Fusarium Ear Blight is a destructive fungal disease of cereals including wheat and can contaminate the crop with various trichothecene mycotoxins. This investigation has produced a new β-glucuronidase (GUS) reporter strain that facilitates the quick and easy assessment of plant infection. The constitutively expressed gpdA:GUS strain of Fusarium graminearum was used to quantify the overall colonisation pattern. Histochemical and biochemical approaches confirmed, in susceptible wheat ear infections, the presence of a substantial phase of symptomless fungal growth. Separate analyses demonstrated that there was a reduction in the quantity of physiologically active hyphae as the wheat ear infection proceeded. A simplified linear system of rachis infection was then utilised to evaluate the expression of several TRI genes by RT-qPCR. Fungal gene expression at the advancing front of symptomless infection was compared with the origin of infection in the rachis. This revealed that TRI gene expression was maximal at the advancing front and supports the hypothesis that the mycotoxin deoxynivalenol plays a role in inhibiting plant defences in advance of the invading intercellular hyphae. This study has also demonstrated that there are transcriptional differences between the various phases of fungal infection and that these differences are maintained as the infection proceeds. 1. Introduction Fusarium Ear Blight (FEB) disease also referred to as Fusarium head scab (http://www.scabusa.org/) is a destructive fungal disease that has the potential to devastate wheat, barley, rye, oat, or maize crops just weeks before harvest. All of the major wheat producing countries (http://faostat.fao.org/) have reported serious and repeated FEB outbreaks in the past decade, making the impact of Fusarium infections a global issue. This worldwide re-emergence is believed to be driven by changes in climate and agronomic practices. CIMMYT describes FEB as a major limiting factor to wheat production across the world [1]. Over the last 10 years, the average FEB incidence in UK wheat fields was 39% (http://www.cropmonitor.co.uk/). Fusarium graminearum (teleomorph Gibberella zeae) is a filamentous ascomycete and one of the main causal agents of FEB throughout Europe, Asia, and the Americas [1, 2]. Besides dramatically reducing yield, grain quality is affected through the selective loss of albumin and gluten proteins [3]. The grains harvested from an infected crop are also contaminated with various fungal mycotoxins often making them unsuitable and/or unsafe for human consumption, animal
References
[1]
H. J. Dublin, L. Glichrist, J. Reeves, and A. McNab, Fusarium Head Scab: Global Status and Prospects, CIMMYT, Mexico, 1997.
[2]
M. McMullen, R. Jones, and D. Gallenberg, “Scab of wheat and barley: a re-emerging disease of devastating impact,” Plant Disease, vol. 81, no. 12, pp. 1340–1348, 1997.
[3]
D. Boyacio？lu and N. S. Hettiarachchy, “Changes in some biochemical components of wheat grain that was infected with Fusarium graminearum,” Journal of Cereal Science, vol. 21, no. 1, pp. 57–62, 1995.
[4]
M. Kimura, T. Tokai, N. Takahashi-Ando, S. Ohsato, and M. Fujimura, “Molecular and genetic studies of Fusarium trichothecene biosynthesis: pathways, genes, and evolution,” Bioscience, Biotechnology and Biochemistry, vol. 71, no. 9, pp. 2105–2123, 2007.
[5]
F. Wu and G. P. Munkvold, “Mycotoxins in ethanol co-products: modeling economic impacts on the livestock industry and management strategies,” Journal of Agricultural and Food Chemistry, vol. 56, no. 11, pp. 3900–3911, 2008.
[6]
S. Hook, R. Williams, C. Edwards, and G. Dodgson, “Guidelines to minimise risk of Fusarium mycotoxins in cereals,” HGCA, London, UK, 2007.
[7]
W. R. Rittenour and S. D. Harris, “An in vitro method for the analysis of infection-related morphogenesis in Fusarium graminearum,” Molecular Plant Pathology, vol. 11, no. 3, pp. 361–369, 2010.
[8]
D. W. Parry, P. Jenkinson, and L. McLeod, “Fusarium ear blight (scab) in small grain cereals - A review,” Plant Pathology, vol. 44, no. 2, pp. 207–238, 1995.
[9]
C. Jansen, D. Von Wettstein, W. Sch？fer, K. H. Kogel, A. Felk, and F. J. Maier, “Infection pattern in barley and wheat spikes inoculated with wild-type and trichodiene synthase gene disrupted Fusarium graminearum,” Proceedings of the National Academy of Sciences of the United States of America, vol. 102, no. 46, pp. 16892–16897, 2005.
[10]
C. Pritsch, G. J. Muehlbauer, W. R. Bushnell, D. A. Somers, and C. P. Vance, “Fungal development and induction of defense response genes during early infection of wheat spikes by Fusarium graminearum,” Molecular Plant-Microbe Interactions, vol. 13, no. 2, pp. 159–169, 2000.
[11]
W. M. Wanjiru, Z. S. Kang, and H. Buchenauer, “Importance of cell wall degrading enzymes produced by Fusarium graminearum during infection of wheat heads,” European Journal of Plant Pathology, vol. 108, no. 8, pp. 803–810, 2002.
[12]
N. A. Brown, M. Urban, A. M. L. van de Meene, and K. E. Hammond-Kosack, “The infection biology of Fusarium graminearum: defining the pathways of spikelet to spikelet colonisation in wheat ears,” Fungal Biology, vol. 114, no. 7, pp. 555–571, 2010.
[13]
J. C. Guenther and F. Trail, “The development and differentiation of Gibberella zeae (anamorph: Fusarium graminearum) during colonization of wheat,” Mycologia, vol. 97, no. 1, pp. 229–237, 2005.
[14]
R. H. Proctor, T. M. Hohn, S. P. McCormick, and A. E. Desjardins, “Tri6 encodes an unusual zinc finger protein involved in regulation of trichothecene biosynthesis in Fusarium sporotrichioides,” Applied and Environmental Microbiology, vol. 61, no. 5, pp. 1923–1930, 1995.
[15]
R. H. Proctor, T. M. Hohn, and S. P. McCormick, “Reduced virulence of Gibberella zeae caused by disruption of a trichothecene toxin biosynthetic gene,” Molecular Plant-Microbe Interactions, vol. 8, no. 4, pp. 593–601, 1995.
[16]
A. Cuzick, M. Urban, and K. Hammond-Kosack, “Fusarium graminearum gene deletion mutants map1 and tri5 reveal similarities and differences in the pathogenicity requirements to cause disease on Arabidopsis and wheat floral tissue,” New Phytologist, vol. 177, no. 4, pp. 990–1000, 2008.
[17]
M. Kimura, I. Kaneko, M. Komiyama et al., “Trichothecene 3-O-acetyltransferase protects both the producing organism and transformed yeast from related mycotoxins. Cloning and characterization of Tri101,” Journal of Biological Chemistry, vol. 273, no. 3, pp. 1654–1661, 1998.
[18]
O. J. Desmond, J. M. Manners, A. E. Stephens et al., “The Fusarium mycotoxin deoxynivalenol elicits hydrogen peroxide production, programmed cell death and defence responses in wheat,” Molecular Plant Pathology, vol. 9, no. 4, pp. 435–445, 2008.
[19]
I. N. Roberts, R. P. Oliver, P. J. Punt, and C. A. M. J. J. Van den Hondel, “Expression of the Escherichia coliβ-glucuronidase gene in industrial and phytopathogenic filamentous fungi,” Current Genetics, vol. 15, no. 3, pp. 177–180, 1989.
[20]
T. Tokai, H. Koshino, N. Takahashi-Ando, M. Sato, M. Fujimura, and M. Kimura, “Fusarium Tri4 encodes a key multifunctional cytochrome P450 monooxygenase for four consecutive oxygenation steps in trichothecene biosynthesis,” Biochemical and Biophysical Research Communications, vol. 353, no. 2, pp. 412–417, 2007.
[21]
R. B. Dyer, R. D. Plattner, D. F. Kendra, and D. W. Brown, “Fusarium graminearum TRI14 is required for high virulence and DON production on wheat but not for DON synthesis in vitro,” Journal of Agricultural and Food Chemistry, vol. 53, no. 23, pp. 9281–9287, 2005.
[22]
K. Y. Seong, M. Pasquali, X. Zhou et al., “Global gene regulation by Fusarium transcription factors Tri6 and Tri10 reveals adaptations for toxin biosynthesis,” Molecular Microbiology, vol. 72, no. 2, pp. 354–367, 2009.
[23]
R. P. Oliver, M. L. Farman, J. D. G. Jones, and K. E. Hammond-Kosack, “Use of fungal transformants expressing β-glucuronidase activity to detect infection and measure hyphal biomass in infected plant tissues,” Molecular Plant-Microbe Interactions, vol. 6, p. 521, 1993.
[24]
H. Buerstmayr, T. Ban, and J. A. Anderson, “QTL mapping and marker-assisted selection for Fusarium head blight resistance in wheat: a review,” Plant Breeding, vol. 128, no. 1, pp. 1–26, 2009.
[25]
C. A. Cuomo, U. Güldener, J. R. Xu et al., “The Fusarium graminearum genome reveals a link between localized polymorphism and pathogen specialization,” Science, vol. 317, no. 5843, pp. 1400–1402, 2007.
[26]
M. Urban, S. Daniels, E. Mott, and K. Hammond-Kosack, “Arabidopsis is susceptible to the cereal ear blight fungal pathogens Fusarium graminearum and Fusarium culmorum,” Plant Journal, vol. 32, no. 6, pp. 961–973, 2002.
[27]
J. Sweigard, F. Chumley, A. Carroll, L. Farrall, and B. Valent, “A series of vectors for fungal transformation,” Fungal Genetics Newsletter, vol. 44, p. 52, 1997.
[28]
M. Urban, E. Mott, T. Farley, and K. Hammond-Kosack, “The Fusarium graminearum MAP1 gene is essential for pathogenicity and development of perithecia,” Molecular Plant Pathology, vol. 4, no. 5, pp. 347–359, 2003.
[29]
R. H. Proctor, T. M. Hohn, and S. P. McCormick, “Restoration of wild-type virulence to Tri5 disruption mutants of Gibberella zeae via gene reversion and mutant complementation,” Microbiology, vol. 143, no. 8, pp. 2583–2591, 1997.
[30]
R. A. Jefferson, The GUS Gene Fusion System (A Laboratory and Reference Manual), 1989.
[31]
S. E. Ruzin, Plant Microtechniques and Microscopy, Oxford University Press, New York, NY, USA, 1999.
