1. Fusarium and mycotoxin analysis AVEQ - WP 5

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 P15Agrotest Fyto, Ltd., Kroměříž (Czech Republic) Ivana Polisenska

3.  Methods of analysis? WP 5: Task and questions  Which Fusarium species?  Which mycotoxins? Analyse mycotoxins and identify Fusarium species  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  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 Harvest years 2008, 2009; each year:

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

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

9. The least contaminated genotypes, 2008 28,529,5GartonSupreme 31,324,7Typhon 36,723,0Joanette 22,320,0Miku 15,315,0Atego 36,710,3Samuel 15,0 6,7Rauhhafer aus Neustadt Mean rank T-2 Mean rank DON  Ranking at three locations, 103 genotypes

10. Standards - inoculation, 2008, DON  

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

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

13. Standards - natural infection, 2008, DON  

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

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

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

17. Mycotoxin concentration is dependant on:  Region- Mainly impact of weather - (cropping practices)  Cereal crop - Impact of host resistance to different fusarium pathogens  Year - Impact of weather 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.  Bologna, 2010 Dissemination of the results  SIGA meeting 2010 (Italian Society of Agricultural Genetics)  Tulln (Austria), Worldwide Mycotoxin Reduction in Food and Feed Chains - ISM Conference 2009  Krakow (Poland), 2009  Brno (CZE), Czech and Slovak conference about Plant Protection 2009  EUCARPIA meeting, Cambridge, April 2010  EOC Ystad, 2010