|VARGA, ELISABETH - University Of Natural Resources & Applied Life Sciences - Austria|
|WIESENBERGER, GERLINDE - University Of Natural Resources & Applied Life Sciences - Austria|
|HAMETNER, CHRISTIAN - Vienna University Of Technology|
|DONG, YANHONG - University Of Minnesota|
|SCHOFBECK, DENISE - University Of Natural Resources & Applied Life Sciences - Austria|
|STUCKLER, ROMANA - University Of Natural Resources & Applied Life Sciences - Austria|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 11/12/2014
Publication Date: 11/12/2014
Citation: Varga, E., Wiesenberger, G., Hametner, C., Ward, T.J., Dong, Y., Schofbeck, D., McCormick, S.P., Broz, K.L., Stuckler, R., Kistler, H.C. 2014. Discovery and toxicity assessment of a novel type A trichothecene produced by US isolates of Fusarium graminearum [abstract]. World Mycotoxin Forum.
Technical Abstract: The filamentous fungus Fusarium graminearum shows a widespread occurrence across temperate regions of the world and can produce several mycotoxins on almost every cereal. A large-scale survey of F. graminearum (sensu stricto) on wheat in the northern United States was conducted to investigate the population dynamics. Some isolated strains produced no deoxynivalenol (DON), nivalenol, or acetylated derivatives thereof even though they were classified as 3-acetyl-DON-chemotype based on molecular markers. Further chemical analysis of rice cultures of these strains showed no contamination with other known trichothecenes. Since the strains caused normal symptoms of Fusarium Head Blight on wheat and the volatile trichothecene precursor trichodiene could be detected, we investigated whether they might produce a novel trichothecene toxin. In LC-MS measurements the extracts of rice cultures of these isolates were compared to those of the previously characterised F. graminearum PH-1 and revealed a candidate compound (named NX-2) in significant amounts. By LC-HRMS/MS the sum formula was determined to be C17H26O6 and the typical fragmentation pattern of trichothecenes was observed. To elucidate the structure, NX-2 was purified from inoculated rice cultures by normal phase and subsequent reversed phase chromatography and 1D- and 2D-NMR measurements were performed. NX-2 is identical to 3-acetyl-DON with the exception that it lacks the keto group at C-8 and hence is classified as type A trichothecene. Similar to 3-acetyl-DON, we infer that in planta NX-2 is rapidly deacetylated and hence we produced the deacetylated form, named NX-3, by alkaline hydrolysis, purified it, and could confirm the structure by NMR. Wheat ears inoculated with the isolated strains revealed a ten-fold higher contamination with NX-3 (up to 500 mg/kg) compared to NX-2. The toxicities of the novel mycotoxins were evaluated in two in vitro translation assays using plant and mammalian ribosomes. Compared to DON, NX-3 has a slightly decreased potency to inhibit protein biosynthesis, whereas NX-2 is far less toxic and similar to 3-acetyl-DON. The main genetic difference of known type A trichothecene producing Fusarium strains and type B trichothecene producing strains is located in the TRI1 gene, encoding for a cytochrome P450 monooxygenase. Genetic analysis revealed a different TRI1 allele in our isolates and we could verify that its gene product is responsible for the hydroxylation of C-7 alone. Concluding, we have discovered a novel, previously undetected type A trichothecene produced by F. graminearum strains, elucidated its biosynthetic pathway and assessed its toxicity regarding the ability to inhibit protein synthesis.