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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 CysteineH2S; 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