Cultural Characteristics of Microorganisms on Basal media (Nutrient agar and Nutrient broth)

Isolation of microorganisms in pure culture is necessary to study the relationship between an organism and a disease. Also pure culture isolation is necessary to study about the biochemical properties as well as the susceptibility of the organisms which are essential in diagnosis and prognosis of various diseases.  For this, microorganisms are artificially cultivated on various media in the laboratory. A media usually provides an artificial environment for the growth and multiplication of microbes.

Basal medium or the general purpose media are the ones one which provides all essential nutrients to support the growth of most of the nonfastidious microorganisms.  It will not promote or suppress the growth of one particular organism over the other.  Egs : Nutrient agar, Nutrient broth,  Trypticase Soy agar and Brain Heart Infusion agar.  Nutrient agar and Nutrient broth are the most widely used basal media and the only difference between them is that the former contains agar as the solidifying agent.  The presence or absence of agar in the media will not affect the growth of the organisms as agar has no nutritive value.

Different other types of media are also used in microbiology laboratories based on the requirements. They include special media like enriched media, enrichment media, selective media, differential media, indicator media, sugar media and transport media.

  

Aim  

To demonstrate the cultural characteristics of microorganisms on basal or supportive media for identification and classification of microorganisms into different taxonomic groups.   

Principle

Different growth patterns and cultural characteristics are produced by microorganisms on cultivation on different media which will be useful in their identification. The cultural characteristics for all the microorganisms are well described in Bergey’s Manual of Systematic Bacteriology. Usually cultural characteristics are determined by cultivating the microorganisms on general purpose media like nutrient agar slants and plates, nutrient broth etc.

  

Nutrient agar plates

Microorganisms produce discrete and well defined colonies on nutrient agar plates. The following characteristics can be evaluated from the agar plate cultures:

1. Size: Pinpoint, small, moderate or large.

2. Pigmentation: Color of colony

3. Form: The shape of the colony

     a) Circular: unbroken peripheral edge

     b) Irregular: Indented peripheral edge

     c) Rhizoid: Root like spreading growth

4. Margin: The appearance of the outer edge of the colony is described as follows:

     a) Entire: Sharply defined, even

     b) Lobate: Marked indentations

     c) Undulate: Wavy indentations

     d) Serrate: Toothlike appearance

     e) Filamentous: Thread like spreading edge

5. Elevation: The degree to which colony growth is raised on the agar surface is described as follows:

     a) Flat: No marked elevation from the surface of the medium

     b) Raised: Slightly elevated

     c) Convex: Dome shaped elevation

     d) Umbonate: Raised with elevated convex central region

Nutrient broth cultures

In broth cultures microorganisms do not produce discrete colonies. Increase in turbidity of the broth is the indication of microbial growth. Microorganisms show characteristic growth patterns in nutrient broth which is also considered helpful in the identification of organisms.  They are as follows:

1. Uniform fine turbidity: Finely dispersed growth throughout.

2. Flocculent: Flaky aggregates dispersed throughout

3. Pellicle: Thick, padlike growth on the surface

4. Sediment: Concentration of growth at the bottom of the broth culture which can be granular, flaky or flocculent

Requirements

24 hr nutrient broth cultures of species Bacillus, Streptococcus, Staphylococcus, Pseudomonas and E.coli.

Nutrient agar plates, nutrient broth tubes, Bunsen burner, inoculating loop, glass marking pencil etc.

Procedure

1.  Using sterile techniques, inoculate each of the appropriately marked media as follows:

        a) Nutrient agar plates: Using a sterile loop, prepare a streak plate inoculation of each of the organisms provided on the agar plates for the isolation into discrete colonies.

        b) Nutrient broth tubes: With a sterile loop, inoculate each organism into the tube containing nutrient broth and shake the loop few times to dislodge the inoculum.

2.  Incubate all cultures at 37^oC for 24 hr to 48 hrs.

Results

On nutrient agar plates:

Bacillus sp. - produced irregularly round, raised, dull, opaque and grayish white colonies.

 Streptococcus sp. – produced circular, small, semitransparent and low convex disc like colonies.

Staphylococcus sp. – produced circular, convex, smooth, shiny, opaque and slightly yellow pigmented colonies.

 Pseudomonas sp. – produced large opaque and irregular colonies.

 E.coli.- produced large, thick, moist, white, smooth opaque or partly translucent colonies.

In nutrient broth tubes:

Bacillus sp. – Flocular deposit with little or no turbidity.

Streptococcus sp. – granular turbidity with a powdery deposit. No pellicle.

Staphylococcus sp. – Uniform turbidity.

Pseudomonas sp. – Dense turbidity with a surface pellicle.

E.coli.- General turbidity and a heavy deposit, dispense completely on shaking.