2005 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Fresh fruit are attacked by fungal diseases that rot fruit after harvest. The fungal diseases that rot fruit after harvest cause losses of about 5% of California's $3 billion fresh fruit production. The control of the disease-causing fungi is important, and is usually accomplished using chemical fungicides and sanitizers, but issues of pest resistance to these chemicals, the dietary safety of their presence foods, and their impact on the environment has made the search for safer replacements important. Fungicides used to control these diseases interfere with the export of citrus fruit because some countries will not accept residues of the fungicides on the fruit. Three fungicides are approved for citrus fruit use in California, sodium ortho-phenyl phenate, imazalil, and thiabendazole. Sodium ortho-phenyl phenate is classified as a probable human carcinogen and imazalil as a possible human carcinogen by the U.S. Environmental Protection Agency. We propose to use 'reduced-risk' compounds, such as lime-sulfur solution, sodium bicarbonate, or ethanol, to control these fungi that will minimize the dietary risk posed by fungicides present in fruit purchased by consumers. This work will not only benefit consumers, and produce growers and distributors will benefit because chemical residues on products are prohibited by some importing countries and in domestic markets where organic produce is sold. 2.List the milestones (indicators of progress) from your Project Plan. Year 1 (FY 2004) Conduct laboratory trials to control postharvest green mold and sour rot on citrus fruit, and postharvest disease control trials with table grapes to control gray mold. Develop of inoculation and culture methods for large-scale powdery mildew inoculation and disease quantification. Conduct anatomical studies of berries surfaces and skin of selections with gray mold resistance. (FY 2004 and 2005) Initiate planned heat treatments of stone fruit and lemons; measure quality and phytotoxicity effects; extract collected samples for soluble sugars. Develop database relating fruit color, soluble solids/titratable acidity, ethanol and navel orange fruit acceptability. Assay antioxidant response to heat treatment and relate to subsequent injury. Year 2 (FY 2005) Conduct large-scale laboratory trials to control postharvest green mold and sour rot on citrus fruit, and postharvest disease control trials with table grapes to control gray mold. Include rigorous fruit quality evaluation of effective approaches, conduct grower/processor demonstration tests, seek industry input and collaboration, inquire about registration issues with regulatory agencies, and establish the category of regulation applied to the process. (FY 2004 and 2005) Initiate planned heat treatments of stone fruit and lemons; measure quality and phytotoxicity effects; extract collected samples for soluble sugars. Develop database relating fruit color, soluble solids/titratable acidity, ethanol and navel orange fruit acceptability. Assay antioxidant response to heat treatment and relate to subsequent injury. Begin large-scale powdery mildew inoculation and disease quantification, establish in which organs resistance is expressed (leaves, rachis, or fruit) and when. Determine if laboratory assessment of resistance among young vines can predict vineyard resistance to powdery mildew. Complete anatomical studies of berries surfaces and skin of selections with gray mold resistance. Year 3 Conduct pilot-scale trials with naturally infected fruit in collaborator’s facilities. Refine treatment protocols to maximize their effectiveness and practicality. Address patent issues. Participate in compliance of registration needs with regulatory agencies and sponsoring registrant. Continue large-scale powdery mildew inoculation and disease quantification trials as in Year 2. Begin analysis of selections where resistance and susceptibility have been established to determine what features are associated with resistance. Prepare a manuscript on the subject of anatomical studies of berries surfaces and skin of selections with gray mold resistance. (FY 2003-2004) Purify samples and determine sugars by HPLC; prepare report. Include additional variables such as waxing and handling into navel orange fruit acceptability study. Continue antioxidant assays and identify compounds present in lemon peel that are responsible for antioxidant response. Year 4 Conduct pilot-scale trials with naturally infected fruit in collaborator’s facilities with several important cultivars. Refine treatment protocols to maximize their effectiveness and practicality. Participate in compliance of registration needs with regulatory agencies and sponsoring registrant. (FY 2003-2004) Purify samples and determine sugars by HPLC; prepare report. Include additional variables such as waxing and handling into navel orange fruit acceptability study. Continue antioxidant assays and identify compounds present in lemon peel that are responsible for antioxidant response. Continue and confirm results from prior seasons of large-scale powdery mildew inoculation and disease quantification trials begun in Year 2. Continue analysis of selections where resistance and susceptibility have been established to determine what features are associated with resistance. Year 5 Continue pilot-scale trials if necessary. Refine treatment protocols to maximize their effectiveness and practicality. Prepare manuscript describing accomplishments. Conclude analysis of selections where resistance and susceptibility have been established to determine what features are associated with resistance, and prepare a manuscript on this subject. Analyze and summarize data; prepare manuscripts for review and publication. 4a.What was the single most significant accomplishment this past year? A method was developed to enhance activity of the fungicide imazalil to control green mold, caused by Penicillium digitatum, while reducing its rate of use by 50% or more using sodium bicarbonate. Reducing fungicide use is valuable to reduce industry costs and residues in the citrus fruit ingested by consumers. The technique also accomplished partial control of imazalil-resistant isolates of the fungal pathogen Penicillium digitatum, which causes most of the postharvest losses in California. With collaborators William Goodwine of Janssen Pharmacuetica and David Sorenson of Sunkist Fruit Grower’s Supply, a series of laboratory, pilot scale, and commercial experiments were conducted to discover and optimize the combination treatment so it could be incorporated into commercial use. A publication describing this work was issued in 2005, and some version of the method is now used in most California packinghouses. 4b.List other significant accomplishments, if any. Freeze injury caused economic losses to citrus growers and packers and the industry needs a simple and inexpensive test for determining freeze injury on quality of oranges. Scientists of the Commodity Protection & Quality Research Unit in collaboration with support of the Citrus Research Board conducted tests of freezing temperatures on volatile emissions from navel oranges. Ethanol and three other chemical compounds were found in frozen oranges while none were detected from unfrozen fruits. The findings could provide the citrus industry a rapid method to differentiate marketable quality fruit from injured fruit. The California citrus industry needs to market the highest quality fruit in order to stay competitive in the marketplace. In a collaboration between David Obenland (ARS-Parlier) and Mary Lu Arpaia (University of California, Kearney) various quality parameters were evaluated during fruit maturation and following different industry postharvest treatments and comparisons made to the sensory acceptability of the fruit as determined by taste panels. Taste was found to be most closely linked to the sweetness of the fruit and an equation to predict taste from the soluble solids concentration was developed from the data. This information may be used to support efforts to change the minimum maturity requirements that govern when oranges may be harvested in California. A rapid method to predict the powdery mildew resistance of mature grapevines using a laboratory inoculation method applied young grapevines was developed in 2005. The purpose for this work is to identify and quantify mildew resistant grapes as early as possible in young vines of several months in age to accelerate the development of resistant vines, which previously required 5 years or more. The method was developed in our laboratory in collaboration with ARS scientist David Ramming of this location. Mildew resistant vines should reduce the fungicide applications growers apply; currently, more pounds of fungicide are applied to table grapes than any crop in California. Postharvest decay of citrus and table grapes could be reduced as much as 90% by exposure of lemons to fungal volatiles from cultures of the biological control fungus Muscodor albus applied after harvest during storage of the fruit. Control of postharvest decay losses of these products by means other than fungicide applications are of value to reduce chemical inputs and fungicide residues in the fruit. In collaboration with Dr. Julien Mercier of AgriQuest Inc. in Davis, California, these applications were developed in a series of experiments primarily conducted in our laboratory, although preparation and formulation of M. albus was done in Davis. Postharvest biological control by M. albus is promising approach for citrus and table grape industries to manage postharvest decay losses, and efforts to obtain regulatory approval by our commercial collaborator are in progress. 4c.List any significant activities that support special target populations. None. 5.Describe the major accomplishments over the life of the project, including their predicted or actual impact. Our major accomplishments, with credit shared with collaborators, are the continued discovery, development, and transfer of technologies to replace or reduce synthetic fungicide use by the citrus industry, and the identification of disease resistance in new grape selections. These accomplishments are in concordance with the objectives 1, 2 and 3 of Project 5302-43000-032-00D “Emerging Technologies to Maintain Postharvest Quality and Control Decay of Fresh Commodities”, and with the Action Plans of National Programs 303 and 306. Use of heated imazalil aqueous solutions of the fungicide, that enable its rates to be reduced by 50% or more, is in common commercial use now as a result of this project. Immersion of fruit in bicarbonate and carbonate solutions, which avoids fungicide use entirely, or in combination with low rates of the fungicides, has also become popular. Promising technologies that are yet to be implemented include “biofumigation” with Muscodor albus and integrated treatments incorporating biological control and thermal curing. In addition to reducing fungicide use, these techniques have improved management of postharvest citrus and table grape diseases, particularly for high value export fruit during long transit. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? In 2005, a new technology, first implemented in the prior year, in citrus industry packinghouses in which we participated was the use of high volume, low pressure overhead drench application of dilute heated imazalil or thiabendazole fungicide solutions. In some facilities, the fungicide is combined with sodium bicarbonate, which allows an even lower rate to be used, and some control of fungicide resistant strains of the green mold pathogen was also observed. The implementation of most of the 'reduced risk' and 'certified organic' approaches is rapid because they may not require formal EPA registration, such as bicarbonate solutions alone or in combination with fungicides, heat treatments, or the use of generally recognized as safe buffers to optimize the pH of already approved fungicides. Other technologies that require formal regulatory approval, such as the postharvest use of ethanol on table grapes, biological control agents such as Muscodor albus or Pantoea agglomerans, or new EPA classified “reduced risk” postharvest fungicide such as pyrimethanil for citrus, have commercial advocates that are petitioners for registration and this process takes several years. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Obenland, D., Influence of maturity and handling on navel orange volatiles and their relationship to quality. Presented to the University of California, Plant Sciences Department, Pest Control Management Seminar at Santa Barbara. April 4, 2005. Smilanick, J.L., Influence of pH and sodium bicarbonate on the performance of the postharvest fungicide imazalil. Presented to the University of California, Plant Sciences Department, Pest Control Management Seminar at Santa Barbara, April 6, 2005. Smilanick, J.L., Practical alternatives to fungicides for citrus packinghouse managers. Presented to the packinghouse manager’s meeting sponsored by Sunkist Fruit Grower’s Supply at Visalia, California, June 2, 2005. Review Publications Smilanick, J.L. 2004. Use of ozone in storage and packing facilities. Meeting Abstract. Proceedings 99th Meeting of the Washington State Horticultural Association. p. 131-139. Smilanick, J.L., Mansour, M., Margosan, D.A., Gabler, M., Goodwine, W.R. 2005. Influence of ph and nahco3 on the effectiveness of imazalil to inhibit germination of spores of penicillium digitatum and to control postharvest green mold on citrus fruit. Plant Disease. 89:640-648. Smilanick, J.L., Mlikota, F.G. 2005. Predicting the field resistance to powdery mildew of grapevine selections based on the laboratory testing of young vines. American Phytopathological Society Annual Meeting. 95:S71. Mercier, J., Smilanick, J.L. 2005. Control of green mold and sour rot of stored lemons by biofumigation with muscodor albus. Journal of Biological Control. 32:401-407. Smilanick, J.L. 2005. Eliminating or minimizing the use of fungicides to control postharvest diseases of fresh fruit. Phytopathology. 95:S140.