microbiology

• Gram negative bacteria


  Lipopolysaccharide layer provides protection from attack by lysozyme and penicillin antibiotics, making them more difficult to control
• Growing microbes


  1. Cultured in Petri dishes or larger containers

  2. Generally grown on agar, a jelly-like substance, to which required resources can be added
• Nutrients required for microbial growth

  • Carbon source (organic compounds like glucose)
  • Nitrogen source (organic or inorganic compounds)
  • Growth factors (water, vitamins, mineral salts)
• Optimum temperature


  Should be considered when growing microbes, generally not 37°C (body temperature) unless growing human pathogens
• Optimum pH


  Should be considered when making growth media, can be adjusted with acid or alkali
• Oxygen requirements


  • Obligate aerobe
  • Facultative anaerobe
  • Obligate anaerobe
• Aseptic technique


  1. Sterilise Petri dishes and nutrient agar before pouring

  2. Sterilise inoculating loop before and after use

  3. Minimise opening Petri dish lid

  4. Secure lid with adhesive tape after inoculation

  5. Incubate at 25°C for 24-48 hrs

  6. Sterilise plates and equipment after use
• Gram positive bacteria


  • Thick peptidoglycan cell wall that retains crystal violet stain, appear violet or purple
• Gram negative bacteria


  • Lipopolysaccharide layer washes away crystal violet stain, appear pink or red with safranin
• Gram staining method


  1. Smear slide with bacterial sample

  2. Stain with crystal violet

  3. Treat with Lugol's iodine

  4. Decolourise with alcohol

  5. Counter stain with safranin
• Bacterial shapes


  • Cocci (spherical)
  • Bacilli (rod-shaped)
  • Spirillum (spiral or corkscrew)
• Counting microbes


  1. Dilute original sample to give 20-200 colonies

  2. Count colonies and multiply by dilution factor to find number in original sample
• Total count


  Living and dead cells
• Viable count


  Living cells only
• Serial dilution


  Dilute sample in steps, e.g. x100, x10,000