I was issued an unknown bacterial sample (number 14) and inoculated a TSA (Trypticase Soy Agar) plate with it. The media on this "streak plate" is not selective in that it does not differentiate between different types of bacteria (most bacteria will grow on it). The streak method is a way of spreading the bacteria so that when it is allowed to incubate, individual colonies can be observed and collected for the differential tests.
This was done on Friday the 25th. The plate was left to incubate at room temperature over the weekend. On Monday morning I observed the plate and something was definitely amiss... There was bacterial growth on the plate but portions of it were missing, there were very odd small "channels" etched throughout the surface of the media, and odd specks of a white material approximately 1 to 2 mm in length.
On the right half of the plate in the above picture, the vertically oriented hazy lines are bacterial growth. The lower left section shows very little bacteria remaining and the upper left hardly any bacteria at all. Notice too, the upper left section contains many more of the small white "flecks" than other regions. I (without unsealing the plate) viewed this under a dissecting microscope at 35x magnification in hopes of determining what the contaminate on the plate could be. I immediately saw that the flecks were small wormlike organisms that were indeed moving and happily munching my bacteria! The channels I had observed were the trails left by this organism. I conferred with Matt, Josh and Robin Cotter, and we all agreed that this could possibly be Planaria, a flat worm, but it was hard to distinguish under the relatively low power of the dissecting scope.
We need more power! I observed the organism with a stronger microscope at 100x and 400x magnification.
Shown here are two examples of the organism. It is a little hard to see but the organism is segmented (definitely easier to see this under locomotion). This ruled out Planaria, and the general consensus was that it was most likely Drosophila - fruit fly larvae. At this point it was decided that this plate would remain sealed and that I would prepare another sequence of streak plates: one using the same unknown sample, one with the same unknown from a different source, and one TSA plate from the stack that I obtained the original plate from with nothing on it. This was an attempt to narrow down the possible sources of contamination. These plates were placed in the same location as the original to incubate at room temperature.
On Tuesday, the next day, I dropped in to check on the growth on the plates...
and look at that, a lone fruit fly was in the plate (that had remained sealed the entire time). I should also note that the channels I described earlier are much more easily seen in this photo. I named him George. The bacteria inoculated plates were beginning to show signs of growth, and the "blank" TSA plate showed none.
Wednesday I was able to begin differential testing. I first observed the bacterial colony morphology.
The individual colonies were:
Pale Yellow
Lustrous - they had a sheen of reflected light across the surface
Circular
Translucent - allowed some light to pass through but were not clear
Smooth edges
Could possibly be seen as a convex profile or umbonate profile (like a side view of a fried egg)
Next I conducted the first differential test, the Gram stain. This revealed that the bacterium was Gram +. Indicating that it has a cell wall composed of thick layers of peptidoglycan. Also during this test I could see that the individual bacterial cells were cocci - they were shaped like small round balls.
The above image is the Gram stain results as seen at 400x magnification showing the cocci shape. The purple color is the result of Crystal Violet staining the peptidoglycan in the cell walls. A Gram - bacteria would have stained pink due to the counterstain Safranin.
The next differential test to be performed was the catalase test. By adding Hydrogen Peroxide to a sample the presence of the enzyme catalase can be seen. If catalase is there, bubbles will be vigorously produced. This is the result of catalase breaking the toxic Hydrogen Peroxide down into Oxygen and water. This tells us that the bacteria can tolerate the presence of Oxygen. The unknown is catalase positive.
I did start more differential tests, but I will wait until I have results to post before describing them, and upon interpreting the results I will have a positive ID on the unknown.
On a side note, Josh, Matt and I had a very fruitful discussion about further research studies, and I am happy to say that I am very excited about it! Good things to come! I will share the topics after I conduct some preliminary research - I need to more firmly grasp the scope of what I will be doing before I try to relate it here.
