Chap 6

• The primary objective of cultivating microorganisms, specifically bacteria, is to maintain viable populations of these organisms under controlled laboratory environments
• Cultivating microorganisms is a challenging process due to highly specific nutritional and environmental requirements and the diversity of these requirements among different species
• All-purpose medium


  Tryptic soy broth (TSB)
• Specialised media


  Used in the identification of bacteria and are supplemented with dyes, pH indicators, or antibiotics

  
• Enriched media


  Contains growth factors, vitamins, and other essential nutrients to promote the growth of fastidious (fussy) organisms, organisms that cannot make certain nutrients and require them to be added to the medium
• Chemically defined medium


  Complete chemical composition of a medium is known
• Complex media


  Contain extracts and digests of yeasts, meat, or plants, the precise chemical composition of the medium is not known
• Complex media


  • Nutrient broth, TSB, brain heart infusion (BHI)
• BHI broth


  Made by combining an infusion from boiled bovine or porcine heart and brain with a variety of other nutrients, often used in food safety, water safety, and antibiotic sensitivity tests
• Selective medium


  Inhibit the growth of unwanted micro-organisms and support the growth of the organism of interest by supplying nutrients and reducing competition

  
• Selective medium


  • MacConkey agar that contains bile salts and crystal violet, which interfere with the growth of many gram-positive bacteria and favour the growth of gram-negative bacteria, particularly the Enterobacteriaceae

  
• Enterobacteriaceae species (enterics) reside in the intestine that adapted to bile salt
• Lobe is a part of an organ that appears to be separate in some way from the rest
• Lactose-fermenting organisms, such as E. coli and Klebsiela spp, grow as pink to red colonies with or without a zone of precipitated bile

  
• Lactose-non-fermenting organisms, such as Salmonella, Shigella and Proteus spp., form colourless or clear colonies

  
• Enrichment medium


  General purpose enrichment agar which can nourish and support the growth of gram-positive as well as gram-negative bacteria

  
• Enrichment medium


  • Blood agar (5% defibrinated mammalian blood - horse, sheep, human) is added to the autoclaved basal media (TSA or Columbia Agar)

  
• Why enriched?


  Supports the growth of fastidious bacteria and inhibits the growth of some bacteria like Neiseria and Haemophilus

  
• Chocolate agar


  Nonselective, enriched growth medium used for isolation of pathogenic bacteria, a variant of the blood agar plate, containing red blood cells that have been lysed by slowly heating to 80 °C

  
• 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

  
• Differential medium


  Slightly inhibits the growth of Gram-positive bacteria and provides a color indicator distinguishing between organisms that ferment lactose (e.g., E. coli) and those that do not (e.g., Salmonella, Shigella)

  
• Differential medium


  • Eosin Methylene Blue (EMB) agar
• Certain lactose-fermenting bacteria produce flat, dark colonies with a green metallic sheen (most E. coli strains)

  
• Other lactose fermenters produce larger, mucoid colonies, often purple only in their centre
• Non-lactose fermenters demonstrated colourless colonies
• Mueller Hinton Agar (MHA)


  Commonly used for the routine susceptibility testing of non-fastidious microorganism by the Kirby-Bauer disk diffusion technique