Progress 11/01/08 to 10/31/13
Outputs
Target Audience: Both Scientists and growers were reached by public presentations and by publishing in journals. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? The project provided opportunities for 4 undergraduates and 2 graduate students to participate in research. How have the results been disseminated to communities of interest? Results have been presented in publications as previously listed and in meetings as follows: 2013. Trail, F. New strategies to control head scab in wheat. Talk presented to the Wheat College, East Lansing Michigan, June 2013. 2013. Trail, F. The life cycle of a head blight pathogen, Fusarium graminearum, and its importance to agriculture. European Fusarium Seminar, Bordeaux, France, May. 2013. Trail, F. Genetics of forcible spore discharge in fungi. Invited talk given at Institut non lin’eare de Nice, CNRS Université Nice, France, May. 2013. Trail, F, U Sikhakolli, K Fellows, N Lehr, JP Townsend. Comparative transcriptomics identifies new genes for perithecium development. Fungal Genetics Conference, Asilomar, CA, March. 2013. Trail, F. Unraveling the genetics of perithecium form and function in Fusarium graminearum. Fungal Genetics Conference, Fusarium meeting, Asilomar, CA, March. 2013. Trail, F. The secret lives of fungi: the good, the bad and the ugly. Given to the local chapter of the Wild Ones native plant society. February. 2012. Trail, F. Dissecting the machinery of the fungal cannon. Department of Applied Mathematics. Harvard University, Cambridge MA. December 12. 2012. Trail, F. The Impact of the Fusarium graminearum Genome Sequence on the Quest for Control of Head Blight. National Fusarium Head Blight Forum, Orlando, FL. December. 2012. Trail, F. and Drew Afton. Host colonization leading to sporulation in Fusarium graminearum. Special Session at the American Phytopathological Society Meetings. Providence RI. August. 2012. Lessons learned from genomic studies of toxigenic Fusarium species. Invited talk. EU Mycored Conference. Ottawa, Canada, June. 2012. Understanding Fusarium head blight of wheat through the fungal life cycle. Bowling Green University, Ohio. March. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Goal 1: We have progressed in understanding the mechanism of forcible ascospore discharge by studying gene expression during perithecium development in F. graminearum. We have generated more than 30 mutants of genes thought to be involved in fruiting body development and spore firing. We have identified several processes that are involved in the function of firing. Goal 2. We have performed field trials to with a product that inhibits ascospore discharge in the apple scab pathogen. The product has been highly variable in its ability to reduce disease. We are currently trying to optimize the use of this product. Goal 3.We are currently trying to optimize the use of this product by testing variants of the formulation and differing concentrations on the apple scab pathogen. Goal 4. Because of stumbling blocks in Goals 2 and 3, we have not moved forward with this goal.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
Cavinder, B., Sikhakolli, U., Fellows, K. M., Trail, F. 2012. Sexual Development and Ascospore Discharge in Fusarium graminearum. J. Vis. Exp. 61: e3895, DOI: 10.3791/3895.
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
Cavinder, B. and Trail, F. 2012. The role of Fig1, a component of the Low Affinity Calcium Uptake System (LACS), in growth and sexual development of filamentous fungi. Eukaryotic Cell, 11(8):p. 987-988. DOI: 10.1128/EC.00007-12
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
Sikhakolli, U.R., L�pez-Gir�ldez, F., Li, N., Common, R., Townsend, J.P., and Trail, F. 2012. Transcriptome analyses during fruiting body formation in Fusarium graminearum and F. verticillioides reflect species life history and ecology. Fungal Genetics and Biology, 49:663-673
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
OConnell, R.J., MR Thon, S Hacquard, SG Amyotte, J Kleeman, MF Torres, U Damm, EA Buiate, L Epstein, N Alkan, J Altmuller, L Alvarado-Balderrama, CA Bauser, C Becker, BW Birren, Z Chen, J Choi, JA Crouch, JP Duvick, MA Farman, P Gan, D Heiman, B Henrissat, RJ Howard, M Kabbage, C Koch, B Karcher, Y Kuba, AD Law, M-H Lebrun, Y-H Lee, I Miyara, N Moore, U Neumann, K Nordstrom, DG Pannacione, R Panstruga, M Place, RH Proctor, D Prusky, G Rech, R Reinhardt, JA Rollins, S Rounsley, CL Schardl, DC Schwartz, N Shenoy, K Shirasu, UR Sikhakolli, K Stuber, SA Sukno, JA Sweigard, Y Takano, H Takahara F Trail, HC van der Does, LM Voll, I Will, S Young, Q Zeng, J Zhang, S Zhou, MB Dickman, P Schulze-Lefert, EVL van Themaat, L-J Ma, LJ Vaillancourt. 2012. Life-style transitions in plant pathogenic Colletotrichum fungi deciphered by genome and transcriptome analyses. Nature Genetics 44:1060-1065.
- Type:
Book Chapters
Status:
Published
Year Published:
2013
Citation:
Trail, F. 2013. Sex and Fruiting in Fusarium. In, Fusarium: genomics, molecular and cellular biology. Daren Brown and Robert Proctor, eds. Horizon Scientific Press and Caister Academic Press, Norwich, UK.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Ma, L-J, DM Geiser, RH Proctor, AP Rooney, K ODonnell, F Trail, DM Gardiner, JM Manners, K Kazan. Fusarium Pathogenomics, Invited review to Annual Review of Microbiology
- Type:
Book Chapters
Status:
Accepted
Year Published:
2013
Citation:
Trail, F, DM Gardiner. Application of genomics to the study of pathogenicity and development in Fusarium. Invited review to The Mycota series.
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: Our work falls into two categories: 1. Basic research in Fusarium graminearum on how the ascospore discharge mechanism works. 2. Application of a discharge inhibitor to control of the apple scab disease. PARTICIPANTS: Participation: Individuals: Drew Afton- graduate student who assisted in setup and processing field trial experiments Brad Cavinder- Graduate student who designed and implemented experiments on ascospore discharge. Chris Wright- graduate student and technician who investigated ascospore discharge in Venturia inaequalis. Frances Trail-PD in Michigan Kerik Cox- PD in New York trials Amy Irish-Brown- advisory for field trials in Michigan Jeffrey Townsend- collaborated in transcriptomic comparisons between Neurospora and Fusarium. Partner Organizations: Michigan State University Cornell University Yale University TARGET AUDIENCES: Target Audience Growers of apples, corn, wheat and barley Scientists, particularly fungal biologists PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
To identify genes involved in ascus formation, we have embarded on a comparative transcriptomics project. We previously analyzed gene expression during 6 stages of fruiting body in Fusarium graminearum and F. verticillioides. By comparing gene expression profiles at each stage between these two species and between these species and three Neurospora species (data collected by collaborate Jeffrey Townsed, Yale University), we identified genes that substantially change expression just in F. graminearum or just in the Fusarium species. The genes that had the greatest shifts in expression in F. graminearum were used as targets for knockouts. A large proportion of these genes are involved in the generation of ascospores. We performed another trial in apple orchards in Michigan at Clarkesville to test the ability of an experimental product Earth-Tec Mg to inhibit spore firing from overwintering fruiting bodies in the leaves on the orchard floor. This year was unusual in that we had a very early period of warm weather, which greatly accelerated flowering and initiated spore production in the apple scab fungus. We performed the test, and found significant disease reduction, although it was reduced over previous years, probably due to the difficulty in predicting the development of the ascospores in advance.
Publications
- Sikhakolli, U.R., Lopez-Giraldez, F., Li, N., Common, R., Townsend, J.P., and Trail, F. 2012. Transcriptome analyses during fruiting body formation in Fusarium graminearum and F. verticillioides reflect species life history and ecology. Fungal Genetics and Biology, 49:663-673.
- Wang, Z., Lehr, N., Trail, F. and J.P. Townsend. 2012. Differential impact of nutrition on developmental and metabolic gene expression during fruiting body development in Neurospora crassa. Fungal Genetics and Biology 49:405-413.
- Cavinder, B. and Trail, F. 2012. The role of Fig1, a component of the Low Affinity Calcium Uptake System (LACS), in growth and sexual development of filamentous fungi. Eukaryotic Cell, 11(8):p. 987-988. DOI: 10.1128/EC.00007-12..
- Cavinder, B., Sikhakolli, U., Fellows, K. M., Trail, F. 2012. Sexual Development and Ascospore Discharge in Fusarium graminearum. J. Vis. Exp., e3895, DOI: 10.3791/3895.
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: Our work falls into two categories: 1. Basic research in Fusarium graminearum on how the ascospore discharge mechanism works. 2. Application of a discharge inhibitor to control of the apple scab disease. Work in Fusarium graminearum this year included generation of a system using RNA interference (gene silencing) of essential genes during the process of ascus and perithecium maturation. Some genes, when mutated, arrest the development of fruiting bodies altogether. This prohibits our ability to understand the role of these genes specifically during ascus function. We developed a technique to induce gene silencing which can be applied to any stage of development to prohibit gene expression during that stage. This will be used for understanding the mechanism of forcible ascospore discharge. Work in apple orchards in Michigan and New York was done to test the ability of an experimental product Earth-Tec Mg to inhibit spore firing from overwintering fruiting bodies in the leaves on the orchard floor. Work in Michigan suggests that the susceptibility of the fungal fruiting bodies to the product may depend on fruiting body maturity. Work in New York suggests the product is effective in reducing disease across the season. Dissemination: Trail, F. 2011. A whole plant/ whole fungus look at the head blight disease. Plenary talk, Canadian Workshop on Fusarium Head Blight. Winnepeg, Canada. November. Trail, F. 2011. Head Blight of Wheat: Entwining of two life cycle. The Ohio State University, Wooster and Columbus, OH. February. F. Trail, D. Afton and K. Cox. 2011. Control of Apple Scab through reduction of inoculum dispersal. Fruit and Vegetable Exp, Grand Rapids, MI. December. PARTICIPANTS: Individuals: Drew Afton- graduate student who assisted in setup and processing field trial experiments Brad Cavinder- Graduate student who designed and implemented experiments on ascospore discharge. Chris Wright- graduate student and technician who investigated ascospore discharge in Venturia inaequalis. Frances Trail-PD in Michigan Kerik Cox- PD in New York trials Amy Irish-Brown- advisory for field trials in Michigan Partnew Organizations: Michigan State University Cornell University TARGET AUDIENCES: Target Audience Growers of apples and wheat and barley Scientists, particularly fungal biologists Efforts Work presented at the Canadian Workshop on Fusarium Head Blight, Michigan Fruit and Vegetable Expo. Both of these involve growers and scientists. Also presented at the Fungal Genetics Conference, which is mainly scientists. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Change in knowledge: We have developed a functional technique to help understand the role of essential genes in development of spores in Fusarium graminearum. We used this information to understand the role of myosin proteins in spore development. We explored the efficacy of EarthTec Mg to inhibit dispersal of ascospores of Venturia inaequalis from the infected fallen leaves and initiate a new disease cycle in the spring.
Publications
- Cavinder, B., Hamam, A., Lew, R.R. and F. Trail. 2011. Mid1, a mechanosensitive calcium ion channel, affects growth, development, and ascospore discharge in the filamentous fungus Gibberella zeae. Eukaryotic Cell 10: 832-841. COVER.
- Cavinder, B., and F. Trail. 2011. Using Next generation sequencing to characterize a complex mutational event in Gibberella zeae. Fungal Genetics Reports 58 (Supple): 84.
- Sikhakolli, U., Townsend, J. and F. Trail. 2011. Transcriptomic foundation of sexual development in Fusarium species. Fungal Genetics Reports 58 (Supple): 85.
- Lehr, N., Wang, Z., Lopez-Giraldez, F., Farre, M., Trail, F., and J. P. Townsend. 2011. Fruiting body development and transcriptomics in Neurospora species. Fungal Genetics Reports 58 (Supple): 110.
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: Activities include conducting and analyzing experiments or surveys, assessments, facilitating, teaching, or mentoring. We researched ascospore discharge in two systems: Venturia inaequalis on apple and Fusarium graminearum on wheat and barley. One PhD student, Brad Cavinder, one technician, Jon Hoerger, and one undergraduate, Nick Harrison worked on these projects. Results were presented at the Great Lakes Fruit Expo in December and at the Gordon Research Conference on Cellular and MOlecular Mycology in June. PARTICIPANTS: We collaborated with Kerik Cox at Cornell University in the field trials. Brad Cavinder, Jon Hoerger and Nick Harrison are in the Trail laboratory and all worked on this project and received training in research and professional development. TARGET AUDIENCES: We hope the new product will move towards commercial development in the coming years. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
A field study was conducted in the spring and summer of 2010 to determine the efficacy of a novel compound for control of apple scab in an research orchard at the New York State Agricultural Experiment Station in Geneva, NY. In 2010, scab disease pressure was relatively mild, but a significant effect was seen in reduction of scab lesions on McIntosh cluster leaves.
Publications
- Cavinder, B., Hallen-Adams, H., Trail., F. 2010. Calcium signaling in ascospore discharge. Gordon Research Conference on Cellular and Molecular Mycology. New Hampshire, June.
- Hallen-Adams, H., Guenther, J., Trail, F. The role of lipids in successful overwintering and subsequent perithecium production by Fusarium graminearum. Mycological Society Meeting, Lexington, KY. July 2010.
- Trail, F. and Cox, K. 2010. Reducing disease pressure by controlling ascospore discharge in Venturia inaequalis. Great Lakes Fruit Expo, Grand Rapids, MI, December.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: We have been investigating the production and dissemination of inoculum of the wheat blight fungus Fusarium graminearum and the apple scab fungus Venturia inaequalis. We are investigating ion channel genes that are important for ascospore discharge and have identified 2 that are essential. These are being localized in the ascus using fluorescent tags. We have performed chemical analysis to determine the components of the fluid surrounding the ascospores in V. inaequalis. It appears based on these analysis that the mechanism is quite similar between the 2 fungi. PARTICIPANTS: Jonathan Hoerger is an undergraduate in the lab who has gained important research experience in this project. Brad Cavinder is a graduate student who has implemented new techniques to study the spore discharge process. He has gained experience in genomics, physiology, advanced microscopy and molecular genetics. TARGET AUDIENCES: BOth diseases are real problems for producers in the midwest. We are disseminating the information we gain to growers groups and to other researchers. They can use the information to enhance disease control. We hope to use this work to develop new disease controls that result in reduced fungicide applications PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
This work is directed towards finding new, life-cycle based controls for these two diseases. The diseases rely on forcible ascospore discharge to initiate disease cycles in the spring. We are investigating whether controls can be developed to arrest initial spore discharge.
Publications
- Guenther, JC, Hallen-Adams, HE, Bucking, H, Shachar-Hill, Y and F. Trail 2009. Triacylglyceride metabolism by Fusarium graminearum during colonization and sexual development on wheat. 2009. Molecular Plant-Microbe Interactions 22:1492-1503.
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: We have been studying the basis of forcible spore discharge in fungal plant pathogens. In the wheat scab fungus, Fusarium graminearum, forcible ascospore discharge initiates the disease cycle in the spring. Our goal is to understand the physiological and genetic basis of forcible ascospore discharge and use this information to design appropriate controls. This year, we have shown that Mid1, a stretch activated ion channel, is essential to ascospore discharge in Fusarium graminearum. Genetic disruption of the Mid1 gene results in strains that can no longer discharge ascospores. The mutants also have compromised hyphal growth, but pathogenic capabilities are not altered. The apple scab pathogen, Venturia inaequalis, is also dependent on forcible ascospore discharge and we have begun investigating the similarities between the mechanism of forcible ascospore discharge between these two pathogens. Towards this end, this year we developed an assay for V. inaequalis to test the effects of ascospore discharge inhibitors that were effective on F. graminearum. Trials of these inhibitors is underway. Results of this work have been presented at the Great Lakes Fruit, Vegetable and Farm Market Expo, and at the Fusarium Head Blight Forum. PARTICIPANTS: The Michigan Apple Committee provided funding for the research on Venturia inaequalis. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Our research is aimed towards addressing the distribution of initial inoculum for a group of diseases caused by ascomycetous fungi that shoot their sexual spores (ascospores)from cellular cannons (called asci). This year we have begun to compare the mechanism in two fungi from different phylogenetic groups that form asci of different morphologies. We are working towards identifying controls that arrest forcible ascospore discharge and will work on a range of species that shoot spores.
Publications
- Hallen, H., and F. Trail. 2008. The L-type calcium ion channel, Cch1, affects ascospore discharge and mycelial growth in the filamentous fungus Gibberella zeae (anamorph Fusarium graminearum). Eukaryotic Cell. 7:415-424.
- F. Trail, R. Rodriguez. 2008. Chemical control of Venturia inaequalis, the apple scab pathogen. Provision Patent application.
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Progress 01/01/07 to 12/31/07
Outputs
Fusarium graminearum produces head blight disease on wheat. The sexual cycle is a crucial component of head blight epidemiology, as forcibly-discharged ascospores serve as the primary inoculum. Using GeneChips, a developmental time course was performed in culture, from vegetative hyphae to mature perithecia with multiseptate ascospores. Time-points represent the development of the major cell-types comprising the mature perithecium. The majority of the 17,830 G. zeae probesets, 78%, were expressed during at least one of the developmental stages; 12% of these are likely to be specific to sexual development. Analysis of the 162 predicted ion transporter genes is reported in detail, due to their association with perithecium function. Expression patterns of the MirA-type siderophores, chloride channels, P-type ATPases and potassium transporters show some specialization in regard to developmental stage. This is the first whole-genome analysis of differential transcript
accumulation during sexual development in a filamentous fungus. We also performed a time-course of wheat stem colonization by Fusarium grminearum by inoculation of the head and following fungal progression down the stem. Using this relatively uniform plant tissue, it was possible to separate the hyphae at the front, which are mainly vegetative, from those further back which begin to colonize radially and finally develop perithecium initials at the epidermis. The time-course of gene expression roughly parallels that of sexual development in culture. Cch1, a putative voltage-gated calcium ion channel, was investigated for its role in ascus development in F. graminearum. Gene replacement mutants of CCH1 were generated and found to have asci which did not forcibly discharge spores, although morphologically ascus and ascospore development in the majority of asci appeared normal. Additionally, mycelial growth was significantly slower and sexual development was slightly delayed in the
mutant; mutant mycelia showed a distinctive fluffy morphology; and no cirrhi were produced. Wheat infected with cch1 mutants developed symptoms comparable to wheat infected with the wild-type, however, the mutants showed a reduced ability to protect the infected stalk from colonization by saprobic fungi. Transcriptional analysis of gene expression in mutants using the Affymetrix Fusarium microarray showed 2449 genes with significant, two-fold or greater changes in transcript abundance across a developmental series. This work extends the role of CCH1 to forcible spore discharge in F. graminearum, and suggests that this channel has subtle effects on growth and development.
Impacts
This is the first time the genes unique to spore production have been identified in any filamentous fungi. Filamentous fungi constitute the vast majority of plant disease organisms. Further understanding of the function of these genes is essential to developing novel controls for plant disease. The work also comprises the first identification of genes involved in regulation of ascospore discharge. Numerous fungal pathogens important to US agriculture require forcible ascospore discharge to disperse the primary disease inoculum. Understanding how spores are launched will lead to the identification of chemicals that may be used to controling the dispersal of inoculum.
Publications
- Cuomo, C., Gueldener, U., Xu, J.R., Trail, F. et al. (45 authors). 2007. The Fusarium graminearum genome reveals a link between localized polymorphism and pathogen specialization. Science 317: 1400-1402.
- Hallen, H., Huebner, M., Shiu, S.-H., Guldener, U., and F. Trail. 2007. Gene expression shifts during perithecium development in Gibberella zeae (anamorph Fusarium graminearum), with particular emphasis on ion transport proteins. Fungal Genetics and Biology 44: 1146-1156.
- Trail, F. 2007. Fungal cannons: Explosive spore discharge in the Ascomycota. FEMS Microbiology Letters 276:12-18. Invited review.
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Progress 01/01/06 to 12/31/06
Outputs
Lipid accumulation and storage is vital to survival of all organisms. Stored lipids are then used for development. In fungi, lipids are stored in vegetative hyphae and spores as lipid bodies. The wheat pathogen, Gibberella zeae, stores lipids mainly as triacylglycerides (TAG) in anticipation of sexual development. We have characterized the process of lipid accumulation and utilization in association with perithecium development in culture and leading up to perithecium development in planta. We characterized and quantified lipids and water soluble sugars during the initiation and development of perithecia. We next examined gene expression patterns for genes associated with lipid biosynthesis and degradation using data collected from Affymetrix GeneChips. Information gathered from these studies indicates an essential role for lipids in the formation of perithecia. Prior to this study it had been observed that large amounts of fats accumulated immediately following
perithecium induction in culture. 1H-NMR analysis of neutral lipids from induced cultures collected at the early stages of perithecium development indicated that TAG were the major stored metabolite in dikaryotic hyphae (those that will produce the perithecia). We examined the transcript accumulation for a discrete set of metabolic genes representing pathways important to lipid accumulation and degradation. Genes representing enzymes related to glycolysis, pentose phosphate pathway, tricarboxylic acid cycle, fatty acid biosynthesis, TAG synthesis and β-oxidation were identified. The normalized transcript data output was used to establish apparent patterns of expression for TAG storage and degradation through developmental time courses in planta and in culture. In general, accumulation of transcripts for genes associated with TAG generation at the initiation of sexual development was opposite the accumulation for genes associated with TAG breakdown, which occurred during
perithecium development.
Impacts
The results of this study clearly show that the head blight fungus must accumulate significant lipid resources from the host in order to produce the following year's inoculum. The accumulation of lipid is most likely due to the transfer of sucrose from the host to the fungus during pathogenesis and is substantially complete by the time the plant is senesced. This is contrary to widespread belief that the fungus invades as a saprophyte and then can produce perithecia, an approach which probably plays little role in the epidemiology of the disease.
Publications
- Qi, W., Kwon, C. and F. Trail. 2006. Microarray analysis of transcript accumulation during perithecium development in Gibberella zeae (anamorph Fusarium graminearum). Molecular Genetics and Genomics 276:87-100. Goswami, R., Xu, J.R., Trail, F. Hilburn, K. and H. C. Kistler. 2006. Genomic analysis of host-pathogen interaction between Fusarium graminearum and wheat during early stages of disease development. Microbiology.
- Gueldener, U., K.-Y. Seong, J. Boddu, S. Cho, F. Trail, J.-R. Xu, G. Adam, H.-W. Mewes, G.J. Muehlbauer, and H. C. Kistler. 2006. Development of a Fusarium graminearum Affymetrix GeneChip For profiling fungal gene expression in vitro and in planta. Fungal Genetics and Biology, 43 (5): 316-325.
- Gaffoor, I., and F. Trail. 2006. Characterization of two polyketide synthase genes involved in zearalenone biosynthesis in Gibberella zeae. Appl. and Environ. Microbiol. 72:1793-1799.
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Progress 01/01/05 to 12/31/05
Outputs
Worldwide, one of the most devastating pathogens of small grains is the head blight fungus, Gibberella zeae. Ascospore-laden perithecia of this fungus develop on mature cereal crops and crop debris and provide the primary inoculum of the disease. This year we finished a study to elucidate the process of colonization of wheat tissue, which leads to perithecium production. Stems were systemically and extensively colonized following inoculation of the wheat head. Haploid mycelia moved down the vascular system and pith and then colonized the stem tissue radially. Dikaryotic hyphae developed at two distinct stages: in the xylem, in support of radial hyphal growth and in the chloremchyma, in support of perithecium development. Perithecium formation was initiated in association with stomates and silica cells. Vascular occlusions prevented mycelia from colonizing the stem in 25% of inoculated plants. Vascular occlusions are an important component of resistance for many plants
that are hosts to vascular pathogens and may also be an important component of resistance to FHB for wheat. In addition, we elucidated the biomechanical aspects of ascospore discharge. Since wind speed drops to zero at a surface, forced ejection should facilitate spore dispersal. But for tiny spores, with low mass relative to surface area, high ejection speed yields only a short range trajectory, so pernicious is their drag. Thus, achieving high speeds requires prodigious accelerations. In G. zeae, we determined the launch speed and kinetic energy of ascospores shot from perithecia, and the source and magnitude of the pressure driving the launch. We asked whether the pressure inside the ascus suffices to account for launch speed and energy. Launch speed was 34.5 m s-1, requiring a pressure of 1.54 MPa and an acceleration of 870,000 g-- the highest acceleration reported in a biological system. This analysis allows us to discount a major component of the epiplasmic fluid, mannitol, as
having a key role in driving discharge, and supports the role of potassium ion flux in the mechanism.
Impacts
We have shown that colonization of wheat vegetation prior to grain harvest in the field is an important step in perithecium development. Vegetation may become colonized by means of stem-base infections and head infections, allowing the head blight pathogen to establish itself prior to saprophytic invasion by other organisms. Strategies to control or eliminate inoculum in the field should focus on slowing or reducing the colonization of wheat vegetation and reducing sporulating structures on debris surfaces. An obvious target for the reduction of vegetative colonization is Type II resistance mechanisms. Reducing sporulation on debris will require a further understanding of the factors that initiate sporulation. An understanding of these processes will aid in measures to reduce potential inoculum production and survival by this fungal plant pathogen.
Publications
- Trail, F., Gaffoor, I., and Vogel, S. 2005. Ejection mechanics and trajectory of the ascospores of Gibberella zeae (anamorph Fusarium graminearum). Fungal Genetics and Biology: 42:528-533
- Guenther, J. and Trail, F. 2005. The development and differentiation of Gibberella zeae (anamorph: Fusarium graminearum) during colonization of wheat. Mycologia 97 (1): 232-240.
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Progress 01/01/04 to 12/31/04
Outputs
This year we submitted a manuscript which is a significant contribution to our understanding of fungal development in planta, particularly in regard to function of dikaryotic hyphae. Staining with acridine orange revealed that wide (dikaryotic) hyphae develop in two different paths, depending on their location in the plant. They may give rise to perithecia or give rise to monokaryotic hyphae that continue the colonization process. This result supports our previous finding that wide hyphae are a stage of development unique from thin-infecting hyphae and fungal fruiting bodies. We proposed to identify the major fungal lipids that accumulate prior to perithecium development. This objective has been completed in culture for both wild type parent (PH-1) and the developmental mutant 123C-44, which does not accumulate lipids. In planta, the objective has been completed for PH-1 with results pending for 123C-44. The major fatty acids stored in the wide hpyae in culture were
C18:2, C18:1 and C:18 saturated, mainly as triacylglycerides. Cultures induced to produce wide hyphae had nearly twice the fatty acids per dry weight mycelium that uninduced cultures had. The same profile was found with hyphae grown in the wheat stems. No fatty acids were produced in planta by the mutant. My laboratory generated a microarray based on the ESTs generated by Trail, and colleagues. We have completed experiments using these arrays to identify genes showing differential patterns of expression during development. The array represents about 18% of the genes in the genome. Compared with vegetative mycelia, 493, 450, and 326 cDNAs were differentially expressed in 4-day, 5-day and 6-day perithecia, respectively. 109 cDNAs were up-regulated in all 3 perithecial samples compared to vegetative mycelia, in which 70% were specific to the perithecia library. Based on their putative identities, cDNAs that were up-regulated in all three perithecia samples represented genes involved in
lipid catabolism, amino acid metabolism and transportation, protein transportation, post-translationally modification, and genes encoding cell wall proteins. Up-regulated genes involved in lipid catabolism included those important for fatty acid elongation, fatty acid oxidation, and metabolisms of membrane phospholipids. These studies give us a handle on genes used for nutritional support for perithecium development.
Impacts
We are moving closer to our goal of understanding inoculum formation and dispersal. we have identified candidate genes, which, if inhibited will reduce the primary inoculum in the field. We have identified a resistance mechanism. These should result in important targets for fungus control.
Publications
- Guenther, J. and Trail, F. 200-. The development and differentiation of Gibberella zeae (anamorph: Fusarium graminearum) during colonization of wheat. Mycologia, In press.
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Progress 01/01/03 to 12/31/03
Outputs
Gibberella zeae causes head blight of wheat and barley and stalk rot and ear rot of corn. As conventional control measures have not produced effective control of this devastating pathogen, a deeper understanding of the life cycle and biolgy of the fungus is necessary. Our long-term goal is to understand the production and spread of inoculum for this fungal disease. Toward that end, this year we have completed the characterization of fungal growth and sexual development in planta. We now know that the fungus infects and spreads using distinct hyphal types through the various plant tissues. We have identified an effective plant resistance response to the fungus that stops spread within the plant. We have characterized this phenomenon in a susceptible variety. We obtained funding for the genomic sequence of the fungus last year and this year we have begun to annotate it and use it to identify genes important to spore dispersal. 10 genes associated with this function have
been disrupted in the fungus and one of them is a proven pathogenicity factor. We have established a microarray based on sequenced genes (ESTs) and used it to identify approximately 100 genes uniquely expressed in the sexual stage. We have just obtained funding for a genomic microarray chip and funds to use it to explore the fungus-host interaction and fungal sexual development.
Impacts
We are moving closer to our goal of understanding inoculum formation and dispersal. we have identified candidate genes, which, if inhibited will reduce the primary inoculum in the field. We have identified a resistance mechanism. These should result in important targets for fungus control.
Publications
- Trail, F., Xu, J.-R., San Miguel, P., Halgren, R. G. and Kistler, H. C. 2003. Analysis of Expressed Sequence Tags from Gibberella zeae (anamorph Fusarium graminearum). Fungal Genetics and Biology 38:187-197.
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Progress 01/01/02 to 12/31/02
Outputs
Fungi have evolved many ways to disperse their spores. Spore dispersal is particularly important to the fungu Gibberella zeae which causes head blight of wheat and barley. To initiate the disease, this fungus fires it sexual spores (the ascospores) into the air, where they are carried to the highly susceptible flowers. We are examining the mechanism of forcible discharge of ascospores in this fungus. We have examined the role for the sugar-alcohol, mannitol, in generating the force to fire these spores. This year we have isolated and characterized the enzyme that produces mannitol, mannitol dehydrogenase and cloned the gene. Preliminary experiments to specifically mutate this gene in the fungus resulted in a partially active enzyme. We are attempting therefore, to isolate a larger clone of the gene from a genomic library. We had previously isolated a mutant strain that does not forcibly discharge it's spores, but develops normally. We have isolated a portion of the
mutated gene from this strain and sequence comparisons with the genetic sequences in GenBank indicate that it is not highly homologous with any known genes. We are in the process of cloning the entire gene.
Impacts
Gibberella zeae is dependent on the forcible discharge of its ascospores to initiate a new disease cycle each spring. The identification of possible modes of elimination of this new inoculum source would greatly reduce the impact of the head blight disease. This research is geared towards understanding the phenomenon of spore dispersal with the goal of developing new modes of control.
Publications
- Trail, F. and Xu, H. 200- Purification and characterization of mannitol dehydrogenase and isolation of the corresponding cDNA from the head blight fungus, Gibberella zeae (Fusarium graminearum). Phytochemistry, in press.
- C. Kwon and F. Trail. 2002. The mechanism of forcible ascospore discharge in Gibberella zeae. Phytopathology 92 (6) S 43.
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Progress 01/01/01 to 12/31/01
Outputs
Gibberella zeae (anamorph Fusarium graminearum) causes head scab of wheat and barley and stalk rot and ear rot of corn. As conventional control measures have not produced effective control of this devastating pathogen, a deeper understanding of the life cycle and biology of the fungus is necessary. Our long-term goal is to understand the production and spread of inoculum for head scab, and the role of the two types of spores (sexual and asexual) in completing the disease cycle. Perithecia, containing the sexual spores, form on crop debris left in the field. We have shown that specific cell types of the wheat plant are infected by mycelia, which the give rise to the perithecia. We have isolated mutants that do not produce perithecia. Using strains with mutations involved in initial stages of perithecium production or lacking perithecia, we can begin the genetic and physiological analysis of this early phase on inoculum production. Our approach is twofold: to combine
basic, laboratory research with field research aimed at the ultimate control of inoculum production. Our research combines studies of the basic biology of G. zeae with behavior of the fungus in the field. These two approaches will result in novel approaches to interruption of the disease cycle.
Impacts
Studies on the development and mechanisms of dispersal of the inoculum will provide valuable insight into effective control procedures for this disease. An understanding how field colonized vegetative host tissue supports perithecium production over the course of 1 year may lead to a change is screening for plant resistance. Identification of genes and gene products involved early in perithecium initiation and development will provide targets for control.
Publications
- Trail, F., Xu, H., Loranger, R. and Gadoury, D. 2002. Physiological and environmental aspects of ascospore discharge in Gibberella zeae. Mycologia 94:181-189.
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Progress 01/01/00 to 12/31/00
Outputs
Gibberella zeae (anamorph, Fusarium graminearum), the head blight pathogen of wheat and barley, infects during crop flowering. Therefore studies focused on the formation and distribution of the inoculum for flower infection will lead to novel and effective means of control. Ascospores, formed and forcibly discharged from perithecia, are the primary inoculum of this disease. We have focused our efforts on understanding the production and distribution of these propagules. We collected wheat and corn stubble from commercial fields year-round from 1997 to 2000 to evaluate the timing of perithecium formation in the field. Results indicate that perithecium formation is limited by average daily temperatures below 9 C and that corn stubble may be the predominant substrate for perithecium formation in G. zeae in Michigan. The data did not show a limiting high temperature for perithecium formation. We have generated 5000 tagged mutants in G. zeae by insertional mutagenesis
using a plasmid conferring hygromycin resistance. These mutants were screened for loss of the ability to forcibly discharge ascospores from perithecia. One mutant has been recovered that forms morphologically normal perithecia, but is discharge minus. Genetic analysis indicates this mutation is tagged with the plasmid. We are currently isolating the gene that has been mutated to determine its function.
Impacts
Information on weather conditions that affect generation of the primary inoculum for the head blight disease can be used in development of prediction systems for head blight epidemics. An understanding of the conditions under which inoculum is formed and disseminated is critical to developing effective means of control for this devastating pathogen.
Publications
- Andries, C., Jarosz, A., and Trail, F. 2000. Effects of rainfall and temperaturee on production of peritehcia by Gibberella zeae in field debris in Michigan. In, Proceedings of the 2000 National Fusarium Head Blight Forum, Dec 10-12, Erlanger, KY. Wheat Scab and Barley Initiative, pp.118-123.
- Trail, F. and R. Common. 2000. Perithecial development by Gibberella zeae: A light microscopy study. Mycologia 92:130-138.
- Andries, C. 2000. Ascospore discharge and perithecium development in Gibberella zeae. Master's Thesis. Michigan State University.
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Progress 01/01/99 to 12/31/99
Outputs
We are studying the production and distribution of the primary inoculum head blight of wheat and barley. Gibberella zeae, the causal agent of this disease, produces perithecia in the field on crop debris. Ascospores are produced in fruiting bodies, called perithecia, on the crop debris left in the field after harvest. The spores are forcibly discharged into the air where they are carried to infect the next year's flowers. Little is known of the process of formation of perithecia on crop debris, the timing of formation, or the mechanism of forcible discharge of ascospores. We had three objectives to study these issues. Our first objective was to characterize the pattern of colonization of stalk tissue in the mature infected plant, as this tissue will become the debris and eventually yield perithecia. Our second objective was to continue the characterization of timing and appearance of mature perithecia on field debris. Our final objective was to screen 5000 random
mutants of G. zeae, created in our lab, for loss of the ability to discharge ascospores. An understanding of the production and distribution of inoculum is vital to designing strategies for disease control. We have collected stalks from plants showing symptoms in naturally infected and inoculated fields. Tissue samples have been removed the nodes and internodal regions of these plants, fixed and embedded in paraffin for histological examination. Preliminary data suggests that mycelia that colonize the epidermal cells before harvest may be important overwintering tissue for the fungus and may give rise to the perithecia in the spring. The embedded samples will be sectioned and examined microscopically to formulate a picture of the infection pattern of the tissue that will become the crop debris. The pattern of appearance of the perithecia on wheat and corn over the last 3 years. In the last 2 years flowering has been quite early, before our collections showed perithecium production in
the field. In all three years the disease incidence has been low. We will continue to collect monthly through at least 2 more years. Data will be added as the samples are analyzed. We have generated over 5000 insertional mutants of G. zeae and screened over 3500 mutants to date for loss of discharge. Twenty of the isolates have shown a loss of discharge in preliminary trials. These putative mutants need to be tested for stability of the phenotype through meiosis. We are in the process of this analysis and continue to screen other mutants for possible loss of discharge. We have identified mannitol as the sole simple sugar found in the ascus exudates, discharged from the perithecia along with the ascospores. We hypothesize that mannitol is crucial in generation of hydrostatic pressure within the ascus. We are currently attempting to genetically disrupt the gene for mannitol dehydrogenase, the mannitol-generating enzyme.
Impacts
Gibberella zeae is the causal agent of the economically devastating wheat and barley scab disease. My research will result in novel methods for control of the primary inoculum of this disease.
Publications
- Trail, F. and R. Common. 2000. Development of perithecia in Gibberella zeae: A light microscopy study. Mycologia 92:130-138.
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