Inoculum production for field experiments with artificial Fusarium inoculation M. Herrmann

Material: Oat cv. Jumbo or Pergamon, or other (wheat or triticale may also be suitable); about 15kg (3 kg per Fusarium isolate)

Fusarium isolates: (from the former BBA-collection – now JKI)

Transfer the cultures to SNA-agar for multiplication of the different isolates.

Preparation of seeds for inoculum production Soaking a few minutes and boiling of oat seeds for 15 min with water. Transferring the boiled seeds with less water into autoclaving bags and autoclaving it two times. Cool off the bags to room temperature. Under sterile conditions: Put some peaces of agar with the Fusarium strain into the autoclaved seeds in the bag. Do not close the bag completely, to permit some gas exchange.

Let the fungus grow under dark conditions and room temperature until the whole seedlot is completely occupied by the fungus. Put the seeds with mycelium to cleaned basins for a slowly drying (over a few weeks). Seeds with mycelium should be covered by cloths, to slow down speed of drying. Mixing the samples by hand all two days.

Dryed samples can be stored in refrigerator for several years Counting of spore density after washing the inoculum substrate in de-ionised water and tween-20 Adjust the spore amount for each Fusarium isolate to get a inoculum density of 1 Mio spores per ml

Spore counting and calculation in 2008: SpeciesNumber of spores found for each isolate Fusarium sporotrichioides Fusarium avenaceum Fusarium culmorum Fusarium langsethiae Fusarium graminearum In Romania and Germany 60l inoculum (3 dates with each 20l) with about Sporen/ml

Analysis of the mycotoxin content of the 2008 inoculum mixture (eurofins, LC/GC; HPLC-MS/MS): Vomitoxin (DON), NIV, 3-ADON, 15-ADON, DAS, FX, <20μg/ kg Zeralenone (ZON)6000μg/ kg HT-28500μg/ kg T280000μg/ kg Neosolaniol600μg/ kg Are the Fusarium strains multiplied mycotoxin producer?

Conclusions: DON- and NIV-producing Fusarium strains need to be involved. Inoculum production method have to be improved.

Many thanks to the sponsors: The European Commissions ( Council Regulation (EC) no. 870/2004), Peter Koelln KGaA, Emco spol. s r. o., Gemeinschaft zur Förderung der privaten deutschen Pflanzenzüchtung e.V. (GFP) Meeting additionally supported by the Association of the Friends and Promotors of the Julius Kühn-Institute (GFF) and Peter Koelln KGaA