Progress 12/29/01 to 12/13/05
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? The coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae) is a serious pest of coffee throughout the world. Endemic to Central Africa, it has now spread to most coffee growing regions in the world. The female adult bores a hole into the coffee berry where she deposits 20- 50 eggs; larvae feed on the seed, thereby reducing yields and/or lowering the quality of the final product. In the past, growers had relied on the use of pesticides, but resistance has been reported. The objective of our research program is to develop innovative biological control methods to manage the coffee berry borer. Approaches include the inoculation of coffee plants with fungal insect pathogens in an attempt to provide systemic control of the insect. All research findings will be of use to
coffee scientists, mycologists, entomologists, and the coffee industry. This research falls within National Program 304 (Crop and Commodity Pest Biology, Control and Quarantine). The proposed research addresses the following program components (1) identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. 2. List by year the currently approved milestones (indicators of research progress) Year 1 (FY2001) - Establishment of Beauveria bassiana in coffee plants through radicle inoculations; coffee endophytes sequencing completed. - Development of B. bassiana-specific markers for detection in planta. Year 2 (FY2002) - Spraying of B. bassiana on coffee flowers in the field. -
Foreign exploration for natural enemies. - Molecular detection of B. bassiana in sprayed plants. Year 3 (FY2003) - Sampling of coffee berries in different countries for B. bassiana detection. - Molecular detection of B. bassiana in berries from different countries. Year 4 (FY2004) - Optimization of B. bassiana inoculation methods for its establishment as a coffee endophyte. - Molecular detection of B. bassiana in inoculated seedlings. Year 5 (FY2005) - Optimization of B. bassiana inoculation methods for its establishment as a coffee endophyte. - Foreign exploration for natural enemies. 4a List the single most significant research accomplishment during FY 2006. Coffee Endophytes surveyed: Our laboratory research has shown that it is possible to introduce into coffee seedlings a fungus (Beauveria bassiana) that is pathogenic to the coffee berry borer. In order to determine whether other fungi known as fungal endophytes - occur inside healthy coffee plants in the field, we conducted a
survey of coffee leaves and berries in Colombia, Hawaii, Mexico, and Puerto Rico. The survey yielded over 700 fungal endophytes. The research addresses the following program components: (1) identification and classification of agriculturally and environmentally important insects and microbes and (2) development of new and improved pest control technologies NP304 component 5. 4b List other significant research accomplishment(s), if any. Beauveria bassiana A Beauveria-specific primer pair to the nuclear ribosomal internal transcribed spacer region for detection of B. bassiana in planta was developed for conventional and real-time PCR. The research addresses the following program components: (1) development of new and improved pest control technologies. NP304 component 5. 4d Progress report. Subordinate Projects see attached files. 5. Describe the major accomplishments to date and their predicted or actual impact. The research addresses the following program components (1)
identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. All the followiong findings will be of use to coffee scientists, mycologists, entomologists, and the coffee industry: (1) As part of the investigations aimed at understanding the contribution of microorganisms to the coffee berry borer's ability to exploit the coffee bean as a food source, two yeasts were isolated from the insect and molecularly identified as Pichia burtonii and Pichia guilliermondii. The yeasts seem to be involved in the production of degradative enzymes used in food utilization, but do not play a role in caffeine breakdown. (2) A sampling for natural enemies of the insect was conducted in Mexico
in collaborative research with investigators at El Colegio de la Frontera Sur (ECOSUR) in Tapachula and resulted in the discovery of a new nematode species that reduced the fecundity and increased the mortality of the coffee berry borer. In a collaborative research project Oregon State University, the new nematode species was described as Metaparasitylenchus hypothenemi. This is the first nematode species found in the Americas as a parasite of the coffee berry borer, and could potentially become an important coffee berry borer biocontrol agent. (3) Also in collaboration with ECOSUR (Mexico) and ARS-Peoria a new species of Penicillium, described as P. brocae, was discovered in coffee berry borers from Mexico. This fungus might be involved in a mutualistic association with the insect, where the fungus provides nutrients needed for insect survival. (4) In collaboration with investigators at ARS-Peoria and the Hawaii Agricultural Research Center, a new endophytic species isolated from a
coffee plant in Hawaii was described as Penicillium coffeae. The endophytic role for this fungal species remains unknown. (5) In collaboration with Cenicafe (Colombia) and Purdue University, a phylogenetic study was conducted on coffee berry borers from 17 countries on three continents; the results revealed that only one species is present. This implies that an effective control strategy should work wherever the insect is present. (6) Twenty-two genera of endophytic bacteria were isolated from adult coffee plant leaves, various parts of the berry, and leaves, stems and roots of seedlings. The highest number of bacteria among the berry tissues sampled was isolated from the seed, and includes Bacillus, Burkholderia, Clavibacter, Curtobacterium, Escherichia, Micrococcus, Pantoea, Pseudomonas, Serratia, and Stenotrophomonas. This is the first survey of the endophytic bacteria diversity in various coffee tissues, and the first study reporting endophytic bacteria in coffee seeds. The
possible role for these bacteria in the biology of the coffee plant remains unknown. (7) Fifty Beauveria bassiana strains isolated from the coffee berry borer were used to develop a novel screening method aimed at selecting strains with the highest biocontrol potential. The screening method is based on percent insect mortality, average survival time, mortality distribution, percent spore germination, fungal life cycle duration, and spore production on the insect. Based on these parameters, only 11 strains merit further study. The use of a sound scientific protocol for the selection of promising fungal entomopathogens should lead to more efficient use of time, labor, and financial resources in biological control programs. (8) In collaboration with scientists at Rutgers University and ARS-Peoria, several endophytic Penicillium species were identified in coffee, including some that tested positive for ochratoxin A; (9) A coffee berry borer parasitoid, Prorops nasuta, was found to be
carrying the fungus Aspergillus westerdijkiae which tested positive for ochratoxin A, thus indicating that the parasitoid might be a vector of an ochratoxin- producing fungus. All these findings have increased the knowledge on the basic biology of the coffee berry borer and its associated organisms and will be helpful in developing biological control programs against the insect. 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? No technologies have yet been transferred. 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). Articles in Popular Press: None. Presentations: Vega, F. Coffee Origins: the Trail of Coffee
before the Roast - National Coffee Association Fall Educational Conference, Brooklyn, NY, November 3-5, 2005. Vega, F. An Introduction to Coffee Agronomy Annual Meeting of the Specialty Coffee Association of America, Charlotte, NC, April 7-10, 2006. Vega, F. Coffee Endophytes Annual Meeting of the Mycological Society of America, Quebec City, Canada, July 29 August 2, 2006. Rehner, S. Deconstructing the natural history of the entomopathogen Beauveria bassiana- Invited presentation to the Department of Plant Pathology, The Pennsylvania State University, State College, PA, May 1, 2006. Articles written about research: Gant, J. 2006. La broca del grano de cafe or You got more (or less) than ground in your coffee, pal. The Specialty Coffee Chronicle, May/June 2006, p. 18-21. Milius, S., 2006. Theyre all part fungus. Science News 169:231-232.
Impacts
(N/A)
Publications
- Peterson, S.W., Vega, F.E., Nagai, C. 2004. Penicillium coffeae, a new endophytic species isolated from a coffee plant and its phylogenetic relationship to P. fellutanum, P. thiersii and P. brocae based on parsimony analysis of multilocus DNA sequences. Mycologia. 97(3): 659-666.
- Vega, F.E., Pava-Ripoll, M., Buyer, J.S. 2005. Endophytic bacteria in coffea arabica l.. Journal of Basic Microbiology. 45:371-380.
- Posada, F.J., Vega, F.E. 2006. Inoculation of cocoa seedlings (theobroma cacao) with the fungal entomopathogen beauveria bassiana (ascomycota: hypocreales) and its establishment as an endophyte.. Mycologia. 97:208- 213.
- Vega, F.E., Posada, F.J., Gianfagna, T.J., Chaves, F.C., Peterson, S.W. 2006. An insect parasitoid carrying an ochratoxin producing fungus.. Naturwissenschaften. 93:297-299.
- Vega, F.E., Posada, F.J., Peterson, S.W., Gianfagna, T.J., Chaves, F. 2006. Penicillium species endophytic in coffee plants and ochratoxin a production.. Mycologia. Vol.98 pg. 31-42
- Posada, F., Vega, F.E. 2005. A new method to evaluate the biocontrol potential of single spore isolates of fungal entomopathogens.. Journal of Insect Science. 5:37.
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Progress 10/01/04 to 09/30/05
Outputs
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? The coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae) is a serious pest of coffee throughout the world. Endemic to Central Africa, it has now spread to most coffee growing regions in the world. The female adult bores a hole into the coffee berry where she deposits 20- 50 eggs; larvae feed on the seed, thereby reducing yields and/or lowering the quality of the final product. In the past, growers had relied on the use of pesticides, but resistance has been reported. The objective of our research program is to develop innovative biological control methods to manage the coffee berry borer. Approaches include the inoculation of coffee plants with fungal insect pathogens in an attempt to provide systemic control of the insect. This research falls within National Program 304 (Crop and
Commodity Pest Biology, Control and Quarantine). The proposed research addresses the following program components (1) identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. 2. List the milestones (indicators of progress) from your Project Plan. This project will terminate this year and a new project has not yet been approved via the OSQR process. The objectives listed in the current project statement are: A) Investigate the contribution of microorganisms to the insects' ability to exploit the coffee bean as a food source; B) Develop innovative biological control methods aimed at the coffee berry borer. 3a List the milestones that were scheduled to be addressed in FY
2005. For each milestone, indicate the status: fully met, substantially met, or not met. If not met, why. 1. Develop innovative biological control methods aimed at the coffee berry borer. Milestone Fully Met 3b List the milestones that you expect to address over the next 3 years (FY 2006, 2007, and 2008). What do you expect to accomplish, year by year, over the next 3 years under each milestone? There are no milestones for future years. 4a What was the single most significant accomplishment this past year? Beauveria bassiana as a coffee endophyte The coffee berry borer is the most serious pest of coffee throughout the world, and methods to control this insect are hard to achieve due to the insect spending most of its life cycle inside coffee berries. We have developed methodologies that are effective in introducing a naturally occurring fungal pathogen of the insect (Beauveria bassiana) inside coffee plants, with the goal of controlling the insect. This new methodology, if
proven to be effective against the coffee berry borer in the field, would provide a new mechanism against this insect not dependent on chemical insecticides, thus allowing small growers throughout the world to protect their crops against the coffee berry borer in a sustainable and environmentally friendly manner. 4b List other significant accomplishments, if any. Beauveria bassiana Novel molecular markers, targeting intergenic regions of nuclear DNA, were developed for investigation of genetic relationships in B. bassiana. In an analysis of African and Neotropical B. bassiana pathogens infecting coffee berry borer, other co-occurring insects and endophytic strains of B. bassiana from coffee, it was determined that African and Neotropical isolates from coffee berry borer share recent common ancestry, although the geographic origin of the lineage, whether African or Neotropical is not altogether clear. However, in the Neotropics, this group occurs throughout Central and South America
where it also was isolated from other insect taxa. Additionally, the genetic variation in the Neotropical group was much greater than that in Africa, indicating that the group has been established in this region for some time. Endophytic strains of B. bassiana from coffee are intermixed with coffee berry borer pathogens indicating that pathogens may alternate as endophytes, which suggests that coffee plants may serve as reservoirs of pathogens against their insect pests. 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. (1) As part of the investigations aimed at understanding the contribution of microorganisms to the coffee berry borer's ability to exploit the coffee bean as a food source, two yeasts were isolated from the insect and molecularly identified as Pichia burtonii and Pichia guilliermondii. The yeasts seem to be involved in the production of degradative enzymes used in food utilization, but do not play a role in
caffeine breakdown. (2) A sampling for natural enemies of the insect was conducted in Mexico in collaborative research with investigators at El Colegio de la Frontera Sur (ECOSUR) in Tapachula and resulted in the discovery of a new nematode species that reduced the fecundity and increased the mortality of the coffee berry borer. In a collaborative research project Oregon State University, the new nematode species was described as Metaparasitylenchus hypothenemi. This is the first nematode species found in the Americas as a parasite of the coffee berry borer, and could potentially become an important coffee berry borer biocontrol agent. (3) Also in collaboration with ECOSUR, Tapachula, Mexico and the ARS laboratory in Peoria, IL, a new species of Penicillium, described as P. brocae, was discovered in coffee berry borers from Mexico. This fungus might be involved in a mutualistic association with the insect, where the fungus provides nutrients needed for insect survival. (4) In
collaboration with investigators at ARS, Peoria and HARC, Hawaii, a new endophytic species isolated from a coffee plant in Hawaii was described as Penicillium coffeae. The endophytic role for this fungal species remains unknown. (5) In collaboration with Cenicafe, Colombia and Purdue University, a phylogenetic study was conducted on coffee berry borers from 17 countries on three continents; the results revealed that only one species is present. This implies that an effective control strategy should work wherever the insect is present. (6) Twenty-two genera of endophytic bacteria were isolated from adult coffee plant leaves, various parts of the berry, and leaves, stems and roots of seedlings. The highest number of bacteria among the berry tissues sampled was isolated from the seed, and includes Bacillus, Burkholderia, Clavibacter, Curtobacterium, Escherichia, Micrococcus, Pantoea, Pseudomonas, Serratia, and Stenotrophomonas. This is the first survey of the endophytic bacteria
diversity in various coffee tissues, and the first study reporting endophytic bacteria in coffee seeds. The possible role for these bacteria in the biology of the coffee plant remains unknown. (7) Fifty Beauveria bassiana strains isolated from the coffee berry borer were used to develop a novel screening method aimed at selecting strains with the highest biocontrol potential. The screening method is based on percent insect mortality, average survival time, mortality distribution, per-cent spore germination, fungal life cycle duration, and spore production on the insect. Based on these parameters, only 11 strains merit further study. The use of a sound scientific protocol for the selection of promising fungal entomopathogens should lead to more efficient use of time, labor, and financial resources in biological control programs. These findings have increased the knowledge on the basic biology of the coffee berry borer and its associated organisms and will be helpful in developing
biological control programs against the insect. 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? No technologies have yet been transferred. 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). Articles in Popular Press: None. Presentations: Posada, F., and F. E. Vega. Inoculation of coffee plants with Beauveria bassiana, Annual Meeting of the Entomological Society of America, Salt Lake City, Utah (November 2004). Vega, F. E. Coffee research at USDA. Graduate Biosciences Society, University of Virginia (January 2005). Rehner, S. A. The potential for and evidence of sexual reproduction in Beauveria bassiana, a mitotic
insect pathogenic fungus. Society for Fungal Genetics, Asilomar, California (March 2005). Vega, F. E. Biological control of the coffee berry borer. Central American Coffee Quality Initiative: Agronomy Group. CATIE, Turrialba, Costa Rica (April 2005). Rehner, S. A. Cryptic speciation and the potential and evidence for sex in the insect pathogen, Beauveria bassiana. Department of Plant Pathology, Cornell University (April 2005). Posada, F., and F. E. Vega. Sustainable biocontrol? The fungus Beauveria bassiana as a coffee endophyte to control the coffee berry borer. BARC Poster Day, Beltsville, MD (May 2005). Vega, F. E. Biological control of insect pests. Imagine Excellence Program, BARC, USDA, ARS, Beltsville, MD (July 2005). Peterson, S. W., F. E. Vega, and F. Posada. Ochratoxin-A production and DNA based identification of an Aspergillus species isolated from the parasitoid wasp Prorops nasuta in coffee berries. International Union of Microbiological Societies Annual
Meeting, San Francisco, California (July 2005). Articles written about research: S. Durham. Stopping the coffee berry borer from boring into profits. Agricultural Research, November 2004, pp. 10-11. Research summary written for ARS News Service by Sharon Durham. Picked- up by: Info Plagas (Argentina), Consuma Seguridad (Spain), The New Farm (The Rodale Institute), Business Line (India), U. S. Department of State (Global Issues website, International Information Programs website, Embassy of the United States in Japan website).
Impacts
(N/A)
Publications
- Ortiz, A., Ortiz, A., Vega, F.E., Posada, F.J. 2004. Volatile composition of coffee berries at different stages of ripeness, and their possible attraction to the coffee berry borer hypothenemus hampei (curculionidae: scolytinae).. Journal of Agricultural and Food Chemistry. 52, 5914-5918
- Vega, F.E. 2004. The coffe berry borer, hypothenemu, hampei (ferrari) (coleoptera:scolytidae). Encyclopedia of Entomology 1: 575-576.
- Vega, F.E., Dowd, P.F. 2005. The role of yeasts as insect endosymbionts. In: Vega, F.E. Blackwell, M., editors. Insect-Fungal Assocations: Ecology and Evolution. Oxford University Press, New York, p.211-243
- Poinar, G.O., Vega, F.E., Castillo, A., Chavez, I.E., Infante, F. 2004. Metaparasitylenchus hypothenemi sp. n. (allantonematidae: nematoda), a parasite of the coffee berry borer, hypothenemus hampei (ferrari) (curculionidae: scolytinae).. Journal of Parasitology. 90:1106-1110
- Vega, F.E., Blackwell, M. 2005. Conclusion: symbioses,biocomplexity, and metagenomes.. Oxford University Press. pp293-295
- Perez, J., Infante, F., Vega, F.E. 2005. Does the coffee berry borer (coleoptera: scolytidae) have mutualistic fungi?. Annals of the Entomological Society of America. 98:483-490
- Rehner, S.A. 2005. Phylogenetics of the insect pathogenic genus beauveria. Insect-Plant Relationships International Symposium Proceedings, pp. 3-27.
- Castillo, A., Infante, F., Guillermo, L., Trujillo, J., Kirkendall, L., Vega, F.E. 2004. Laboratory parasitism by phymastichus coffea upon non- target bark beetles associated with coffee plantations. Florida Entomologist 87: 272-277.
- Blackwell, M., Vega, F.E. 2005. Introduction: seven wonders of the insect- fungus world. Oxford University Press, pp. xiii-xvii.
- Benavides, P., Vega, F.E., Severson, J.R., Bustillo, A.E., Stuart, J.J. 2005. Biodiversity and biogeography of an important inbred pest of coffee, the coffee berry borer, hypothenemus hampei (ferrai).. Annals of the Entomological Society of America. 98:359-366
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Progress 10/01/03 to 09/30/04
Outputs
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? The coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae) is a serious pest of coffee throughout the world. Endemic to Central Africa, it has now spread to most coffee growing regions in the world. The female adult bores a hole into the coffee berry where she deposits 20- 50 eggs; larvae feed on the seed, thereby reducing yields and/or lowering the quality of the final product. In the past, growers had relied on the use of endosulfan, but resistance has now been reported. The objective of our research program is to develop innovative biological control methods to manage the coffee berry borer. Approaches include the inoculation of coffee plants with fungal insect pathogens in an attempt to provide systemic control of the insect. This research falls within National Program 304 (Crop
and Commodity Pest Biology, Control and Quarantine). The proposed research addresses the following program components (1) identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. 2. List the milestones (indicators of progress) from your Project Plan. This project will terminate this year and a new project has not yet been submitted via the OSQR process. Research objectives in the current Project Plan are (a) to investigate the contribution of microorganisms to the insects' ability to exploit the coffee bean as a food source, and (b) to develop innovative biological control methods aimed at the coffee berry borer. 3. Milestones: A. The milestones listed below were
scheduled to be completed in 2004. All milestones were completed. Inoculation and detection of coffee seedlings with virulent fungi selected in bioassays. B. A new Project Plan with milestones has not yet been submitted via the OSQR process (current CRIS expires on August 31, 2005). However, anticipated achievements for the next three years are: 1. Optimize inoculation of coffee plants with the fungal entomopathogen Beauveria bassiana in order to introduce it as a fungal endophyte for systemic control of the coffee berry borer. 2. Conduct a search for parasitoids, fungi, and nematodes of the coffee berry borer in various countries in the Americas. 3. Determine the role of the bacterium Wolbachia in the coffee berry borer and rear the bacterium in coffee berry borer cell lines. 4. Determine mechanism(s) of genetic recombination among B. bassiana strains that parasitize the coffee berry borer. 4. What were the most significant accomplishments this past year? A. Single most significant
accomplishment during FY 2003: Beauveria bassiana as a coffee endophyte The coffee berry borer is the most serious pest of coffee throughout the world, and methods to control this insect are hard to achieve due to the insect spending most of its life cycle inside coffee berries. We have developed methodologies that are effective in introducing a naturally occurring fungal pathogen of the insect (Beauveria bassiana) inside coffee plants, with the goal of controlling the insect. This new methodology, if proven to be effective against the coffee berry borer in the field, would provide a new mechanism against this insect not dependent on chemical insecticides, thus allowing small growers throughout the world to protect their crops against the coffee berry borer in a sustainable and environmentally friendly manner. B. Other Significant Accomplishment(s), if any: Coffee endophytes An enormous diversity of fungi has been discovered living inside coffee tissues, including the seeds. The
role that these fungi might be playing in the coffee plant is not known, but many of the genera that have been identified are known to produce metabolites that could be detrimental to human health. We are studying the possibility that some of these toxigenic fungi might already be 'embedded' in the plant, challenging present concepts involving post-harvest storage as the main focus for infection with these fungi. Beauveria bassiana To determine whether genetic exchange occurs naturally among individuals of the entomopathogenic fungus Beauveria bassiana, gene sequencing approaches were developed that facilitate species discovery and identification. Additionally, 11 highly variable microsatellite markers were developed for analyzing patterns of reproduction within these species. Gene sequencing revealed significant species diversity within B. bassiana, and several species are pathogens of coffee berry borer. Results of analyses of microsatellite variation indicate that B. bassiana is
capable of genetic recombination, although the precise mechanism of recombination is presently uncertain. These results suggest that release of B. bassiana for biocontrol of pest insects such as the coffee berry borer may impact conspecific indigenous populations of these fungi. Further investigation of the frequency of recombination is needed to assess the likelihood and potential risks of genetic exchange between applied and native B. bassiana. C. Significant Activities that Support Special Target Populations. The project does not support special target populations. D. Progress Report for Subordinate Projects. 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. 1) As part of the investigations aimed at understanding the contribution of microorganisms to the coffee berry borer's ability to exploit the coffee bean as a food source, two yeasts, were isolated from the insect and molecularly identified as Pichia burtonii and
Pichia guilliermondii. The yeasts seem to be involved in the production of degradative enzymes used in food utilization, but do not play a role in caffeine breakdown. 2) A sampling for natural enemies of the insect was conducted in Mexico in collaborative research with F. Infante, J. Perez, and A. Castillo at El Colegio de la Frontera Sur in Tapachula and resulted in the discovery of a new nematode species that reduced the fecundity and increased the mortality of the coffee berry borer. In a collaborative research project established with L. Carta (Nematology Laboratory, USDA, ARS, Beltsville, MD), the new nematode species was placed in the genus Sphaerulariopsis. This is the first nematode species found in the Americas as a parasite of the coffee berry borer, and could potentially become an important coffee berry borer biocontrol agent. 3) Also in collaboration with the Mexican scientists and S. Peterson at the ARS laboratory in Peoria, IL, a new species of Penicillium (P. brocae)
was discovered in coffee berry borers from Mexico. This fungus might be involved in a mutualistic association with the insect, where the fungus provides nutrients needed for insect survival. These findings have increased the knowledge on the basic biology of the insect and its associated organisms and will be helpful in developing biological control programs against the coffee berry borer. 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? No technologies have yet been transferred. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. Articles in Popular Press: None
Impacts
(N/A)
Publications
- Posada, F.J., Vega, F.E., Rehner, S.A., Blackwell, M., Weber, D.C., Suh, S. , Humber, R.A. 2004. Syspastospora parasitica on beauveria bassiana attacking the colorado potato beetle.. Journal of Insect Science. Vol.#4, Pg. 24
- VEGA, F.E., ROSENQUIST, E., COLLINS, W.W. THE GLOBAL COFFEE CRISIS AND ITS EFFECT ON COFFEE RESEARCH.. SCIENCE MAGAZINE. 2003.
- Perez, J., Infante, F., Vega, F.E., Holguin, F., Macias, J., Valle, J., Nieto, G., Peterson, S.W., Kurtzman, C.P., O Donnell, K. 2003. Mycobiota associated with the coffee berry borer hypothenemus hampei (ferrari) (coleoptera: scolytidae) in chiapas, mexico. Mycological Research.
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Progress 10/01/02 to 09/30/03
Outputs
1. What major problem or issue is being resolved and how are you resolving it? The coffee berry borer (Hypothenemus hampei (Ferrari); Coleoptera: Scolytidae) is the most serious pest of coffee throughout the world. Endemic to Central Africa, it has spread to most coffee growing regions in the world. The female adult bores a hole into the coffee berry where she deposits 20-50 eggs; larvae feed on the seed, thereby reducing yields and/or causing abscission of the fruit. The objective of our research program is to develop innovative biological control methods to manage the coffee berry borer. Approaches include the inoculation of coffee plants with fungal insect pathogens, and research on bacteria, nematodes and fungal endophytes in an attempt to provide systemic control of the insect. 2. How serious is the problem? Why does it matter? As an example of the seriousness of the coffee berry borer as an insect pest, we can use Colombia as a case study. The insect was
first reported in Colombia in 1988, and has now spread to more than 600,000 hectares of Colombian coffee. Yield losses of up to 60% have been reported, and the resulting low yields have prompted the abandonment of coffee plantations in many regions. The situation is similar wherever the insect is found. As more that 100 million people throughout the world depend on coffee for their subsistence, the coffee berry borer can have a direct impact on their well being. Use of effective biological control agents should counter reduced coffee yields, threatened survival of coffee plantations, and detrimental effects on the livelihood of small land holders that grow coffee. 3. How does it relate to the National Program(s) and National Program Component(s) to which it has been assigned? The coffee berry borer research program falls within National Program 304 (Crop and Commodity Pest Biology, Control and Quarantine). The proposed research addresses the following program components (1)
identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. To achieve our objectives, collaborative projects have been established with "El Colegio de la Frontera Sur" in Mexico (F. Infante), the "Centro Nacional de Investigaciones de Cafe" in Colombia (G. Cadena), Purdue University (J. Stuart), the Milwaukee Public Museum (A. Young), and Rutgers University (T. Gianfagna). In addition, collaborative research is routinely conducted with ARS scientists in Peoria (C. Kurtzman, S. Peterson, P. Dowd) and Beltsville (S. Rehner, L. Carta, J. Buyer). 4. What were the most significant accomplishments this past year? A. Single most significant accomplishment during FY 2003: There
is an urgent need to develop cost efficient and biologically safe pest management methods aimed at the coffee berry borer, such as establishing the fungal insect pathogen Beauveria bassiana within coffee plants to provide systemic protection against the coffee berry borer. Sampling for microorganisms present inside coffee plant tissues was done in Hawaii, Colombia, Mexico, and Puerto Rico. Yeasts, fungi and bacteria were found in the sampled tissues. These results will provide a better understanding of the myriad organisms co-occurring within coffee plants and will increase the chances of success in inoculating the fungal insect pathogen B. bassiana into coffee. B. Other Significant Accomplishment(s), if any: There is a strong need in the field of insect pathology to develop techniques aimed at differentiating among strains of fungal insect pathogens isolated from diseased insects. Various molecular techniques were used to determine if they could differentiate among Beauveria
bassiana strains isolated from coffee berry borers collected in Ivory Coast, Togo, Brazil, Colombia, El Salvador, and Mexico. Nuclear genes revealed that all the B. bassiana strains are very closely related, but the use of microsatellites, which are more sensitive genetic markers than nuclear genes, show that the African and American Beauveria groups are genetically distinct from one another. These molecular techniques will allow for the differentiation among fungal insect pathogen strains, thus reducing the number of strains that need to be tested in evaluation of efficacy bioassays and for the monitoring of strains once they are released in the field. C. 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. 1) As part of the investigations aimed at understanding the contribution of microorganisms to the coffee berry borer's ability to exploit the coffee
bean as a food source, two yeasts, were isolated from the insect and molecularly identified as Pichia burtonii and Candida fermentati. The yeasts seem to be involved in the production of degradative enzymes used in food utilization, but do not play a role in caffeine breakdown. 2) A sampling for natural enemies of the insect was conducted in Mexico in collaborative research with F. Infante, J. Perez, and A. Castillo at El Colegio de la Frontera Sur in Tapachula and resulted in the discovery of a new nematode species that reduced the fecundity and increased the mortality of the coffee berry borer. In a collaborative research project established with L. Carta (Nematology Laboratory, USDA, ARS, Beltsville, MD), the new nematode species is being described. This is the first nematode species found in the Americas as a parasite of the coffee berry borer, and could potentially become an important coffee berry borer biocontrol agent. 3) Also in collaboration with the Mexican scientists and
S. Peterson at the ARS laboratory in Peoria, IL, a new species of Penicillium (P. brocae) was discovered in coffee berry borers from Mexico. This fungus might be involved in a mutualistic association with the insect, where the fungus provides nutrients needed for insect survival. 4) New guidelines have been established for selecting potential fungal entomopathogens in a bioassay experiment testing 50 B. bassiana strains against the coffee berry borer. Traditional methods have relied on concentrations that kill 50% of the population. Our assessment has been based on mortality distribution, average survival time, spore germination, spore production, and life cycle in the insect. 5) Collaborative research with P. Benavides and J. Stuart (Purdue University) has revealed there is just one coffee berry borer species throughout the world, after sampling insects from 17 countries. These findings are directly related to the objectives in National Program 304: identification and
classification of agriculturally and environmentally important insects and microbes; investigation of the biology of pest and beneficial organisms; understanding pest/host interaction and economic or environmental impact; development of new and improved pest control technologies; and integration of component technologies into IPM systems. 6. What do you expect to accomplish, year by year, over the next 3 years? Year 1. Initiate field and laboratory inoculation experiments aimed at establishing Beauveria bassiana as a fungal endophyte. Continue coffee endophytes molecular identification. Year 2. Continue endophyte research. Develop microarrays to conduct comparative analyses of gene expression among strains of Beauveria that differ in virulence against coffee berry borer. Year 3. Continue research initiated in years 1 and 2. 8. List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: This does not
replace your peer-reviewed publications listed below). Presentations: Vega, F. E. "Coffee at origin: from seed to boat," National Coffee Association Fall Educational Conference, New York City (October 2002). Benavides, P., Stuart, J., Vega, F. E., Romero-Severson, J., and Bustillo, A. "Biodiversity and biogeography of the coffee berry borer, Hypothenemus hampei." The Annual meeting of the Entomological Society of America, Ft. Lauderdale, November 17-20, 2002. (Poster presentation). Vega, F. E. "Coffee endophytes." The Second Cacao Endophyte Working Group meeting, Rutgers University, April 14-15, 2003.
Impacts
(N/A)
Publications
- Dowd, P. F., Vega, F. E. Autodissemination of Beauveria bassiana by sap beetles (Coleoptera: Nitidulidae) to overwintering sites. Biocontrol Science and Technology. 2003. v. 15 p. 69-79.
- Vega, F. E., Jackson, M. A., Mercadier, G., Poprawski, T. The impact of nutrition on spore yields for various fungal entomopathogens in liquid culture. World Journal of Microbiology and Biotechnology. 2003. v. 19 p. 363-368.
- Vega, F. E., Blackburn, M. B., Kurtzman, C. P., Dowd, P. F. Identification of a coffee berry borer associated yeast: doest it break down caffeine? Entomologia Experimentalis et Applicata. 2003. v. 107 p. 19-24.
- Peterson, S. W., Perez, J., Vega, F. E., Infante, F. Penicillium brocae, a new species associated with coffee berry borer in Chiapas, Mexico. Mycologia. 2003. v. 95 p. 141-147.
- Posada, F., Vega, F. E., Rehner, S. A. Beauveria as a posible coffee endophyte. Society for Invertebrate Pathology. 2003. p. 47.
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Progress 10/01/01 to 09/30/02
Outputs
1. What major problem or issue is being resolved and how are you resolving it? The coffee berry borer (Hypothenemus hampei; Coleoptera: Scolytidae) is a serious pest of coffee throughout the world. Endemic to Central Africa, it has now spread to most coffee growing regions in the world. The female adult bores a hole into the coffee berry where she deposits 20-50 eggs; larvae feed on the seed, thereby reducing yields and/or lowering the quality of the final product. In the past, growers had relied on the use of endosulfan, but resistance has now been reported. The objective of our research program is to develop innovative biological control methods to manage the coffee berry borer. Approaches include the inoculation of coffee plants with fungal insect pathogens, bacteria or nematodes in an attempt to provide systemic control of the insect. 2. How serious is the problem? Why does it matter? As an example of the seriousness of the coffee berry borer as an insect
pest, we can use Colombia as a case study. The insect was first reported in Colombia in 1988, and has now spread to more than 600,000 hectares of Colombian coffee. Yield losses of up to 60% have been reported, and the resulting low yields have prompted the abandonment of coffee plantations in many regions. Approximately 250,000-300,000 hectares of coffee are expected to be abandoned. The insect has developed resistance to endosulfan, the most commonly used insecticide to control the insect. Use of endosulfan has also resulted in more than 100 poisonings per year in Colombia, and several deaths. Thus, it is clear that the insect has important implications on the socioeconomic conditions of Colombia. The Colombia example also applies to most countries where the insect is present. Use of effective biological control agents should counter reduced coffee yields, threatened survival of coffee plantations, and detrimental effects on the livelihood of small land holders that grow
coffee. 3. How does it relate to the national Program(s) and National Program Component(s) to which it has been assigned? This research falls within National Program 304 (Crop and Commodity Pest Biology, Control and Quarantine). The proposed research addresses the following program components (1) identification and classification of agriculturally and environmentally important insects and microbes; (2) investigation of the biology of pest and beneficial organisms; (3) understanding pest/host interaction and economic or environmental impact; (4) development of new and improved pest control technologies; and (5) integration of component technologies into IPM systems. 4. What was your most significant accomplishment this past year? A. Single most significant accomplishment during FY 2002: In order to determine whether a bacterium known to cause a high number of females in insect populations was present in the coffee berry borer, we used molecular techniques designed to detect this
bacterium. Scientists of the IBL at BARC, in collaboration with S. O'Neill of Yale University who initiated the project, and P. Benavides and J. Stuart (Purdue University) who did the Wolbachia cloning and sequencing, detected the bacterium in coffee berry borers from 16 countries. This finding increases the knowledge on the basic biology of the most important pest of coffee throughout the world and provides an explanation for the skewed sex ratio observed in insect populations. Elimination of Wolbachia from field populations could result in lower number of females being produced and thus, reduced population growth. B. Other Significant Accomplishment(s), if any: To understand the genetic diversity among strains of the fungal entomopathogen Beauveria bassiana that attack the coffee berry borer, more than 60 B. bassiana strains obtained from coffee berry borers from Africa (Ivory Coast, Togo), South America (Brazil, Colombia), Central America (El Salvador, Mexico), and Asia (India)
were genetically analyzed. These strains were sequenced by S. Rehner of the Insect Biocontrol Laboratory at BARC using two distinct regions in the genome, and were found to cluster the Beauveria strains into two groups with very minor genetic differences. We hypothesize that the close genetic similarity among the pathogenic strains on different continents is because these fungal strains may have originated in Central Africa and have co dispersed with the insect during the global transport of coffee from its ancestral range in Africa. This information reveals a markedly narrow genetic diversity of a common insect pathogen attacking coffee berry borers throughout the world and suggests that there might be a narrow range of B. bassiana strains that can be effectively used in biological control programs aimed against the insect in the field. C. Significant Activities that Support Special Target Populations. None 5. Describe your major accomplishments over the life of the project,
including their predicted or actual impact? 1) As part of the investigations aimed at understanding the contribution of microorganisms to the coffee berry borer's ability to exploit the coffee bean as a food source, two yeasts, were isolated from the insect and molecularly identified as Pichia burtonii and Pichia guilliermondii. The yeasts seem to be involved in the production of degradative enzymes used in food utilization, but do not play a role in caffeine breakdown. 2) A sampling for natural enemies of the insect was conducted in Mexico in collaborative research with F. Infante, J. Perez, and A. Castillo at El Colegio de la Frontera Sur in Tapachula and resulted in the discovery of a new nematode species that reduced the fecundity and increased the mortality of the coffee berry borer. In a collaborative research project established with L. Carta (Nematology Laboratory, USDA, ARS, Beltsville, MD), the new nematode species was placed in the genus Sphaerulariopsis. This is the
first nematode species found in the Americas as a parasite of the coffee berry borer, and could potentially become an important coffee berry borer biocontrol agent. 3) Also in collaboration with the Mexican scientists and S. Peterson at the ARS laboratory in Peoria, IL, a new species of Penicillium (P. brocae) was discovered in coffee berry borers from Mexico. This fungus might be involved in a mutualistic association with the insect, where the fungus provides nutrients needed for insect survival. These findings have increased the knowledge on the basic biology of the insect and its associated organisms and will be helpful in developing biological control programs against the coffee berry borer. 6. What do you expect to accomplish, year by year, over the next 3 years? 2003. Continue research aimed at determining the role of yeasts in food utilization by the coffee berry borer, and determine virulence of different fungal entomopathogens on the coffee berry borer using standard
bioassay techniques. Initiate endophyte research. 2004. Continue yeast and endophyte research initiated in year 1. Develop microsatellite markers for conducting population genetic analyses of B. bassiana infecting coffee berry borers. 2005. Continue research initiated in years 1 and 2. Develop microarrays to conduct comparative analyses of gene expression among strains of Beauveria that differ in virulence against coffee berry borer. 7. What technologies have been transferred and to whom? When is the technology likely to become available to the end user (industry, farmer other scientist)? What are the constraints, if known, to the adoption durability of the technology? No technologies have yet been transferred. 8. List your most important publications and presentations, and articles written about your work (NOTE: this does not replace your review publications which are listed below) Articles in Popular Press: Vega, F. E., Rosenquist, E. USDA's Coffee Research Program. The Coffee
Reporter. 2001. v.6. p.6. Vega, F. E., Rosenquist, E. The coffee berry borer and coffee research at the United States Department of Agriculture. Proceedings of the World Coffee Conference, London, UK. 2001. Available at: www.ico.org. Presentations: "State of the Art Review on Coffee Technology, Genetics, Diseases, Organic Coffee and Sustainable Agriculture - Discussant," World Coffee Conference sponsored by the International Coffee Organization, London (May 2001). "Coffee Agriculture", National Coffee Association Fall Educational Conference, New York City (October 2001). Articles written about research: Wolbachia research was featured in the front page of the January 2002 issue of "Industrial Bioprocessing"; in the "ARS News and Information" web site under "Coffee pest may harbor its weakest link" by Sharon Durham; and in Spanish in three web sites: "RedNet News," "Agroandino.com," and "Noticias e Informacion de ARS."
Impacts
(N/A)
Publications
- Quimby, Jr., P.C., Mercadier, G., Meikle, W., Vega, F.E., Fargues, J., Zidack, N. Enhancing biological control through superior formulations: a worthy goal but still a work in progress. Vurro, M., Gressel, J., Butt, T., Harman, G., Nuss, D., Sands, D., St. Leger, R., editors. IOS Press, The Netherlands. Enhancing Biocontrol Agents and Handling Risks - NATO Science Series Vol. 339: Life and Behavioural Sciences. 2001. p. 86-95.
- Fargues J., Smits, N., Vidal, C., Vey, A., Vega, F.E., Mercadier, G., Quimby, Jr., P.C. 2001. Effect of liquid culture media on morphology, growth, propagule production, and pathogenic activity of the Hyphomycete, Metarhizium flavoviride. Mycopathologia. 2001. v. 154. p. 127-138.
- Vega, F.E., Benavides, P., Stuart, J., O'Neill, S.L. Wolbachia infection in the coffee berry borer (Coleoptera: Scolytidae). Annals of the Entomological Society of America. 2002. v. 95. p. 374-378.
- Mercadier, G., Saethre,M.-G., Vega, F.E. First report of the fungal entomopathogen Paecilomyces in Norway. Norwegian Journal of Entomology. 2002. v.49. p. 71-73.
- Vega, F.E., Franqui, R.A., Benavides, P. The presence of the coffee berry borer, Hypothenemus hampei, in Puerto Rico: fact or fiction? Journal of Insect Science. 2002. v. 2. Available online: insectscience.org/2.13
- Castillo, A., Infante, F., Barrera, J., Carta, L., Vega, F.E. 2002. First field report of a nematode (Tylenchida: Sphaerularioidea) attacking the coffee berry borer, Hypothenemus hampei (Ferrari) (Coleoptera: Scolytidae) in the Americas. Journal of Invertebrate Pathology. 2002. v. 79. p. 199- 202.
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