Progress 02/06/06 to 02/05/11
Outputs
Progress Report Objectives (from AD-416) The overall objective of this project is to apply the Pichia anomala yeast product to pistachio orchards early in the season prior to June 15 to be followed by 'atox' technology. Measure the reduction of recoverable Aspergillus flavus spores in treated plots or use some other appropriate measure of reduced colonization of natural substrates. Will develop commercially viable methods for control of fungal and insect pests which contribute to pre-harvest aflatoxin contamination of tree nuts. The methods are to be environmentally benign and not harmful to humans. To control fungi, there are two main objectives. The first objective for this project is to control mycotoxin-producing fungi using bacteria. The second biocontrol effort is well underway and involves the biological control of A. flavus in tree nut orchards using the saprophytic yeast Pichia anomala. The yeast will also be examined to determine its effectiveness against other fungal pathogens, e.g., Fusarium spp., Penicillium spp. The third objective of this project is also an ongoing research effort. This objective entails development of semiochemical- based low-risk control strategies against key insect pests of tree nuts. Insect feeding damage is associated with the invasion of microbial pathogens and mycotoxin contamination. Approach (from AD-416) Develop mentods to control insect pests and toxic fungi of tree nuts. Insects include naval orangeworm, codling moth and peach twig borer. Feeding damage by these insects leads to infection by aflatoxigenic aspergilli. Control methods for insects are to be environmentally benign and employ semiochemicals to disrupt insect behavior. Control of toxic fungi focuses on biological control using competitive or antagonistic microorganisms. These microorganisms include either yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post harvest environments. Replacing 5325-42000-031-00D (2/06). This is the final report for the project 5325-42000-036-00D terminated January 2011. There was a period between the beginning of FY2011 and the termination date for the project in fall and winter. All planned experimentation for this project was completed. Some of this work served as a springboard for further research in a new project salient to the efforts of this project. The continuing efforts resulting from this project are outlined in the newly numbered project 5325-42000-038-00D. PMR scientists developed methods for insect and microorganism control with regard to the aflatoxin contamination problem. MU scientists identified a number of host-plant volatiles (HPVs) that, as a blend, attract female and male navel orangeworm (NOW) at a higher rate than current commercial lures under field conditions. MU scientists also developed a new method for collecting HPVs. Much of this work was done under a CRADA with a major tree nut stakeholder in California. MU scientists discovered a strain of yeast, Pichia anomala, that is a viable biocontrol microorganism against the fungi that make aflatoxin. This yeast shows a great deal of viability in tree nut orchards, has no human pathogenicity, is not phytotoxic, and can be sprayed directly onto the tree nut canopy. PMR scientists optimized the culture medium and are able to produce vast quantities of this organism. Field trials to examine the efficacy of bacterial biocontrol strains for controlling aflatoxin and fumonisin on corn showed that bacterial treatments had no effect on mycotoxin levels in treated corn. Accomplishments 01 Using bacteria to control fungal infection in crops. Food safety will improve by developing methods to reduce mycotoxin contamination of food commodities. Research focused on developing methods using biological control to control mycotoxin-producing fungi. ARS scientists at Albany, demonstrated reduction of fungal populations in soil co-inoculated with biocontrol bacteria, leading to reduced spore production and potential f reduced infection of corn via windborne spores. The use of such bacteria could lower the need for fungicides, which may be harmful to the environment. 02 Safe, natural yeast reduces aflatoxin contamination. Aflatoxin is a natural carcinogen that can contaminate tree nuts. Reducing this contamination is necessary to reduce economical losses due to tight controls on exports to other countries. ARS scientists in Albany, CA, in collaboration with CRADA partners, discovered one yeast species, Pichia anomala that reduces infection of tree nuts by aflatoxin-producing fungi They have developed production methods for this yeast, as well. In addition, they are using molecular techniques to identify other species yeast that are natural residents of tree nut orchards. These other yeast may prove to be good biocontrol agents, as well. Effective biocontrol wi be a safe way to enhance food safety. 03 Control of pest insects of tree nuts using natural lures. Insect damage of tree nuts allow contamination by aflatoxin. ARS scientists in Albany CA, in collaboration with CRADA partners, have developed methods for collecting, identifying and bioassaying natural volatiles that influence behavior of insect pests of tree nuts. They found blends of natural volatiles that can be used to lure these pests to traps; two in particul include codling moth (CM) and navel orangeworm (NOW). One lure, the pear ester, enhances pheromone mating disruption of CM and pear ester microencapsulated (PE-MEC) formulation disrupts normal behavior of larva CM. Several blends of volatiles show promise for control of NOW. Some of these blends show better activity in the field, as lures, than currently available commercial products. Use of natural lures to control tree nut insect pests can reduce the risk of loss due to insect damage and concomitant contamination of aflatoxin.
Impacts
(N/A)
Publications
- Palumbo, J.D., O Keeffe, T.L., Mcgarvey, J.A. 2011. Incidence of fumonisin B2 production within Aspergillus section Nigri populations isolated from California raisins. Journal of Food Protection. 74:672-675.
- Hua, S.T., Brandl, M., Hernlem, B.J., Eng, J.G., Sarreal, S.L. 2010. Fluorescent viability stains to probe the metabolic status of aflatoxigenic fungus in dual culture of Aspergillus flavus and Pichia anomala. Mycopathologia. Available:
- Light, D.M., Beck, J.J. 2010. Characterization of microencapsulated pear ester, (2E,4Z)-ethyl-2,4-decadienoate: a kairomonal spray-adjuvant against neonate codling moth larvae. Journal of Agricultural and Food Chemistry. 58(13):7838-7845.
- Roitman, J.N., Merrill, G.B., Beck, J.J. 2011. Survey of ex situ fruit and leaf volatiles from several Pistacia cultivars grown in California. Journal of the Science of Food and Agriculture. 91(5):934-942.
- Palumbo, J.D., O Keeffe, T.L., Vasquez, S.J., Mahoney, N.E. 2011. Isolation and identification of ochratoxin A-producing Aspergillus section Nigri strains from California raisins. Letters in Applied Microbiology. 52:330-336.
|
Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) The overall objective of this project is to apply the Pichia anomala yeast product to pistachio orchards early in the season prior to June 15 to be followed by 'atox' technology. Measure the reduction of recoverable Aspergillus flavus spores in treated plots or use some other appropriate measure of reduced colonization of natural substrates. Will develop commercially viable methods for control of fungal and insect pests which contribute to pre-harvest aflatoxin contamination of tree nuts. The methods are to be environmentally benign and not harmful to humans. To control fungi, there are two main objectives. The first objective for this project is to control mycotoxin-producing fungi using bacteria. The second biocontrol effort is well underway and involves the biological control of A. flavus in tree nut orchards using the saprophytic yeast Pichia anomala. The yeast will also be examined to determine its effectiveness against other fungal pathogens, e.g., Fusarium spp., Penicillium spp. The third objective of this project is also an ongoing research effort. This objective entails development of semiochemical- based low-risk control strategies against key insect pests of tree nuts. Insect feeding damage is associated with the invasion of microbial pathogens and mycotoxin contamination. Approach (from AD-416) Develop methods to control insect pests and toxic fungi of tree nuts. Insects include naval orangeworm, codling moth and peach twig borer. Feeding damage by these insects leads to infection by aflatoxigenic aspergilli. Control methods for insects are to be environmentally benign and employ semiochemicals to disrupt insect behavior. Control of toxic fungi focuses on biological control using competitive or antagonistic microorganisms. These microorganisms include either yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post harvest environments. Replacing 5325-42000-031-00D (2/06). PMR scientists have made significant progress in developing methods that involve insect and microorganism control with regard to the aflatoxin contamination problem. MU scientists identified a number of host-plant volatiles (HPVs) that, as a blend, were as active at attracting navel orangeworm (NOW) females as the current commercially used almond press cake. Moreover, these volatiles also attracted male NOW equivalent to and better than current NOW pheromone bait traps. PMR scientists have made substantial progress in this area by developing a new method for collecting VOCs in situ and using electro-antennagram (EAG) and wind tunnel bioassays to determine which VOCs have bioactivity. This work is being done under a CRADA with a major tree nut stakeholder, Paramount Farms, in California. These VOCs that attractd NOW are now being tested in field studies by our collaborator. The second approach by the scientists in the project for controlling aflatoxin contamination is use of competitive micro-organisms against the fungus that makes aflatoxin. In this regard, there are two approaches. PMR scientists have discovered a strain of yeast, Pichia anomala, that is a viable biocontrol microorganism against the fungi that make aflatoxin. This yeast shows a great deal of viability in tree nut orchards, has no human pathogenicity, is not phytotoxic, and thus can be sprayed directly onto the tree nut canopy. PMR scientists have optimized the culture medium for the yeast and are able to produce large quantities. The research is being done in collaboration with scientists in the Dept. of Pomology, Univ. of California, Davis. A second biocontrol effort identified a number of natural soil bacteria in tree nut orchards and vineyards that show promise as biological control agents against fungi in soil. Accomplishments 01 Host-plant-volatiles (HPVs) affect navel orangeworm (NOW) behaviors. NO is the major insect pest of the California tree nut industry requiring a great deal of pesticidal control. Improved and more efficient means of control are needed. One approach would be to develop lures and monitorin methods based on host volatiles that are attractive to these insects. Researchers at the Plant Mycotoxin Research Unit in Albany, California developed new instrumentation for collection of volatile compounds from almond and pistachio orchards. Nearly 200 compounds were identified as candidate attractants of NOW based on laboratory and field methods, with some blends being more active as attractants than current, commercially available lures. Discovery of any effective natural lures for the contro of NOW would have a major impact on California agriculture in that this insect destroys millions of dollars of tree nuts annually. 02 Culture medium for mass production of a biocontrol agent of aflatoxin producing fungi. Aflatoxin is a natural cacinogen made by fungi and its contamination of tree nuts is a major economic and food safety issue. P harvest control aflatoxin-producing fungi would greatly assist the industry. Biocontrol using competing microorganism is an economic and sa approach to controlling aflatoxin-producing fungi. Researchers at the Plant Mycotoxin Research Unit in Albany, California have found a method for maintaining the viability of a promising biocontrol agent, Pichia anomala, by developing a routine procedure for analyzing yeast cell viability. This led to development of a stable liquid formulation of P. anomala for application in field plots for the control of aflatoxin- producing fungi. 03 Attracticide developed against codling moth (CM), a major pest of pome fruits and tree nuts. CM is a major insect pest of pome fruits (apples, pears, etc.) and tree nuts and improved control measures that reduce pesticide usage are greatly needed. Researchers at the Plant Mycotoxin Research Unit in Albany, California discovered a novel attract and kill (AK) control tactic against CM larvae using a volatile from pears. This volatile was encapsulated in small beads and co-applied in sprays that included various insecticides. This combination enhanced the effectivene of insecticidal usage against CM allowing for lower, less costly, and mo environmentally safe rates of insecticides to be used for CM control.
Impacts
(N/A)
Publications
- Dragull, K.D., Beck, J.J., Merrill, G.B. 2010. Essential oil yield and composition of Pistacia vera "Kerman" fruits, peduncles, and leaves grown in California. Journal of the Science of Food and Agriculture. 90(4):664- 668.
- Beck, J.J. 2010. Letter to the Editor: Styrene-producing microbes in food- stuff. Journal of Food Science. 75(1):X.
|
Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) The overall objective of this project is to apply the Pichia anomala yeast product to pistachio orchards early in the season prior to June 15 to be followed by 'atox' technology. Measure the reduction of recoverable Aspergillus flavus spores in treated plots or use some other appropriate measure of reduced colonization of natural substrates. Will develop commercially viable methods for control of fungal and insect pests which contribute to pre-harvest aflatoxin contamination of tree nuts. The methods are to be environmentally benign and not harmful to humans. To control fungi, there are two main objectives. The first objective for this project is to control mycotoxin-producing fungi using bacteria. The second biocontrol effort is well underway and involves the biological control of A. flavus in tree nut orchards using the saprophytic yeast Pichia anomala. The yeast will also be examined to determine its effectiveness against other fungal pathogens, e.g., Fusarium spp., Penicillium spp. The third objective of this project is also an ongoing research effort. This objective entails development of semiochemical- based low-risk control strategies against key insect pests of tree nuts. Insect feeding damage is associated with the invasion of microbial pathogens and mycotoxin contamination. Approach (from AD-416) Develop methods to control insect pests and toxic fungi of tree nuts. Insects include naval orangeworm, codling moth and peach twig borer. Feeding damage by these insects leads to infection by aflatoxigenic aspergilli. Control methods for insects are to be environmentally benign and employ semiochemicals to disrupt insect behavior. Control of toxic fungi focuses on biological control using competitive or antagonistic microorganisms. These microorganisms include either yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post harvest environments. Significant Activities that Support Special Target Populations Aflatoxin is a highly carcinogenic compound produced by certain fungi that can infect a wide variety of edible agricultural products, such as peanuts, corn, tree nuts, and in some countries, wheat. Aflatoxin contamination is a major food safety issue and is strictly regulated, resulting in it also being a significant international trade issue involving US agricultural products. The research in this project mainly involves development of environmentally friendly methods of preventing aflatoxin contamination. USDA scientists in the Plant Mycotoxin Research Unit, Albany, CA, are tackling this problem by developing methods that involve insect and microorganism control, namely methods that fall under the general umbrella of what is termed "biological control." MU scientists clearly showed that insect feeding damage can lead to aflatoxin contamination. This was shown in experiments involving the navel orangeworm (NOW), the major insect pest of almond. NOW was able to carry spores of aflatoxin producing fungi on hairs that cover its body. MU scientists are attempting to control pest insects by identifying host- plant volatiles (HPVs), also termed volatile organic compounds (VOCs). These HPVs could be used as lures to traps or as a means of confusing or distracting the insects from locating the host-plant. MU scientists have made substantial progress in this area by developing a new method for collecting VOCs in situ and using electro-antennagram (EAG) and wind tunnel bioassays to determine which VOCs have bioactivity. This work is being done under a CRADA with a major tree nut stakeholder, in California. Efforts are also underway to use combinations of lure VOCs and pesticides in order to create an "attracticide." This strategy would function by using the volatiles to lure just the target insects to a specific trap containing a pesticide. This would serve as an alternative to a widely broadcasted pesticide spray. The second approach by the scientists in the project for controlling aflatoxin contamination is use of competitive micro-organisms against the fungus that makes aflatoxin. In this regard, there are two approaches. MU scientists have discovered a potentially promising strain of yeast, Pichia anomala. This yeast shows a great deal of viability in tree nut orchards, has no human pathogenicity (unlike current atoxigenic strains of the fungus that produces aflatoxin that are currently being promoted as the best biological control agents), is not phytotoxic, and thus can be sprayed directly onto the tree nut canopy. The research is being done in collaboration with scientists in the Dept. of Pomology, Univ. of California, Davis. A second biocontrol approach involves identification of natural soil bacteria in tree nut orchards that could show promise as biological control agents against fungi in soil. In this project, a number of bacteria have been identified and are currently being tested in corn fields by collaborators in the Crop Genetics and Production Research Unit, Stoneville, Mississippi. Technology Transfer Number of Active CRADAS: 1 Number of New Patent Applications filed: 1
Impacts
(N/A)
Publications
- Beck, J.J., Merrill, G.B., Higbee, B.S., Light, D.M., Gee, W.S. 2009. In Situ Seasonal Study of the Volatile Production of Almonds (Prunus dulcis), var. 'Nonpareil' and Relationship to Navel Orangeworm. Journal of Agricultural and Food Chemistry. 57(9):3749-3753.
- Palumbo, J.D., O Keeffe, T.L., Abbas, H.K. 2008. Microbial interactions with mycotoxigenic fungi and mycotoxins. Journal of Toxicology Toxins Reviews.27(3):261-285.
- Beck, J.J., Merrill, G.B., Palumbo, J.D., O Keeffe, T.L. 2008. Strain of Fusarium Oxysporum Isolated From Almond Hulls Produces Styrene and 7- Methyl-1,3,5-Cyclooctatriene as the Principal Volatile Components. Journal of Agricultural and Food Chemistry. 56(23):11392-11398.
|
Progress 10/01/07 to 09/30/08
Outputs
Progress Report Objectives (from AD-416) The overall objective of this project is to apply the Pichia anomala yeast product to pistachio orchards early in the season prior to June 15 to be followed by 'atox' technology. Measure the reduction of recoverable Aspergillus flavus spores in treated plots or use some other appropriate measure of reduced colonization of natural substrates. Will develop commercially viable methods for control of fungal and insect pests which contribute to pre-harvest aflatoxin contamination of tree nuts. The methods are to be environmentally benign and not harmful to humans. To control fungi, there are two main objectives. The first objective for this project is to control mycotoxin-producing fungi using bacteria. The second biocontrol effort is well underway and involves the biological control of A. flavus in tree nut orchards using the saprophytic yeast Pichia anomala. The yeast will also be examined to determine its effectiveness against other fungal pathogens, e.g., Fusarium spp., Penicillium spp. The third objective of this project is also an ongoing research effort. This objective entails development of semiochemical- based low-risk control strategies against key insect pests of tree nuts. Insect feeding damage is associated with the invasion of microbial pathogens and mycotoxin contamination. Approach (from AD-416) Develop methods to control insect pests and toxic fungi of tree nuts. Insects include navel orangeworm, codling moth and peach twig borer. Feeding damage by these insects leads to infection by aflatoxigenic aspergilli. Control methods for insects are to be environmentally benign and employ semiochemicals to disrupt insect behavior. Control of toxic fungi focuses on biological control using competitive or antagonistic microorganisms. These microorganisms include either yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post harvest environments. Significant Activities that Support Special Target Populations Aflatoxin is a highly carcinogenic compound produced by certain fungi that can infect a wide variety of edible agricultural products. Aflatoxin contamination is a major food safety issue and is strictly regulated, resulting in it also being a significant international trade issue. USDA scientists in the Plant Mycotoxin Research Unit, Albany, CA, are tackling this problem by developing methods that involve insect and microorganism control, namely methods that fall under the general umbrella of what is termed "biological control." MU scientists clearly showed that insect feeding damage can lead to aflatoxin contamination. This was shown in experiments involving the navel orangeworm (NOW), the major insect pest of almond. NOW was able to carry spores of aflatoxin producing fungi on hairs that cover its body. Moreover, when NOW created feeding wounds on almond kernels, these spores were transferred to these wounds, increasing the potential for aflatoxin contamination. MU scientists are attempting to control pest insects by identifying host-plant volatiles (HPVs), also termed volatile organic compounds (VOCs). These HPVs could be used as lures to traps or as a means of confusing or distracting the insects from locating the host-plant. MU scientists have made substantial progress in this area by developing a significant HPV lure to the codling moth, a major pest of walnuts and pome fruits. MU scientists are also attempting to identify HPVs that could be used against the navel orangeworm (NOW), one of the most destructive insect pests to the tree nut industry. Development of a lure to NOW has been elusive, to date. In this regard, MU scientists have developed new methods of trapping test volatiles in the field and are using sophisticated electro-physiological tools to hone- in on the HPVs of most significance. This work is being done under a CRADA with a major tree nut player, Paramount Farms, in California. The second approach by the scientists in the CRIS for controlling aflatoxin contamination is use of competitive micro-organisms against the fungus that makes aflatoxin. In this regard, there are two approaches. MU scientists have discovered a potentially promising strain of yeast, Pichia anomala. This yeast shows a great deal of viability in tree nut orchards, has no human pathogenicity (unlike current atoxigenic strains of the fungus that produces aflatoxin that are currently being promoted as the best biological control agents), is not phytotoxic, and thus can be sprayed directly onto the tree nut canopy. The research is being done in collaboration with scientists in the Dept. of Pomology, Univ. of California, Davis. A second biocontrol approach involves identification of natural soil bacteria in tree nut orchards that could show promise as biological control agents against fungi in soil. In this project, a number of bacteria have been identified and are currently being tested in corn fields by collaborators in the Crop Genetics and Production Research Unit Stoneville, Mississippi. This work supports NP 108 Component 2.1.5. Technology Transfer Number of Active CRADAS: 1
Impacts
(N/A)
Publications
- Palumbo, J.D., O Keeffe, T.L., Abbas, H.K. 2007. Isolation of maize soil and rhizosphere bacteria with antagonistic activity against Aspergillus flavus and Fusarium verticillioides. Journal of Food Protection. 70(7) :1615-1621.
- Beck, J.J., Higbee,B.S., Merrill, G.B., Roitman, J.N. 2008. Comparison of volatile emissions from undamaged and mechanically damaged almonds. Journal of the Science of Food and Agriculture.88(8):1363-1368
- Chou, S., Everngam, M.C., Beck, J.J. 2008. Allelochemical phenolic acids from Gypsophila paniculata. Journal of Undergraduate Chemistry Research. 7(2):40-42.
- Beck, J.J., Smith, L., Merrill, G.B. 2008. In Situ Volatile Collection, Analysis, and Comparison of Three Centaurea Species and Their Relation to Biocontrol with Herbivorous Insects. Journal of Agriculture and Food Chemistry.56(8):2759-2764.
- Wilson, K.A., Beck, J.J. 2007. Complete Proton and Carbon Assignment of Triclosan via One- and Two- Dimensional Nuclear Magnetic Resonance Analysis. The Chemical Educator.12(5):338-342
|
Progress 10/01/06 to 09/30/07
Outputs
Progress Report Objectives (from AD-416) The overall objective of this project is to apply the Pichia anomala yeast product to pistachio orchards early in the season prior to June 15 to be followed by 'atox' technology. Measure the reduction of recoverable Aspergillus flavus spores in treated plots or use some other appropriate measure of reduced colonization of natural substrates. Will develop commercially viable methods for control of fungal and insect pests which contribute to pre-harvest aflatoxin contamination of tree nuts. The methods are to be environmentally benign and not harmful to humans. To control fungi, there are two main objectives. The first objective for this project is to control mycotoxin-producing fungi using bacteria. The second biocontrol effort is well underway and involves the biological control of A. flavus in tree nut orchards using the saprophytic yeast Pichia anomala. The yeast will also be examined to determine its effectiveness against other fungal pathogens, e.g., Fusarium spp., Penicillium spp. The third objective of this project is also an ongoing research effort. This objective entails development of semiochemical- based low-risk control strategies against key insect pests of tree nuts. Insect feeding damage is associated with the invasion of microbial pathogens and mycotoxin contamination. Approach (from AD-416) Develop mentods to control insect pests and toxic fungi of tree nuts. Insects include naval orangeworm, codling moth and peach twig borer. Feeding damage by these insects leads to infection by aflatoxigenic aspergilli. Control methods for insects are to be environmentally benign and employ semiochemicals to disrupt insect behavior. Control of toxic fungi focuses on biological control using competitive or antagonistic microorganisms. These microorganisms include either yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post harvest environments. Replacing 5325-42000-031-00D (2/06). Accomplishments Discovery of Safe Bacteria for the Biological Control of Fungi that Produce Aflatoxin. Aflatoxin contamination of agricultural crops, such as corn, tree nuts, peanuts and cotton seed, can result in serious economic losses on an annual basis. One approach that shows some promise of success in controlling contamination is use of other microbes as biological control agents against aflatoxin producing fungi. The discovery of any isolates that show promise as biocontrol agents could be fruitful for future biocontrol programs. Strains of the bacteria Pseudomonas chlororaphis and Pseudomonas fluorescens were discovered that reduced aflatoxin producing fungi by 10 to 100 fold in soil from corn fields. Experiments conducted by scientist in the Mycotonins Research Unit in Albany for 16 days, demonstrating persistence of bacterial populations and the extent of the resulting biocontrol. Above accomplishment falls under National Program 108 (Food Safety), Component, 2.1.5 (Biocontrol) Following growth of fungi that normally produce aflatoxin in this soil for 16 days, with or without bacterial co-inoculation, there was no extractable aflatoxin present, suggesting that environmental and nutritional conditions in this soil were not conducive to aflatoxin production. Experiment conducted by scientist in thr Mycotonins Research Unit in Albany CA in collaboration with ARS scientist in Stoneville, MS for field trials on corn. Above accomplishment falls under National Program 108 (Food Safety), Component, 2.1.5 (Biocontrol) Field Trials Using a Safe Yeast Shows Promise for the Biological Control of Aflatoxin Producing Fungi. Aflatoxin contamination of agricultural crops, such as corn, tree nuts, peanuts and cotton seed, can result in serious economic losses on an annual basis. One approach that shows some promise of success in controlling contamination is use of other microbes as biological control agents against aflatoxin producing fungi. A few years ago, USDA scientists in the Plant Myctoxin Research Unit, Albany, CA, discovered a natural, safe yeast, Pichia anomala, that showed promise in lab experiments as a potential biological control agent. Large scale field tests using the yeast were conducted in 2005 and 2006 resulted in significant reductions in cluster loss and significant increases in harvestable yield of pistachios, and the yield for the yeast treatment was 12% greater than for the control. The yeast also effectively controlled a waterborne bacterial contaminant, and may be useful as a control measure against human pathogenic bacterial contaminants in the field. Genetic experiments also showed that the yeasts caused a reduction in the activity of genes involved in aflatoxin biosynthesis (omtA, O- methyltransferase A; omtB, O-methyltransferase B; aflR, aflatoxin biosynthesis regulator; and aflJ, adjacent to aflR). This research shows that this yeast may be a promising biological approach to controlling aflatoxin producing fungi directly on pistachios, without harming the quality of the harvested nuts. Above accomplishment falls under National Program 108 (Food Safety), Component, 2.1.5 (Biocontrol) Use of Natural Chemical Lures and Viruses to Control Insect Pests of Tree Nuts. Insect feeding damage results in significant losses in harvest to the tree nut industries and impact plant health resulting from insect feeding which causes wounds in the protective layers surrounding nut kernels that allows infection by microbes, including fungi that produce aflatoxin. Aflatoxin contamination of foods is a significant food safety issue and USDA scientists in the Plant Mycotoxin Research Unit, Albany, CA, are finding ways of controlling these insect pests by using natural chemical lures that the insects use to find the nuts. By combining these lures with pesticides or specific insect viruses, these pest insects can be controlled effectively without contaminating the environment or inadvertently having harmful effects on non-pest organisms. Chemists within the unit are using instruments that can detect different chemical compounds from the host-plant while simultaneously measuring how the insects respond to the chemical by focusing on the navel orangeworm, (NOW) , the most significant insect pest to the almond and pistachio and codling moth (CM), the most significant insect pest to walnut. To date, a virus has been discovered that is effective for control of both CM and NOW, in addition to use of a discovered lure to CM in combination with pesticides that has been found to be quite effective in the control of CM. Above accomplishment falls under National Program 108 (Food Safety), Component, 2.1.5 (Biocontrol). Technology Transfer Number of New CRADAS and MTAS: 1 Number of Invention Disclosures submitted: 2 Number of Patent Applications filed: 1 Number of Web Sites managed: 1 Number of Non-Peer Reviewed Presentations and Proceedings: 9 Number of Newspaper Articles,Presentations for NonScience Audiences: 1
Impacts
(N/A)
Publications
- Chang, P.-K., Hua, S.S.T. 2007. Nonaflatoxigenic Aspergillus flavus TX9-8 Competitively Prevents Aflatoxin Accumulation by A. flavus Isolates of Large and Small Sclerotial Morphotypes. International Journal of Food Microbiology. 114:275-279.
- Chang, P.-K., Hua, S.T. 2007. Molasses Supplementation Promotes Conidiation but Suppresses Aflatoxin Production by Small Sclerotial Aspergillus flavus. Letters in Applied Microbiology. 44(2):131-137.
- Hua, S.T., Tarun, A.S., Pandey, S.N., Chang, L.Y., Chang, P. 2007. Characterization of AFLAV, a Tfl/Sushi retrotransposon from Aspergillus flavus. Mycopathologia. 163(2):97-104.
- Bayman, P., Baker, J.L. 2006. Ochratoxins: a global perspective. Mycopathologia. the Netherlands: Kluwer Academic Publishers 162: 215-223
|
Progress 10/01/05 to 09/30/06
Outputs
Progress 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? Why does it matter? Mycotoxin contamination of agricultural commodities can result in major food safety and foreign trade issues. Aflatoxin contamination of tree nuts (almonds, pistachios and walnuts) is a significant mycotoxin-related problem now. Tree nuts are valued at approximately $2 billion/yr. Aflatoxin is reported to be one of the most potent natural carcinogens known. Action levels for aflatoxin set by a number of major importing nations are quite strict at 4 ppb. About 50-70% of domestically produced tree nuts are exported and action levels for aflatoxin set by importing nations threaten the tree nut industry. Biological control of organisms associated with aflatoxin contamination of tree nuts is a promising approach to curtailing this problem. Biological control is an environmentally safe
approach that does not entail use of toxic substances that may adversely affect the environment or human health. Two approaches are used, one involves control of the fungi producing mycotoxins, the other involves controlling insects. Insect feeding creates wounds in protective layers surrounding nut kernels. These wounds are major avenues through which fungal spores can infect the kernel. This project's goals are to develop new, effective and innovative approaches to controlling both microbes and insect pests in orchard environments and improve other pre-harvest approaches to reducing aflatoxin contamination of tree nuts. The research undertaken falls under National Program 108 (Food Safety). The research specifically addresses the issues concerning mycotoxin contamination as outlined in goals 2.1.4 (Breeding resistant crops) and 2. 1.5 (Biocontrol) as described in the National Program Action Plan. Mycotoxin contamination of food commodities is becoming an increasing problem. In
particular, aflatoxin contamination of tree nuts intended for human consumption has become a serious food-safety and marketing issue. Strict threshold levels for aflatoxin create constraints on marketing and pose serious problem for growers and processors. Meager contamination levels found in tree nut exports have resulted in rejection of almost 100 tons of domestically produced tree nut commodities over the past few years. Consumers are very sensitive to the issue and negative public perceptions threaten the industry. 2. List by year the currently approved milestones (indicators of research progress) Year 1 (FY 2006) Collection of bacterial isolates from almond and corn field samples; development of biocontrol screens. Evaluate mycotoxin on corn and spore number on wounded nut fruits treated with yeast. Influence of medium composition on biomass of P. anomala. Bioassay screening of yeast effective on controlling fungal pathogens. Preliminary hybridization of P. anomala cDNA to S.
cerevisiae microarray. GC/MS intact leaves, hulls, nuts, fruits; Develop database, Develop bioassays Now. GC/MS leaves, hulls nuts, fruits; Continue develop CM MD PEK enhancement trials. Develop adult attracticide stations and bait-sprays, Kill station and bait-spray designs. Year 2 (FY2007) Identification and characterization of active bacteria; publication of results; initiation of corn field trials. Determine number and time of spray, check aflatoxin level in nuts. Select the best medium, check viability of wet yeast paste, formulation of yeast cells. Green house and field trials of the selected yeast strains on strawberry, corn and fruits. Design experiments to identify genes enhancing biocontrol efficacy Collect & GC/MS infested/infected HPVs, Examine effects; Chemoreception studies. Collect & GC/MS infested/infected HPVs; Bioassay concs, fractions & cmpnds NOW & OFM. Tech. transfer with CRADA partner &continue field trials & develop protocols. Year 3 (2008) Continuation of
corn field trials; continued collection and isolation of bacterial candidates from field samples. Molecular characterization of bacterial biocontrol phenotypes. Test formulation in field trials. Repeat field trials. Continue to isolate yeasts from a variety of crops. Osmotic and heat stress experiments using microarray. Identify genes important for heat and water stress. Continue GC/MS; Field test attraction, oviposition effects. Bioassay cmpnds, Design traps - protocols; Chemoreception interaction studies. Initiate attracticide Tech. transfer, initiate large-scale validation; Chemoreception studies. Year 4 (2009) Continuation of corn field trials. Continuation of molecular characterization of bacterial biocontrol phenotypes; publication of results. 10 acre trials on corn and 2-5 acre trials on pistachio, collect mycotoxin data. Feed back from field trials, develop new types of formulations. Scale-up field trials. Collect efficacy data. Define P. anomala genes of in response to A.
flavus through micoarray gene profiling. Validate behaviors; Develop monitoring throughout CA. Tech. transfer CRADA/ Patent; Initiate MD for NOW, OFM & PTB. Implement attracticide, Tech. Transfer by broad regional demonstration trials. Year 5 (2010) Analysis of corn field trial data; publication of results. Continuation of molecular characterization of bacterial biocontrol phenotypes; publication of results. Test new formulations in field, patent application, tech transfer, Repeat field trials, determine efficacy, patent, Tech transfer, Repeat field trials, determine efficacy, patent, Tech transfer, Summarize data, attempt to use the new knowledge for improving biocontrol efficacy of P. anomala.,. Tech. Transfer and Initiate control development of insects using developed lures. 4a List the single most significant research accomplishment during FY 2006. Discovery of Potential Bacterial Biocontrol Agents. The accomplishment of this research is identified with National Program 108
component 2.1.5 (Biocontrol). Aflatoxin contamination to agricultural crops, such as corn, tree nuts, peanuts and cotton seed, can result in serious economic losses on an annual basis. One approach that shows some promise of success is use of other microbes as biological control agents against aflatoxin producing fungi. In addition, other types of mycotoxin producing fungi, such as those producing fumonisins (Fusarium spp.), may also potentially be controlled with such a strategy. Bacteria from Mississippi corn field soil samples (199 isolates) and corn rhizosphere samples (223 isolates) have been characterized for antifungal phenotypes against Aspergillus flavus, and screened for antifungal phenotypes against Fusarium verticillioides, and some have been speciated using 16s rRNA gene sequence analysis and Biolog carbon utilization analysis. Selected strains have been sent to Dr. Hamed Abbas (USDA-ARS, Stoneville, MS) for field trials against A. flavus and F. verticillioides on corn.
The discovery of any isolates that show promise as biocontrol agents could be fruitful for future biocontrol programs. 4b List other significant research accomplishment(s), if any. The accomplishment of this research is identified with National Program 108 component 2.1.5 (Biocontrol). Aflatoxin contamination is well documented to be associated with infection of wounding by Aspergillus flavus in corn, peanuts, cotton and tree-nuts before harvest. The biocontrol yeast, Pichia anomala WRL-076 has been applied as foliar spray and was exposed to environmental stress such as temperature fluctuations, water availability and UV-irradiation. In laboratory experiments, PEG (polyethylene glycol) 8000 was used to adjust the culture medium aw to 0. 96, which mimicked a water stress condition of 5.62 MPa and P. anomala WRL-076 was found to grow at this low water activity (aw). The yeast cells formed a film and inhibited the growth of A. flavus inoculated to the medium. Field experiments
conducted by Tom Isakeit of Texas A&M University indicated that the application of P. anomala significantly reduced the level of pre-harvest aflatoxin in corn as much as 70% at one location. Experiments were conducted in a commercial pistachio orchard in collaboration with D. E. Parfitt and Brent Holtz, University of California Davis. The data show that sprays of P. anomala WRL-076 reduced the frequency of A. flavus colonization by 4 to 10 times and decreased the total propagules of A. flavus by 80 to 99% on the pistachio nut-fruits. 4d Progress report. This CRIS project was restructured from the old one (5325-42000-031-00D) to add the research on biocontrol of microorganisms that contribute to mycotoxin contamination. The prior project had mainly focused only on insect control. There are no subordinate projects currently associated with the new CRIS project at this time. However, it is foreseen that there will be some in the next year. 5. Describe the major accomplishments to date
and their predicted or actual impact. All accomplishments of this research align with National Program 108 component 2.1.5 (Biocontrol), except where noted. New natural plant lure is new tool for control of codling moth. Codling moths are one of the most destructive agricultural pests to walnut, apple and pear. A natural plant odor rivaling the moth's own pheromones but also attracts female moths is now being used in formulations that include the lure and pesticides. This method of control will only kill the codling moths, does not kill other organisms, including beneficial ones, and reduces risk of people being exposed to pesticides. The chemical identity of natural volatiles from pistachio and walnut was determined. This is the first comparative analysis of the natural volatiles of Pistache (pistachio) and Juglans (walnut) species. The volatiles discovered in nut trees serve as a basis in the search for lures for the navel orangeworm, a major pest of pistachio and codling moth.
Aflatoxin is converted to a carcinogen by enzymes in the human liver. Discovered two compounds, natural constituents of common spices, significantly inhibit this reaction. Eating foods which contain these spices could lower risk of aflatoxin associated cancer of the liver to mycotoxins in general. (component 2.1.1) Larvae of the navel orangeworm (NOW) and codling moth (CM) are commonly found in tree nut kernels highly contaminated with aflatoxin. NOW produced three aflatoxin B1 (AFB1) biotransformation products, chiefly aflatoxicol (AFL), and minor amounts of aflatoxin B2a (AFB2a) and aflatoxin M1 (AFM1). With AFL as a substrate, NOW larvae produced AFB1 and aflatoxicol M1 (AFLM1). Neither NOW nor CM produced AFB1-8,9-epoxide (AFBO), the principal carcinogenic metabolite of AFB1. In comparison, metabolism of AFB1 by chicken liver yielded mainly AFL, whereas mouse liver produced mostly AFM1 at a rate eight-fold greater than AFL. Mouse liver also produced AFBO. This study revealed
the biochemical and evolutionary basis by which these insects can avoid deleterious effects of fungal toxins. (component 2.1.1) 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? The new codling moth lure developed with a CRADA partner has been licensed. The product is now commercially available as lures and traps. Together with the CRADA partner, we are developing monitoring, trapping and attracticide systems using semiochemicals. These systems will be used as a method to control insects with minimal impact to the environment. A new CRADA is under development with a new partner, Paramount Farms, for the development of a lure for navel orangeworm. CRADA Agreement with Trece, Inc., Salinas, CA. Discovery and Development of Host-Plant Volatiles for the
Control of the Codling Moth. August 1999-July 2003. Light DL, Henrick, CA. 2003. Bisexual attractants, aggregants and arrestants for adults and larvae of codling moth and other species of Lepidoptera. US Patent No. US 6,528,049 B2 Issued Mar. 4, 2003. The yeast strain of P. anomala was first proven to be quite effective at controlling A. flavus infection of tree nuts, especially pistachio. Field tests in test pistachio orchards indicate that use of this yeast shows promise. What remains to be determined is how often it needs to be applied, since its viability under field conditions has not been fully determined. This organism may prove to be a useful alternative to using strains of A. flavus, that do not produce aflatoxin, as biocontrol agents and which may find some problems being accepted for use in California. 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). Fruit Perfume Lures Female Codling Moths. 2001. Agricultural Research June 2001. p. 10-12. Controlling Interest. California Farmer. 2002. v. 285 (2). p. 12-18. Using pear ester to monitor codling moth in sex pheromone treated orchards. Oregon State University Extension Bulletin EM8904, February 2006. 10 pp.
Impacts
(N/A)
Publications
- Palumbo, J.D., Baker, J.L., Mahoney, N.E. 2006. Isolation of bacterial antagonists of aspergillus flavus from almonds. Microbial Ecology. DOI:10. 1007/s00248-006-9096-y. URL http://dx.doi.org/10.1007/s00248-006-9096-y.
- Chang, P., Ehrlich, K., Hua, S.T. 2006. Cladal relatedness among Aspergillus oryzae isolates and Aspergillus flavus S and L morphotype isolates. International Journal of Food Microbiology. 108:172-177.
- Knight, A.L., Light, D.M. 2005. Factors affecting the differential capture of male and female codling moth (Lepidoptera: Tortricidae) in traps baited with ethyl (E,Z)-2,4-Decadienoate. Environmental Entomology. 34(5):1161- 1169.
- Knight, A.L., Light, D.M. 2005. Seasonal flight patterns of codling moth (Lepidoptera: Tortricidae) monitored with pear ester and codlemone-baited traps in sex pheromone-treated apple orchards. Environmental Entomology. 34(5):1028-1035.
- Knight, A.L., Light, D.M. 2005. Timing of egg hatch by early-season codling moth (Lepidoptera: Tortricidae) predicted by moth catch in pear ester and codlemone-baited traps. Can. Entomol. 137(6):728-738.
- Knight, A.L., Light, D.M. 2005. Developing action thresholds for codling moth (Lepidoptera: Tortricidae) with pear ester and codlemone-baited traps in apple orchards treated with sex pheromone mating disruption. Can. Entomol. 137(6):739-747.
- Hua, S. S. 2006. Progress in prevention of aflatoxin contamination in food by preharvest application of a yeast strain, pichia anomala wrl-07. In: Mendez-Vilas, A., editor. Modern Multidisciplinary Applied Microbiology: Exploiting Microbes and Their Interactions. Weinheim, Germany: Wiley-VCH Verlag. p. 323-327.
- Hua, S.T., Mcalpin, C.E., Ly, S.B. 2006. Population of aspergillus flavus on pistachio buds and flowers. In: Mendez-Vilas, A., editor. Modern Multidisciplinary Applied Microbiology: Exploiting Microbes and Their Interactions. Weinheim, Germany:Wiley-VCH Verlag. p.440-445.
- Knight, A.L., Hilton, R., Vanbuskirk, P., Light, D.M. 2006. Using pear ester to monitor codling moth in sex pheromone treated orchards. Oregon State University Agricultural Experiment Station. Oregon State University, Extension Service Publication EM 8904, February 2006. 8 p.
|
|