10.12: Activity 5 - Winogradsky Columns

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Section: _____________________________

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Adapted from HHMI BioInteractive activity “Winogradsky Columns: Microbial Ecology in the Classroom”

Introduction

Microorganisms play an essential role in cycling elements that make the planet habitable for all other types of organisms. The diversity of these simple life forms is evident in the myriad of ecological niches they inhabit, from hydrothermal vents to the acidic lining of your stomach.

In this activity, you will observe a sampling of the diverse microorganisms that live in your local environment. Winogradsky columns will be set up-simple devices for constructing a stratified ecosystem-that illustrate different types of microbial metabolism in a colorful way.

Samples will be set up at the beginning of the semester and you will be required to make observations four times throughout the the spring. Answers to the questions below are due in Lab 10: Soil Ecology so ensure that you have completed these by that lab period.

Materials

• 4 clear containers
• 1-4 disposable containers (plastic baggies) for mixing soil
• 1 trowel for digging sediment sample
• 4 small labels
• Water
• 1 bucket container large enough to hold 6-10 cups of soil
• Large measuring cup for measuring soil and mixture
• A carbon source (organic forest litter); appx 1 cup loosely packed
• 6-10 cups soil
• Sulfur source; raw egg yolk is best
• Large mixing spoon
• 1 funnel

Procedure

Four Winogradsky columns will be set up: a control column and columns that contain added carbon, added sulfur, and carbon and sulfur. Make sure to set aside a few minutes on each of the assigned observation weeks during the experimental period to make observations and take photos of your columns.

Collect the Soil Sample

1. Identify a soil source in your area-anywhere with soil and water is appropriate, such as a stream, creek, lake beach (sand) or even back yard.
2. Take 2-3 photographs of your sample site to illustrate the location where your sediment was collected.
3. Collect approximately 10 cups of sediment in the bucket. The sample should be wet. Avoid or pick out rocks, sticks and leaves; include some additional water from the sample site if possible.

Assemble the Winogradsky Columns

1. Label 4 containers: “Control,” “Carbon,” “Sulfur,” and “Carbon and Sulfur.”
2. Label 4 disposable containers corresponding to the plastic containers and add appx 1.5 cups soil to each. Do not cross-contaminate samples.
3. For “Control” column, skip to step 5
4. For the other three containers,
   1. For “Carbon” column: Add ½ cup shredded newspaper (loosely packed) to soil and mix with spoon or trowel
      • The newspaper contains cellulose, a source of carbon
   2. For the “Sulfur” column: Add the yolk of a raw egg to the soil and mix. If using hardboiled eggs, crumble the yolks.
      • Egg yolk is a source of calcium sulfate in the column
   3. For the “Carbon and Sulfur” column: Add both nutrients as described in steps a and b.
5. Mix each of the samples thoroughly. Make sure to remove any large debris such as leaves, rocks or sticks. Slowly mix in water until the mixture has a milkshake consistency.
6. Using a large spoon, slowly add appx 1 cup of the mixture to the appropriately labeled plastic column. As you add the sample, tap the column on the counter to release any trapped air in the column.
7. Add water on top of the soil until there is a 2 cm layer of water on the surface, and air at the top of the column.
8. Place the lid on each column and turn a half turn. Do not tighten the lid!
9. Place all 4 columns in a well-lit place (window).
10. Using a camera or phone, take a photograph of the columns to document week 0 of experiment.

Make and report observations

A. Weekly observations

You will observe your Winogradsky columns throughout the semester during labs 3, 5, 8 and 10. Spend a few minutes on assigned dates recording your visual observations and take a photograph (remember to label photos with the week the photo was taken).

Consider the following questions when recording observations:

• Has the color of the soil changed?
• Has the color of the water changed?
• What differences between the columns do you observe?
• Does the soil appear to have shifted or settled?
• Do you see any layers forming in the sediment? In the water?
• How does the thickness of the layers change from week to week?
• Are there any differences between the side facing the light and the side away from the light? Be sure to keep the same orientation relative to the light if you must move the columns.
• Sketch the columns, in color, three times over the 8-week experiment.

Date	Observations during the week of:	Control	Carbon	Sulfur	Carbon & Sulfur
	0
	Lab 3
	Lab 5
	Lab 8
	Lab 10

Questions

1. How do your columns differ? How are the similar?
2. Did you observe changes in the control column? If so, explain why they occurred
3. Winogradsky columns form oxygen concentration gradients. Predict the distribution of oxygen throughout the column. Consider the soil, water and air
4. Winogradsky columns form sulfide concentration gradients as well. In the columns that contain egg yolk, predict how sulfide will be distributed throughout the column.
5. Purple sulfur bacteria and green sulfur bacteria are two types of bacteria that use sulfide to support photosynthesis. In general, green sulfur bacteria tolerate higher levels of sulfide than purple sulfur bacteria do. Predict where the green and purple sulfur bacteria would be in relation to each other.
6. If samples were extracted from various layers of all the columns, where would you find photosynthetic organisms such as cyanobacteria and algae? Why?
7. Explain how Winogradsky columns illustrate the diversity of microorganisms found on Earth today in terms of the diversity of niches they occupy.