TYPES OF CULTURE MEDIA-I

TYPES OF CULTURE MEDIA-I


Alkaline Peptone Water

  • Alkaline Peptone Water is a broth medium for the enrichment of Vibrio species from food, water and clinical samples.
  • Shread, et al. formulated peptone water for use as a nonselective enrichment broth for the cultivation of Enterobacteriaceae from foodstuffs and other specimens.
  • Alkaline Peptone Water is a modification of peptone water with the pH adjusted to 8.4.
  • The alkaline pH of this medium allows the growth of Vibrio organisms while inhibiting the growth of commensal intestinal bacteria.
  • Enrichment of specimens in Alkaline Peptone Water prior to inoculation of plated media has been shown to increase the recovery rate of Vibrio spp.

USES: 

  • Alkaline Peptone Water is commonly used for the enrichment of Vibrio cholera and Vibrio species from food, water and clinical samples.
  • The broth can also be used for direct microscopic examination of samples using the hanging drop method.
  • It is used as both enrichment and transport media.
  • Initially, it was used as a non-selective enrichment broth for the cultivation of Aeromonas.


Chocolate Agar

  • Chocolate agar is essentially the same as blood agar except that during preparation the red blood cells are lysed when added to molten agar base. 
  • As a result, the cell lysis releases intracellular nutrients such as hemoglobin, hemin (“X” factor), and the coenzyme nicotinamide adenine dinucleotide (NAD or “V” factor) into the agar which is utilized by fastidious bacteria. 
  • Red blood cell lysis gives the medium a chocolate-brown coloration when prepared from which the agar gets its name. 
  • The most common bacterial pathogens that require this enriched medium for growth include Neisseria gonorrhoeae and Haemophilus species.

LIMITATIONS

  • It is recommended that additional biochemical and/or serological tests should be performed on isolated colonies from the pure culture in order to complete identification.
  • Chocolate Agar is an enriched medium, thus non-pathogenic organisms may overgrow pathogenic bacteria. If isolation of  N. gonorrhoeae is desired, a selective medium such as Thayer Martin Agar, Modified should be used in parallel with this non-selective formula.
  • Precipitated hemoglobin may appear as dark spots on or in the media and does not affect the performance of the media.
  • The presence or absence of  N. gonorrhoeae in a specimen does not rule out the possible presence of other pathogenic organisms.

Buffered Charcoal Yeast Extract (BCYE)

  • Feeley et al in 1978 developed a medium to isolate Legionella species which was later modified by substituting yeast extract for casein hydrolysate and beef extract, and replacing starch with activated charcoal and named it as Charcoal Yeast Extract (CYE) Agar.
  • A further modification was made by Pasculle et al in 1980 by the addition of ACES (N-2-acetamido-2-aminoethane sulfonic acid) buffer in order to maintain the proper pH for optimal growth designated as BCYE for Buffered Charcoal Yeast Extract. In 1981, 
  • Edelstein et al further modified the medium by addition of the potassium salt of alpha-ketoglutaric acid which increased the sensitivity of the medium. 
  • It is recommended for use in the cultivation and primary isolation of Legionella spp. in water and other samples suspected of harboring the bacteria.
  1. It is recommended that biochemical, immunological, molecular, or mass spectrometry testing be performed on colonies from pure culture for complete identification.

  2. It is also recommended that more than one type of medium is used for isolating Legionella species and that non-selective and selective BCYE Agar plates be inoculated in parallel.

  3. Due to nutritional variation, some strains may be encountered that grow poorly or fail to grow on this medium.

  4. The Centers for Disease Control and Prevention (CDC) recommend incubation of environmental samples with 2.5% CO2; however,  L. gormanii is the only known species with enhanced growth under this condition.

  5. Colonies of Legionella that develop on white membrane filters may have a different appearance to those that develop against a black or dark background filter.

  6. When handling Legionella species, it is important to avoid aerosol formation.

Exam Important

  • Alkaline Peptone Water is commonly used for the enrichment of Vibrio cholera and Vibrio species from food, water and clinical samples.
  • The most common bacterial pathogens that require this enriched medium of choclate agar for growth include Neisseria gonorrhoeae and Haemophilus species.
  • BCYE is recommended for use in the cultivation and primary isolation of Legionella spp. in water and other samples suspected of harboring the bacteria.
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