Diagnostic of Medically important
Gram Negative Bacteria
Representative Gram Negative
Bacteria
• Gram-negative Cocci
– Neisseria gonorrhoea
– Neisseria meningitis
• Gram-negative rods
– Enterobacteriaceae family
• Escherichia, Enterobacter, Salmonella, Shigella,
Klebsiella, Proteus, Morganella
– Pseudomonaceae family
• Pseudomonas
Identification
+
Pseudomonaceae
Neisseriaceae
+
Neisseriaceae
—
Pseudomonaceae
Fermentation
of glucose
Oxydase
+
—
Enterobacteriaceae
Fermentation
of lactose
McConkey
—
Enterobacter
Escherichia
Klebsiella
Serratia
Proteus
Salmonella
Shigella
Morganella
Identification of Oxidase Negative
Gram Negative Rods
• MacConkey
– Selective and
differential medium
– Prevents growth of
Gram +
– Lactose fermentation
• Pink colonies
IMViC Tests
• Indole, Methyl Red, Voges-Prosakaur, Citrate
(IMViC) :
– These four tests include an important series of
determinations which are collectively called the
IMViC reaction series
– The IMViC reaction series allows the
discrimination
of
bacteria
of
the
Enterobacteriaceae family.
IMViC: Indole Test
• Principal
– Some microorganisms can metabolize tryptophane
by the tryptophanase
Tryptophane
Tryptophanase
Indole + acide Pyurvic + NH3
Kovac’s reagent
Red color
IMViC Test
Methyl Red-Voges Proskauer
• Methyl Red Test :
– Fermentation with accumulation of acids:
• Glucose  pyruvate  lactic and/or acetic acid + CO2
-
+
-
+
• Voges Proskauer Test
– Fermentation with accumulation of butanediol
– Glucose  pyruvate  acetoine  2 butanediol + CO2
IMViC Test : Citrate Utilization
• Unique carbon source
– Citrate
• Indicator
– Bromthymol blue
• Citrate utilization generates alkaline
end products
– Changes from green to blue
Positive
Klebsiella, Enterobacter
Negative
E. coli
TSI — Three Sugars and Iron
• Three sugars
– Glucose (limiting)
– Sucrose
– Lactose
• Proteins
– Cysteine
• Indicator
– Phenol red
SIM — H2S, Indole and Motility
• Semi-solid medium
– Allows to visualize motility
• Cystein metabolism
CysteineH2S; H2S+ FeSO4 Black precipitate
• Tryptophan metabolism
(A) Tryptophan Indole + NH4 + Pyruvate
(B) Indole + Kovac reagent Red
Non inoculated
Non-motile
+
H2S and motile
Indole
-
Multi Test: Enteropluri-test
• Strips of multiple metabolic tests
–
–
–
–
Use of a constant inoculum
Quick
Reading can be automated
Preparations and inconsistencies are normalized