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Based on colony morphology
- Assessment of gross colony characteristics is usually performed by visually inspecting growth on surface of agar plates. Plates are examined under bright, direct illumination so that light reflected from different angles. The use of hand lens or dissecting microscopes to assist in the detection of tiny or immature colonies.
Characteristics of colonies used in identification of Bacteria
- The following terms are used to describe the colony characteristics .
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Size
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Diameter in mm.
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Form
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Punctiform (pinpoint), circular, filamentous, irregular, rhizoid, spindle shape
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Elevation
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Flat, raised, convex, pulvinate, umbonate, umbilicate.
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Margin
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Entire, undulant, lobate, erose, filamentous, curled.
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Color
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White, yellow, black, buff, orange, etc
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Surface
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Glistening, dull, other.
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Density
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Opaque, translucent, transparent, other.
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Consistency
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Butyrous, viscid, membranous, brittle, other.
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Reaction in agar media used in the identification of Bacteria
- Hemolysis on blood agar
- Alpha : Partial clearing of blood around colonies with green discoloration of the medium,outline of red blood
cells intact.
- Beta : Zone of complete clearing of blood around colonies due to lysis of the red Blood cells.
- Gamma : No change in the medium around the colony; no lysis or discoloration of the red blood cells.
- Alpha prime : Halo of incomplete lysis immediately surrounding colonies with a second zone of complete
hemolysis at the periphery.
- Pigment production in agar media
- Water soluble pigments discoloring the medium eg.pyocynin
- Fluorescent pigments
- Nondiffusible pigments confined to the colonies
- Reaction in egg yolk agar
- Lecithinase : Zone of precipitate in medium surrounding colonies.
- Lipase : Pearly layer an iridescent film in and immediately surroundings colonies , visible by reflected light.
- Proteolysis : clear zone-surrounding colonies.
- Based odour production
- Although difficult to describe specifically, odors produced by the action of certain bacteria in plating media. Liquid media can be very helpful in the tentative identification of the microorganisms involved. Examples of micro organisms exhibiting distinctive odours include
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Pseudomonas species
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grape juice
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Proteus species
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burned chocolate
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Alkaligenes faecalis
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freshly cut apples
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Corynebacterium species
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fruity
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Nocardia and Streptomyces species
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musty basement
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Clostridium species
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faecal, putrid
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Pasteurella multocida
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pungent (indole) like
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- Change in differential medium
- Various dyes, pH indicators, and other ingredients are included in differential medium to serve as indicators of enzymatic activities and aids in identifying bacterial isolates.
- By assessing the described colonial characteristics and action on media, the microbiologist is able to make a preliminary identification of different bacteria isolated by primary culture. The more commonly encountered colonial types, the groups of bacteria to suspect for each and additional tests required for definite identification are illustrated in table.
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Colonial type
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Bacterial Group
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Additional tests
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Convex, entire edge,2mm to 3mm, creamy, yellowish, zone of hemolysis
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Staphylococcus sp
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Catalase ,Coagulase, DNase, Mannitol utilisation, Tellurite reduction, Novobiocin resistance
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Convex, or pulvinate, translucent, pinpoint in size, butyrous, wide zone of & hemolysis
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Streptococcus sp
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- Catalase
- 6.5% NaCI tolerance
- Bile-esculi
- CAMP test
- Hippurate hydrolysis
- L-pyrrolidonlysis
- L-pyrrolidonly- & Naphthylamid(PyR)
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Umbilicate or flat, translucent, butyrous or mucoid, broad zone of a-hemolysis
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Pneumococcus
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Pulvinate, semiopaque, gray, moist to somewhat dry, B- hemolysis may or may not be present
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Escherichia coli and other Enterobacteriaceae
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- Multiple tests
- Indole
- Methyl red
- Voges-Proskauer reaction
- Citrate
- Decarboxylases
- Urease
- Phenylalanine
- Carbohydrate fermentations
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Flat, gray; spreading as thin
Film over agar surface;
Burned chocolate odor
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Proteus
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- Phenylalanine deaminase
- Urease
- Lysine deaminase
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Flat, opaque, gray to greenish, margins erose or
Spreding, gree-blue pigment, grapelike oder
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Pseudomonas
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- Cytochrome oxidase
- Fluorescence
- Of carbohydrate assimilations Denitrification
- DNase
- Hydrolysis of acetamide
- Growth at 420 C
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Participants