CORNELL Composting - Observing Compost Microorganisms

Observing Compost Microorganisms

Introduction

Observe the microbial communities in your compost over the course of several weeks or months as the compost heats up and then later returns to ambient temperature. Can you identify differences in microbial communities at various stages of the composting process?

Materials

compound microscope
.85% NaCl (physiological saline)
methylene blue stain (Prepare stain by adding 1.6 g methylene blue chloride to 100 ml of 95% ethanol, then mixing 30 ml of this solution with 100 ml of 0.01% aqueous solution of KOH)

Procedure

Make a wet mount by putting a drop of water or physiological saline on a microscope slide and transfering a small amount of compost to the drop. Make sure not to add too much compost or you will not have enough light to observe the organisms.
Stir the compost into the water or saline (the preparation should be watery) and apply a cover slip.
Observe under low and high power. You should be able to find many nematodes (they should be very wiggly), flatworms, rotifers (notice the rotary motion of cilia at the anterior end of the rotifer and the contracting motion of the body), mites, springtails and fast-moving protozoans. Pieces of fungi mycelia can be seen, but might be difficult to recognize. Bacteria can be seen as very tiny, roundish particles, which seem to be vibrating in the background.
If you want to observe the bacteria directly, you can prepare a stained slide and observe the slide using a 100X oil immersion lens. To prepare a stained slide, mix a small amount of compost with a drop of physiological saline on a slide. Spread with a toothpick. Let the mixture air dry until you see a white dried film on the slide. Next fix the bacteria to the slide by passing the slide through a hot flame a few times. Stain the slide using methylene blue stain. Flood the slide with the methylene blue stain for one minute and then rinse with distilled water and gently blot dry using blotting or filter paper.
Fungi and actinomycetes may be difficult to recognize with the above technique because the entire organism (including the mycelium, reproductive bodies and cells) will probably not remain together. Fungi and actinomycetes will be observed best if you can find fungal growth on the surface of the compost heap. The growth looks fuzzy, powdery, or like a spiderweb. Lift some compost with the sample on top, and and prepare a slide with cover slip to view under the microscope. You should be able to see the fungi well under 100X and 400X. The actinomycetes can be heat-fixed and gram-stained to view under oil immersion at 1000X.
To separate nematodes, rotifers, and protozoans, a continuous column of water leading from the compost to the collection vial is necessary, and the following adaptation of the above method should be used: The compost is put into a beaker with the screen stretched across the top and taped in place. The beaker is then turned over into the funnel. Plastic tubing is placed at the end of the funnel stem and a screw clamp is placed a few inches below the end of the funnel stem on the pastic tubing.
The plastic tubing should lead into a collection vial or small beaker. The clamp is closed and water is poured into the funnel until the beaker is about 1/2 filled. After a few days the clamp is slightly and slowly opened and organisms which have concentrated at the end of the tubing should fall into the vial.

Techniques for Detailed Study of Compost Microorganisms

This protocol was written by Elaina Olynciw, biology teacher at A. Philip Randolph High School, New York City, while working in the laboratory of Dr. Eric Nelson at Cornell University as part of the Teacher Institute of Environmental Sciences.


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