Progress 06/01/15 to 01/31/17
Outputs
Target Audience:Flowerbulb Brokers ex. Planthaven, Inc. Santa Barbara, CA Flowerbulb Distributors ex. Gardenstate Bulb, Inc. Vineland, NJ Fertilizer developer involved in phytoremediation, such as Xplosafe Changes/Problems:We askedfor extension of grant until June 2017 so we can complete one more season.The most significant change was trying to isolate and sequence the CaYMV genome. This was problematicbecause we obtained sequence data that suggested there are more than one virus species. We reported anddeveloped diagnostic primers for the one species and just this week identified the second species. Both are badnaviruses that exist in canna plants. However through the summer we revised our PCR diagnostic test to detectthe genome we isolated but could not make sense of the remaining PCR data suggesting another species mightexist. We failed to isolate this other species until November 2016 but have succeeded in January 2017. This slowed down our ability to report newfindings to the scientific community and to consumers/purchasing agents. The cost of tissue culture production of cannas along with its low success in producing the quality of rhizomes that we need, we decided to shelve this approach and focus on optimizing other advances that would benefit production and increase product. In particular we spent three months working out the best conditions for using immunostrips and the cost for immunostrips to see if we could migrate from PCR based diagnosis to this alternative approach. This turns out to be quite promising. We launched a set of experiments in the summer of 2016 to learn if CaYSV, BYMV, CaYMV can be transmitted from plant-to-plant via contaminated soil, by rhizomes touching each other, or by insects. This became complicated because the diagnostic PCR test for CaYMV did not match the genome sequence that we isolated. Since we managed to clone a second virus in January, 2017 we now must revisit our data and see if we can make better conclusions. The work was not delayed but the decision making based on the work was slowed. What opportunities for training and professional development has the project provided?This project was instrumental in launching Austin Gimondo to graduate studies in the field of horticulture and greenhouse management. This program helped to provide MSc training for Dulanjani Wijasekara. Hannah Jones is an undergraduate pre med student and training in virology has helped her to prepare for MCATS. Funding also helped to pay temporary salaries for at least 1 worker at the Horn Canna Farms. VF Canna LLC is housed on campus at Oklahoma State University. This SBIR subcontracted to Dr. Verchot, who is also the leader for VF Canna LLC and was the basis for the greenhouse program for screening cannas is essentially a production greenhouse. We have databases that track every individual plant that goes through the program, disease ratings based on visual observation, and diagnostic tests. Students who were employed were trained in greenhouse management, disease management and advanced database for greenhouse production. Graduate students at Oklahoma State University also visited Horn Canna Farms to also assist in training on how to identify disease and segregate healthy stocks. One-on-one students work with farm workers by walking the fields, answering questions, and sharing knowledge. Students learn about farm production, concerns, disease management, while sharing their own knowledge from the greenhouse experience. Hannah Jones provided Horn Canna Farms with a report on how to successfully use immunostrips for virus detection in field grown cannas. How have the results been disseminated to communities of interest?2015 Verchot provided information table for Botanical Garden. Dustin Snow has attended trade shows, There is one published extension report, one scientific report, and another scientific report in preparation. Jan 20, 2017 updated VF Canna LLC website based on conversations with LARTA to better represent the contributions of this company to the industry and advertise opportunities to work with other companies. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
We developed and published a multiplex RT-PCR assay to detect three viruses in cannas (Chauhan et al 2015). For in house work, we modified this method by substituting the species specific primers with genus level primers to maximize the number of viruses we could detect in a single assay. This allows us to detect five virus genomes, at pictogram quantities, in total nucleic acids isolated from canna leaves. We reported first full genome sequence for CaYMV, which we call CaYMV-0028, and developed new diagnostics that are improved over prior methods. At the same time we were concerned that there might be another virus present in cannas related to CaYMV-0028. The reason was that when we used primers that can detect any badnavirus, and then used primers detecting only CaYMV-0028 we would get differential results. We also identified an alternative host for canna yellow streak virus and in our experiments testing mechanical inoculation, we had evidence to suggest another virus may be present. In December we carried out RNA sequencing project and recovered a more complete transcript. Using this information, we are in the process of cloning the entire length of a second virus species related to CaYMV-0028. This will further inform our multiplex PCR tests. As of now, the multiplex testing of cannas is routine and we have produced more than 10,000 clean cannas of four varieties in the greenhouse and have transferred these to the field, completing one year of rhizome expansion. After one more season of expanding these stocks, we will be ready in 2018 to introduce these into the market as scientifically tested clean canna varieties. The second goal was to expand tissue culture production of cannas. We made several attempts which were successful, but we determined that this process was expensive and required specialized skills in the laboratory. This approach did not yield high quality, large rhizome foundation stocks within the first year. After several attempts to mainstream tissue culture production of cannas into the operational process we decided to not pursue this further because the rhizomes are too small and not productive enough for introducing into the fields. It is not in the best interest, financially or otherwise, for Horn Canna Farms to manage a tissue culture program, but it makes better sense with regard to time spent, costs and level of success, to focus on the greenhouse sanitization by screening plants. In the last quarter of the project, we determined an even more robust, cost and time saving approach to speed production of clean cannas (See below). The third objective was to produce 10,000 clean canna rhizomes that could be introduced into production. We focused on Red Futurity, Burning Ember, Red King Humbert, and Striped Beauty. We exceeded the goal of producing 5,000 virus free rhizomes by reaching 10,000 virus free rhizomes. Importantly, in the last 3 months we assessed the performance of the Agdia potyvirus immunostrip testing for use in fields to assess the health of the crop. Immunostrips are a trademarked lateral flow test that has antisera detecting viruses. Simply place a plant sample in a clear plastic bag, macerate the sample and dip the immunostrip test into the plant sap. Within 15 minutes the results appear on the strip. We wrote a protocol now for on farm use of this kit to detect potyviruses and cucumoviruses, which represent four of the five viruses infecting cannas. The cost is comparable or higher to RT-PCR, the time spent testing is much less, and any person can be trained to use these tests. For Phase II we identified a company that will sell us the equipment and supplies to print our own immunostrips and we estimated the costs to be ~$1.00 per strip and we can manage two tests per strip. We plan to use genus specific antisera that detects all potyviruses and cucumoviruses and combine these into one test. We also plan to make antisera to CaYMV which we may be able to incorporate as an alternative to cucumoviruses, or as a third line on a printable test strip. This new approach will speed virus detection, segregation of healthy and infected stocks, and will easily become a routine part of operational functions at the farm and could transform production in a single season. In summary, we established 10,000 clean planting stocks of elite plant cultivars. We implemented new protocols for sanitization of farm and greenhouse equipment. We collected data on the sanitization methods and demonstrated an 80% reduction of disease from season to season in the greenhouse if we implemented these protocols. These were transferred to use on farm equipment. The method includes using Virkon for cleaning floors, and machinery in the processing equipment after harvest of rhizomes and before packaging. In Phase II we will propose new transformative projects that will allow us to market the use of scientific data for quality assurance. The impact of Phase I was to transform the production system by introducing new clean plants.. Dr Verchot has reported the genome sequences for CaYSV and CaYMV. Through this cooperation Horn Canna Farm is working to transform its reputation for using scientific data in their production. This project has attracted new cooperative partners. First, VF Canna LLC worked with LARTA on developing a constuency. VF Canna LLC introduced Horn Canna Farms and Plant Haven LLC. Plant Haven is an agent that helps breeders bring new varieties to market. We are working with Dr Verchot and Plant Haven to develop and release two new varieties of canna into the market. This project has also attracted a partnership with Xplosafe LLC. This company has proprietary solvent for cleaning up nitrogen and phosphorus from lakes, ponds, and storm water. Pilot projects by other countries determined that canna is a terrific crop for phytoremediation. Canna consumes large amounts of nitrogen and phosphorus and is being implemented in grey water recycling programs in India and other developing countries. Xplosafe is piloting a project with VF Canna LLC to test the use of their saturated product as a fertilizer for cannas. It is expected that the rhizome biomass should improve with application of the solvent to growing rhizomes. If the pilot project succeeds we will look for ways of using these products on the farm. This could cut costs for chemical fertilizer while aiding the environment.
Publications
- Type:
Journal Articles
Status:
Other
Year Published:
2017
Citation:
Two species of Canna yellow mottle virus infect Cannas
- Type:
Other
Status:
Published
Year Published:
2015
Citation:
Verchot J and Webb A (2015) Greenhouse propagation of ornamental cannas grown from rhizomes or seeds. EPP-7333
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Progress 06/01/16 to 01/31/17
Outputs
Target Audience:Flowerbulb Brokers example Planthaven, Inc. Santa Barbara, CA. Negotiating with PlantHaven for Phase II to launch 2 new canna varieties. Screening of germplasm for viruses will be carried out by an independent entity known as VF Canna LLC. They provide testing and quality assurance of material before planting in the field by Horn Canna Farms. Other target audiences includeFlowerbulb Distributors, for example Gardenstate Bulb, Inc. Vineland, NJ Commercial greenhouses Home gardeners This project has also attracted parternship involving VF Canna LLC and Xplosafe LLC. This company has proprietary solvent for cleaning up nitrogen and phosphorus from lakes, bonds, and storm water. Pilot projects by other countries determined that canna is a terrific crop for phytoremediation. Canna consumes large amounts of nitrogen and phosphorus and is being implemented in grey water recycling programs in India , etc. Xplosafe is piloting a project with VF Canna LLC to test the use of their saturated product as a fertilizer for cannas. It is expected that the rhizome biomass should improve with application of the solvent to growing rhizomes. If the pilot project succeeds we will look for ways of using these products on the farm. This could cut costs for chemical fertilizer while aiding the environment. Changes/Problems:We are asking for extension of grant until June 2017 so we can complete one more season. The most significant change was trying to isolate and sequence the CaYMV genome. This was problematic because we obtained sequence data that suggested there are more than one virus species. We reported and developed diagnostic primers for the one species and just this week identified the second species. Both are badnaviruses that exist in canna plants. However through the summer we revised our PCR diagnostic test to detect the genome we isolated but could not make sense of the remaining PCR data suggesting another species might exist. We failed to isolate this other species until November 2016. This slowed down our ability to report new findings to the scientific community and to consumers/purchasing agents. We launched a set of experiments in the summer of 2016 to learn if CaYSV, BYMV, CaYMV can be transmitted from plant-to-plant via contaminated soil, by rhizomes touching each other, or by insects. This became complicated because the diagnostic PCR test for CaYMV did not match the genome sequence that we isolated. We used PCR primers for both species until we could make sense of the results. The work was not delayed but the decision making based on the work was slowed. What opportunities for training and professional development has the project provided?This project was very instrumental in launching Austin Gimondo to graduate studies in the field of virology. VF Canna LLC is housed on campus at Oklahoma State University. This SBIR subcontracted to Dr. Verchot, who is also the leader for VF Canna LLC. This subcontract provided training to students in Horticulture and Plant Pathology. The greenhouse program for screening cannas is essentially a production greenhouse. We have databases that track every individual plant that goes through the program, disease ratings based on visual observation, and diagnostic tests. Students who were employed were trained in greenhouse management, disease management and advanced database for greenhouse production. Graduate students also assist in training growers at Horn farms on how to identify disease and segregate healthy stocks. One-on-one students work with farm workers by walking the fields, answering questions, and sharing knowledge. Students learn about farm production, concerns, disease management, while sharing their own knowledge from the greenhouse experience. How have the results been disseminated to communities of interest?Mostly through social media and direct mail. We also have opportunities to visit directly with our customers about the bright future that cannas have because of this research. We get to help them understand that not all negavtive symptoms in cannas or plants in general are a result of virus. Knowledge sharing is via several websites: Verchot.okstate.edu, vfcanna.com, and cannas.net. VF Canna LLC sponsored an exhibit at the OSU botanical gardens providing clean cannas and information sheets. What do you plan to do during the next reporting period to accomplish the goals?We are placing a great deal of focus on expansion the number of varieties that are virus free. Through outreach efforts we're going to work to encourage growers to change to our newest PCR diagnostic technology to reduce false positives. Create more documentation of conditions that could lead to false positives results during diagnostics.
Impacts
What was accomplished under these goals?
We reported first full genome sequence for CaYMV and developed new diagnostics that are improved over prior methods. In the last weeks we believe that we have identified at least one new species related to CaYMV and we are cloning the full genome and will develop and implement diagnostic tests over the next 6 months. Reduction in false positives, detects less plant sequences, and higher specificity for detecting CaYMV, CaYSV, and BYMV simultaneously. This is proprietary, but hoping to make available in the next year to diagnostic labs who may want to purchase protocol kit. We have consistently worked with three varieties of canna with success in past years. This year we have added two new clean varieties of which have green foliage foliage the three varieties from past years had bronze foliage and variegated foliage. We are pleased to have clean green foliage which adds needed depth to our inventory. We exceeded the goal of producting 5,000 virus free rhizomes by reaching 10,000 virus free rhizomes. We tested the performance of the Agdia potyvirus immunostrip test for use in fields to assess the health of the crop. We are writing a protocol now for onfarm use of this kit. We are also contacting filter paper manufacturers such as 3M to obtain the equipment needed to prepare our own immunostrip test for inhouse use. This will be cheaper than purchasing Agdia kits. Working with several sources around the country to identify adequate starting material. Learned the best and not so good sources of cannas. We established 10,000 clean planting stocks of 6 elite plant cultivars. We implemented new protocols for sanitization of farm and greenhouse equipment. We collected data on the sanitization methods and demonstrated an 80% reduction of disease from season to season in the greenhouse if we implemented these protocols. These were transferred to use on farm equipment. The method includes using Virkon for cleaning floors, and machinery in the processing equipment after harvest of rhizomes and before packaging. Green house produced cannas were transferred to Horn Canna Farms production, bulked up, and monitored for health for 2 seasons. Impacts were transformation of the production system by introducing new clean plants. We also launched a new iniative with Plant Haven LLC. Plant Haven is an agent that helps breeders bring new varieties to market. We are working with Dr Verchot and Plant Haven to release 2 new varieties of canna into the market. Dr Verchot has reported the genome sequences for CaYSV and CaYMV. Through this cooperation Horn canna farms has gained a reputation for using scientific data in their production and this is important for reputation. Horn has been renting land from neighbors and has now returned to their own fields for production. Rotating crops between fields is likely to reduce the incidence of virus disease. Soybean is becoming the preferred rotation crop. VF Canna LLC worked with LARTA on developing a constuency. Together they determined that a role for VF Canna LLC is to maintain the greenhouse clean canna production program and attract breeders and agents who want to launch new varieties and want performance testing done. This will be contractual work. VF Canna will also play a role for consulting and training producers, agents, breeders to ensure success of the industry. A second goal is to expand to other markets beyond cannas. This project has also attracted parternship with Xplosafe LLC. This company has proprietary solvent for cleaning up nitrogen and phosphorus from lakes, bonds, and storm water. Pilot projects by other countries determined that canna is a terrific crop for phytoremediation. Canna consumes large amounts of nitrogen and phosphorus and is being implemented in grey water recycling programs in India , etc. Xplosafe is piloting a project with VF Canna LLC to test the use of their saturated product as a fertilizer for cannas. It is expected that the rhizome biomass should improve with application of the solvent to growing rhizomes. If the pilot project succeeds we will look for ways of using these products on the farm. This could cut costs for chemical fertilizer while aiding the environment.
Publications
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2016
Citation:
Canna yellow mottle virus and persistent subviral DNA associate with potyvirus infection in Canna
Journal: Virus Research
- Type:
Other
Status:
Published
Year Published:
2015
Citation:
Verchot J and Webb A (2015) Greenhouse propagation of ornamental cannas grown from rhizomes or seeds. EPP-7333
|
Progress 06/01/15 to 05/31/16
Outputs
Target Audience:Flowerbulb Brokers ex. Planthaven, Inc. Santa Barbara, CA Flowerbulb Distributors ex. Gardenstate Bulb, Inc. Vineland, NJ Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Hugely helpful for Austin Gimondo which has launched to him to graduate studies. How have the results been disseminated to communities of interest?Mostly thorugh social media and direct email. (Twitter and Facebook are the primary tool in social media for us) What do you plan to do during the next reporting period to accomplish the goals? Expand the number of varieties. Through outreach efforts we're going to work to encourage growers to change to our newest PCR diagnostic technology to reduce false positives. Further documentation of conditions that could lead to false positive results during diagnostics.
Impacts
What was accomplished under these goals?
We reported first full genome sequence for CaYMV and developed new diagnostics that are improved over prior methods. Reduces false positives, detects less plant sequences, and higher specificity for dectecting CaYMV, CaYSV, and BYMV simultaneously. This is proprietary, but hoping to make available in the next year to diagnostic labs who may want to purchase a protocol/kit. We've consistently worked with 3 varieties of canna with success in past years. This year we have added two new clean varieties of which have green foliage the three varieties from past years had bronze foliage and variegated. We are pleased to have clean green foliage which adds needed depth to our inventory. We exceeded the goal of producing 5000 virus free rhizomes by reaching 8,000 clean rhizomes.
Publications
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2015
Citation:
Canna yellow mottle virus and persistent subviral DNA associate with potyvirus infection in Canna
Journal: Virus Research
|