Identification of unknown bacterium PRACTICAL Identification of unknown bacterium

Why species level identification required? To know which species is involved in the particular condition To know the subspecies To further know the types Immunotypes, serotypes, chemotypes etc Epidemiological studies

Basic steps to follow

Pure culture follow up-Day 1

Pure culture follow up-Day 1

First day – preliminary findings Growth characteristics Gram stain Catalase Oxidase Motility

Presumptive follow up

Members of enterobacteriaceae: oxidase Negative Non enterobacteriaceae Oxidase positive Escherichia Klebsiella Citrobacter Salmonella Shigella Proteus etc Vibrios Pseudomonas etc

Members of enterobacteriaceae: oxidase Negative(example) LF (ONPG) NLF Escherichia Salmonella Klebsiella Shigella Citrobacter Proteus

A gram negative rod on BA/Macconkey agars Non hemolytic, moist, irregular, flat, centrally raised, 3-5 mm colonies, Lactose fermenting, Flat,irregular centrally raised, moist 2-4 mm colonies

Tests to read on second day Check whether the culture is pure Repeat catalase, oxidase,gram and motility to confirm previous day’s findings

Biochemical tests So it is Escherichia coli TSI: A/A, Gas +, H2S Negative SIM: H2S,Neg, Indole: Positive., Motile Motility; (PW) Motile It could be ESCHERICHIA, CITROBACTER,OR ENTEROBACTER Glucose : Positive (Acid and Gas) Lactose : Positive Sucrose : Negative / Positive Mannitol : Positive Indole : Positive MR : Positive VP : Positive Citrate : Negative Urease : Negative Macconkey: LF Gram: Gram Negative rods Catalase: Positive Oxidase ; Negative LAO : +-+ Melonate: - KCN ; _ So it is Escherichia coli One of the members of Enterobacteriaceae

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