Partial Characteriztion of the biodegrading ability of the fungi Xylaria
sp. and its four mutants on natural rubber, chicken feathers and
1. to determine if Xylaria sp mutants and wildtype can
degrade/consume/ assimilate natural rubber as a carbon source
2. to determine if Xylaria sp mutants and wildtype can
degrade/consume/ assimilate chicken feathers as a carbon and
3. to determine if Xylaria sp mutants and wildtype can
degrade/consume/ assimilate polyurethane as a carbon source
4. to compare the degradation capability of the wild type and the
Outline of methodology
I. Preparation of Inoculum
Isolate the Xylaria sp. by culturing it in a Potato Dextrose Agar
(PDA) medium. Adjust to pH 5 and incubate at 25˚C. After 2-3 days,
transfer the fungi into test media.
II. Preparation of Pollutants
- Cut 1x1 cm squares from plastic cups.
B. Chicken feather
- Obtain fresh feathers from Gallus gallus sp. Wash and
sterilize in an autoclave.
C. Natural Rubber
- Obtain a unused rubber latex glove. Cut 1x1 cm of the
III. Preparation of Test Media*
A. For plates:
Prepare 2 sets of plates containing 20 ml PDA in triplicate
(6 plates per mutant/wild type, per pollutant tested). Add 0.5%
glucose in set A and add 0.5% of the liquid pollutant in set B.
Adjust to pH 5 by adding small amounts of either 0.1M NaOH or
0.1M HCl. Inoculate the fungi using agar discs from the PDA
plate described in I. Agar discs will be obtained by cutting a 2-3
day-old inoculum on the peripheral area of a colony using a 5-8
mm diameter cork borer. The agar discs will be transferred to
the PDA plates of sets A and B by using a sterile toothpick.
Incubate at 25˚C.
Observe the colony growth in set B and compare it always
with set A. Set A will be the control group and it would indicate
whether the fungi transferred is active. Then measure the colony
growth by its diameter.
B. For flasks and test tubes:
Prepare 2 sets of flasks containing 50 ml Mineral Medium
each, in triplicate. Add 0.5% glucose in set A and B. Adjust to pH
5 by adding small amounts of either 0.1M NaOH or 0.1M HCl.
Then add the solid pollutant in set B only. Inoculate the fungi by
using ………….. When all the glucose has been used up and the
fungi had grown into a considerable mass as examined visually,
add another MM + 0.5% glucose in set A only, leaving the set B
flasks to utilize the solid pollutants as the sole carbon source.
The extent of colonization should be carefully examined every
day until rate of colony growth can be predicted (growth in
mm/day). But if otherwise, continue adding the MMG (mineral
medium+0.5%glucose) to both sets of flask until the fungi has
grown and thrived. Incubate for 50-80days, with the flasks in a
room with more or less 250C in temperature, under shaking
conditions, while the test tubes in an incubator with 25 0C in
temperature as well.
IV. Remove solid pollutants from the culture medium and examine
under a scanning electron microscope (SEM).
* - this step is intended for each mutant and for the wild type. Since we
have 4 mutant strains and a wild type, this step will be repeated five times
multiplied with the number of pollutants to be used.