Title:

The Blight Fungus in American Chestnut Trees

Author:
Stevie Hall
Science Research I
Mod 3
May 4, 2018
Abstract:
So in my investigation I went out and took two separate samples of Chestnut trees. I took
samples of the chestnut trees growing in the front of our school and samples of the chestnut trees
in our own little chestnut orchid. The procedure was pretty simple, I took a 96 well plate and
took 4 samples of each tree canker, North, South, East, and a West sample. We took samples
from 3 trees out front and 3 trees out in the orchid. We used a borer to take small circular
samples. We then placed them in the wells and put tape over them to keep them in place. Once
we took them and labeled them we put them into the fridge until we were ready to inoculate.
Once we inoculated our samples, we set them out at room temperature to grow their fungus. For
my first sample from the front of the school, all my samples grew a secondary fungus. Trees A,
and C both grew green and brown fungus. Tree C had some yellow fungus as well. What really
stood out was tree B. Tree B grew brown fungus. The North and West samples grew close
together but never touched. This meant that there are two different strains of blight fungus
growing in one tree.
Introduction:
The focus of my experiment was to see if our American chestnut trees had any
different types of strains of the blight fungus. The American chestnut tree is a large
monoecious deciduous tree of the beech family. (Carpel late flowers are always found on
the same plant, falling off at maturity). It was native to Eastern North America. Before the
species was devastated by the chestnut blight, it was one of the most important forest trees
throughout its range, and it was considered the finest chestnut tree in the world. It was an
essential component of the eastern U.S. forest ecosystem. They stood up to 100 feet tall,
and numbered in billions. As a late flowering, reliable, and extremely productive tree, it
was unaffected by seasonal frosts, making it the single most important food source for a
wide variety of wildlife. Many rural communities depended on it for its nut harvest as an
annual cash crop to feed livestock. It was straight grained, easily worked, lightweight, and
highly rot resistant. This made it ideal to work with. The blight fungus was accidentally
imported from Asia to the United States at the beginning of the 20th century. It was first
detected in New York in 1904. The blight being a wound pathogen, it enters the tree
through an injury in the bark. It spreads to the underlying plant tissue and wood, killing the
tissues as it advances. The flow of nutrients is eventually choked off to and from sections
of the tree above the infection, killing the tree above ground. There is more than one strain
of blight affecting our chestnut trees. This means they are vegetatively compatible.

Materials and Method/Procedure:

 Chestnut trees
 96 well plates (holds samples)
 Borer (takes small samples from trees)
 Perti dishes (where samples grow their fungus)
 Bleach (used to sterilize samples)
 Tape (holds samples in well plates)
 Tweezers (used to safely grab samples)
 Isopropyl alcohol (sterilize tweezers)
 Lighter (sterilize tweezers)
 Logbook (take notes, record data)
 Millipore water (sterilize samples)
 Goggles (protect eyes)
 100 ml beaker (hold bleach)
 Taking samples of Chestnut trees:
1. Collect all materials needed to get samples from chestnut trees; Borer, 96 well plate,
tape.
2. Go out to chestnut trees and find cankers in trees.
3. Take samples of North, South, East, West sides of cankers.
4. Put them in 96 well plate,
5. Cover with tape to keep from moving

Inoculating your samples:

1. Wash your hands
2. Get your samples
3. Put on your goggles
4. Submerge your samples in your bleach solution for ten minutes
5. Flush them with water and let that sit for two minutes
6. Once they have sat, remove them from the water and put them in your petri
dishes, by N, S, E, and W.
7. Make sure to sterilize your tweezers each time with alcohol and a lighter.
8. Once finished put the lids on and tape them together
9. Leave out at room temperature for a few days to grow its fungus.

Results:

This is from tree A. You can see not all my samples grew. My East sample grew the best and
there was a secondary fungus growing in my sample. (Sample is from out front of school).
This is from tree B. you can see clearly here that my samples from North and West grew close
together but never touched. This means that they are two different strains of the blight fungus.
(Sample is from out front of school).

This is tree C. A secondary fungus has grown, but you can see that all my samples grew more
fungus. Nothing grew close together enough to show whether they are different types of the
blight fungus. (Sample is from out front of school).
This is tree A from my Chestnut orchid sample, here you can see the small dots and the trail of
liquid from my northern sample and the eastern and western sample.

This is tree B from the chestnut orchid, this one grew the most fungus and I believe has a
secondary fungus.
This is tree C from the chestnut orchid. What really interested me about this sample is the bright
red fungus growing from the southern sample. Not sure what it is but it’s really cool.

Discussion:
So looking at my results there was at least one tree that showed there was vegetative
compatible fungus growing in our trees. That was tree B, from the front of our school. The
sample that I took from the northern side and the sample that I took from the western side grew
close together but never touched. This thin line between the two growing fungi meant that there
is more than one type or strain of the blight fungus growing in one tree. When looking at my
samples from the chestnut orchid in the back of the school you can see that tree C has a bright
red spot by the southern sample. Not sure what exactly it is but looks very interesting. Tree A
from the Chestnut orchid looked like water got into my sample, or some type of liquid formed at
my north, east and western samples. Not sure what this is either but it yet again looks interesting.
Tree b was the one that grew the most fungus from the orchid. It had possibly a secondary
fungus which may have been the reason for the copious amount of fungi in the petri dish.
Sources of error could be cross contamination from not sterilizing all my tools properly. I could
also have cross contaminated my samples when I was taking the samples from the trees. I could
have forgotten to sterilize the borer when I moved to the next spot or tree. Ways to improve the
investigation could be disposable tools that way you could throw away after one use that way
you wouldn’t have to sterilize over and over after each sample was taken.

Conclusion:
From my experiment I found that more than one type of fungus can grow in one tree.
Tree B from the sample I took out front of the school proved that when the fungus grew close
together but never touched each other. Tree C from the orchid sample grew a bright red fungus
which I thought was interesting. Tree A from that set of samples looked like water had gotten in
the petri dish or some type of liquid fungus grew. Some future areas of investigation could
include testing these samples even further to see what kind of strain of the blight fungus they are.
Another way you could further the investigation is to see how the fungus effects other trees and
how they tolerate it.

References:
“The American Chestnut Tree.” About the Tree, www.acf.org/the-american-chestnut/.