Methods

 

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Materials Used
Flags
Soil core sampler
bags
Black Permanent Marker
Ruler
Commercial food Wrap
60 Uhlig extractor
120 Petri dishes
Nylon mesh
Black hefty bag                             
Plastic Saran wrap
White canvas
White grinders
Balances
Filter paperGlass funnels
Ring stand
Microscope slides
Microscope
Capillary tubes
Disposable Pipettes
Methyl Green staining Solution
Slide covers
Distilled water

 

Procedure

  1. Cut 3, 15 by15 cm squares of plastic cling wrap.

  2. Do the same for nylon mesh, black plastic, black hefty plastic bag and commercial food wrap.

  3. Find 3 areas in site 4 which are exposed to the greatest amount of sunlight.

  4. Select a 1 and half meter square plot in every location and make sure to clean out all kinds of excess stuff over the soil including all detritus and leaves. However make sure not to erode any soil.

  5. For  site 1, make 5 different plots 15 by 15 cm in measure adjacent to each other and mark them Degree 1 to Degree 5 using flags.

  6. Then dig the soil core sampler into the soil 20 cm deep directly under the 1st degree of UV exposure plot.  (click for video) Separate first 10 cm of the soil and store it in the bag marked accordingly 0 - 10 "before" cm for the particular plot and site. Then put the rest of the soil in the bag marked 10 - 20 "before" for the particular site. Do the same with the 4 other degrees of UV exposures as well as for Site 2 and Site 3.

  7. After taking all the uncontrolled soil samples cover the site with the plastic cling wrap on the plot for degree 1 securing it on the sides using the flags. 

  8. Do the same with the saran wrap degree 2, mesh (degree 3),white canvas,(degree 4) and black plastic hefty bag (degree 5). Then do the same with site 2 and site 3.

  9. Leave all the plots and sites untouched for 1 day and the next day go and take soil samples as shown in step 6 except put them in the bags that say "after" instead of "before".

  10. Then test for the levels of protozoa in all the different samples, before controlling and after controlling.

  11.  Place sample of soil into the bottom of a clean, empty Petri dish; and allow to dry completely.

  12. Sift 9-10 g of the soil into a 2nd clean Petri dish using a 1 mm2 nylon screen or mesh.

  13. Add 20 ml of distilled water to saturate the soil

  14. Cover the Petri dish with its lid and allow to sit for 7 hours.

  15. Place the soil sample in a modified Uhlig extractor containing 30 ml of distilled water for 24 hours.

  16. Remove the filtrate and filter a 2nd time using 12.5 cm qualitative filter paper.

  17. Using a capillary tube, deposit 7 µl of methyl-green stain on a clean microscope slide (1 µl = 1 drop from the capillary tube). Examine under a light microscope at 40X (or 100X for qualitative observations) of the various protozoa living in the soil.

  18. Use the following equation to determine the population density of protozoa in the soil sample:

            [(# per field of view at 40X) • (total ml of water used) • 747] ¸ (grams of sifted soil ) = # of protozoa per gram of soil

***The protozoa extraction process used above complies to the Kate Brock Meyer method.