biochemical test
TRANSCRIPT
Basic Microbiology laboratory for Dental Students
By LecturerYasemin Al-Shboul
Spring2011-2012
Biochemical Activities of Microorganisms (Part 2)
• IMViC Reaction: Indole Test, Methyl Red Test, Voges-Proskauer Test, and Citrate Test.
• Indole Test:• The Indole test is a biochemical test performed on bacterial species to determine the ability of the
organism to split Indole from the amino acid tryptophan. This division is performed by a chain of a number of different intracellular enzymes, a system generally referred to as tryptophanase .
• Tryptophan
• Kovac's reagent (Yellow) + Indole --------- Red Color
Procedure.
Interpretation: Pink-red ring: Positive / Yellow ring: Negative
• Indole test
Biochemical Activities……
MR-VP Tests:Divide Enterobacteriaceae into two groups based on the way the
metabolize glucose.
Methyl Red (MR) TestDetermines the ability of an organism to produce acids as final products from glucose fermentation. Fermentation
Glucose Lactic Acid, Acetic Acid, Formic Acid
Red Color MR indicator + Acidic pH
fermentation
MR TEST• Methyl red is a pH indicator; it is red in pH under 4.4, yellow in pH over 6.2
Biochemical Activities..……
Procedure.
Interpretation:
Red color: Positive / Yellow color: Negative
Voges-Proskauer (VP) Test
Determines the ability of an organism to produce Acetoin as a final product from glucose fermentation.
Glucose Acetoin
Acetoin + α-Naphthol Diacetyl
fermentation
40% KOH
Oxidation
• VP TEST
Biochemical Activities..……
Diacetyl + Peptone (from medium Pink Red
Procedure.
InterpretationPink Red color: Positive / Yellow color: Negative
Citrate Test:Determines the ability of an organism to use citrate as the only source of carbon.
Citrate Organic Acids + CO2
CO2 + Na (from conjugate base salt) Na2CO3
α-Naphthol
Citrate
Utilization
• Citrate test
Biochemical Activities..……
Bromothymol Blue Indicator (Green) Blue
Procedure InterpretationPink Red color: Positive / Yellow color: Negative
Urease Test:Determines the presence of Urease enzyme.
Urea NH4 (Ammonia) + CO2
Phenol Red Indicator (Yellow) Pink-Red
Na2CO3
Alkaline pH
Urease
NH4
Alkaline pH
• Urease test
Biochemical Activities..……
Procedure.InterpretationPink Red color: Positive.
Yellow color (no change): Negative.
Motility Test:
Many bacteria are capable of movement.Most motile bacteria use structures called Flagella.
Flagella rotate in a counterclockwise direction, thus giving motility.
Biochemical Activities..……
Flagella Types:Monotrichous: single polar flagellum.
Lophotrichous (tufts): multiple polar or bipolar flagella.
Peritrichous: flagella distributed over the entire cell.
Procedure.
Interpretation:
Turbid media: Positive / Clear around stab line: Negative.
Triple Sugar Iron (TSI) Agar: Differentiates bacteria based on their ability to ferment glucose, lactose, and
sucrose to acid end products, +/- gas and to hydrogen sulfide (H2S).
Kligler Iron Agar (KIA): similar to TSI Agar but without sucrose.
• Motility test
Biochemical Activities..……TSI agar media contents:
0.1% Glucose.
1% Lactose.
1% Sucrose .
Phenol Red pH Indicator: Yellow (Acid), Red (Base).
Sodium Thiosulfate: for hydrogen sulfide production.
Ferric Ammonium Sulfate: as H2S indicator, which forms ferric sulfide
(FeS) Black Precipitate.Procedure.
Interpretation.
Biochemical Activities..……
A=Acidic (Yellow color) / K-Alkaline (Red color)
Slant / Butt
K/A: Only Glucose fermentation
A/A: Glucose & Lactose fermentation.
K/K or K/No Change: No G. or L. Fermentation
A/K: Will never happen.
Black Precipitate: H2S Production.
Cracking in media: Gas Production (CO2, H2).
• KIA AGAR
Example KIA IMViC Urease Motility
Escherichia coli A/A/G - - + +-+Kelbsiella pneumonia
A/A/G+ + - -+-Enterobacter
aerogenes A/A/G+ + - --+
Salmoella typhi K/A/G/H2S + - + --+Proteus volgarisK/A/G/H2S+ - + -++Shigella sonneiK/A- - + +--Pseudomonas
aeruginosaK/K+ - - --+
GOOD LUCK