Selective and Differential Media

Selective and Differential Media

Objective

• Identify bacterial species based on their physical and metabolic differences.

Key Concepts

• Selective Media

  • Contains chemicals that inhibit the growth of unwanted bacterial species while allowing the desired species to grow.
  • Examples of selective chemicals: antibiotics, dyes, salts.

• Differential Media

  • Used to identify bacterial species based on differences in metabolism, often linked to the presence or absence of specific enzymes.
  • The growth allows for visual differentiation between species, typically in color.

Mannitol Salt Agar (MSA)

• Selective Agent: 7.5% NaCl

  • Inhibits non-salt tolerant microbes, allowing for the growth of salt-tolerant Gram-positive halophilic species such as Staphylococcus species, which thrive on human skin.

• Differential Agent: Mannitol

  • A carbohydrate energy source that, when fermented, produces acid end-products lowering the pH.
  • pH Indicator: Phenol red
  • Yellow: pH < 6.8 (acidic)
  • Red: pH 6.8-7.4
  • Pink: pH > 7.4 (alkaline)

• Observations:

  • Most Staphylococci do not ferment mannitol (red), but Staphylococcus aureus ferments it (yellow).

MacConkey Agar (MAC)

• Selective Agents: Bile salts and crystal violet

  • Inhibit growth of Gram-positive bacteria and allow growth of Gram-negative enteric bacteria (Enterobacteriaceae).

• Differential Agent: Lactose

  • Fermentation produces acid end-products that lower pH, indicating fermentation.
  • pH Indicator: Neutral red
  • Pink: pH < 6.8 (acidic)
  • Clear/Yellow: pH > 6.8 (alkaline)

• Observations:

  • Lactose fermenters (coliforms) produce acid and appear red.

Bacterial Classification

• Domain: Bacteria, Archaea, Eukarya
• Kingdom: Fungi, Plant, Animal, Protist
• Phylum:
• Class:
• Order:
• Family: Enterobacteriaceae
• Genus: Escherichia
• Species: Escherichia coli

Enterobacteriaceae Family

• Characteristics:
  • Bacillus-shaped, Gram-negative bacteria known as enterics due to their intestinal habitat (Greek "enteron" = intestine).
  • Facultative anaerobes that typically ferment glucose.
• Normal Microbiome Members:
  • Includes Enterobacter, Klebsiella, Proteus, and most Escherichia coli strains.
• Pathogenic Members:
  • Diarrheal diseases: Shigella, Salmonella enterica, and pathogenic strains of E. coli (e.g., E. coli O157:H7).
  • Life-threatening conditions caused by:
  • Salmonella enterica serotype Typhi (typhoid fever).
  • Yersinia pestis (bubonic and pneumonic plague).

Coliforms

• Enteric bacteria that ferment lactose, used as water fecal pollution indicators (e.g., E. coli, Enterobacter aerogenes).
• Presence indicates possible fecal-borne pathogens in water.

Eosin Methylene Blue Agar (EMB)

• Selective Agents: Eosin and methylene blue dyes

  • Inhibit most Gram-positive microbes.

• Differential Agent: Lactose

  • Fermentation lowers pH, causing color changes due to dye interaction:
  • Dark purple/black: lactose fermentation.
  • Metallic green: E. coli lactose fermentation.
  • Light purple/pink: non-lactose fermenters.

Bile Esculin Agar (BEA)

• Selective Agent: Bile (oxgall)

  • Selects for Group D streptococci and enterococci.

• Differential Action: Ability to hydrolyze esculin via esculinase

  • Reaction: Esculin → Glucose + Esculetin.
  • Esculetin forms a dark brown/black precipitate with ferric citrate, darkening the medium.
  • Identifies esculinase-positive species, such as Enterococcus faecalis, Enterococcus casseliflavus, and Group D Streptococcus species.

Diagram of Bile Esculin Hydrolysis Reaction

• Highlights the site of hydrolysis and the cleavage that results in the formation of a dark precipitate.

Conclusion

• Selective and differential media are important in identifying and isolating bacterial species, enhancing our understanding of microbiological profiles and their implications for health and safety.