Culture Media & Culture Methods

Prepared by: Ricci Alyssa S. Tumampil, SMT

History

• Bacteria need to be grown (cultured) for identification.
• They must be grown separately (isolated) on culture media as pure samples for study.

Early Culture Media

• Urine/Meat Broth: Original media used by Louis Pasteur.
• Liquid Medium: Results in diffuse growth.
• Solid Medium: Produces discrete colonies.
• Colony: Visible collection of bacteria originating from a single cell.
• Cooked Cut Potato: Introduced by Robert Koch as the earliest solid medium.
• Gelatin: Not satisfactory because it liquefies at 24°C.

Agar

• Introduced by Frau Hesse.
• Used for solid medium preparation.
• Obtained from seaweeds with no nutritive value.
• Melts at 98°C and sets at 42°C.
• 2% agar is used in solid media.

Types of Culture Media

1. Based on Consistency

   • Solid Medium: 2% agar; used for colony morphology and pigmentation studies. Examples: Nutrient agar, Blood agar.
   • Liquid Medium: No agar; used for inoculum preparation and isolation. Example: Nutrient broth.
   • Semi-Solid Medium: 0.5% agar; used as motility medium.

2. Based on Constituents/Ingredients

   • Simple/Basal Media: Nutrient broth (peptone, meat extract, NaCl) and Nutrient agar.
   • Complex Media: Contains added ingredients for special nutrients.
   • Synthetic/Defined Media: Pure chemical substances with known composition. Example: Peptone Water.

3. Based on Special Media

   • Enriched Media: Contains added nutrients like blood or serum. Examples: Blood agar, Chocolate agar.
   • Enrichment Media: Used to isolate pathogens with inhibitory substances. Examples: Selenite F Broth, Alkaline Peptone Water.
   • Selective Media: Inhibitory substances to suppress unwanted organisms. Examples: Mac Conkey’s Medium, TCBS Agar.
   • Indicator Media: Contains indicators for bacterial growth results. Example: Blood agar for hemolysis.
   • Differential Media: Distinguishes between bacteria. Example: MacConkey’s medium.

4. Based on Oxygen Requirement

   • Aerobic Media
   • Anaerobic Media

Special Media Details

• Sugar Media: Contains fermentable substances like glucose.
• Transport Media: Used for transporting samples. Examples: Stuart’s medium, Buffered glycerol saline.
• Anaerobic Media: For anaerobic organisms. Examples: Robertson’s cooked meat medium, Thioglycollate medium.

Culture Methods

• Streak Culture: Isolation of bacteria in pure culture using a wire.
• Lawn Culture: Uniform surface growth, used for typing and sensitivity testing.
• Stroke Culture: Made in tubes for diagnostic tests.
• Stab Culture: Demonstrates gelatin liquefaction and oxygen requirements.
• Pour Plate Method: For bacterial count estimation.
• Liquid Culture: Used for blood culture and continuous methods.
• Anaerobic Culture Methods: Different approaches to culture anaerobic bacteria, such as McIntosh-Fildes’ Anaerobic Jar and Gaspak.

Biochemical Reaction Media

• Oxidase Test: Detects oxidase enzyme. Positive = purple color. Example: Pseudomonas.
• Triple Sugar Iron (TSI) Agar: Tests sugar fermentation and gas production.
• Indole Test: Detects indole production. Positive = pink ring.
• Citrate Utilization: Detects citrate usage. Positive = blue color.
• Urease Test: Detects urease production. Positive = pink color.

Rapid vs. Slow Urease Producers

• Rapid Urease (TPUNCH): Trichophyton, Proteus, etc.
• Slow Urease (CKEYS): Citrobacter, Klebsiella, etc.