LAB: 4Culture of microorganisms from environment
- Microorganisms are found throughout the environment. For better study of these microorganisms pure culture is needed.Pure culture: a single kind of microorganism growing alone in a protected environment.There are two methods used widely for preparation of pure culture from mixed population.1. streak plate method.2. pour plate method.Streak plate method ( Quadrant plate)
This method is used for the isolation of pure culture of bacteria from mixed population e.g. sputum, urine, stool or from pus of infected wound or abscess.
• Technique of Streak plate method :
• Sterilize the loop in a benzene flame, cool it and streak the specimen over an area-A.• Re-sterilize the loop, cool it, then streak over an area-B from the distal part of area-A.
• Continue the streak in the same manner for areas C and D.
• Incubate the plates at 37°C for 24 hours.
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• A subculture is done after 24 hours incubation from one of the colonies needed for study on a sterile medium following the same technique mentioned previously. The culture produced is a pure one.
Pour plate method
-This method involved serial dilution of specimen and mixed with melted media. Then pour the content into sterile plate plates.Cultural characteristics:
Cultural characteristics involve both :1. Growth requirement.
2. Colonial morphology.
It is used to study the macroscopic characteristics of a pure culture on a solid medium.
Colonial morphology
The following points have to be considered in describing a colony:1. Size: measured by mm.
2. Shape: rhizoid ,circular, filamentous, irregular.
3. Elevation: flat, convex, raised, umbonate.
4. margin: entire lobulated ,filamentous
5. Consistency: dry, mucoid.
6. Surface texture: rough, smooth.
7. Color or pigmentation.
8. Optical density : opaque, translucent, glistening.
9. changes in the inoculated medium e.g heamolysis.
• 10. Odour : bad or sweat musty odour.
Stock culture :-Means preservation of a microorgansim in a culture medium for future study. It can be kept usually for 1 month at 4 °C.
• Technique of stock culture:
• Use a universal bottle containing medium in slanted position ( to provide a wide surface for inoculation and good nutrition).• Using a sterile loop inoculate the surface of the medium with one or few colonies of a pure culture.
• Incubate the inoculated medium at 37 °C for 24 hours, then store at 4 °C.