Pathogenic Cocci Lab Overview

Pathogenic Cocci Monday (Day 1):  Start Pathogenic Cocci Lab

Selective and Differential Media and Tests Introduction, Inoculation and Gram Staining

Our bacteria:

    • M. luteus (Ml)
    • S. epidermidis (Se)
    • S. aureus (Sa)
    • E. faecalis (Ef)
    • E. coli (Ec):  gram negative control for gram stain

Work flow

  1. Gram Stain: Make two slides, each with three circles.
    • Slide #1: Se, Ml, and Ec
    • Slide #2:  Sa, Ef and Ec
    • Ec should be on both slides as a control for proper gram staining
    • Use 2 loopfuls of bacteria, set aside to DRY completely
  2. Inoculation:  It's a lot of bacteria so we are going to divide and conquer.
    • Groups 1, 3, and 5: Inoculate with Ec, Se, and Ml
    • Groups 2, 4, and 6:  Inoculate with Ec, Sa and Ef
      • For TSA and PEA:  divide the plate into three sections and inoculate with each of their three assigned organisms with a simple back and forth streak
      • For MSA and Blood Agar:  divide the plate into two halves and inoculate only with the cocci, (Se and Ml OR Sa and Ef) in half-moon shape
      • For the BEA slants, inoculate only with the cocci (Se and Ml OR Sa and Ef) by zig-zagging up the media
    • Be sure that you label the plates and tubes very clearly!! They will be shuffled around and grouped together during the next lab, so it is critical that the labels can read by everyone!
      • your group #, date, Dr. Y AM or PM

  3. Smears should be dry, so HEATFIX, then Gram Stain
  4. Clean up all liquids
  5. Observe your gram stains under the microscope
    • 3 additional slide stations are set up with: N. gonorrhea, S. pyogenes, and Spn
  6. Clean up microscopes and put away

** There is a lot to do during this lab.  If it feels overwhelming, you can complete the gram stains, store the slides in your cubbies, and look at them during the next lab class

 

overview of pathogenic cocci lab

 

Pathogenic Cocci Wednesday (Day 2):  Results and Dichotomous Keys

Collect data/results from Pathogenic Cocci lab

  • Each lab bench is a station: 
    • TSA station with all but one set of TSA plates
    • PEA station with all PEA plates
    • MSA station with all MSA plates
    • Blood Agar Station with all blood agar plates
    • BEA station with all BEA slants
    • One Catalase station that contains
      • one set of TSA plates
      • a couple of bottles of H2O2
      • a box of glass slides
      • a lighter and bunsen burner
    • Microscope stations where you can view your gram stains if you ran out of time on Monday

Work flow

  • Rotate around the stations to collect the data for each different type of media in your lab report.  You should look at all of the samples (3-4 of each), but you only need to record one, representative sample.  
  • When you are finished, fill in a pathogenic cocci data summary table Download cocci data summary table(using just "+" for positive tests and "-" for negatives) which is hugely helpful when making the dichotomous key and for the unknown experiment (coming up in a couple of weeks)
  • You will need all of this data for the unknown experiment, so it is important to take good notes and keep track of the lab reports and summary tables!  

Clean up: 

  • Glass BEA culture tubes and their lids should be disposed of in their respective bins near the incubators.  
  • All petri dishes should be disposed of in the autoclave garbage 
  • Slides should be disposed of in the slide waste bin near the incubator