9.1

• MacConkey-Positive, Oxidase-Negative

  • Enterobacteriaceae
  • Acinetobacter
  • Streptophomonas

  
• True intestinal pathogens

  • Salmonella
  • Shigella
  • Yersinia
  • Escherichia

  
• Opportunistic pathogens

  • Escherichia
  • Citrobacter
  • Proteus
  • Edwardsiella
  • Morganella
  • Hafnia
  • Providencia
  • Enterobacter
  • Serratia
  • Klebsiella

  
• MacConkey-Positive, Oxidase-Positive

  • Pseudomonas
  • Burkholderia
  • Achromobacter
  • Chryseobacterium
  • Sphingobacterium
  • Alcaligenes
  • Bordetella (Non-Pertussis)
  • Comamonas
  • Vibrio
  • Aeromonas
  • Plesiomonas
  • Chromobacterium

  
• MacConkey-Negative, Oxidase-Positive


  • Sphingomonas paucimobilis
  • Moraxella
  • Eikenella
  • Pasteurella
  • Actinobacillus
  • Kingella
  • Cardiobacterium
  • Capnocytophaga

  
• MAC-Negative, Oxidase-Variable

  • Haemophilus
• Optimally recovered in special media

  • Bartonella
  • Hafnia
  • Campylobacter
  • Acrobacter
  • Helicobacter
  • Legionella
  • Brucella
  • Bordetella Pertussis And B. Parapertussis
  • Francisella
  • Streptobacillus Moniliformis
  • Spirillum Minus
• Selective and differential media selectively isolate Gram-negative and enteric bacteria and differentiate them based on lactose fermentation

  
• Lactose fermenters turn red or pink on MacConkey agar, and nonfermenters do not change color

  
• Family ENTEROBACTERIACEAE

  • Gram negative bacilli/coccobacilli
  • Facultative anaerobe
  • Non-spore forming
  • Motile with peritrichous flagella except Tatumella (polar flagella), Shigella & Klebsiella (nonmotile)
  • Predominantly facultative flora of the human bowel/gastrointestinal tract
  • Some are agents of zoonoses

  
• Endotoxins are responsible for much of the morbidity and mortality resulting from infections associated with these bacteria
• Symptoms caused by endotoxins


  • Fever
  • Chills
  • Hypotension
  • Granulocytosis
  • Thrombocytopenia
  • Disseminated intravascular coagulation
• Colonies on BAP are luxurious, moist, smooth, gray, shiny, entire, convex and opaque
• Categories of Enterobacteriaceae based on clinical infections

  • Opportunistic pathogens
  • Primary pathogens

  
• Primary pathogens include S. enterica, Shigella spp., and Yersinia spp.

  
• Members of this family may be responsible for a large number of opportunistic infections when introduced into inappropriate body sites
• MAC-Positive characteristics

  • Primary selective and differential agar
  • Contains crystal violet dye
  • pH indicator is neutral red
  • Bacterial fermentation of lactose results in acid production

  
• Oxidase-Negative means cytochrome oxidase participates in electron transport and in the nitrate metabolic pathways of certain bacteria
• 1% tetramethyl-p-phenylenediamine dihydrochloride is the reagent used for oxidase testing
• CULTURE MEDIA

  • Eosin Methylene Blue (EMB)
  • McConkey agar
  • Salmonella Shigella agar (SSA)
  • Hektoen Enteric agar (HEA)

  
• Indicators in Eosin Methylene Blue (EMB)

  • Eosin Y
  • Methylene blue
• Organism characteristic colonies on Eosin Methylene Blue (EMB)


  • Escherichia coli (pink/purple colonies with greenish metallic sheen)
  • Klebsiella (pink mucoid colonies)
  • Enterobacter (“fish eye colonies”)

  
• McConkey agar characteristics

  • Gram(+) bacteria inhibitor: Crystal violet
  • Bile salts
  • CHO: lactose
  • pH indicator: Neutral red

  
• Color of colony on McConkey agar

  • Lactose fermenter: pink
  • Non-fermenter: colorless

  
• Salmonella Shigella agar (SSA) characteristics

  • Selective & differential for Salmonella & Shigella
  • CHO: lactose
  • Gram (+) bacteriostatic agent: Bile salts
  • pH indicator: neutral red

  
• Organisms on Salmonella Shigella agar (SSA)

  • Salmonella (NLF, H2S (+))
  • Shigella (NLF, H2S (-))

  
• Hektoen Enteric agar (HEA) characteristics

  • Gram (+) bacteriostatic agent: bile salts
  • CHO: lactose, sucrose & salicin
  • pH indicator: bromthymol blue

  
• H2S (+) organisms on Hektoen Enteric agar (HEA)


  • Colonies with black center

  
• Biochemical tests

  • Triple Sugar Iron (TSI)
  • Kligler’s Iron Agar (KIA)
  • Lysine Iron Agar (LIA)
  • IMViC
• Triple Sugar Iron (TSI) is the initial step in Enterobacteriaceae ID
• Media components of Triple Sugar Iron (TSI)

  • CHON - peptone
  • CHO source (lactose, sucrose, glucose)
  • pH indicator: phenol red
  • H2S indicator: ferrous sulphate
  • Source of sulphur: Na thiosulfate
• Quality Control for TSI

  • Escherichia coli – A/A with gas
  • Salmonella typhi – K/A H2S+
  • Pseudomonas aeruginosa – K/NC

  
• Formation of H2S requires an acidic environment
• Lysine Iron Agar (LIA) components

  • Small amt of CHON
  • CHO source (0.1% glucose)
  • Lysine (1% lysine)
  • pH indicator: Bromcresol purple
• Quality Control for LIA

  • Salmonella typhimurium – Alkaline slant and butt, H2S positive
  • Shigella flexneri – Alkaline slant, acid butt
  • Proteus vulgaris – Red slant, acid butt
• IMViC tests

  • Indole
  • Methyl red
  • Voges-Proskauer
  • Citrate
• Indole test determines the ability of the microbe to degrade tryptophan

  
• Tryptophan in medium is converted to indole by enzyme
• H2S production in LIA

  • Not sensitive to H2S production of Proteus
• Lysine decarboxylation


  1. Produces cadaverine

  2. Purple (gihapon)

  3. Glucose fermented produces acid (yellow)