Unknown Report Bio 211
The study of microbiology requires not only an academic understanding of the microscopic world but also a practical understanding of lab techniques and procedures in order to to identify and control microorganisms(MOs). The proper identification of a microorganism is not only important in a microbiology lab but also in the medical, industrial, and pharmaceutical fields. In this lab report, lab techniques and procedures I learned during this course were performed to assess my practical knowledge in microbiology. The goal of this lab report is to explain the tests performed on the isolated unknown that led to the identification of the unknown and to give a background on the characteristics and pathogenicity.
An unknown labeled as “Unknown #28” was given out by the lab instructor. The methods that have been learned thus far for identifying bacteria have been applied to this unknown. Procedures were followed by Terrye Light as stated in the course laboratory manual unless otherwise noted. The first procedure that needed to be done was to streak the unknown out on a Trypticase Soy Agar plate(TSA), using the Streak Plate Method as described in the lab manual. A single loopful of the culture(Unknown #28) was spread four times in the first quadrant. The loop was flamed and allowed to cool for 10 seconds. Five streaks were made from quadrant one into quadrant two. Following flaming and cooling of the loop, six streaks were made from quadrant two into quadrant three, using up all of the uninoculated space on the plate. Finally, the loop was flamed and placed aside. This needed to be done in order to test the purity of the unknown.
After the plates were incubated and grown, a Gram stain was performed. This technique separates bacteria into two groups, Gram positive and Gram negative, based on differences in the structure of the cell wall. Gram negative cell walls contain a thin peptidoglycan layer (without techoic acids) that is surrounded by a thick plasma membrane. Gram positive bacteria will stain purple because of their thick peptidoglycan cell wall. First, the slide was labeled with the name of the Unknown. A Sterilized loop was used aseptically adding one loopful of water to the center of a clean slide. A sterilized needle was dipped into obtained specimen and culture was mixed with the water and spread over the slide. Heat-fixed smear was prepared by passing the smear three times through the flame of the Bunsen Burner and let air dry for 5 minutes. Once dry, the slide was flooded with primary stain crystal violet for one minute (Both Gram positive and Gram negative cells were colored deep purple at this point) and then rinsed. Iodine was added as a mordant for one minute. (The Iodine created an insoluble complex within the thick peptidoglycan walls of the Gram-positive cells) The slide was then rinsed. Cells were decolorized with ethyl alcohol for 8 seconds and then rinsed. (Dissolved lipids found in...