Procedure
The Purpose of the Experiment
In our experiment, we sought to study the relationship between ferric iron, active aluminum, and mold in the soil.
Choosing the Test Sites
In order to perform this experiment, you will need to choose three 20m by 20m sites. Try to choose sites that represent different microenvironments. Divide each site into four equally sized, square quadrats. It is customary for quadrat #1 to be in the northwest section of your site, quadrat #2 to be in the northeast, quadrat #3 in the southeast, and quadrat #4 in the southwest. Within each quadrat, randomly pick three areas from which to take soil samples. (You will have 36 soil samples altogether).
Once you have collected all of your soil samples (by following the Taking the Soil Samples procedure below), test your samples for Active Aluminum and Ferric Iron using the procedure described below (Chemical Testing).
Now, look at your data and identify the quadrats with the highest aluminum levels. Then, identify the quadrats with the lowest aluminum levels. Finally, identify a medium-high quadrat, a medium quadrat, and a medium-low quadrat.
Next, repeat this process, this time considering iron levels. The 5 aluminum quadrats and the 5 iron quadrats that you have identified are the ones you will retest for both aluminum and iron. You will also test them for mold.
Taking the Soil Samples
Materials: 15cm metal soil cores; plastic bags; permanent marker
1. Determine which sites to take samples from, based on previous data concerning active aluminum and ferric iron in the soil. An ideal mix of sites would contain one very low, one low, one medium, one slightly high, and one high amount for each area of testing (the number of samples may sometimes have to be altered for practical purposes).
2. With the permanent marker, label plastic bags with these sites data Site, Quadrat, and Sample Letter so that they may be identified later.
3. Using a 15 x 2 cm3 soil corer, retrieve a sample from each sample site you selected. Place the soil core over the spot where you wish to take the sample, and push it into the ground until it reaches the first dashed line, or the 15 cm mark (see Figures 1 and 2).
4. Remove the soil from the soil core sampler and place it in its corresponding bag. Seal tightly to avoid contamination of the sample.
5. Repeat steps 3 and 4 with each site you have selected. Be sure to wash the soil core between each sample to avoid contamination.
Chemical Testing
(to be
preformed simultaneously with the Mold tests)
Materials: goggles; latex (or similar non-latex) gloves; closed-toed shoes; LaMotte Combination Soil test kit (model STH-14); soil samples; notebook and pen or other way to record data
1. Before beginning to test the soil samples, make sure you are wearing safety goggles, gloves, and closed-toed shoes.
2. Assemble the materials for the soil extraction procedure, and follow the directions for the extraction procedure. Since you will not be doing multiple tests, fill the tube with only 7 mL of Universal Extracting Solution and one level measure of soil (see figure 3). Be sure not to pack the soil into the measure. Please see the diagram to the right (Figure 12) for instructions on preparing the filter paper that you will use when making the general soil extract. Also see figures 4 and 5.
3. Follow the directions contained in the LaMotte Combination Soil Test Kit to test the soil extract for Active Aluminum and Ferric Iron (see figures 5 and 6).
4. Be sure to record the results for the tests. Don't forget to check the directions for the ppm levels that correspond to the relative levels (Low, Medium, etc.) that you will see on the Active Aluminum color chart. Irons results, which the test gives in pounds per acre, should be converted to parts per million by halving the lbs/acre.
5. Follow steps 2-4 for each soil sample collected.
Testing for
Mold
(to be preformed
simultaneously with the Chemical Tests)
Materials: Twenty-five (25) 15mL culture tubes; 1cc scoop; sterile water (you can use a jug of distilled water); notebook or other way to record data; twenty-five (25) 3M PetrifilmTM Yeast and Mold Count Plates; five (5) soil samples
1. Set out plastic test tubes in sets of five, one set for each serial dilution you will be performing.
Label the tubes 100, 10-1, 10-2, 10-3, and 10-4.
2. Fill the 100 tube in each set with 10 mL of distilled water (see figure 7), and the rest with 9 mL.
3. Take 1cc of soil from the desired sample and place the soil into the 100 tube. Cap the tube and shake vigorously for at least 25 seconds (see figure 8).
4. Transfer 1 mL of the soil solution from this tube into the 10-1 tube (see figure 9). Cap and shake vigorously for at least 25 seconds.
5. Repeat Step 3 with the remaining tubes, using the 10-1 tube to transfer to the 10-2 tube, the 10-2 tube to the 10‑3 tube, and the 10-3 tube to the 10-4 tube.
6. Set out the Petrifilm plates and label them according to Site, Quadrat, Sample Letter, and Dilution.
7. Deposit 100΅L (100 micro-litres) of each solution onto its respective plate (by site, quadrat, sample letter, and dilution). See figures 10 and 11.
8. Repeat steps 2-6 with all soil samples.
9. Collect all plates and place them in a room temperature area, out of direct sunlight, for at least two days. Sterilize your work area to ensure the prevention of disease.
10. When the prescribed waiting time is up, set out the plates. With the magnifying glass, count the molds contained in the lowest dilution of a sample at which mold are visible. Be careful not to confuse molds with yeasts the former are fuzzy blue-green patches, while yeasts are small, defined dots.
11. To estimate the amount of mold in the original 1cc soil sample, use the following equations:
# of mold visible on the lowest dilution 102 = # of
mold in the dilution tube
# of mold in dilution tube 10[ number of dilutions]
= estimated # of mold/cc
12. Record the data in your lab notebook.
13. Sterilize the work area and deposit the mold plates in a biohazard container. If possible, wash hands with antibacterial soap.
Analyzing the Data
After you have recorded the results for your chemical tests and your mold counts, examine your data. What relationship do you see between the mold levels and the iron and aluminum levels? As mold levels increase, what happens to iron and aluminum levels? Are low mold levels usually accompanied by high iron and aluminum levels?
Having trouble conducting this experiment? See our troubleshooting page, or contact us! (See the buttons at the top left of this page).
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This site was
created by
Erin Cheek, Lauren Malishchak, Katrina Szabo, and Sophia
Uddin as
part of the E.S.S.R.E.
Program at Roland Park Country School, 2005.
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Figure 1 Taking a soil sample with a soil core sampler. Click to enlarge.
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Figure 2 A 15cm high by 2cm in diameter cylindrical soil sample. Click to enlarge.
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Figure 3 Pouring 7mL of the Universal Extracting Solution into the correct test tube. Click to enlarge.
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Figure 4 Preparing the general soil extract: the filtering stage (with filter paper, funnel, and the proper tube). Click to enlarge.
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Figure 5 Mixing with a clean stirring rod. Notice how the liquid is in the LARGE depression on the spot plate. Also notice the general soil extract draining through the funnel and filter paper. Click to enlarge.
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Figure 6 Notice the Ferric Iron color chart, the .05g scoop, and the Iron testing powder and liquid. Click to enlarge.
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Figure 7 Measuring distilled water to put in the culture tubes. Click to enlarge.
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Figure 8 Shaking a culture tube to mix the soil and water. Click to enlarge.
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Figure 9 Transferring 1 mL of soil solution from one tube to the next. Click to enlarge.
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Figure 10 Taking soil solution out of a culture tube with a micropipette in preparation to plate it. Click to enlarge.
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Figure 11 Placing 100΅L of diluted solution onto a Petrifilm plate. Click to enlarge.
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Figure 12
