Winogradsky Columns

Sergei Winogradsky was among the first microbiologists to investigate the organisms found in complex biofilm communities.

Working in the late decades of the last century he was a pioneer in selective enrichment techniques. One of the strategies that Winogradsky employed to isolate organisms from nature was a miniature model pond cross section which has since been called a Winogradsky column.

Photo courtesy of Michael Cox, Anaerobe Systems, San Jose, CA.

The column is constructed of soil or mud from virtually any source and water from the same or a different source. To these natural components, are added supplemental carbon and sulfur.

Above the soil is a layer of water and the column is usually covered to retard evaporation. The entire column is then illuminated to encourage the growth of phototrophs. The resulting growth of microoraganisms can be quite spectacular and colorful.

The most obvious inhabitants of the column are aerobic and anaerobic phototrophs, but a vast array of heterotrophs is also found and the routine presence of methanogens makes this a useful exercise in expanding the microbial diversity of the introductory microbiology laboratory.

The column can be constructed in virtually any roughly cylindrical container and I have seen them made in vessels as diverse as olive jars, two liter pop bottles and twenty liter carboys.

There are also flat plate columns which give large surface areas for biofilms to develop and are excellent for photographic record keeping (see the Anarobe Systems web site).

Instructions for constructing Winogradsky columns can be found in many laboratory manuals, in an informative Carolina Tips (Sept. 1, 1978) and in the Microcosmos Curriculum Guide (Building a Microbial City by Fred Stein).

Source of materials:

Mud or soil can be collected from nearly any source, and the time of year appears not to be significant.

Pond mud collected in the middle of winter has produced beautiful columns. Stones and sticks should be removed by screening or by hand. Water may be collected from the same source but distilled water or tap water will also serve.

Students gathering materials for a column at Prince Gallitzen State Park, Cambria Co. PA. This and the following pictures are of students building a column in a 20 L carboy.

Carbon source:

Vegetable materials such as shredded hay, grass clippings, shredded newspaper, sawdust, corn starch, oatmeal, will work fine, use your imagination. These materials release carbon relatively slowly. Fast release carbon sources include sodium carbonate, sodium bicarbonate or calcium carbonate.
Prince Gallitzan State Park
Cambria Co, PA

Sulfur source:

Once again use your imagination, elemental sulfur, calcium sulfate, magnesium sulfate, raw or hard boiled eggs, or cheese have all been used with success.

Procedure:

Mix the mud/soil with the desired carbon and sulfur sources. Add some water to the container and add the soil mix a little at a time, using a glass rod or stick to pack the material in the bottom of the column. Air bubbles should be excluded as much as possible although any oxygen remaining trapped in the mud is probably consumed by facultative organisms very rapidly.

Add water and soil mixture until the column is about three fourths full. Use a wash bottle to rinse excess mud from the container above the soil level and then add additional water to produce a water column (the pond).

Leave some air space above the surface of the water and cover the column with plastic wrap or parafilm.

A NOTE ON SAFETY: There is no easy way to determine if the mud students collect is contaminated with sewage, farm waste, heavy metals or other toxic materials. I have students ware latex or vinal gloves when mixing materials for and packing a column.

Column structure:

The column contains at least two steep gradients.

The water column at the surface is in contact with the atmosphere and is therefore aerobic but it becomes increasingly anaerobic with depth. The surface layer of the column may produce an aerobic liquid air biofilm (pellicle) that can be sampled by dipping a coverslip into the column and lifting a portion of the film from the water.

In the highly anaerobic base of the column, decomposition and the activity of sulfate reducing bacteria results in the production of hydrogen sulfide gas. This hydrogen sulfide gradient decreases toward the top of the column. These two gradients acting in opposite directions create a great range of habitats selective for a variety of microorganisms.


The column is illuminated by sunlight or by tungsten bulbs. In a perfectly uniform column one might expect a variety of photoautotrophs to be selected in specific zones within the column, but such uniformity is rarely obtained.

Typically the lower portions of the column are colonizied by photoautotrophic green and purple sulfur bacteria. The aerobic surface of the column is occupied by oxygenic cyanobacteria. Just below the surface phototrophic purple non-sulfur bacteria predominate.

A great variety of heterotrophs also can be found in these columns including obligate anaerobes such as clostridia and methanogenic bacteria.

The majority of the bacteria are located in a thin film between the soil/mud substrate and the container wall. When using plastic containers such as two liter pop bottles, these bacteria can be sampled by inserting a needle through the container wall. Cylindrical plastic columns such as those provided by Carolina Biological can be frozen and the entire contents can be extruded under pressure.

The electro-chemical gradient that develops within the column can be measured with a standard VOM if one insulated wire with the insulation removed from the last 1-2 cm is inserted to the bottom of the column and a similar, shorter wire is inserted into the mud water interface at the top of the column. The voltage generated varies with time, but one Winogradsky column in a 20 L carboy as been generating between 0.3-0.5 volts continuously for three years.