culturing microorganisms

• Culturing bacteria

  In order to study bacteria, we often need to grow them in a laboratory setting.
• The cultured bacteria can be used to investigate the action of disinfectants and antibiotics.
• Factors affecting the speed of bacterial growth:

  • Temperature
  • most grow fast in warm environments.
  • Nutrient availability
  • Good supply of nutrients to grow rapidly.
  • Moisture
  • Most Grow fast in moist conditions.
  • Oxygen
  • different types of bacteria either need the presence or absence of oxygen for growth.
• Bacteria need a mixture of nutrients in order to reproduce, including:

  • carbohydrates (for energy)
  • nitrogen compounds (for protein synthesis)
  • vitamins
  • minerals
• Nutrient broth and agar are two nutrient-rich substances used to grow

  bacteria.

  • Often referred to as 'mediums' or 'media'.

  
• Nutrient broth


  A liquid medium (like water)

  
• Agar:


  A gel medium (like firm jelly)

  
• Agar plate:


  A plastic dish containing a thin layer of agar.

  
• An individual bacterium is invisible to the naked eye, but will multiply rapidly when placed in a growth medium.

  • The resulting group of bacterial cells is called a bacterial colony.
  • Each colony is visible to the naked eye, containing millions of bacteria.

  
• Bacteria grow on agar plates as colonies.

  
• Aseptic techniques:

  A set of procedures used to make sure a culture medium is not contaminated (the presence of unwanted microorganisms), when attempting to culture microorganisms.
• Examples of aseptic techniques:

  • Cleaning surfaces with disinfectant
  • Washing hands with antiseptic
  • Sterilising all instruments, solutions, & mediums.
  • Creating a sterile field using a Bunsen burner
  • Growing bacteria in incubators set to a maximum of 25°C.
• Examples of aseptic techniques:

  Sterilising all instruments, solutions, and mediums before they are used.

  • Sterilisation involves heating objects to a temperature at which all contaminating microorganisms are destroyed.
  • So only desired bacteria are transferred to the loop.
• Examples of aseptic techniques:

  Creating a sterile field using a Bunsen burner.

  • Try to work within the sterile field, and minimise the time that cultures and growth media are open to the environment.
• Sterile field:


  A sterilised area created by the updraft of the flame. 
• Examples of aseptic techniques:

  Growing bacteria in incubators set to a maximum of 25°C.

  • To prevent the growth of harmful pathogens, which prefer body temperature (37°C).
• Inoculation:


  The process of transferring bacteria from a broth to an agar plate.

  
• Agar plates are commonly used to investigate the effect of antiseptics

  or antibiotics on bacterial growth.

  By taking filter paper discs, soaking them in different antiseptic or antibiotic solutions.

  • Then placing them on an agar plate, we can find out how good the different solutions are at killing bacteria.
• Bacteria that are not affected by a substance will grow in the agar plate.

  • bacterial growth will be visible close to the paper discs.
• inhibition zone:


  A clear area surrounding the paper discs, if the Bacteria were affected by the substance & died.

  • A larger zone of inhibition means the substance is more effective at killing the bacteria.

  
• The inhibition zone is calculated using:

  area = π r2
• The effectiveness of each antibiotic at killing the bacteria:

  • Antibiotic A is not effective - there's no zone of inhibition.
  • Antibiotic B is somewhat effective - there's a small zone of inhibition.
  • Antibiotic C is very effective - there's a large zone of inhibition.

  
• Steps to take to culture microorganisms found in different places in the classroom:

  • Swab different areas
  • Sterilise the area & the loop
  • Only remove the lid of the petri dish slightly
  • Spread bacteria onto the agar jelly
  • Place tape on two sides
  • Incubate for at least three days at 25 Celsius.
• Reasons why this test does not prove that toothpaste kills all bacteria on teeth:

  • The test only uses one species of bacteria
  • There're still bacteria on the agar plate.
  • The test is not done on teeth
  • The conditions in the mouth are different
  • Toothpaste is only used on teeth for a short time