synthetic cultivars to improve production of desired steviol glycosides in stevia. Acta Horticulturae.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: S. R. Mentreddy, DeAnthony Price, S. Kumar, Khadejah Scott, and Trang Pham. 2022. Specialty Crops Research at Alabama A&M University. Presentation at the Kuwait Institute for Scientific Research, Kuwait City, Kuwait, October 15, 2022.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Kaur P, Bahmani K, and Warner RM. Identification of QTL for leaf yield component traits in Stevia. Poster presented at the 31st International Horticultural Congress (Symposium 1 -International Symposium on Breeding and Effective Use of Biotechnology and Molecular Tools in Horticultural Crops) in Angers, France, August 14-20, 2022.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2022 Citation: Warner RM, Abeli PJ, and Beaudry RM. Development of synthetic cultivars to improve production of desired steviol glycosides in stevia. Poster presented at the 31st International Horticultural Congress (Symposium 1 -International Symposium on Breeding and Effective Use of Biotechnology and Molecular Tools in Horticultural Crops) in Angers, France August 14-20, 2022.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Kaur P, Bahmani K, and Warner RM. Identification of QTL for leaf yield component traits in Stevia. Oral presentation at the annual meeting of the American Society for Horticultural Science, Chicago, IL, July 30-August 3, 2022.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Biswas, B. 2022. Stevia Production Practices and Various Glycosides Found in Stevia Leaves Grown at the Fort Valley state University. Presented at 10th World Convention, Stevia Tasteful 2022,World Stevia Organization, Lisbon, Portugal.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Cho, S. Stevia, A Natural Non-Caloric Sweetener. Presented at 2022 KFN (The Korean Society of Food Science and Nutrition) International Symposium and Annual Meeting (October 21, 2022), Jeju Island, South Korea
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Cho, S. Consumer perception and response to next-generation stevia sweeteners (Rebaudioside D and M. STAR (Science-Telling About Revolution in Sensory Nudges) (October 20, 2022), International Virtual Conference hosted by the University of Arkansas Sensory Service Center with a collaboration with the University of Parma (Parma, Italy) and Sejong University (Seoul, South Korea)


Progress 09/01/20 to 08/31/21

Outputs
Target Audience:Stevia growers, breeders, processors and consumers. The ag-chemical industry, as well as state and local extension agents. Changes/Problems:No major changes, though a no-cost extension was pursued and granted as some research activities were slowed due to covid-19. What opportunities for training and professional development has the project provided?Seven graduate students completed their degree programs. How have the results been disseminated to communities of interest?Results were disseminated primarily through conference presentations, journal articles, popular press,and online bulletins. A hybrid format stakeholder meeting was planned and will take place in October 2021. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Characterized Septoria isolates sampled from symptomatic stevia plants in native stevia plants in Paraguay and performed a phylogenetic and phylogeographic analysis of Paraguayan isolates of S. steviae with other isolates using a multilocus DNA sequence analysis. Phylogentic trees indicate that worldwide populations of Septoria steviae are derived from populations in Paraguay. Preliminary studies indicate that the pathogen may be moving of debris associated with seed. Evaluate the efficacy of anaerobic soil disinfestation for the management of stevia diseases in NC, Mexico and Paraguay and found that anaerobic soil disinfestation may be effective for small farm and organic production. Evaluated the efficacy of organic fungicides applications on reducing disease intensity of stevia diseases. Generated data in support submission of fungicides effective in controlling soil-borne pathogens for label approval. Worked in coordination with IR-4 program towards registration of multiple herbicide registrations. Field experiments were conducted to determine the i) effects of three organic manures, chicken manure, cow manure, and vermicompost compared to no-manure control on stevia growth, biomass and steviol glycosides; ii) effects of 0, 100, 200, 300 Kg/ha of N, and no fertilizer control of N on the growth biomass, and steviol glycosides of stevia; iii) effects of 0, 100, 150, 200 Kg of P2O5 /ha and no-fertilizer control on growth, biomass, and steviol glycosides of stevia; and iv) effects of 0, 100, 150, 200 Kg K2O and no fertilizer control on the growth, biomass, and steviol glycosides of stevia. Identified QTL for steviol glycoside production, vegetative development rate and other leaf-yield component traits (leaf size, branch production) in stevia using two years of field phenotyping data for a stevia mapping population (MSU18-02) at two locations in MI. Performed consumer sensory analysis of ice creams sweetened with Reb A, D, and M and sucrose as a control. Found that the ice creams sweetened Reb D and M were significantly preferred by regular ice cream consumers than the Reb A ice cream, showing also significantly higher purchase intent. Performed E-tongue analysis of pure Reb A, D, and M products and also various commercial stevia sweeteners using a new protocol that we developed last year. These samples were well separated on a taste map, confirming that the new protocol worked well for this study. Continued the activities for introduction of Stevia as a crop and food additive to students ranging from 4th grade to college level with FVSU revolving Life on the Farmer tours. During these visits students could handle and taste stevia grown in experimental plots.

Publications

  • Type: Theses/Dissertations Status: Published Year Published: 2021 Citation: Bahmani, K. (2021). Genetic Analysis of Important Metabolites in Fennel and Stevia (Doctoral dissertation, Michigan State University).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Kaur P, Bahmani K and Warner RM. 2021. Identification of QTL regions for studying development rate in stevia. Poster presented at the Annual meeting of the American Society for Horticultural Science, Denver, CO, August 5-9.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Vallejo VA and Warner RM. Identifying quantitative trait loci for steviol glycoside production and agronomic performance in Stevia rebaudiana using transcriptome-derived SSRs. Industrial Crops and Products 161:113176
  • Type: Theses/Dissertations Status: Published Year Published: 2020 Citation: Tao, R. (2020). Discrimination and Sensory Characterization of Steviol Glycosides (Rebaudioside A, D, and M) by Consumers and Electronic Tongue. M.Sc. Thesis. Michigan State University.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Rogers, LW, Koehler, AM, and Shew, HD. 2020. Evaluation of foliar fungicides for management of Septoria leaf spot on stevia in North Carolina, 2019. Plant Disease Management Reports, 14(V143).
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Rogers, LW and Koehler, AM. 2021. Nondestructive Sampling to Monitor Macrophomina Phaseolina Root Colonization in Overwintering Stevia. Plant Health Progress 22:151-153.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Sanabria-Velazquez, A. D. and Shew, H. D. 2020. APS OIP Global Experience Award: Get Funded for Your Globally Collaborative Ideas. The American Phytopathological Society (APS).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Sanabria-Velazquez, A.D.; Enciso-Maldonado, G.A. and Shew H. D. 2020. Identificacion de areas prioritarias de investigacion de la estevia en Paraguay mediante el analisis multidimensional de preferencias. III Jornadas Internacionales de Estad�stica Apl�cada.
  • Type: Journal Articles Status: Submitted Year Published: 2021 Citation: Warner RM. Influence of environmental parameters, pinching, and ethephon application on growth and branching of potted stevia. HortScience
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Huber, B. M. and T. C. Wehner. 2021. Performance of 16 Stevia rebaudiana seed cultigens for glycosides and yield in North Carolina. Scientia Hort. 277.
  • Type: Journal Articles Status: Accepted Year Published: 2021 Citation: Sanabria-Velazquez, A.D.; Cubilla, A.; Flores-Giubi, M.E., Barua, J.E. Romero-Rodr�guez, MC, Thiessen, LD; and Shew, H.D. 2021.First Report of Charcoal Rot Caused by Macrophomina euphorbiicola in Stevia Production Fields in Paraguay. Plant Disease
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Sanabria-Velazquez, A.D. ; Barua-Chamorro, J.E.; Enciso-Maldonado, G.A.; Maidana-Ojeda, M.; and Shew H. D.2021. Plant Disease Management Workshop:Stevia, in the quest of sweetness& Plant diseases appear! The American Phytopathological Society (APS). (Abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Sanabria-Velazquez, A.D.; Enciso-Maldonado, G.A.; Maidana-Ojeda, M.; Diaz-Najera, J.F. and Shew H. D. 2021. Exploring alternative management of plant pathogens of stevia in North Carolina, Mexico, and Paraguay. The American Phytopathological Society (APS).


Progress 09/01/19 to 08/31/20

Outputs
Target Audience:Stevia growers, breeders, processors and consumers. The ag-chemical industry, as well as state and local extension agents. Changes/Problems:No major changes in approach, but COVID-19 has delayed or prevented some research and outreach activities from taking place, including field trials, fielddays, eye-tracking reserach data collection and in-person taste panels. Weanticipate completing the project objectives, but will likely need some period of no-cost extension to complete project activities. What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?Results were disseminated primarily through conference presentations and online bulletins. No in-person meetings were held this year due to COVID-19. What do you plan to do during the next reporting period to accomplish the goals?We will utilize the genotyping and phenotyping to develop a genetic linkage map for stevia and identify QTL for steviol glycoside production, plant development rate, and other biomass production associated traits. Several field trials that could not be performed in 2020 due to COVID-19 will performed in 2021. We will verify the draft cost of production numbers by meeting with additional farmers in North Carolina. We will develop a capital cost budget in the next reporting year.

Impacts
What was accomplished under these goals? Conducted fungicide trials for management of S. steviae on stevia in 2019 in Rocky Mount and Laurinburg, NC, and in Delaware for IR 4 labels. Screened M. phaseolina isolates (collected from stevia) for the presence of QoI fungicide resistance mutations within the cytb gene. Screened M. phaseolina isolates (collected from stevia) for the presence of QoI fungicide resistance mutations within the cytb gene. There are five herbicide projects currently moving through the IR-4 Project. Evaluated effects of organic manures, chicken manure, vermicompost, and composted cow manure relative to no manure control on biomass and steviol glycoside content. Phenotyped the MSU18-02 stevia population at two field sites in MI and one site in GA. Employed genotyping-by-sequencing to genotype the MSU18-02 population and parents for SNP identification for linkage map generation and QTL identification. Utilized RNAseq data comparing high- and low-reb D-producing stevia genotypes to identify differentially expressed genes and utilized Weight Gene Co-expression Network Analysis to identify gene modules associated with reb D content. Conducted online survey of consumer preferences driving purchasing decisions for ice cream and cola products containing different types of sweeteners. Performed a consume-based characterization study to describe taste attributes of different sweeteners. Developed a new protocol for E-tongue analysis using the most updated sensor array #6 (Alpha MOS). Using the new protocol, E-tongue successfully discriminated steviol glycosides and stevia leaf samples.

Publications

  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Tao,R. and Cho, S.. Consumer-based sensory characterization of steviol glycosides (rebaudioside A, D, and M). 2020. Foods, 9, 1026. doi: 10.3390/foods9081026
  • Type: Book Chapters Status: Published Year Published: 2020 Citation: " Koehler AM, Rogers LW, Shew HD. 2020. Characterization of the cytb gene structure in Macrophomina phaseolina to assess fungicide resistance risk. In: Deising HB; Fraaije B; Mehl A; Oerke EC; Sierotzki H; Stammler G (Eds), "Modern Fungicides and Antifungal Compounds", Vol. IX, pp. 63-68. Deutsche Phytomed. Gesellschaft, Braunschweig.
  • Type: Other Status: Published Year Published: 2020 Citation: Rogers, Layne & Koehler, Alyssa. (2020). Evaluation of foliar fungicides for management of Septoria leaf spot on stevia in Delaware, 2019. Plant Disease Management Reports. Biological and Cultural Tests.
  • Type: Other Status: Published Year Published: 2020 Citation: Rogers, Layne & Koehler, Alyssa & Shew, David. (2020). Evaluation of foliar fungicides for management of Septoria leaf spot on stevia in North Carolina, 2019. Plant Disease Management Reports. Biological and Cultural Tests.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Rogers L.W., Koehler A.M., and Shew H.D. 2020. Recovery of Macrophomina phaseolina from stevia roots following fumigation and foliar fungicide treatments. APS Potomac Division Meeting, virtual meeting.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Sanabria-Velazquez, A.D., H.D. Shew. 2020. The origin of the problem: Phylogeographic analysis of Septoria steviae, causal agent of Septoria leaf spot of stevia. In: APS Annual Meeting 2020, Denver, Colorado, USA
  • Type: Journal Articles Status: Under Review Year Published: 2020 Citation: Huber, B. and T. C. Wehner. 2020. Performance of 25 stevia (Stevia rebaudiana) cultigens for yield, glycosides and other plant traits in North Carolina. Agron. J.
  • Type: Journal Articles Status: Under Review Year Published: 2020 Citation: Huber, B. and T. C. Wehner. 2020. Heritability and genetic variance estimates for agronomic traits and glycoside yield in four stevia breeding populations. Crop Sci
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Bahmani K, Vallejo VA and Warner RM. Interrelationships among and between some phytochemical and morphological traits in stevia. Poster presented at American Society for Horticultural Science annual conference (virtual) August 10-13, 2020
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Bahmani K, Vallejo VA and Warner RM. Utilizing RNA seq in stevia to identify components of the genetic control of rebaudioside D biosynthesis. Poster presented at American Society for Horticultural Science annual conference (virtual) August 10-13, 2020
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Warner RM, Beaudry RM, and Vallejo, VA 2019. Utilizing selection, genetics and genomics to improve and understand synthesis of minor steviol glycosides. Paper presented at 9th World Stevia Organization Convention, Amsterdam, The Netherlands, Oct. 24-25.
  • Type: Journal Articles Status: Under Review Year Published: 2020 Citation: Vallejo VA and Warner RM. Identifying quantitative trait loci for steviol glycoside production and agronomic performance in Stevia rebaudiana using transcriptome-derived SSRs. Industrial Crops and Products


Progress 09/01/18 to 08/31/19

Outputs
Target Audience:Stevia growers, processors and consumers. The ag-chemical industry, as well as state and local extension agents. Changes/Problems:Dr. Sungeun Cho, Co-PI started a new position at Auburn University on September 1, 2019. However, she will continue to be a part of this project, and her portion of the award and E-Tongue (mostly purchased on this grant) will be transferred to Auburn University. Dr. Cho's master student, Ran Tao will be continuing her degree working on this project at MSU under Dr. Cho's guidance. What opportunities for training and professional development has the project provided?Several graduate students were mentored in areas of plant science and food science. These students also engaged in professional development through presentations at conferences, our stakeholder meeting, and field day workshops. How have the results been disseminated to communities of interest?Results have been deisseminated online through a stevia web portal (https://stevia.ces.ncsu.edu) and social media accounts. Additionally, a meeting with our stakeholder advisory board was held in East Lansing, MI in August 2019. This consistedof two days of presentations and discussions. Two field days were held, one at NCSU and one at FVSU, with 55 and 75 attendees, respectively,including stevia and ag-chemical industry, state and local extension, and growers. What do you plan to do during the next reporting period to accomplish the goals?Several multi-year trials will continue into the next reporting period. We will assemble and annotate the Stevia rebaudiana genome sequence, and phenotype the stevia mapping population for vegetative development rate, steviol glycoside accumulation and other important agronomic traits in a multi-state replicated trial. SNP markers will be generated for the mapping population to facilitate linkage map construction and QTL discovery. We will verify the draft cost of production numbers by meeting with additional farmers in North Carolina. We will continue to collect data on yields and price. The capital budget will estimate the total equipment investment farmers will need to make as part of the feasibility evaluation of stevia.

Impacts
What was accomplished under these goals? Obj. 1) Continued study on the effect of fungicides used for foliar diseases on overwintering of stevia and the diversity of root infecting fungi in stevia roots. Completed year two trial of fumigant effects on suppression of root-infecting pathogens and yield. Identified a new leaf disease of stevia caused by a Phoma spp. Completed an initial study on efficacy and crop safety data on a new fungicide for stem rot control. A field experiment was established to determine the effects of three levels of nitrogen on growth, biomass production, and steviol glycosides of stevia. Obj. 2) Evaluated steviol glycoside profile of stevia genotypes with contrasting glycoside profiles and utilized RNAseq to identify differentially expressed genes between three high- and three low-Reb D-producing genotypes. Generated 10X Genomics- and Oxford Nanopore-based genome DNA reads to facilitate assembly of a stevia genome sequence. Evaluated field performance of half-sib families and clones at Whiteville, Clinton and Oxford. NC. Traits were measured on the accessions at 3 locations during the growth season, and included plant growth habit, yield, and glycoside content. Data were used to rank the accessions, and to estimate the heritability of the traits. Obj. 3) Conducted an online survey to investigate how different label attributes would affect purchase behavior of stevia sweetened ice cream compared to sugar sweetened ice cream. Used electronic tongue to analyze multiple types of commercial high intensity sweeteners (aspartame, monk fruit, stevia, sucralose), pure rebaudiosides (Reb M, Reb D, and Reb A), and dried stevia leaf samples. Recruited two sets of 50 participants for Magnitude Estimation to develop iso-sweet concentrations of sucrose, a commercial blend of steviol glycosides, and Reb M. Obj. 4) interviewed Stevia farmers growing under a different production system than the 2018 farmer interviewed and used information to develop cost of production estimates. Obj. 5) NCSU established Facebook and Twitter feeds to disseminate information. A field day was held at NCSU with 55 attendees. Presented stevia cost of production information at the Fort Valley State University Stevia Field Day with approximately 75 attendees.

Publications

  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Koehler A.M. and Shew H.D. 2019.�Evaluation of soil fumigants for management of soilborne pathogens in North Carolina stevia fields, 2018. Plant Disease Management Reports. 13:N044.�
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Koehler A.M. and Shew H.D. 2019. Evaluation of Priaxor for Septoria leaf spot control on stevia in North Carolina, 2018. Plant Disease Management Reports.�Plant Disease Management Reports. 13:V018.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Koehler A.M., Larkin M.T., Rogers L.W., Carbone I., Cubeta M.A., Shew H.D. 2019.�Identification and characterization of�Septoria steviae�as the causal agent of Septoria leaf spot disease of stevia in North Carolina. Mycologia 111:456-465.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Koehler A.M. and Shew H.D. 2019.�Effects of fungicide applications on root-infecting microorganisms and overwintering survival of perennial stevia. Crop Protection. 120:13-20.
  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2019 Citation: Koehler A.M., Rogers L.W., and Shew H.D. 2019. Characterizing�Macrophomina phaseolina�CYTB�gene structure to assess resistance risk and document baseline fungicide sensitivities. Proceedings of�the 19th�International Reinhardsbrunn Symposium
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: V.C. Anche, S. Ekke; M. Pudota; M. Bomma; K. Bhat; A. Byars; S. Kumar; and S.R. Mentreddy. 2018. The Effects of Three Organic Manures on Growth, Biomass, Light Interception, Mineral Content, Antioxidants, and Steviol Glycosides of Stevia rebaudiana. Poster presentation at the 20th Biennial Symposium of the Association of Research Directors, Jacksonville, FL. March 29-April 3, 2019.


Progress 09/01/17 to 08/31/18

Outputs
Target Audience:Stevia growers, processors and consumers. The ag-chemical industry, as well as state and local extension agents. Changes/Problems:We are not able to find an optimal combination of Reb A, D, and M using an electronic tongue as planned in the proposal. This is due to difficulty in obtaining sufficient quanitities of pure Reb D and M to work with. We purchased analytical standards of Reb A and D from Sigma-Aldrich, but we could not source Reb M; we need about 5 grams of each of Reb A, D, and M to conduct Response Surface Methodology. Thus, we will need to change our objective from "finding an optimal combination of Reb A, D, and M using E-Tongue, descriptive sensory analysis and consumer testing" to "finding the best tasting glycoside profiles of stevia leaves using E-Tongue and descriptive sensory analysis." What opportunities for training and professional development has the project provided?Graduate students were trained in agronomy, phytochemistry, plant breeding, plant pathology and food science. Additional research training opportunities were provided to several undergraduate students. How have the results been disseminated to communities of interest?Results have been deisseminated online through a stevia web portal (https://stevia.ces.ncsu.edu) and social media accounts. Additionally, a meeting with our stakeholder advisory board was held in Raleigh, NC in August 2018. This included one day of presentations and discussions, followed by a field day to see ongoing experiments. The field day was open to the broader community, and over 70 people, including stevia and ag-chemical industry, state and local extension, growers, and people from 6 states, attended. What do you plan to do during the next reporting period to accomplish the goals?Field trials from 2018 will be replicated in 2019. We will assemble and annotate the Stevia rebaudiana genome sequence, and phenotype the stevia mapping population for vegetative development rate, steviol glycoside accumulation and other important agronomic traits in a multi-state replicated trial. SNP markers will be generated for the mapping population to facilitate linkage map construction and QTL discovery. The 24 stevia accessions obtained will be distributed as seed to the cooperators for use in 2019 multistate field evaluations. We will conduct consumer testing to find how label attributes (sweetener information, claims, price, etc.) on beverages and ice cream products will affect sensory perception. Dried stevia leaves [from breeding lines] will be analyzed using HPLC to create glycoside profiles. We will screen stevia leaf samples that show differences in the glycosides profiles and then E-tongue and the descriptive panel will analyze them. Verify the draft cost of production numbers by meeting with additional farmers in the South East. Since transplants make up the largest cost we will evaluate different plant populations as well as production systems. We will develop a capital cost budget in the next reporting year.

Impacts
What was accomplished under these goals? Obj. 1) Conducted field trials to assess: 1. the response of stevia to three organic manures i.e., cow manure, chicken manure, and vermicompost; 2. the efficacy of multiple soil fumigants for management of nematodes and soilborne pathogens of stevia; 3. efficacy of multiple fungicides in the management of Septoria leaf spot and enhanced overwintering of stevia; 4. efficacy, residue, and crop safety for pre-plant, post-transplant, and management of overwintering weeds. Obj. 2) A Stevia rebaudiana genetic mapping population was developed between parental selections displaying variation in vegetative development rate and steviol glycoside production . One of the parents of this population was chosen as the genotype for which a reference genome is currently under development. Additionally, 24 stevia accessions were collected. These accessions were evaluated for field performance and heritability of several growth and yield traits were determined. Obj. 3) Conducted an online survey to investigate how different label attributes would affect purchase behavior of stevia sweetened ice cream compared to sugar sweetened ice cream. Acquired and installed an Electronic Tongue in April, 2018. Analyzed various types of caloric (sugar, honey, maple syrup) and non-caloric sweeteners (aspartame, monk fruit, stevia, sucralose) with analytical standards of Rebaudioside A and D. Designed descriptive sensory analysis and developed training methods for stevia samples. Recruited descriptive sensory panel. Obj. 4) Personal interviews with farmers were conducted to collect cost of production data of North Carolina farms. Obj. 5) A web portal and social media accounts were developed to provide baseline stevia production information and keep stakeholders aware of progress with project objectives.

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Koehler A.M. and Shew H.D. 2018. Field efficacy and baseline sensitivities of fungicides used for the management of Septoria leaf spot of stevia. Crop Protection. 109:95-101.