Biochemical Tests for Enterobacteriaceae
Zhawaan Szepietowski
Esherichia Coli
Esherichia 0157:H7
Strains of E. Coli
Enterotoxigenic (ETEC)
Enteropathogenic (EPEC)
Enteroinvasive (EIEC)
Enterohemorrhagic (0157:H7) (EHEC)
Enteroaggregative (EAEC)
Strains of E. Coli
Enterotoxingenic
adults diarrhea in tropics; travelers diarrhea; high infectious dose
ETEC
Enteropathogenic
usually daycare centers and hospital nurseries
EPEC
Enteroinvasive
dysentery with destruction of mucosa; similar to shigella, but infectious dose is far higher
EIEC
Enterohemorrhagic
O157:H7;
watery diarrhea to bloody diarrhea;
HUS hemolytic uremic syndrome
EHEC
Enteroaggregative
Part of enteroadherent, adhere to intestinal mucosa
EAEC
Triple Sugar Iron Agar
Slant: Acid (A)
Butt: Acid (A)
Gas: Pos
H2S: Neg
TSI agar
Rapid Spot Indole Test
Rapid Spot Indole Test
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
MacConkey Agar
E.Coli 0157:H7 shows colorless on MacConkey Agar, other strains will be pink
SMAC- Enterohemorrhagic E. coli
E. coli 0157:H7: Colorless on SMAC
Other E. coli strains are pink
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Media Test
Motility Media
Oxidative fermentative Test
Oxidative Fermentative
Nitrate Test
Nitrate Test
Shigella
- gray entire non hemolytic smooth colonies on blood agar
- catalse pos
- colonies w/o yellow coloration on he agar
- Shigella colonies appear green with the color fading to the periphery of the colony.
- inert biochemical rxns
- bacillary dysentery
- characterized by blood and pus in stool
- S. sonnei (most common in US)
- S. flexneri
- S. dysenteriae
- S. boydii
- Very low id50
Triple Sugar Iron Agar
- Slant:Alkaline(K)
- Butt:Acid (A)
- Gas:Neg
- H2S:Neg
TSI Agar
Rapid Spot Indole
Positive or Negative Result for groups A B and C
sonnei - negative result
Rapid Spot Indole
Rapid Cytochrome Oxidase
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
- good growth
- lack of red color
- non lactose fermenter
- produce colorless or transparent colonies with rare exceptions
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Media Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Salmonella
Salmonella
- gray entire nonhemolytic smooth colonies on blood ag
- oxidase neg
- catalse pos
- black colonies with a red halo on xld agar
- alk/acid rxn on kia w/h2s
- lysine pos
- common diseases
- • Gastroenteritis
- • Bacteremia or septicemia
- • Enteric fever
- • Carrier state
Triple Sugar Iron Agar
Slant: alkaline (K)
Butt: acid (A)
Gas: Pos
H2S: Pos
TSI Agar
Rapid Spot Indole Test
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
- good growth
- lack of red color
- non lactose fermenter
- produce colorless or transparent colonies with rare exceptions
Voges Proskauer Test
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Media Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Citrobactor
slow/weak lactose fermenter
Citrobactor
Triple Sugar Iron Test
TSI Agar
Rapid Spot Indole Test
Positive or Negative Result
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
may appear colorless or slightly pink after 24 hours
MacConkey Agar Apperance
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Urea Test
Positive and Negative Result
Urea Test
Motiltity Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Triple Sugar Iron Agar
Slant: Acid (A)
Butt: Acid (A)
Gas: Pos
H2S: Neg
TSI Agar
Rapid Spot Indole Test
kliebsiella pneumoniae - neg
klebsiella oxytoca - pos
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
Strong rxn, produces pink colonies with pink/red pigment diffsusing surrounding agar
MacConkey Agar Apperance
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Triple Sugar Iron Agar
Slant: Acid (A)
Butt: Acid)9A)
gas pos
h2s neg
TSI Agar
Rapid Spot Indole Test
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar AppAppearance
MacConkey Agar Apperance
Pink colored after 24 hr incubation; pale-colored colonies turn pink after further 24 hr incubation.
mucoid colonies but smaller than Klebsiella.
Voges Proskauer Test
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Urease Test
Positive or Negative Result
Urea Test
Motility Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Triple Sugar Iron Agar
Slant alkaline
butt acid
gas neg
h2s neg
TSI Agar
Rapid Spot Indole Test
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
growth appears red w/o pigment color leaching into surrounding agar
MacConkey Agar Apperance
Voges Proskauer Test
Voges Proskauer
Citrate Utilization
Citrate Utilization
Motility Test
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Proteus
- oxidase negative
- catalse pos
- black pigment
- h2s pos colonies on xld agar w/o surrounding halo
- pad tda pos
Triple Sugar Iron Agar
slant alkaline
butt acid
gas pos
h2s pos
TSI Agar
Rapid Spot Indole Test
certain strains may give false negatives with this test
proteus mirabilis - neg
proteus vulgaris - pos
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance Test
MacConkey Agar Apperance
- Spreading colonies on blood agar
- no swarming on MacConkey agar specifically
- colorless or transparent colonies with rare exceptions
- foul smell
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Positive or Negative Result
Citrate Utilization
Motility Test
Motility Media
Oxidative Fermentative test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Triple Sugar Iron Agar
TSI Agar
Rapid Spot Indole Test
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
Flat colorless non-lactose fermenters
MacConkey Agar Apperance
Voges Proskauer
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Test
Motility Media
Oxidative-Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Yersinia
- relatively small flat irreugular gray/yellow smooth colonies on blood agar
- optimal growth at 25C
- oxidase neg
- catalase pos
- indole pos
- lysine neg
- ornithine pos
Yersinia
Triple Sugar Iron
TSI Agar
Rapid Spot Indole Test
postive and negative Result
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
MacConkey Agar Apperance
- Colorless to peach pigmented colonies
- nonlactose fermenter
MacConkey Agar Apperance
Voges Proskauer Test
Voges Proskauer
Citrate Utilization Test
Citrate Utilization
Motility Media
Motility Media
Oxidative Fermentative Test
Oxidative Fermantative
Nitrate Test
Nitrate Test
Test Explanations
- what each test actaully tests for
- what a postive and negative demonstrates
Test Explantions
Triple Sugar Iron Agar
- Alkaline slant/no change in butt (K/NC) i.e Red/Red = glucose, lactose, and sucrose non-fermenter
- Alkaline slant/alkaline butt (K/K) i.e Red/Red = glucose, lactose, and sucrose non-fermenter
- Alkaline slant/acidic butt (K/A); Red/Yellow = glucose fermentation only, gas (+ or -), H2S (+ or -)
- Acidic slant/acidic butt (A/A); Yellow/Yellow = glucose, lactose and/or sucrose fermenter gas (+ or -), H2S (+ or -).
TSI Agar
Rapid Spot Indole Test
- The Indole Test is a test that demonstrates the ability of certain bacteria to decompose the amino acid tryptophan to indole, which accumulates in the medium. This test is important in the identification of Enterobacteria. The test is performed by adding Kovac’s reagent to a broth culture containing tryptophan. If indole is present, the solution will turn from yellow to cherry red
- designed to detect Gram-negative bacilli (Oxidase-negative).
Rapid Spot Indole
Rapid Cytochrome Oxidase Test
Rapid Cytochrome Oxidase
Voges Proskauer Test
Voges Proskauer
- The Voges-Proskauer (VP) Test is a biochemical test used in microbiology to determine if an organism produces acetylmethyl carbinol from glucose fermentation. This test was first observed by Voges and Proskauer in 1898
- Positive test:
- The medium changes color from yellow to pink-red, indicating that the organism can produce acetylmethyl carbinol1.
- Negative test:
- The medium remains yellow or fails to turn pink-red within 48 hours, indicating that the organism cannot produce acetylmethyl carbinol.
- identification of several genera and species of Enterobacteriaceae, including Enterobacter, Klebsiella, Serratia
Urea Test
Urea Test
- used in microbiology to determine if an organism can produce the enzyme urease. This enzyme hydrolyzes urea into ammonia and carbon dioxide.
- Positive test:
- The medium changes color from light orange to magenta, indicating that the organism can produce urease12.
- Negative test:
- The medium remains light orange, indicating that the organism cannot produce urease12.
- The Urease Test is used to differentiate organisms based on their ability to hydrolyze urea with the enzyme urease1. This test can be used as part of the identification of several genera and species of Enterobacteriaceae, including Proteus, Klebsiella, and some Yersinia and Citrobacter species
Motility Media test
Motility Media
- The Motility Test is a method used in microbiology to determine if an organism can move by itself. This ability is mostly demonstrated in a semi-solid agar medium. In semi-solid agar media, motile bacteria ‘swarm’ and give a diffuse spreading growth that is easily recognized by the naked eye.
- Positive test:
- Diffuse, hazy growths that spread throughout the medium rendering it slightly opaque.
- Negative test:
- Growth that is confined to the stab-line, with sharply defined margins and leaving the surrounding medium clearly transparent.
- used particularly with proteus, morganella and providencia
Nitrate Test
Nitrate Test
- Positive test: The medium changes color from yellow to red, indicating that the organism can produce nitrate reductase1.
- Negative test: The medium remains yellow, indicating that the organism cannot produce nitrate reductase1.
- The Nitrate Reduction Test is used to differentiate organisms based on their ability to produce nitrate reductase1. This test can be used as part of the identification of several genera and species of Enterobacteriaceae, including Proteus, Morganella, and Providencia2.