Dirty Deeds: A look into our filthy microbe's cell wall structure

Our next series of experiments concern the cell wall structure of our unknown bacterium. Bacterial cell walls are typically classified as gram-positive or gram-negative, a characteristic that can be determined by a process referred to as gram-staining. A stain known as crystal violet is used to dye the bacterium's cell wall. Gram-positive cell walls are extremely thick in comparison to those of gram-negative cells and consist of a single layer of a compound known as peptidoglycan. Gram-negative cell walls are bit more complex. They are composed of an inner plasma membrane, a thin peptidoglycan layer and an outer membrane that is covered by a lipopolysaccharide layer. The thick layer of peptidoglycan in gram-positive cells is able to retain the crystal violet dye, causing the cells to appear as purple following the gram-staining process. Gram-negative cells are not able to retain the dye and appear pink following this process.

So why does the structure of something that you can't even see impact your life?? The differences in cell wall structure between gram-positive and gram-negative cells affect the ways in which they are able to infect your body. The lipopolysaccharide (LPS) that covers the outside of gram-negative cells is able to act as an endotoxin that causes inflammation, high fever and even death. The outer membrane layer also allows for resistance to common antibiotics like penicillin, making them more difficult to overcome than gram-positive infections. Basically, don't try to make friends with gram-negative bacteria, they're mean. 

Last week in lab, we did our own gram-staining of the soil microbes we harvested from outside of Subway. After performing gram-stains on the gram-positive control (B.megaterium), gram-negative control (K.pneumonia), and our unknown, we viewed the cells and they looked remarkably like hot dogs. We observed their rod-like shape (bacillus) and dark color. Stains of gram-positive cells are known for their deep purple hue. Therefore, we believed our bacterium was gram-positive because of its similarity in color to our gram-positive control. In order to confirm this prediction, we plated our bacterium and the gram-positive/negative controls on MacConkey agar, which is commonly used to isolate gram-negative bacteria and differentiate between lactose-fermenting/non-fermenting bacteria. Our bacteria would not grow on this media if it were gram-positve.

As can be seen in the photo below, the only growth observed was located in the gram-negative quadrant of the plate. This result is consistent with our prediction regarding the gram-positive nature of our bacterium.

In this week's lab, we will be performing acid-fast staining on our microbe!
Check back next week for more details!

Anne and Austin

http://en.wikipedia.org/wiki/Gram-negative_bacteria#Medical_treatment
http://www.life.umd.edu/classroom/bsci424/BSCI223WebSiteFiles/GramPosvsGramNeg.htm