Have You Ever Wondered How We Culture and Isolate for Oak Wilt?

This week the students at the U of I Plant Clinic learned how to culture and isolate the fungal pathogen, Ceratocystis fagacearum, that causes oak wilt in oak trees.

When someone suspects that their oak tree is dying from oak wilt, they can send a sample to the U of I Plant Clinic. We usually suggest that they sample from areas of the trees that are showing symptoms typical of oak wilt as well as wood that may show streaking or darkening of vascular tissue (see pictures below). We would like the sample to consist of several 1 to 2 foot long branches with at least the diameter the size of a thumb. 

Some oak leaves showing a symptoms of oak wilt.

Dark streaking of the wood caused by the oak wilt fungal pathogen.

Here students at the U of I Plant Clinic:
1.  Take notes on the condition of the oak sample.
2.  Label agar plates with sample number, date, type of agar, and OW (Oak Wilt)
3.  The bark is peeled back from the end of the branch, so that the wood is exposed under a sterile hood.
4.  They flame their knife and notch the wood into tiny wood chips, which remain attached to the branch.
5.  Then, they flame their tweezers and pick off wood chips.
6.  These wood chips are placed, very quickly, (to avoid any unwanted contamination) into PDA -Potato Dextrose Agar, plates under a sterile hood.

The agar plates are stored on a shelf in our lab, with their paperwork.  They are kept there for up to 7 to 10 days to allow time for the fungal isolates to grow.

If we were successful at isolation of the oak wilt fungus, the above pictures show what it would look like growing in our PDA culture plates.

Lastly, we take clear tape and place it on top of the fungus to "catch spores".  This clear tape is placed onto a microscope slide with a drop of water on it.  If the oak tree is infected with oak wilt (oak wilt positive), we will see the chains of spores that is seen in the picture above.  If none of these spores can be found, the oak sample will be considered "oak wilt negative". (Picture above taken by Travis Cleveland)

For more information on oak wilt, you can refer to: http://ipm.illinois.edu/diseases/rpds/618.pdf