Microbiology Media Review- Practical #1

Mannitol Salt Agar

Selective for Halotolerant Organisms

Differential for Mannitol fermentation

Fermentative (+): Yellow Agar

Negative Hektoen Enteric Agar

Selective for Gram (-) sp.

Differential for Lactose, Sucrose, & Salicin

Neutral: Blue/ Green

Positive Hektoen Enteric Agar

Selective for Gram (-) sp.

Differential for Lactose, Sucrose, & Salicin Fermentation (+): Yellow/ Orange,

Blood Agar

Differential for hemolysis of red blood cells

Alpha (α, partial lysis, green/brown)- hemolysis (Beta: full lysis, clear halo, Gamma: no lysis)

MacConkey Agar

Selective for gram (-) sp.

Differential for lactose fermentation

Fermentation (+): Pink Colonies

Beige agar (-): Peptone Utilization

EX: Escherichia coli

Tellurite Agar

Differential for organisms ability to reduce Tellurite → Tellurium

Gram (+) sp. prefer this medium!

Milk Agar

Tests for organisms ability to use casienase to break down casein.

Casein hydrolysis (+) - Clearing around organism

Skim Milk Agar

Tests for organisms ability to use casienase to break down casein.

Inability for casein hydrolysis (-) - No clearing

Starch Agar

Tests for organisms ability to breakdown starch via the amylase enzyme.

Ability to use amylase to breakdown starch (+) - Clearing of organism

DNase Agar

Differentiates organisms based on the ability to produce the DNase enzyme.

Ability to produce DNase (+) - Clearing around organism

Sugar Fermentation Broth

Tests for organism's ability to ferment sugars (i.e., Glucose or Lactose).

Fermentative sp. (+) - Yellow broth

Urea Broth

Tests for organisms ability to use Urease enzyme to hydrolyze.

Positive test: Magenta broth

Arginine Dihydrolase Medium

Tests for the presence of the Arginine Dihydrolase enzyme.

Positive test: Broth turns purple

Lysine Decarboxylase Medium

Differentiates sp. based on their ability to make the Lysine Decarboxylase enzyme.

Production of enzyme (+): Purple broth

No enzyme production (-) test: Yellow broth

Negative Gelatinase Medium

Tests for organisms ability to produce gelatinase to hydrolyze gelatin.

Negative test- Solid medium

Positive Gelatinase Medium

Tests for organisms ability to produce gelatinase to hydrolyze gelatin.

Positive test (+) - Liquid medium

Nitrate Broth

Tests organisms ability to reduce Nitrate to Nitrite (or Nitrogen/ Ammonia).

Reagents: Sulfanilic Acid + α-naphylamine / Zinc for negative Rxn's (colorless after addition of reagents).

Nitrate Broth w/o addition of Zinc: Red

Organism is able to reduce Nitrate to Nitrite.

Nitrate Broth w/ addition of Zinc: Colorless

Reduction of Nitrate to Nitrogen or Ammonia (N2 or NH3)

Nitrate Broth w/ addition of Zinc: Red

Zinc reduces Nitrate NOT the organism.

Positive Catalase test

Microorganism produces catalase enzyme - bubbles form

Negative catalase test

Microorganism does not produce catalase enzyme - no bubbles

Positive oxidase test

Redox gas reduces cytochrome c oxidase - colorless dye turns dark purple

Negative oxidase test

Redox gas does not reduce cytochrome c oxidase - colorless dye remains

Crystal violet is removed after 95% EtOH rinse

Gram (-)

Peptidoglycan layer shrinks and dehydrates, retaining crystal violet/grams iodine complex

Gram (+)

Positive acid fast stain

Carbol fuchsin binds to mycolic-acid - fuchsia color

Negative acid fast stain

Deep blue - mycolic-acid is not present

Positive string test

Gram (-)

Potassium hydroxide breaks down cell walls and releases chromosomal material

Bacterial suspension becomes thick and sticky

Safranin counter stain

Stains both pink

Gram (+): crystal violet + pink = purple

Gram (-): pink

Crystal violet

Stains both purple

Entangles with peptidoglycan layer

Capsule staining - Congo Red

Stains proteins in the background of the cells

Hektoen enteric agar enzyme

Thiosulfate reductase

Gram (-) cell walls

Double phospholipid bilayer surrounding thin peptidoglycan layer

Endospores

Some bacteria grown them - they protect vegetative DNA from extreme environments

Acid fast staining - Methylene Blue

Carbol fuchsin stains bright red, blue stain provides contrast between (+) and (-)

Grams iodine in gram staining

Forms complex w/ crystal violet in peptidoglycan layer

ONLY GRAM (+)

Halotolerant organisms

Capable of surviving and growing in high-salt environments but don’t require them

EX: Bacillus cereus