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Fusarium and mycotoxin analysis

AVEQ - WP 5. Fusarium and mycotoxin analysis. WP 5: Partners. P4 Eurofins WEJ Contaminants GmbH (Germany) Scarlett Biselli, Ole Winkelmann, Simone Staiger P6 (CRA – MAC) Bergamo (Italy) Rita Redaelli

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Fusarium and mycotoxin analysis

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  1. AVEQ - WP 5 Fusarium and mycotoxin analysis

  2. WP 5: Partners • P4 Eurofins WEJ Contaminants GmbH (Germany) Scarlett Biselli, Ole Winkelmann, Simone Staiger • P6 (CRA – MAC) Bergamo (Italy) Rita Redaelli • P8 (CRA-GPG) Fiorenzuola (Italy) Valeria Terzi, Caterina Morcia • P15 Agrotest Fyto, Ltd., Kroměříž (Czech Republic) Ivana Polisenska

  3. WP 5: Task and questions Analyse mycotoxins and identify Fusarium species • Which mycotoxins? • Which Fusarium species? • Methods of analysis? • To dehulle or not to dehulle?

  4. WP 5: Methods I • DON and T-2 (Commission regulation (EC) No. 1881/2006) • ELISA (LOD for DON < 20 ppb, T-2 < 5 ppb) • LC-MS/MS • DON, T-2, HT-2, ZEA, NIV, DAS, 3-ADON, 15-ADON • harmonisation of ELISA laboratories • determination of Fusarium species by PCR (P8)

  5. WP 5: Methods II • samples are not dehulled before analysis COMMISSION REGULATION (EC) No 1881/2006

  6. Methods III - samples Harvest years 2008, 2009; each year: • DON and T-2 by ELISA 100 inoculated accessions from three locations • DON and T-2 by ELISA 100 non inoculated accessions from one location • 11 standards in two replications from all locations, both inoculated and non inoculated • DON, T-2, HT-2, ZEA, NIV, DAS, 3-ADON, 15-ADON by LC-MS/MS for standards from all locations both I and NI) In total: 2200 ELISA, 200 LC-MS/MS, 200 PCR analysis

  7. μg/kg Results - accessions (105) - 2008 - DON - inoculation - CZE, GER,ROM

  8. μg/kg Results - accessions (105) - 2008 - T-2 - inoculation - CZE, GER,ROM

  9. Mean rank DON Mean rank T-2 Rauhhafer aus Neustadt 6,7 15,0 Samuel 10,3 36,7 Atego 15,0 15,3 Miku 20,0 22,3 Joanette 23,0 36,7 Typhon 24,7 31,3 GartonSupreme 29,5 28,5 The least contaminated genotypes, 2008 • Ranking at three locations, 103 genotypes

  10. Standards - inoculation, 2008, DON D C

  11. μg/kg Standards - CZE - 2008, 2009 - DON, T-2 - inoculation - the lowest Mina, Saul - the highest Genziana, Krezus, Evora

  12. Standards - GER μg/kg -2008, 2009 - DON, T-2 - inoculation - Mina and Saul in the middle - the highest Genziana and Evora

  13. Standards - natural infection, 2008, DON D C

  14. Standards - CZE - 2008, 2009 - DON, T-2 - natural infection - low level in 2009 - the lowest Mina and Saul

  15. Standards - IT - 2008, 2009 - DON, T-2 - natural infection - very low in 2009 - Mina and Saul low

  16. Microscopic fungi • inoculum • substrate • environment conditions Perithecium Giberella zeae, 14.5.2009 Different fungi differ in their requirements on environment conditions

  17. Mycotoxin concentration is dependant on: • Year - Impact of weather • Region - Mainly impact of weather - (cropping practices) • Cereal crop - Impact of host resistance to different fusarium pathogens Influence is different for different mycotoxins

  18. Field variability, DON

  19. Field variability, T-2

  20. ELISA vs. LC-MS/MS • verification of ELISA results • analysis of other mycotoxins

  21. LC-MS/MS analysis, natural infection, CZE

  22. Dissemination of the results • EUCARPIA meeting, Cambridge, April 2010 • Bologna, 2010 • Tulln (Austria), Worldwide Mycotoxin Reduction • in Food and Feed Chains - ISM Conference 2009 • Krakow (Poland), 2009 • SIGA meeting 2010 (Italian Society of Agricultural Genetics) • Brno (CZE), Czech and Slovak conference about Plant Protection 2009 • EOC Ystad, 2010

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