Microbiology Lab Practical Organism Profiles

Escherichia coli (E. coli)

• Identification: Gram (-), rod, single rods, 2–3 μm × 0.5 μm, motile (peritrichous flagella), facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), not halotolerant

• Cell Structures: Thin peptidoglycan, outer membrane with LPS (Lipid A endotoxin), porins, fimbriae, pili, no endospores, not acid-fast, capsule not prominent

• Biochemical Characteristics: Lactose fermenter, H₂S negative, mannitol not tested (doesn't grow on MSA), usually γ-hemolytic

• Colony Characteristics: TSA: Cream/off-white, circular, smooth, moist, convex; Broth: Turbid

• Signature Media: EMB (metallic green sheen), MAC (bright pink colonies), HE (salmon/orange colonies)

• Quick ID Clues: Metallic green on EMB, pink on MAC, salmon/orange on HE, pink Gram-negative rod, growth throughout thioglycollate (more at top), motile (peritrichous flagella), LPS = Gram-negative outer membrane, no endospores, not acid-fast

• Memory Sentence: Gram-negative, motile, facultative anaerobic rod that ferments lactose → metallic green on EMB, pink on MAC, salmon/orange on HE, and grows throughout thioglycollate with heavier growth at the top.

Staphylococcus aureus (S. aureus)

• Identification: Gram (+), cocci in clusters, 0.8–1 μm, non-motile, facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), halotolerant (grows in 7.5% NaCl)

• Cell Structures: Thick peptidoglycan, teichoic & lipoteichoic acids, no outer membrane, may produce microcapsule (not emphasized), no endospores, not acid-fast, no flagella

• Biochemical Characteristics: Mannitol fermenter, halotolerant, lactose not tested, H₂S negative, β-Hemolytic (Blood Agar)

• Colony Characteristics: TSA: Cream to golden-yellow, circular, smooth, moist, convex, opaque; Broth: Turbid

• Signature Media: MSA (yellow media), Blood Agar (β-hemolysis)

• Quick ID Clues: Purple cocci in clusters, yellow MSA, β-hemolysis, grows on MSA but not on MAC/EMB/HE, facultative anaerobe, non-motile, thick peptidoglycan + teichoic acids, no endospores, not acid-fast

• Memory Sentence: Gram-positive, non-motile cocci in clusters that tolerate salt, ferment mannitol (yellow MSA), produce β-hemolysis on Blood Agar, grow throughout thioglycollate with heavier growth at the top, and retain crystal violet because of their thick peptidoglycan cell wall.

Staphylococcus epidermidis (S. epidermidis)

• Identification: Gram (+), cocci in clusters, 0.8–1 μm, non-motile, facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), halotolerant

• Cell Structures: Thick peptidoglycan, teichoic & lipoteichoic acids, no outer membrane, produces slime layer/biofilm, no endospores, not acid-fast, no flagella

• Biochemical Characteristics: Mannitol non-fermenter, halotolerant, H₂S negative, usually γ-hemolytic

• Colony Characteristics: TSA: White to cream, circular, smooth, moist, convex, opaque; Broth: Turbid

• Signature Media: MSA (pink/red media)

• Quick ID Clues: Purple cocci in clusters, grows on MSA but media stays pink/red, halotolerant, non-motile, facultative anaerobe, biofilm producer, usually γ-hemolytic, no endospores, not acid-fast

• Memory Sentence: Gram-positive, non-motile cocci in clusters that are halotolerant but do not ferment mannitol, so they grow on MSA while the media stays pink/red. They are facultative anaerobes, usually γ-hemolytic, and are known for forming biofilms on medical devices.

Salmonella spp.

• Identification: Gram (-), rod, single rods, 2–5 μm × 0.7–1 μm, motile (peritrichous flagella), facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), not halotolerant

• Cell Structures: Thin peptidoglycan, outer membrane with LPS, porins, flagella, fimbriae, some serotypes have Vi capsule (not emphasized), no endospores, not acid-fast

• Biochemical Characteristics: Lactose non-fermenter, H₂S positive, usually γ-hemolytic

• Colony Characteristics: TSA: Cream to gray-white, circular, smooth, moist, convex; Broth: Turbid

• Signature Media: HE Agar (green colonies with black centers), MAC (colorless), EMB (colorless)

• Quick ID Clues: Gram-negative motile rod, green colonies with black centers on HE, H₂S positive, colorless on MAC/EMB, facultative anaerobe, growth throughout thioglycollate, LPS-containing outer membrane, no endospores, not acid-fast

• Memory Sentence: Gram-negative, motile, facultative anaerobic rod that does not ferment lactose but does produce H₂S, giving green colonies with black centers on HE agar and colorless colonies on MacConkey and EMB.

Shigella dysenteriae (Shigella)

• Identification: Gram (-), rod, single rods, 2–3 μm × 0.5–0.7 μm, non-motile, facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), not halotolerant

• Cell Structures: Thin peptidoglycan, outer membrane with LPS, porins, no flagella, may have fimbriae, no endospores, not acid-fast, no capsule emphasized

• Biochemical Characteristics: Lactose non-fermenter, H₂S negative, usually γ-hemolytic

• Colony Characteristics: TSA: Small, smooth, circular, gray-white; Broth: Turbid

• Signature Media: HE Agar (green colonies, no black centers), MAC (colorless), EMB (colorless)

• Quick ID Clues: Gram-negative non-motile rod, green colonies on HE without black centers, H₂S negative, colorless on MAC/EMB, facultative anaerobe, growth throughout thioglycollate, LPS-containing outer membrane, no endospores, not acid-fast

• Memory Sentence: Gram-negative, non-motile, facultative anaerobic rod that does not ferment lactose or produce H₂S, giving green colonies on HE agar (without black centers) and colorless colonies on MacConkey and EMB.

Bacillus cereus

• Identification: Gram (+), rod, single rods/chains, 3–5 μm × 1 μm, motile (peritrichous flagella), facultative anaerobe, mesophile (37°C), neutrophile (pH 6.5–7.5), not halotolerant

• Cell Structures: Thick peptidoglycan, teichoic acids, subterminal/central endospore, no capsule emphasized, not acid-fast

• Biochemical Characteristics: Endospore former, usually β-hemolytic, not known for lactose/mannitol fermentation

• Colony Characteristics: TSA: Large, flat, rough, dry, irregular edges, white/cream

• Signature Lab: Endospore Stain (green endospores with pink vegetative cells)

• Quick ID Clues: Gram-positive rod, endospore former, motile, large rough colonies, facultative anaerobe

• Memory: Gram-positive, motile rod that forms endospores.

Bacillus subtilis

• Identification: Gram (+), rod, chains, 4–10 μm, motile, obligate aerobe, mesophile, neutrophile, not halotolerant

• Cell Structures: Thick peptidoglycan, elliptical/subterminal endospores, not acid-fast

• Biochemical Characteristics: Endospore former

• Colony Characteristics: TSA: Large, dry, rough, wrinkled, white/cream

• Signature Lab: Endospore Stain (green spores with pink vegetative cells)

• Quick ID Clues: Gram-positive rod, endospore former, obligate aerobe, motile

• Memory: Aerobic Bacillus that forms endospores.

Clostridium sporogenes

• Identification: Gram (+), rod, single rods, 3–8 μm, motile, obligate anaerobe, mesophile, neutrophile, not halotolerant

• Cell Structures: Thick peptidoglycan, terminal endospore ("drumstick"), not acid-fast

• Biochemical Characteristics: Endospore former, fermentative metabolism, oxygen is toxic

• Colony Characteristics: TSA (anaerobic): Gray-white, irregular, spreading colonies; Broth: Turbid under anaerobic conditions

• Signature Lab: Endospore Stain (green terminal spores), Thioglycollate Broth (growth only at bottom)

• Quick ID Clues: Gram-positive rod, terminal endospore, bottom of thioglycollate, motile, obligate anaerobe

• Memory: Bottom of the tube + terminal spore = Clostridium.

Mycobacterium gordonae

• Identification: Weakly Gram-positive/Gram-variable, rod, single rods, 2–5 μm × 0.2–0.6 μm, non-motile, obligate aerobe, mesophile, neutrophile, not halotolerant

• Cell Structures: Waxy cell wall with mycolic acids, thin peptidoglycan, no endospores, no LPS, capsule not emphasized

• Biochemical Characteristics: Acid-fast positive, slow-growing, does not form endospores

• Colony Characteristics: TSA: Slow-growing, dry, yellow-orange pigment

• Signature Lab: Acid-Fast Stain (bright red/fuchsia rods)

• Quick ID Clues: Acid-fast positive, bright red rods, mycolic acid cell wall, obligate aerobe, non-motile, no endospores

• Memory: Red rod = Mycobacterium.

Micrococcus luteus

• Identification: Gram (+), cocci (tetrads), 0.5–3 μm, non-motile, obligate aerobe, mesophile (37°C), neutrophile (6.5–7.5), salt tolerance variable

• Cell Structures: Thick peptidoglycan, teichoic acids, no outer membrane, no endospores, not acid-fast, no capsule emphasized

• Biochemical Characteristics: Produces yellow carotenoid pigment, obligate aerobe

• Colony Characteristics: TSA: Bright yellow, circular, smooth, convex, entire margin, opaque

• Signature Lab: TSA/Simple Stain (bright yellow colonies, purple cocci)

• Quick ID Clues: Gram-positive cocci, bright yellow colonies, often tetrads, obligate aerobe, non-motile, top growth in thioglycollate, no endospores, not acid-fast, no LPS

• Memory: Yellow colony = Micrococcus.

Saccharomyces cerevisiae (Yeast)

• Identification: Eukaryote (fungus), oval, single cells/budding, 5–10 μm, non-motile, facultative anaerobe, mesophile (25–37°C), neutrophile, salt tolerance not emphasized

• Cell Structures: Nucleus, membrane-bound organelles, cell wall (glucans, mannans, chitin), vacuole, reproduces by budding, no bacterial endospores, not acid-fast

• Biochemical Characteristics: Ferments sugars, produces CO₂ and ethanol, facultative anaerobe

• Colony Characteristics: TSA: Cream-colored, smooth, moist, buttery, convex

• Signature Lab: Simple Stain (large oval budding cells)

• Quick ID Clues: Largest organism viewed, eukaryote, oval cells, budding reproduction, cream-colored colonies, facultative anaerobe, ferments sugars

• Memory: Large budding cells = Yeast.

Serratia marcescens

• Identification: Gram (-), rod, single rods, 0.5–0.8 μm × 0.9–2 μm, motile (peritrichous flagella), facultative anaerobe, mesophile, neutrophile, not halotolerant

• Cell Structures: Thin peptidoglycan, outer membrane with LPS, peritrichous flagella, fimbriae, no endospores, not acid-fast

• Biochemical Characteristics: Variable lactose fermentation (usually non-fermenter), produces prodigiosin (red/pink pigment), facultative anaerobe

• Colony Characteristics: TSA: Smooth, circular, moist, convex, red/pink pigment (especially at room temperature, ~25°C)

• Signature Characteristic: Red/Pink Pigment (prodigiosin)

• Quick ID Clues: Gram-negative rod, red/pink pigment, motile, facultative anaerobe, LPS-containing outer membrane, no endospores, not acid-fast

• Memory: Gram-negative, motile, facultative anaerobic rod famous for producing a red/pink pigment (prodigiosin), especially when grown at room temperature.

High-Yield Comparison Lists

Gram-Positive Organisms

• Cocci: Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus

• Rods: Bacillus cereus, Bacillus subtilis, Clostridium sporogenes, Mycobacterium gordonae (weakly Gram-positive/acid-fast)

Gram-Negative Organisms

• Escherichia coli

• Salmonella enterica

• Shigella dysenteriae

• Serratia marcescens

Eukaryote (Not Gram Positive or Gram Negative)

• Saccharomyces cerevisiae (yeast)

Motile Organisms

• Gram-Positive: Bacillus cereus, Bacillus subtilis, Clostridium sporogenes

• Gram-Negative: Escherichia coli, Salmonella enterica, Serratia marcescens

Non-Motile Organisms

• Gram-Positive: Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus, Mycobacterium gordonae

• Gram-Negative: Shigella dysenteriae

• Eukaryote: Saccharomyces cerevisiae

Oxygen Requirements

• Facultative Anaerobes: Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Bacillus cereus, Salmonella enterica, Shigella dysenteriae, Serratia marcescens, Saccharomyces cerevisiae

• Obligate Aerobes: Bacillus subtilis, Mycobacterium gordonae, Micrococcus luteus

• Obligate Anaerobe: Clostridium sporogenes

Thioglycollate Broth Growth Patterns

Signature Media & Hallmark Features

Exam Memory Tricks

• Metallic green? → E. coli

• Yellow MSA? → S. aureus

• Pink MSA? → S. epidermidis

• Black center on HE? → Salmonella

• Green only on HE? → Shigella

• Green spores? → Bacillus or Clostridium

• Red acid-fast rods? → Mycobacterium

• Bright yellow colony? → Micrococcus

• Large budding oval cells? → Saccharomyces

• Bottom of thioglycollate? → Obligate anaerobe

• Growth only at top? → Obligate aerobe

• Growth throughout with more at the top? → Facultative anaerobe

Additional info:

• All organisms are mesophiles (optimal growth ≈37°C) unless noted.

• All are neutrophiles (optimal pH 6.5–7.5).

• Endospore stain is the "Christmas tree" mnemonic: green = endospore, pink = vegetative cell.

• HE agar is designed for enteric Gram-negative bacteria (E. coli, Salmonella, Shigella).

• Serratia marcescens pigment (prodigiosin) is best produced at room temperature (~25°C).