Winogradsky column

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This picture depicts the initial appearance of three different Winogradsky Columns. They are soil and water samples from a river, the later two columns have been modified with phosphate, nitrate and sulfur additives. These additions promote the growth of various bacteria specific to the anaerobic and aerobic regions of the column.

Shown above as a result of a 7 week period where the columns have been allowed to grow algae, cyanobacteria and other bacterial colonies. Of specific interest are the red regions of the middle column, indicative of purple non-sulfur bacteria(i.e Rhodospirillaceae) Also in column three, the red growth along the side of the column: a purple sulfur bacterium,Chromatium.

The Winogradsky Column is a simple device for culturing a large diversity of microorganisms. Invented by Sergei Winogradsky, the device is a column of pond mud and water mixed with a carbon source such as newspaper (containing cellulose) or egg-shells (containing calcium carbonate) and a sulfur source such as gypsum (calcium sulfate) or egg-yolk. Incubating the column in sunlight for months results in an aerobic/anaerobic gradient as well as a sulfide gradient. These two gradients promote the growth of different micro-organisms such as clostridium, desulfovibrio, chlorobium, chromatium, rhodomicrobium, beggiatoa, as well as many other species of bacteria, cyanobacteria, and algae. The column provides numerous gradients, depending on additive nutrients, from which the variety of aforementioned organisms can grow. The aerobic water phase and anaerobic mud or soil phase are one such distinction. Due to low oxygen solubility in water the water quickly becomes anoxic towards the interphase of the mud and water. Anaerobic phototrophs are still present to a large extent in the mud phase, there is still capacity for biofilm creation and colony expansion, as noted by the images. Algae and other aerobic phototrophs are present along the surface and water of the upper half of the columns. Green growth is often attributed to these organisms. Construction The column is a rough mixture of ingredients and exact measurements are not critical. A tall glass or plastic tube (30cm long, >5cm wide) is filled by a third with pond mud, omitting any sticks, debris or air bubbles. Supplementation of ~0.25% w/w calcium carbonate and ~0.50% w/w calcium sulfate or sodium sulfate is required. Mixed in with some shredded newspaper or hay (for cellulose), ground egg-shell and egg yolk respectively are rich in these minerals. An additional anaerobic layer, this time of un-supplemented mud, brings the container to two thirds full. This is followed by water from the pond to saturate the mud and occupy half the remaining volume. The column is sealed tightly to prevent evaporation of water and incubated for several months in strong natural light. External links Animated tutorial by Science Education Resource Center Carleton College Winogradsky column: perpetual life in a tube Edinburgh University

Using A Winogradsky Column to Analyze Microbial Communities Frances Vandervoort

1991 Woodrow Wilson Biology Institute Objectives In this investigation, students will use easily obtained materials to study ecological succession in a microbiological community. The investigation is appropriate for a variety of age groups. Elementary students will be fascinated by the changes occurring over time in their bottles. First and second year high school students can do microscopic sampling of their communities that will introduce them to diversity among microorganisms. Upper level high school students can use sophisticated sampling techniques of microorganisms and chemicals within the columns for both quantitative and qualitative studies. Background Bacteria, ubiquitous in nature, are responsible for the evolution of the Earth's atmosphere. Photosynthesis occurred first in anaerobic bacteria, which adapted to the Earth's ancient environment by using sunlight, water, and readily available carbon to produce carbohydrates and free oxygen. At least four groups of photosynthetic bacteria abound on Earth today; the green sulfur, the purple sulfur, the purple nonsulfur bacteria, and the brownish nonsulfur Heliobacterium (Sagan and Margulis, 1988). It is possible to establish a culture of photosynthetic bacteria in a "Winogradsky column", a device based on a more elaborate one made by Russian microbiologist Sergei N. Winogradsky, who lived in Russia and France the early part of this century. Materials Clear plastic 1 liter bottle Mud and sand from outdoor source, especially from a marshy area where colorful mats and scums of microorganisms can be found. Saltwater or freshwater marshes can be used. Water from outdoor source, such as a stream, pond, lake, or puddle. Again, use either fresh water or salt water. Calcium sulfate of hard boiled egg yolk (as a sulfur source) Shredded paper (a piece of newsprint about 100 cm2) or cellulose fibers (as a carbon source) Procedure Mix the sand or mud with about a tablespoonful of egg yolk or calcium sulfate. Add a few shredded cellulose fibers or paper. Layer the sand and mud in the plastic bottle. It is easier to fill the bottle if you first remove the narrow neck of the bottle by carefully cutting through the plastic with a single-edged razor blade or paper cutter. Pour the water you have collected over the top of the sand-mud mixture so that there is a small amount of unabsorbed water on top. Cover the bottle with plastic film secured by a rubber band, place it in a sunny window, and a rich culture of photosynthetic bacteria

will develop within a few weeks. A similar column kept in the dark will also develop a bacterial population, but the bacteria will not be photosynthetic. Watch the columns over several months. Color changes indicate that ecological succession is taking place. Discussion Questions Describe the changes you see taking place over a period of several months. Record observations every one to two weeks. Using a long pipette, take samples from various depths within your column and place them on a microscope slide. Examine and draw those that you see. Use a key to microorganisms to identify all you can. How is a Winogradsky column similar to the bottom of a pond? How is it different? References Pigage, Helen K. "The Winogradsky Column: A Miniature Pond Bottom." American Biology Teacher (47/4), April, 1985. Pp. 239-240. Sagan, Dorion, and Margulis, Lynn. Garden of Microbial Delights. Boston: Harcourt, Brace, Jovanovich. 1988.

Results from the Winogradsky Column Study

SES: Microbial Methods In Ecology, 1999
In the Microbial Methods In Ecology Course, students constructed Winogradsky columns from sandy sediments and water that were collected from Little Sippewisset estuary located in Falmouth, MA, except for column 8, which was collected from a freshwater pond. These columns served as 1) a source of microorganisms for labs on general microbial methods, such as bacterial productivity, and 2) model systems to study sediment-water biogeochemistry and microbial biogeochemical diversity. Presented below are some of the student results involving these columns. Columns Columns were constructed from soda bottles (20 cm x 7 cm) and fitted with 5 luer lock sample ports and stopcocks (Cole-Parmer). All columns (except 7) were filled approximately 3/4 full with sediments, and topped with estuarine water, except Col. 8 in which freshwater sediments and water were used (see treatments). Columns were placed in the laboratory near a SW-facing window. Pictures were taken 6 weeks after initialization (9 Sep - 19 Oct 1999).

Column 1

Column 2

Column 3

Column 4

Column 5

Column 6

Column 7

Column 8 Profiles of hydrogen sulfide (H2S) and methane (CH4) versus depth. Depth is measured from the lip of the column down. Methane was measured on a gas Chromatogram and a spectrophotometric method was used to measure H2S concentration.

Column 1

Column 3

Column 4

Observations The following are some notable observations (click on images to obtain higher resolution picture):

Formation of sulfur bacteria (probably Beggiatoa) in the top of column 2 (column that was placed in the dark).

Patchy formation of purple and green sulfur bacteria. It is uncertain why this column developed such patches, and it is interesting that this column also exhibited low concentrations of H2S and CH4 as compared to the other columns.

Column 5, which received glucose, developed a hard mat or biopolymer at the air-water surface. The water in this column became anaerobic and stayed that way during all sampling.

The formation of iron sulfides (FeS) in Column 6 causes the sediments in this column to be black. Only this column was amended with Fe (see Treatments).

Purple and green sulfur bacteria are evident in several columns, but are most pronounced in Column 7, shown here. These anaerobic bacteria are phototrophic, but use H2S as an electron donor instead of H2O. Retrieved from "http://en.wikipedia.org/wiki/Winogradsky_column"