EMRE YÖRÜK, Aylin GAZDAĞLI, GÜLRUH ALBAYRAK

Effects of Growth Conditions on tri4 Gene Expression in Fusarium culmorum

Expression of tri4, found in the tri5 gene cluster, is essential for DON production. In this study, effects of different growth conditions on tri4 expression, as well indirectly on DON production, were investigated in F15 isolate of Fusarium culmorum via qPCR (real time polymerase chain reaction). Control group was grown on potato dextrose agar (PDA) at 25°C (pH 5.6). The effects of pH 3.0 and -7.0 were examined on cultures grown at 25°C. Moreover, 0.5 mM hydrogen peroxide (H2O2) was concurrently added to medium. High quality (A260/280= 1.9-2.0) and quantity (2-3µg/µL) of total RNAs were isolated from all groups. β-tubulin expression was used as internal control and relative quantification values were recorded. tri4 expression was detected in all experiments except F15 grown on pH 3.0. Cp values were calculated as 22.26±1.14-26.84±4.79. tri4 expression levels in experiments were lower than control. Their ΔΔCT and 2-ΔΔCT values were 0-5.54 and 0-0.582, respectively. While maximum tri4 expression was recorded in control, minimum expression was detected in the conditions consisting of pH 5.6 and at 15°C. Findings showed that different pH and temperature values and supplementation of H2O2 resulted in decreasing of tri4 expression. Also, it was detected that acidic pH was a potential repressor for DON production. Findings support the importance of kit development requirement for mycotoxin detection based on gene expression analysis in the field or harvested crops.

Keywords:

Fusarium culmorum, qPCR, tri4 gene expression,

PDF

___

Bai G, Shaner G (2004). Management and resistance in wheat and barley to Fusarium head blight. Annual Review of Phytopathology, 42: 135-161.
Boutigny AL, Barreau C, Atanasova-Penichon V, Verdal-Bonnin MN, Pinson-Gadais L, Richard-Forget F (2009). Ferulic acid, an efficient inhibitor of type B trichothecene biosynthesis and Tri gene expression in Fusarium liquid cultures. Mycological Research, 113: 746-753.
Chandler EA, SimpsonDR, Thomsett MA, Nicholson P (2003). Development of PCR assays to tri7 and tri13 trichothecene biosynthetic and characterisation of chemotypes of Fusarium graminearum, Fusarium culmorum and Fusarium cerealis. Physiological and Molecular Plant Pathology, 62: 355-367.
Desjardins AE, Proctor RH (2007). Molecular biology of Fusarium mycotoxins. International Journal of Food Microbiology, 119: 47-50.
Foroud NA, Eudes F (2009). Trichothecenes in cereal grains, International Journal of Molecular Sciences, 10: 147-173.
Girgin G, Başaran N, Şahin G (2001). Dünya’da ve Türkiye’de insan sağlığını tehdit eden mikotoksinler. Türk Hijyen ve Deneysel Biyoloji Dergisi, 58 (3): 97-118.
Gutleb AC, Morrison E, Murk AJ (2002). Cytotoxicity assay for mycotoxins produced by Fusarium strains. Enviromental Toxicology and Pharmocology, 11: 309-320.
Jennings P, Coates ME, Walsh K, Turner JA, Nicholson P (2004a). Determination of deoxinivalenol- and nivalenol-producing chemotypes of Fusarium graminearum isolated from wheat crops in England and Wales. Plant Pathology, 53: 643-652.
Jennings P, Coates ME, Turner JA, Chandler EA, Nicholson P (2004b). Determination of deoxinivalenol and nivalenol chemotypes of Fusarium culmorum isolates from England and Wales by PCR assay. Plant Plathology, 53: 182-190.
Kimura M, Tokai T, O’Donnell K, Ward TJ, Fujimura M, Hamamoto H, Shibata T, Yamaguchi I (2003). The trichothecene biosynthesis gene cluster of Fusarium graminearum F15 contains a limited number of essential pathway genes and expressed non-essential genes. FEBS Letters, 539: 105-110.
Kimura M, Tokai T, Takahashi-Ando N, Ohsato S, Fujimura M (2007). Molecular and genetic studies of Fusarium trichothecen pathways gene and evolution. Bioscience, Biotechnology, and Biochemistry, 71: 2105-2123.
Lauren DR, Smith WA (2001). Stability of Fusarium mycotoxins nivalenol, deoxynivalenol and zearelenone in ground maize under typical cooking conditions. Food Additives and Contaminants, 18: 1011-1016.
Lee T, Han YH, Kim KH, Yun SH, Lee YW (2002). Tri13 and Tri7 determine deoxynivalenol- and nivalenol- producing chemotypes of Gibberella zeae. Applied and Environmental Microbiology, 68: 2148–2154.
Livak JK, Schmittgen TD (2001). Analysis of relative gene expression data using real time quantitative PCR and the 2-ΔΔCT method. Methods, 25: 402-408.
McDonald T, Brown D, Keller NP, Hammond TM (2005). RNA silencing of mycotoxin production in Aspergillus and Fusarium species. Molecular Plant-Microbe Interactions, 18 (6): 539-545.
Merhej J, Boutigny AL, Pinson-Gadais L, Richard-Forget F, Barreau C (2010). Acidic pH as a determinant of TRI gene expression and trichothecene B biosynthesis in Fusarium graminearum. Food Additives and Contaminants, 27 (5): 710-717.
Özer N, Soran H (1991). Fusarium species of Turkey. Hacettepe Üniversitesi Eğitim Fakültesi Dergisi, 6: 259-271.
Parry DW, Jenkinson P, McLeod L (1995). Fusarium ear blight (scab) in small grain cereals-a review. Plant Pathology, 44: 207-238.
Pinson-Gadais L, Richard-Forget F, Frasse P, Barreau C, Cahagnier B, Richard-Molard D, Bakan B (2008). Magnesium represses trichothecene biosynthesis and modulates Tri5, Tri6, and Tri12 genes expression in Fusarium graminearum. Mycopathologia, 165: 51-59.
Ponts N, Couedelo L, Pinson-Gadais L, Verdal-Bonnin MN, Barreau C, Richard-Forget F (2009). Fusarium response to oxidative stress by H2O2 is trichothecene chemotype-dependent. FEMS Microbiology Letters, 293: 255-262.
Scherm B, Orrù M, Balmas V, Spanu F, Azara E, Delogu G, Hammond TM, Keller NP, Migheli Q (2011). Altered trichothecene biosynthesis in TRI6-silenced transformants of Fusarium culmorum influences the severity of crown and foot rot on durum wheat seedlings. Molecular Plant Pathology, 12 (8): 759-771.
Scherm B, Balmas V, Spanu F, Pani G, Delogu G, Pasquali G, Migheli Q (2013). Fusarium culmorum: causal agent of foot and root rot and head blight on wheat. Molecular Plant Pathology, 14 (4): 323-341.
Sudakin DL (2003). Trichothecenes in the environment: relevance to human health. Toxicology Letters, 143: 97-107.
Wagacha JM, Muthomi JW (2007). Fusarium culmorum: Infection process, mechanisms of mycotoxin production and their role in pathogenesis in wheat. Crop Protection, 26: 877-885.
Wang JH, Li HP, Qu B, Zhang JB, Huang T, Chen FF, Liao YC (2008). Development of a generic PCR detection of 3-acetyldeoxy-nivalenol-, 15-acetyldeoxynivalenol- and nivalenol-chemotypes of Fusarium graminearum clade. International Journal of Molecular Sciences, 9: 2495–2504.
Yli-Mattila T, Rämö S, Hietaniem, V, Hussien T, Carlobos-Lopez AL, Cumagun CJR (2013). Molecular quantification and genetic diversity of toxigenic Fusarium species in Northern Europe as compared to those in Southern Europe. Microorganisms, 1: 162–174.
Yörük E, Albayrak G (2014). Tri4 and tri5 gene expression analysis in Fusarium graminearum and F. culmorum isolates by qPCR. Plant Pathology Journal, 13(2): 133-138.
Yörük E (2014). Quelling of trichothecene production in Fusarium species. IU Graduate School of Science and Engineering, PhD Thesis, Istanbul.

Gaziosmanpaşa Üniversitesi Ziraat Fakültesi Dergisi-Cover

ISSN: 1300-2910
Yayın Aralığı: Yılda 3 Sayı
Başlangıç: 1985
Yayıncı: Tokat Gaziosmanpaşa Üniversitesi Ziraat Fakültesi Dergisi Yayın Ofisi