Culturing Techniques in Laboratory Settings

Importance of Culturing Microbes

• Objective: To grow bacteria in a lab setting for study.
• Challenge: Growing in an artificial environment outside the natural habitat.
• Requirement: Understanding nutrient needs of microbes, which varies between species (some are fastidious).

Culture Medium

• Definition: The "food" for microbes, containing necessary nutrients for growth.
• Forms: Can be solid or liquid.
• Sterility: Must be sterile to prevent contamination.
• Sterilization Methods:
  • Filtration
  • Autoclaving
  • Pasteurization

Inoculation

• Process: Introducing microbes into a growth medium.
• Inoculum: The microbes introduced to the culture medium.

Types of Medium

• Defined Medium:
  • Known exact composition.
  • Example: E. coli growth medium with glucose and salts.
• Complex Medium:
  • Components known but not exact quantities.
  • Consists of extracts (e.g., beef, yeast) and peptones.

Differential and Selective Media

• Selective Media:
  • Inhibits growth of some microbes, supports others.
• Differential Media:
  • Distinguishes microbes based on ability to consume specific nutrients (often visualized by color change).
  • Example: Growth medium can be both differential and selective.

Reducing and Capnophile Media

• Reducing Media:
  • Used for anaerobes (cannot tolerate oxygen).
  • Contains chemicals to neutralize oxygen.
• Capnophile Media:
  • Requires higher CO2 levels.
  • Candle jars can be used to increase CO2 for capnophiles.

Plating Techniques

• Pour Plate Method:
  • Mix bacteria with liquid agar and pour into plate.
  • Swirl to disperse bacteria, aiming for individual colonies.
• Spread Plate Method:
  • Spread bacteria on solidified agar using a spreader.
  • Goal: Separate colonies starting from single cells.

Streak Plate Method

• Objective: Isolate single bacterial colonies to reduce genetic diversity.

• Procedure:

  1. Sterilize loop, inoculate with bacteria.
  2. Streak on petri dish in zigzag motion.
  3. Re-sterilize loop, streak new area touching previous streak.
  4. Repeat for 4-5 streaks to dilute bacteria.
  5. Observe individual colonies in later streaks.

• Outcome: Allows for isolation of pure cultures from mixed populations.