HURO/0901/147 2.2.2 Szeged - Timisoara axis for the safe food and feed SZETISA1

1. HURO/0901/1472.2.2Szeged-Timisoara axisfor the safe food and feedSZETISA1 2nd Progress Meeting Szeged, September 22-23, 2011 DisclaimerThe content of this page does not necessarily represent the official position of the European Union.

2. Sampling from maize genotypes for the 3rd reporting period Csiszár Jolán And members of dept. Of Plantbiology, University of SZEged, Mesterházy Ákos, Toldiné Tóth Éva , Cereal Research Nonprofit Kht, Szeged DisclaimerThe content of this page does not necessarily represent the official position of the European Union.

3. Fusariumearrots • Fusarium ear rots are the most dangerous food and feed safety challenges of maize. • Fusarium and Gibberella ear rots are the two main diseases, but more than ten further Fusarium spp. cause ear rots. • Natural infection is caused by a mixture of the local Fusarium spp., normally one is dominating. • In Germany 13 Fusarium species were isolated from naturally infected maize, the prevalent species changed between years according to weather. Á. Mesterházy, M. Lemmens:Breeding for resistance against Fusarium ear rots in corn - a review

4. FusariumearrotsIn Hungary, 16 Fusariumspp. were identified in 1974 and 1975 In 1974 (humid),554 isolates: 29% were F. graminearum, 27% F. verticillioides, 6% F. sporotrichioides, 4% F. culmorum, 3.6% F. semitectum, 3% F. fusarioides, 1.62% F. poae, and in low amounts further 8 Fusarium spp. In 1975 (dry), 645 isolates: 69% were F. verticillioides (earlier names: F. moniliforme and F. moniliforme var. subglutinans), 28% were F. graminearum and all others were represented by 1-2 isolates only. F. g. F. v. F. c.

5. The Fusarium spp. produce a large number of chemically very different mycotoxins F. acuminatum T2, MON, HT2, DAS, MAS, NEO, BEA F. anthophilum BEA F. avenaceum MON, BEA F. cerealisNIV, FUS, ZEN, ZOH F. chlamydosporum MON F. culmorumDON, ZEN, NIV, FUS, ZOH, AcDON F. equiseti ZEN, ZOH, MAS, DAS, NIV, DAcNIV, FUS, FUC, BEA F. graminearum DON, ZEN, NIV, FUS, AcDON, DAcDON, DAcNIV F. heterosporum ZEN, ZOH F. nygamai BEA, FB1, FB2 F. oxysporum MON, BEA F. poae DAS, NIV, FUS, MAS, T2, HT2, NEO, BEA F. proliferatum FB1, BEA, MON, FUP, FB2, F. sambucinum DAS, T2, NEO, ZEN, MAS, BEA F. semitectum ZEN, BEA F. sporotrichioides T2, HT2, NEO, MAS, DAS F. subglutinans BEA, MON, FUP F. tricinctum MON, BEA F. verticillioides FB1, FB2, FB3 a Fusarium nomenclature according to Nelson et al. (1983). b Bold letters indicate the important mycotoxins. Abbreviations: AcDON – Mono-acetyldeoxynivalenols (3-AcDON, 15-AcDON); AcNIV – Monoacetylnivalenol, (15-AcNIV); BEA – Beauvericin; DiAcDON – Di-acetyldeoxynivalenol (3,15-AcDON); DAcNIV – Diacetylnivalenol (4,15-AcNIV); DAS – Diacetoxyscirpenol; DON – Deoxynivalenol (Vomitoxin); FB1 – Fumonisin B1; FB2 – Fumonisin B2; FB3 – Fumonisin B3; FUP – Fusaproliferin; FUS – Fusarenone-X (4-Acetyl-NIV); FUC – Fusarochromanone; HT2 – HT-2 toxin; MAS – Monoacetoxyscirpenol; MON – Moniliformin; NEO – Neosolaniol; NIV – Nivalenol; T2 – T-2 toxin; ZEN – Zearalenone; ZOH – zearalenols (α and β isomers). (Mesterházy, LemmensBreeding for resistance against Fusarium ear rots in corn)

6. The pathogenicity of the Fusarium spp. • The pathogenicity of the Fusarium spp. in corn is very variable. • F. graminearum and F. culmorum are highly pathogenic; the very aggressive isolates are present in a considerable proportion. F. verticillioides and the other Fusarium spp. display lower pathogenicity. • The Fusarium spp. produce a large number of chemically very different mycotoxins. • The main toxins are: T2- T-2 toxin, DON – Deoxynivalenol , ZEN – Zearalenone, HT2 – HT-2 toxin, Fumonisins

7. The effect of toxins • Fusarium toxins have been shown to cause a variety of toxic effects in both experimentalanimals and livestock, on some occasions have beensuspected to cause toxicity in humans. • T-2-and HT-2 toxin often occur together in infected cereals. T-2 toxin could inhibit synthesis of DNA and RNA, proteins, affected the permeability of cell membranes, caused changes in the phospholipid turnover, and haemolysis of erythrocytes, increased lipid peroxides in the liver of rats, caused apoptosis. • Ascorbic acid, α-tocopherol , selenium, glutathione precursors have a protective effectagainst lipid peroxidation induced by T-2 toxin.

8. Fusariuminfection The severity of natural infection varies strongly fromyear to year • with the locations; • with length of the vegetation period (earlier hybrids normally show less ear rot infection and less toxin contamination . Late-sown or later hybrids tend to be more infected by F. graminearum and contaminated by DON than early ones. • Good agronomy praxis can also decrease the risk of fumonisin contamination . • The weather conditions - may affect Fusarium spp. in different ways. • Maize researchers must use artificial inoculation methods to inoculate the plant material with fungal spores.

9. Fusariumresistance • Until the discovery of the mycotoxins, ear rot diseases were regarded as a minor problem. • At present, the majority of the inbreds and hybrids are susceptible. • Natural infection is caused by a mixture of the local Fusarium spp., normally one is dominating. • There is significant or highly significant resistance differences to the Fusarium spp. used.

10. BreedingforFusariumresistance • New breeding programmes have been started with the aim of breeding more resistant hybrids available for human and animal consumption as food and feed safety problems have emerged and the economic losses due to Fusarium ear rots have multiplied. • Artificial inoculation is used instead of natural infection.

11. MaizeFusariumresistancebreedingprogrammeinCereal Research Nonprofit Kht. (Prof. Mesterházy Ákos, Dr. Tóth Éva) • Testing the kernel resistance of maize hybrids for different Fusarium ssp. By the toothpick method: • The middle of the ear will be inoculated 10 das after midsilking. • The spreding of the infection will be rated at harvest: the infected cob area is estimated as percentage for the about 20 cobs per row infected.

12. Samplingfrommaizegenotypesforelementcontent and toxin analysisCereal Research Nonprofit Kht. • I. sampling: 26.08. 2011., Újszeged • The area was not treated with fungicid • Two selected hybrids: Sarolta: earlier was more resistant to Fusarium Boglár: earlier was more sensitive to Fusarium • Hybrids are sown in 7 m long rows with 20 cm plant distance. Five rows from each: one row was control and the four replicates were treated with F. graminearum, F.culmorum and F. verticillioides isolates.

13. Samplingfrommaizegenotypesforelementcontent and toxin analysisCereal Research Nonprofit Kht. • One parcel for each genotype • The order of inoculted lines: F.g., F.c., C, F.v.1 (4x) The most pathogenic was F. culmorum, followed by F. graminearum and the least pathogenic was F. verticillioides.

14. Samplingfrommaizegenotypesforelementcontent and toxin analysisCereal Research Nonprofit Kht. One cob with the leaf and a piece of stem was cutted out.

15. Samplingfrommaizegenotypesforelementcontent and toxin analysisCereal Research Nonprofit Kht. • Three cobs from each row, in 3 replicates

16. Samplingfrommaizegenotypes

17. Samplingfrommaizegenotypesforelementcontent and toxin analysisDept. of PlantPhysiology The cobs, leaves and stems were separated in the laboratorium

18. Samplingfrommaizegenotypesforelementcontent and toxin analysisDept. of PlantPhysiology The cobs, leaves and stems were separated in the laboratorium

19. Samplingfrommaizegenotypesforelementcontent and toxin analysisDept. of PlantPhysiology Samples from leaves were taken for further physiological investigations, materials are stored on -80 °C.

20. Samplingfrommaizegenotypesforelementcontent and toxin analysisCereal Research Nonprofit Kht. II. sampling: at the harvest 13.09. 2011., Újszeged • Onother 3-3 cobs from each row, in 3 replicates. The samples are stored in 25 °C.

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23. The effect of toxins II • DON inhibits the synthesis of DNA and RNA and protein synthesis at the ribosomal level. Thetoxin has a haemolytic effect on erythrocytes. An acute dose of DON can induce vomiting(emesis) in pigs, whereas at lower concentrations in the diet it reduces growth and feedconsumption (anorexia). Both effects, which are also seen with other trichothecene toxins, arethought to be mediated by affecting the serotonergic activity in the CNS or via peripheral actions on serotonin receptors