Brief Summary of Minutes of Annual Meeting

The meeting was called to order by Chairman Hillman at 9:00 am on September 25, 2009 at the Manchester Inn, Ocean Grove, NJ. As Senior Associate Director for the New Jersey Agriculture Experiment Station and Director of Cooperative Research, Hillman is now the Administrative Advisor for NE-1033.

2010 Business Meeting
Sandra Anagnostakis was elected Chair for 2011. Paul Sisco, chair-elect, stated that the 2010 meeting will be held September 16-19, 2010 at Cataloochee Ranch in Maggie Valley, NC. It's about 45 minutes west of Asheville in a gorgeous location at 5,000 feet adjoining the Great Smoky Mountains National Park. The website is: http://www.cataloocheeranch.com/

Note: Detailed accomplishment report is in the attached Minutes.

Completion of proposed milestones:
2005:

• Characterization of the role of hypovirus p29 in virus RNA accumulation in C. parasitica, and virus transmission through conidia of the fungus:
  Completed early:
  Suzuki, N., Maruyama, K., Moriyama, M. and Nuss, D. L. Hypovirus papain-like protease p29 functions in trans to enhance viral double-stranded RNA accumulation and vertical transmission. J. Virol. 77:11697-11707, 2003.
• Generation of polyclonal antibodies against 5 overlapping regions of hypovirus ORF B, and construction of a C. parasitica EST database.
  Both completed in 2004.

2006:

• Publication of a C. parasitica EST database containing approximately 2500 ESTs.
  Completed early.
  Dawe, A.L., McMains, V.C., Panglao, M., Kasahara, S., Chen, B. and Nuss, D.L. An ordered collection of expressed sequences from Cryphonectria parasitica and evidence of genomic microsynteny with Neurospora crassa and Magnaporte grisea. Microbiology 149:2373-2384, 2003.

2007:

• ORF B polyprotein processing pathway in C. parasitica confirmed, ORF B mature proteins responsible for altering fungal cell signaling pathways mapped and DNA microarray analysis of hypovirus-mediated alteration of fungal gene expression initiated.
  ORF B polyprotein processing pathway not completed. Microarray analysis initiated giving publication in 2003.

  Allen, T.D., Dawe, A.L. and Nuss, D.L. Use of cDNA microarrays to monitor transcriptional responses of the chestnut blight fungus Cryphonectria parasitica to infection by virulence-attenuating hypovirus. Eukaryotic Cell 2:1253-1265, 2003.

2008:

• Polyprotein processing maps completed for hypoviruses CHV1-EP713 and CHV1-Euro 7, and a detailed view compiled of the changes in cellular transcriptional profiles caused by infection of C. parasitica with mild and severe hypoviruses.
  Polyprotein processing map has not been competed. Transcriptional profiles caused by mild and severe hypoviruse have been generated. The C. parasitica EST microarrays have also been used to examine the effect of hypovirus infection on G-protein signaling and to expose a linkage between mitochondrial and viral hypovirulence.

  Allen, T.D. and Nuss, D.L. Specific and common alterations in host gene transcript accumulation following infection of the chestnut blight fungus by mild and severe hypoviruses. J. Virol. 78:4145-4155, 2004.

  Dawe, A.L., Segers, G.C., Allen, T.D., McMains, V.C. and Nuss, D.L. Microarray analysis of Cryphonectria parasitica Ga- and Gbg- signaling pathways reveals extensive modulation by hypovirus infection. Microbiology 150:4033-4043, 2004.

  Allen, T.D. and Nuss, D.L. Linkage between mitochondrial hypovirulence and viral hypovirulence in the chestnut blight fungus revealed by cDNA microarray analysis. Eukaryotic Cell 3:1227-1232, 2004.

Completion of milestones not proposed:

• A proposal to sequence the C. parasitica genome was approved by the Department of Energy Community Sequencing Program in June of 2006.
  The assembled 8.5 X C. parasitica genome sequence was released to the public on September 30, 2008. This is a tremendous resource for future studies on the chestnut blight fungus and its interaction with the chestnut tree.
• Demonstration that hypovirus p29 suppresses RNA silencing in C. parasitica and in heterologous plant system. This is the first report of a mycovirus-encoded suppressor of RNA silencing.
  Segers, G.C., van Wezel, R., Zhang, X., Hong, Y. and Nuss, D.L. Hypovirus Papain-like protease p29 suppresses RNA silencing in the natural fungal host and in a heterologous plant system. Eukaryotic Cell 5:896-904, 2006.
• Demonstrated that RNA silencing serves as an antiviral defense mechanism in C. parasitica (first example for any fungus) against hypoviruses and mycoreoviruses.
  Segers, G.C., Zhang, X., Deng, F., Sun, Q. and Nuss, D.L. Evidence that RNA silencing functions as an antiviral defense mechanism in fungi. PNAS USA 104:12902-12906, 2007.
• First report of the cloning and sequence analysis of mycovirus-derived small RNAs (vsRNAs) generated by RNA silencing. The vsRNAs were shown to be produced, in a dicer dcl-2-dependent manner, from both positive and negative hypovirus RNA strands at a ratio of 3:2 and to be non-randomly distributed along the viral genome. C. parastiica was shown to respond to mycovirus infection with a 10-15 fold increase in dcl-2 transcript acuumulation while the expression of dcl-1 was modestly increased. The expression of dcl-2 was further increased (~35 fold) following infection by a CHV1-EP713 mutant that lacks the p29 suppressor of RNA silencing. A similar response in dicer gene expression following virus infection of plants or animals has not yet been reported. In this regard, it is anticipated that the evolutionary position of fungi relative to animals and plants will provide insights into additional novel mechanisms for the induction and suppression of RNA silencing pathways yet to be revealed in the other organisms.
  Zhang, X., Segers, G.C., Sun, Q., Deng, F. and Nuss, D.L. Characterization of hypovirus-derived small RNAs generated in the chestnut blight fungus by an inducible DCL-2-dependnet pathway. Journal of Virology 82:2613-2619, 2008.
• Virus RNA recombination is an important component of virus evolution that contributes to the emergence of new viruses and the generation of internally deleted mutant RNAs, termed defective interfering (DI) RNAs, that are derived from, and dependent on, the parental viral genomic RNA. We provided the first experimental evidence that a host RNA silencing pathway is required for DI RNA production and virus vector RNA instability for a single-strand, positive sense RNA virus.
  Zhang, X. and Nuss, D.L. A host dicer is required for defective viral RNA production and recombinant virus vector instability for a positive sense RNA virus. Proc. Natl. Acad. Sci. USA. Early Edition, October 13, 2008.

2009

• Determine specific genetic fingerprints of tested cultivars through the use of microsatellite markers and find loci useful for parentage analysis (Fulbright)
• Refine the genetic linkage and genome sequence maps for map-based cloning of fungal vic and pathogenicity genes
• Tom Kubisiak used the C. parasitica Genome sequence generated by the JGI Community sequencing Program to identify 689 simple sequence repeats (SSRs) and designed 141 primer pairs. One hundred and thirty Four of the 141 primer pairs amplified discrete products, with 96 of the 134 showing polymorphism for the JA17 and X17.8 parents of the mapping cross. Allele data were generated for 96 progeny from the mapping cross for 32 polymorphic SSRs and a total of 30 of these markers were placed within the context of the published C. parasitica linkage map (Kubisiak and Milgroom, 2006, FG&B 43:453-463). The new linkage data were used by the JGI finishing group to connect a number of scaffolds in Version 1 of the genome assembly to generate Version 2 that was released July 10, 2009.
• Complete assembly and community manual annotation of the C. parasitica genome sequence
  Nine hundred and ninety five of 11,251 transcripts have been manually curated by the C. parasitica annotation team to date. Version 2 of the genome assembly has been released by the JGI consisting of 26 scaffolds. Five of the new scaffolds contain two teleomers and are of a length consistent with a complete chromosome. Six additional scaffolds contain one teleomer and are in excess of 1 MB in length.
• Use the C. parasitica genome sequence to develop new microarray chip and proteomics platforms for analysis of global gene expression in the blight fungus when challenged by viral pathogens
• Orchard established in WV with advanced, back-cross chestnut trees from VA for assessment of host resistance with hypovirulence in the Cryphonectria parasitica populationreplanted in 2009.

In progress.

2010

• Complete characterization of C. parasitica antiviral RNAsilencing pathways. Two Dicer genes were identified in the C. parasitica genome, cloned and disrupted. Dicer DCL2 was shown to be required for antiviral RNA silencing while Dicer DCL1 was not required (Segers et al., PNAS 2007, 104:12902-12906). Four Argonaute genes were identified in the C. parasitica genome, cloned and disrupted. Only Argonaute AGL2 was required for antiviral RNA silencing (Sun et al., PNAS, IN PRESS). Three RNA dependent RNA polymerases and an orthologue of the QIP exonuclease involved in transgene silencing in Neurospora crassa have been identified in the C. parasitica genome and are currently being cloned and disrupted.

2008-2009 Publications Aguilar, F.X., M.M. Cernusca, and M.A. Gold. 2009. A preliminary assessment of consumer preferences for chestnuts (Castanea spp.) using conjoint analysis. HortTechnology 19: 216-223. Andrade, G.M., C.J. Nairn, H.T. Le and S.A. Merkle. 2009. Sexually mature transgenic American chestnut trees via embryogenic suspension-based transformation. Plant Cell Reports 28:1385-97. Barakat, A., D.S. DiLoreto, Y. Zhang, C. Smith, K. Baier, W.A. Powell, N. Wheeler, R. Sederoff and J.E. Carlson. 2009. Comparison of transcriptome from cankers and healthy stems in American chestnut (Castanea dentata) and Chinese chestnut (Castanea mollissima). BMC Plant Biology, 9:51 11 pages. Cernusca, M.M., M.A. Gold and L.D. Godsey. 2008. Influencing consumer awareness through the Missouri chestnut roast. Journal of Extension 46(6). Published online. 8 p. www.joe.org/joe/2008december/rb7p.shtml Cooper, W.R. and L.K. Rieske. In review. Ecological interactions among Dryocosmus kuriphilus Yasumatsu (Hymenoptera: Cynipidae) community associates. Ecological Entomology, submitted fall 2009. Cooper, W.R. and L.K. Rieske. In review. Chestnut species and jasmonic acid treatment influence development and community interactions of galls produced by Dryocosmus kuriphilus Yasumatsu (Hymenoptera: Cynipidae). Arthropod Plant Interactions, submitted summer 2009. Cooper, W.R. and L.K. Rieske. 2009. Woody stem galls interact with foliage to affect community associations. Environmental Entomology 38, 417-424. Cooper, W.R. and L.K. Rieske. 2008. Differential responses in American (Castanea dentata Marshall) and Chinese (C. mollissima Blume) chestnut (Fagales: Fagaceae) to foliar application of jasmonic acid. Chemoecology 18, 121-127. Cooper, W.R. and L.K. Rieske. 2009. Sustainable Management of the Invasive Asian Chestnut Gall Wasp? Interactions between native and introduced natural enemies. The Nutshell, June 2009. Dawe, A.L. W. A. Van Voorhies, T. A. Lau, A. V. Ulanov and Z. Li. 2009. Microbiology, in press (available online as doi: 10.1099/mic.0.029033-0). Gold, M.A., M.M. Cernusca and L.D. Godsey. 2008. A competitive market analysis of the U.S. shiitake mushroom marketplace. HortTechnology 18: 489-499. Gold, M.A., M.M. Cernusca and L.D. Godsey. 2009. Agroforestry Product Markets And Marketing. Chapter 11. In: Garrett, H.E. (ed). North American Agroforestry: An Integrated Science and Practice. 2nd Edition. Agronomy Society of America, Madison, WI. Gold, M.A. and H.E. Garrett. 2009. Agroforestry Nomenclature, Concepts and Practices. Chapter 3. In: Garrett, H.E. (ed). North American Agroforestry: An Integrated Science and Practice. 2nd Edition. Agronomy Society of America, Madison, WI. Gold, M.A. and M.H. Hall (eds.). 2009. Agroforestry Comes of Age: Putting Science into Practice. 11th North American Agroforestry Conference Proceedings. Columbia, MO. May 31-June 3, 2009. 517 p. Gonzalez, Donis I.R., M. Mandujano, C. Medina-Mora and D.W. Fulbright. 2009. Presence of mycotoxins after 90 days of storage in fresh chestnuts. 4th Annual Chestnut Symposium, Beijing, China, Oct. 2008. Acta Horticulturae (in press). Jacob-Wilk, D., M. Turina, P. Kazmierczak and N.K. Van Alfen. 2009. Silencing of Kex2 significantly diminishes the virulence of Cryphonectria parasitica. Molecular Plant-Microbe Interactions 22:211-221. Kim. M.-J., B.-R. Kwon, S.-M. Park, H.-J. Chung. M.-S. Yan, A.C. L. Churchill, N.K. Van Alfen and D.-H. Kim. 2008. Promoter analysis of the cell surface-abundant and hypoviral-regulated cryparin gene from Cryphonectria parasitica. Mol. Cells 26:496-502. Lan, Xiuwan; Ziting,Yao, Yan Zhou, Jinjie Shang, Haiyan Lin, D.L. Nuss, and B. Chen. 2008. Deletion of the cpku80 gene in the chestnut blight fungus, Cryphonectria parasitica, enhances gene disruption efficiency. Current Genetics 53:59-66. Milgroom M.G., K. Sotirovski, M. Risteski, and M.T. Brewer. 2009. Heterokaryons and recombinants of Cryphonectria parasitica in two clonal populations in southeastern Europe. Fungal Genet. Biol. 46: 849-854. Monteiro-Vitorello, C.B., G. Hausner, D.B. Searles, E.A. Gibb, D.W. Fulbright and H. Bertrand. 2009. The Cryphonectria parasitica mitochondrial ms gene: plasmid-like elements, introns and homing endonucleases. Fungal Genetics and Biology 46:837-848. Sun, Q., G.H. Choi and D.L. Nuss. 2009. Hypovirus-responsive transcription factor gene pro1 of the chestnut blight fungus Cryphonectria parasitica is required for female fertility, asexual spore development and stable maintenance of hypovirus infection. Eukaryotic Cell 8:262-270. Sun, Q., G.H. Choi and D.L. Nuss. 2009. A single Argonaute gene is required for induction of RNA silencing antiviral defense and promotes viral RNA recombination. Proc. Nat. Acad. Sci. (in press). Valdivia, C., L. Zabek, J. Arbuckle, M. Gold and C. Flora. 2009. Human and Institutional Dimensions of Agroforestry. Chapter 13. In: Garrett, H.E. (ed). North American Agroforestry: An Integrated Science and Practice. 2nd Edition. Agronomy Society of America, Madison, WI. Willyerd, K.L. A.M. Kemp and A.L. Dawe. 2009. Controlled gene expression in the plant pathogen Cryphonectria parasitica by use of a copper-responsive element. Appl. Environ. Microbiol. 75:5417-5420. Zhang, X. and D.L. Nuss. 2008. A host dicer is required for defective viral RNA production and recombinant virus vector RNA instability for a positive sense RNA virus. Proc. Nat. Acad. Sci. 105:16749-16754.