Last and final...

Introduction:

The winogradsky column, created by Sergei Nikolaievich Winogradsky, is a simple yet effective device used to culture a large diversity of microorganisms. The column can be created in anything from a soda bottle with a cut off top to a laboratory grade graduated cylinder. You need mud, water, a cellulose or calcium carbonate source, and a calcium carbonate source. The winogradsky column houses creates both an anaerobic and aerobic environment.
For my experiment I made 2 Winogradsky Column's both in 1 L containers but different shapes. On of the columns is in a 1 L graduated cylinder 40 cm tall and 7 cm in width. The second column is in a 1 L container that is 15 cm tall and 11 cm in width.
I used to different containers because I am testing to see if the shape of the container will effect bacterial growth. Will the shorter container with more surface (water) area develop more aerobic bacteria first and be slower to develop anaerobic bacteria? I predict that the taller cylinder will show greater and clearer bacterial growth (in the from of colored layers) before the shorter container.

Materials:

San Francisco Bay mud

San Francisco Bay brackish water = 31 ppt

parafilm

1 L graduated cylinder

1 L (shorter and wider) container

1.5 cups shredded newspaper

2 Tablespoons powdered CaCO3 (crushed antacid tablets - tropical fruit)

2 Tablespoons of CaSO4


Methods

I collected mud from the San Francisco Bay (in Sausalito, CA) during a low tide. I tested the salinity of the water with a refractometer and it was 31 ppt. I scoped both soft mud and water into a bucket. Once home I drained off some of the water (setting it aside). I scooped 2 L of mud into a mixing bowl and followed the nasa protocol (*http://quest.arc.nasa.gov/projects/astrobiology/fieldwork/lessons/demo.html) for mixing the mud and water until it is a "milkshake" like consistency. Once I reached my desired consistency I mixed in 1.5 cups of loosely packed shredded newspaper. Next I mixed in 2 tablespoons of powered Calcium Sulfate (CaCO3) - I crushed antacid tablets with a mortar and pestle to achieve a powered consistency. Lastly I mixed in 2 tablespoons of (CaSO4). I mixed for another couple of minutes and then scooped my mixture into both containers filling them up to 900 ml. I then filled the remaining 100 ml with the brackish water that I had set aside and sealed them both with parafilm. I placed both containers in the same window where they could receive indirect sunlight.

Results:

There was very little predicted growth in either of my columns. The only colors that my columns exhibited were white, which means that sulfur oxidizing bacteria was more prevalent in my columns. See Figure 1 and 2. There was signs of diatoms and cyanobacteria in the upper water portion. See Figure 3 and 4.

Conclusion:

Since my columns were created using brackish water and sediment they would have contained a considerable amount of sulfur because sulfate is found in seawater. Anaerobic microbes use the sulfates a terminal receptor and produce hydrogen sulfide. Since I sealed my columns with parafilm it eventually created an anoxic environment (1). White was the dominate color that I saw throughout both of my columns, though the shorter column showed significantly less growth, which supported my hypothesis. The white color was most likely sulfur oxidizing bacteria. This makes sense because my columns were extra sulfur rich.
In the beginning my column did not lack life. I saw two very common marine organisms, however, they only lasted a few days. I had a large zooplankton, about 5-8mm long. I planned to save and attempt to ID it but it died before I could find the time to catch it. I also had many polychaete worms. See Figure 5.
Due to living restrictions (i.e. a studio apt, with 2 large Maine Coon cats) I had to wedge my columns into a window that unfortunately got very little light. I think that this might have inhibited my column from developing as it should have. Once the cats found the column on March 23rd I had to take them outside. The tall column suffered some bite wounds though the parafilm but this was easily remedied using saran wrap. Once moved outside I was expecting to see more growth and color develop but my column actually did the opposite. The white color that had predominated my columns was disappearing. This could have been the growth of Desulfovibrio, which would have made them darker.
I think that if my columns had more time in a warmer environment they would develop naturally, creating the different colored zones. I think that they were stunted by lack of light and then cold nights (down in the 40’s and 50’s) here in the Bay Area. In the future I would pick a temperature stable environment that had an adequate light source and no disruptions.


1. Willey, J.M., Sherwood, L.M., Woolverton, C.J. 2008. Prescott, Harley, and Klein’s
Microbiology. 7th Ed. McGraw Hill. New York, NY.