Final Winogradsky Column Post 12/7/2008

My Winogradsy experiment began nine weeks ago with the construction of two columns that have flourished into a diverse collection of microbial life. One column consisted of water and sediment from a local pond and the other consisted of water and sediment from a local river, the Paulinskill River, in order to compare the microbial activity.

Water and sediment were collected from the pond and the river on October 5, 2008. Once extraneous debris was removed, both sediment samples were amended with the same amount of grass clippings (about 1 cup) as a carbon source and magnesium sulfate (about 1 tablespoon) as a sulfur source. Water was added to each mixture, and each sample was transferred to a separate column until it was approximately 3/4 full. Water was added to the top of the column, and they were covered with plastic wrap secured by a rubber band. Both columns were placed in the same south facing window for 9 weeks.

Pond Mud Column River Mud Column
Over this time, the two columns experienced considerable changes in both the color of the water and sediment and odor. These changes indicate that observable microbial activity has been taking place. This final report will serve to document progress of both columns over the semester.

The Pond Mud Column


10/18/2008
The pond mud column which began as a visibly homogenous mixture of pond sediment, grass clippings, and magnesium sulfate with a layer of pond water on top quickly changed. Black spots appeared in the sediment which eventually created a thick layer that was completely black. At the water line, an orange layer appeared within a thin brown layer. The water became discolored, and there was a slight rotten egg odor when the plastic was removed. The column showed no significant difference between the side facing toward the sunlight and the side facing away from the sunlight.


11/19/2008
The following changes took place over the course of 4 weeks. The bottom layer of the column remained black but was not as thick. A thin layer of white separated the black layer from it from the layer above. This was a relatively thick light brown layer with swirls of black throughout. Much of this layer had a green hue to it as well which could be evidence of photosynthesis. A thin layer of white could be found just below the water line. The water was clear, however, there was a brown/green film over the water that had gas bubbles trapped inside.

12/7/08
The pictures below depict what the column looks like today. The first picture is the side facing the sunlight, the second is the side facing away from the sun, and the third is a picture from the top.

There have been some changes to the column since the last post over two weeks ago. The black sediment on the bottom of the column is due to the presence of sulfur reducing bacteria. The amount of black sediment on the bottom has not increased or decreased significantly as the anoxic conditions at the bottom of the column have not changed. The white layer seems to have become a bit more pronounced in the last few weeks. It was discussed in a posting that the white layer is a result of the metabolism of sulfur producing bacteria. Since this layer is just above the anaerobic layer of sulfur reducing bacteria, this would make sense.

A significant change in the column since the last posting is the presence of two distinct layers of green that can be seen in the most recent picture. The bottom green layer is most likely the result of green sulfur bacteria while the top green layer may be a result of photosynthetic bacteria like cyanobacteria. The brownish layer between these two green layers has swirls of black throughout which I am unsure about.

The water is clear but may have a green tint to it (it is hard to tell). Any green coloring at the top of the column would be the result of photosynthesis, most likely it is algae in the water, on the film, and on the sides of the column container. As the picture shows, there is a film on top with gas bubble contained in it. Gases from microbial metabolism may have risen to the surface and become trapped by the film.

The River Mud Column


10/19/08

The river mud column was much slower to change than the pond mud column and the changes were not as dramatic. When it was first constructed, the column had one layer of sand with visible grass clippings intermixed. For the first several weeks, the only changes to the column were some black coloring around the grass clippings and a thin layer of black mud with orange splotches at the water line. The water also showed signs of a greenish/brown color and there was some orange debris floating in it. There was no significant odor either.

11/19/08
Over the next four weeks this column showed more change than it had previously. The bottom layer became black throughout. Above the black layer, sand could still be distinguished, however, there was a definite green hue to it. At the water line, the sediment was a deep red color. The water was discolored as well. It was not a green hue like in the pond mud column but rather a brownish-red with the same color debris floating in it. There was brownish-red sediment on the plastic covering as well. There was an odor emitting from the column around the midway point between these posts but it had ceased at this point.

12/7/08

The pictures below depict what the column looks like today. The first picture is the side facing the sunlight, the second is the side facing away from the sun, and the third is a picture from the top.

The bottom layer of the river mud column is black and most likely the result of sulfur reducing bacteria. However, it is not as consistent as in the pond mud column. The sand particles from the river sediment are visible in this layer upon close inspection. Since the pore spaces between sand particles is greater than for silt or clay, I suspect some air was trapped in this column. This may be the reason why the black layer took longer to develop. Something that I find interesting is that the black layer extends all the way up to the water on the side facing the sun.

Green growth can be seen in the sand above the black layer. This is most likely algae or a photosynthetic bacteria. The layer at the sediment/water interface remains a deep rust color, and, while the water has definite green algae growth, the sides of the container and the plastic covering have a brownish-red sediment, perhaps the result of iron oxide. Gases rose to the surface in this container as well, although the gases were not trapped by a film as in the pond mud column. The plastic covering of the container would rise under pressure and often the column would leak. This was evidence of gas being released as a result of microbial metabolism.




Summary

This experiment was fantastic for illustrating microbial activity in an extraordinary way that is sure to gain the enthusiasm of science students. There were significant differences between the microbial activity of the two columns. The pond mud column as expected showed greater changes and, therefore, more diverse and drastic microbial activity. I would like to keep these columns for another semester and perhaps set up two more with different amendments and compare the changes.

Update 11/19/2008

There has been a lot of activity in both of my columns since my last post in October. These changes became evident in late October/early November and have been less pronounced since then. The column containing the pond mud shows some definite layering as shown in the first picture below.

This picture shows the side facing away from the light of the pond mud column. The water appears a reddish-brown from the outside of the column, but, once the plastic is removed from the top, the water itself is clear, the sides of the column in the water are lined in green, and a brown/green film with bubbles floats atop the water.

At the water-sediment boundary there is a thin section that is a whitish color. Below the whitish line, the sediment is light brown with swirls of black throughout. Although it is not apparent by just looking at the column, the picture shows a good amount of green throughout this layer also. At the bottom boundary of the light brown layer, there is a distinct thin layer of white. Finally, the sediment is black at the bottom of the column.

There is no significant difference between the side facing toward the light and the side away from the light other than the location of the layers. As can be seen from the picture, the bottom, black layer is about two-thirds down the column on the side facing away from the light. On the side facing toward the light, the final layer is not even halfway down the column.

It is important to note that, when the column first presented some noticable changes in late October/early November, there was a strong odor emitted from the column when the plastic top was removed. This odor is almost not detectable at this time.

The column constructed of the river mud changed as well (and more than I thought it would). The first picture below is one of the side that faced toward the light. As was the case with the pond mud column, the bottom layer which is also black is much higher for the side facing the light than the side facing away from the light.

The water in this column was distinctly brownish-red both from the outside and when the plastic top was removed as seen in the picture below. There was some brownish-red sediment on the plastic as well.

The layer at the water-sand line was a deep red color. The sand below the water looked much like it did when it was first placed into the column except that the layer had a definite green tint to it (the picture shows this). Finally, as mentioned previously, the bottom layer of the column was black like the bottom layer of the pond mud column. This column also emitted an odor when it was changing a few weeks ago that has lessened as of this update.

Both of my columns leaked several times during the course of this experiment (and ruined two windowsills). During the day, the plastic bellowed upward presumably from gases emitted from the column into the air space and water leaked out creating white, crusty streaks on the outside of the columns. Most recently, this has been occurring with the second column (river mud) much more exclusively.

Update 10/15/08

Pond Mud Column

It has been 10 days since I set up my Winogradsky columns. Over this time, the pond mud column has undergone some significant changes. At first, there were thin streaks of black scatter throughout the brown mud in the column. I think that these streaks initiated where the carbon source (grass clippings) were mixed into the mud. As time progressed, the black color overtook the brown color of the mud in the column which can be seen in the photo above. On top of the black layer of mud below the water line, there is a light brown layer that has a slight orange layer within it. The water which was once clear is now discolored. There is also some debris suspended along the sides of the container in the water layer. At this point, there is not a significant difference between the side facing the sun and the side facing away from the sun.


River Mud Column

The changes to the river mud column are much less significant. The water in the column which was clear is now discolored with a greenish hue. Where the mud meets the water, there is a very thin layer of mud that has turned black with orange splotches. There is also orange colored debris suspended in the water. There has not been a significant change in the mud in the rest of the column.

Winogradsky Column Setup 10/05/2008

For my Winogradsky column experiment, I wanted to compare the microbial activity in the mud of the banks of a pond and a river (seen in the pictures above, respectively). With the help of my 4 year old lab assistant, I collected mud from the north bank of a small pond near my home that is bordered by farmland on one side and a recreational trail on the other. We then collected mud from the north bank of the Paulinskill River. The river is located less than a quarter of a mile from the pond and is bordered by the same recreational trail as the pond. We also collected water from both the pond and the river.

The protocol that I used to set up my Winogradsky column was: http://www.personal.psu.edu/faculty/j/e/jel5/biofilms/winogradsky.html.

I also watched a video demonstration before constructing my column found at: http://quest.nasa.gov/projects/astrobiology/fieldwork/lessons/demo.html.

The pond mud column was constructed first. Small stones and sticks were removed from the pond mud. Then, a generous amount (about 1 cup) of grass clippings were mixed into the mud as a carbon source. Magnesium sulfate (about 1 tablespoon) was mixed into the mud as a sulfur source. Water was added to the mud mixture to give it a slightly smoother consistency (similar to heavy cream). The mud mixture was then added to a 1-liter bottle with the top cut off until the container was about 3/4 full. As the mud mixture was added, I tapped the bottle on the counter to reduce air bubbles as much as possible. Water was added to the top of the column and the bottle was covered with plastic wrap and secured with a rubber band.

Here is the finished pond mud column:

The same procedure was used to construct the river mud column. However, before constructing the column, the river mud had to be filtered through a mesh strainer to remove small stones, shells, and coarse sand particles. The river mud (shown below) was still quite sandy so I am not sure what kind of result this column will provide, if any. The finished column is pictured below.

I set both columns on my window sill which faces south. It gets light from morning until late afternoon. The containers were also marked to indicate which side of the column faces the sunlight.