Grain Spawn Inoculum Protocols
I’ve been following the methods described by Pat Lipps at Ohio State with some
modifications (Phytopathology 88:1078-1086).
1. Calculate the amount of corn grain needed for field inoculation.
Rate = 36 g/sqm or 3.3 g/sqft
We buy the cheaper whole kernel feed corn. It comes in 50 lb bags,
so convert grams to lbs to know how many bags you will need.
2. Determine how many bags you might night need to prepare for your test.
We get our spawn bags from Fungi Perfecti Olympia, WA (800) 780-9126.
We have a very large autoclave and have found that when we use the spawn bags we can double the amount of grain previously needed than when using the mason jars thus dividing the total number of units (bags/jars) needed in half. The bags are quite a bit larger than the jars.
Use 1000 g corn per bag.
Place corn kernels in spawn bags with tap water at a rate of 1.3 g grain/ml water. Allow the grain/water to soak in bags overnight. This step is essential as saturated grain autoclaves better killing more contaminants and the moisture is essential for Fusarium growth.
3. Neatly fold over the top inch of each bag a couple of times and secure with autoclave tape. THIS IS CRITICAL, AS THE BAGS WILL NOT SEAL WELL IF THERE ARE CREASES IN THE PLASTIC.
The bags will become more rigid after autoclaving. If the top part of the bag is not protected, it will crinkle making it difficult to get a good seal. If it doesn’t look like you are getting a good seal, seal it again.
4. Autoclave corn in grain spawn bags for 2 consecutive days.
5. Inoculate bags with F. graminearum. (See F. graminearum culture methods).
6. Seal bags with sealer. It might take two people to do this, one to hold and stretch the bags tight and another to make the seal. Follow the sealers instructions.
7. Sit bags upright on shelf for incubation. In several days, as the fungus begins to colonize the grain and respire, the bags will fill out.
8. Begin to shake the grain in the bags, 2-3 days after inoculating. This helps ensure uniform colonization of the grain in the bag and prevents clumping. Monitor grain colonization daily.
9. When all kernels are colonized take bags to field.
10. Plan to incubate the grain for at least 2 weeks prior to taking
to the field. We keep the jars in the seed house at room temp. The colonized grain
should be covered with white to pinkish mold when it is ready to spread. Shake
bottles daily to keep the kernels from clumping completely.
11. It is very important to keep the colonized grain hydrated after spreading in field.
Initially we wet the grain by running irrigation for 15 minutes. Plan to irrigate 3x
daily (8 am, noon and 8 pm) for a short period of time, especially if dry weather
occurs. Wheat is at GS 7.
When you start to see abundant perithecia forming on grain, return to regular
irrigation schedule for disease development.