microbiology lab midterm

• Potential hazards in the lab

  • Chemicals
  • Infectious agents
  • Fire
• Standard safety practices in the lab

  • Do not eat, drink, smoke, store food, or apply cosmetics during lab
  • Wear closed-toed shoes
  • Tie back long hair
  • Disinfect workspace
  • Wash hands before and after
• Personal protective equipment used in the lab

  • Lab coat
  • Gloves if necessary
  • Safety glasses
• Disposing of various materials in the lab

  Place broken glassware contaminated with microbial cultures or body fluids in the To Be Autoclaved container
• Magnification


  Increase in the apparent size of the image
• Resolution


  Ability to distinguish two different points, to see them as separate
• Contrast


  Ability to distinguish an object from the background
• Wet mount


  Easy way to visualize microorganisms
• Making a wet mount

  1. Suspend infusions by shaking them carefully

  2. Use a pipette to transfer a very small drop of one hay infusion to a slide

  3. Handle the coverslip carefully by its edges and place it on the microscope
• Hanging drop technique


  Used to observe motility
• Performing the hanging drop technique


  1. Obtain a clean hanging-drop slide

  2. Pick up a small amount of petroleum jelly on a toothpick

  3. Pick up a coverslip and carefully touch the petroleum jelly to get a small rim of jelly. Repeat on other three edges

  4. Place the coverslip on a paper towel, with the jelly side up

  5. Transfer a small drop of peppercorn infusion to the coverslip
• True motility


  Ability to move around
• Bacteria are found everywhere
• Sterile environment


  An area that microorganisms cannot be found in
• Factors that affect bacterial growth

  • Temperature
  • Oxygen
  • pH levels
  • Osmotic pressure
• Agar medium


  A nutrient, jelly like solid used to grow bacterial cultures
• Agar plate


  Growth medium solidified with agar
• Colony


  A population of cells that arises from a single bacterial cell
• Don't stab agar when using agar plates to grow bacteria
• Pure culture


  Studying one species without worrying for contamination
• Contamination


  Presence of unwanted microorganisms
• Using aseptic technique to prevent contamination


  Using flame before using tubes
• Culture media and their advantages/disadvantages

  • Broth culture: provides many bacteria in a small area that can be easily transported
  • Agar slant: gives a solid growth surface and easily transportation than Petri dishes
  • Agar deeps: used to grow bacteria that require less oxygen
• Inoculating tools and when to use them


  • Loop: when using liquid media
  • Needle: when using solid media
• Smear


  Thin film of bacteria on a slide
• Purpose of making a smear

  To fix bacteria on a slide and prevent the sample from getting lost during the staining process
• Fixing


  Killing bacteria
• A smear needs to be fixed so they don't wash off the gram staining
• Purpose of staining a smear

  To give them color for the microscopic observation
• Simple stain


  Only using one stain
• Negative stain


  A simple stain that stains the background but leaves bacteria the same
• Differential stains


  Two differently colored stains to clearly contrast cell types
• Purpose of differential stains


  To distinguish between bacteria
• Examples of differential stains

  • Gram stain
  • Acid-Fast stain
  • Endospore stain
• Purpose of gram-staining


  To distinguish between morphology & whether the cell wall is + or -
• Steps of gram-staining


  1. Crystal violet for 30 sec

  2. Rinse w/ sterilized water

  3. Gran iodine for 10 sec

  4. Rinse w/ sterilized water

  5. Decolorizer with ethanol

  6. Rinse w/ sterilized water

  7. Smear w/ safranin for 30 sec

  8. Rinse w/ sterilized water

  9. Blot dry
• Clinical application of gram-staining


  Diagnosis of bacterial infections for antibiotic use
• Failure to flame the tube could lead to contamination when transferring bacteria using aseptic technique
• Airborne bacteria could fall anywhere on this slant and grow to form a colony if the top of the tube is not flamed
• The smear can overheat during heat fix when gram-staining