Culture Media

Streptococcus Selective Agar Composition, Preparation, and Principle

Roantree et al introduced a medium for the isolation of streptococci from group A beta-hemolytic. The medium was enriched with yeast nucleic...

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This article writter by MN Editors on December 09, 2021

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Streptococcus Selective Agar Composition, Preparation, and Principle
Streptococcus Selective Agar Composition, Preparation, and Principle

Roantree et al introduced a medium for the isolation of streptococci from group A beta-hemolytic. The medium was enriched with yeast nucleic acids and maltose boosted the size of colonies and improved the clarity, sharpness and precision of the hemolytic zones created by these organisms. Streptococcus Selective Agar (SAA) is a specific medium suggested for the initial isolation from groups that comprise the A Streptococcus specie (S. Pyogenes) from oral culture as well as respiratory specimens. It’s designed to inhibit staphylococci and gram negative bacilli and thereby facilitate the subculture, isolation, and the identification of pathogenic streptococci including beta-hemolytic streptococci, and S. pneumoniae.

Composition of Streptococcus Selective Agar

IngredientsGrams/L
Casein Peptone15.0 g
Peptic Digest of Soybean Meal5.0 g
Sodium Chloride5.0 g
Nucleic Acid3.0 g
Maltose0.5 g
Neomycin10.0 mg
Polymyxin B10.0 mg
Sheep Blood50.0 ml (5%)
Agar15.0 g

Final pH 7.3 +/- 0.2 at 25ºC.

Principle of Streptococcus Selective Agar

The nutrient source of organic nitrogen, especially amino acids and peptides with long chains that are required by the growth of bacterial cells are provided by the combination of casein and soybean peptones. The combination makes the medium extremely nutritious. Salts of sodium chloride help maintain the Osmotic equilibrium of the medium. Maltose and ribonucleic acid enhance the process of producing hemolysis. The addition of sheep’s blood (5 percent) can provide additional growth factors to microorganisms that are fastidious and assists in the identification of hemolytic reactions. Selective agents like Neomycin as well as Polymyxin B are added to reduce the oral microflora that is common to us which includes coliforms, staphylococci, Micrococcus, Haemophilus and Neisseria species.

Purpose

Selective Streptococcus Agar was designed to aid in the isolating of streptococci from group A of respiratory origin.

Physical Properties of Streptococcus Selective Agar Media

  • Appearance: Cream to yellow homogeneous free flowing powder
  • Gelling: Firm, comparable with 1.5% Agar gel
  • Colour and Clarity of prepared medium:  Light to medium amber-coloured clear, to slightly translucent gel forms within Petri plates
  • Reaction: Reaction of 4.56% w/v aqueous solution at 25°C. pH : 7.4±0.2
  • pH: 7.20-7.60

Preparation of Streptococcus Selective Agar

  1. Add all the ingredients, except lamb blood, maltose solution and antibiotics to deionized/distilled water. Increase volume until 930.0 mg.
  2. Mix thoroughly.
  3. Then gently heat it and heat until boiling.
  4. Autoclave for 15 mins at 15 psi pressure 121 degC.
  5. Cool to 45deg-50degC.
  6. Aseptically, add sterile sheep blood as well as sterile maltose solution and an antibiotic inhibitor sterile.
  7. Mix thoroughly.
  8. Pour into sterile Petri dishes or distribute into sterile tubes.

Result and Interpretation of Streptococcus Selective Agar

  • After 18-48 hours of incubation in an environment rich in carbon dioxide, the plates exhibit isolated colonies in scattered areas, and confluent growth within areas of high inoculation.
  • The streptococci group A ( S. pyogenes) appears as opaque or transparent with a gray to white color. tiny (1 up to 2 mm) colonies that are surrounded by a ring that is beta hemolysis.
  • Pinpoint or tiny colonies of nonhemolytic alpha- and other streptococci that are beta-hemolytic could develop in tiny numbers.
  • Neisseria species, viridans strains of streptococci staphylococci, rods with gram-negative phenotypes, and the majority of beta- streptococci that are not in the groups B and A aren’t inhibited by the medium.

Uses of Streptococcus Selective Agar

  • Streptococcus Selective Agar was developed to be used for the primary separation of streptococci group A of respiratory origin.
  • It is also utilized to determine the identity of all Streptococcus species including groups of streptococcal streptococcal A (S. Pyogenes) and B (S. Agalactiae) C D, F G along with S. pneumoniae particularly by respiratory specimens.

Limitations of Streptococcus Selective Agar

  • Certain strains of Streptococci group A ( S. pyogenes) can be found to not thrive on this medium. The characteristics of the samples as well as the physiological state of the organisms could affect the recovery of the targeted species, and also altering the inhibitory properties of the medium. Thus, it is recommended to examine nonselective controls and comparing them with the selective medium in order to gain additional information and to ensure the best recovery of any pathogens that could be a threat is highly recommended.
  • Further tests for biochemical and serological procedures are recommended to ensure final confirmation.
  • It is important to recognize that the organisms most are susceptible to the antimicrobial agents in a specific medium could be completely or partially blocked based on the amount of the drug, specifics that the particular strain, and the quantity of organisms within the inoculum. Organisms that are typically resistant to the antimicrobial agent must not be hindered. Cultures of samples produced on selective media should be compared to specimens that were grown on nonselective media in order to gain additional information , and to aid in that pathogens are not killed.
  • Hemolytic reactions of certain strains belonging to streptococci group D have been found to be affected due to variations in the blood of animals. The strains involved are beta-hemolytic on human, horse and rabbit blood agars, and alpha-hemolytic in sheep blood agar.
  • The conditions of incubation are well-known to affect hemolytic reactions in beta-hemolytic streptococci. For the best performance, it is recommended to place the blood agar base medium incubation under CO2 levels that are higher (5 to 10 percent) according to established lab procedures.

Quality Control

  • Examine plates according to the instructions under “Product Deterioration.”
  • Take a look at the plates in a visual way to make sure that any physical imperfections will not affect the use.
  • Calculate the pH potentiometrically the room temperature to ensure that it is in line with the standard of 7.3 + 0.2.
  • Take note of the firmness of the plates throughout the inoculation process.
  • Incubate representative plates that have not been inoculated at 35 + 2°C for 72 hours and then check for contamination by microbial species.

Storage Instructions

When you receive the plates, keep them in the dark for 2-8 degrees Celsius. Avoid overheating and freezing. Don’t open the appliance until it is you are ready to use. Reduce exposure to sunlight. The plates that are kept in the original sleeve wrapper at temperatures between 2 and 8 degC for a few hours the time of use could be inoculated until the expiration date, and incubated according to the recommended time of incubation. Let the medium cool to room temperature prior to inoculation.

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Microbiology Notes is an educational niche blog related to microbiology (bacteriology, virology, parasitology, mycology, immunology, molecular biology, biochemistry, etc.) and different branches of biology.

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