microbiology lab

Bile Esculin Agar

Used to distinguish enterococci from streptococci
Enterococci form colonies
Streptococci dont

Mannitol salt agar

Used to culture staphylococcus species
Other bacterial species do not form

Eosin Methylene Blue Agar

Used for presumptive identification of Escherichia Coli
E. Coli Colonies have metallic green sheen

MacConkey Agar

Primary isolation medium for Enterobacteriaceae and other selected aerobic gram-negative bacilli.
Lactose-positve= red colonies
Lactose-negative=colorless colonies

Salmonella-shigella Agar

Used to distinguish Salmonellae from shigellae
Colonies of H2S-positive salmonellae=colonies with black centers
colonies of H2s-negative shigellae=colorless colonies
Sodium thiosulfate=source of sulfur
Ferric Citrate=H2S indicator

Bacitracin Disk

Used to identify streptococcus pyogenes

Optochin disk

Used to identify streptococcus pneumoniae

Mueller-Hinton Agar

Standardized media for kirby-bauer method.
Supplemented with 5% ovin blood.

Brain heart infusion blood agar

Primary isolation medium for obligate anaerobic bacteria
5% ovine blood + brain heart infusion agar

Chocolate Agar

hemin + NAD
Cultivate selected bacteria with fastidious nutritional requirements

Blood Agar

Primary isolation for aerobic bacteria
Tryptose sugar and 5% ovine blood

Motility Test

Soft agar with .5% agar and tetrazolium salts(Indicator)
Obligate aerobic and facultative anaerobic bacteria
Obligate aerobe and motile-adjacent to agar surface
obligate aerobe and nonmotile-adjacent to innoculation line at agar surface
Facultative anaero

Hanging Drop method

Drop of broth culture on a cover glass which is inverted on a glass slide and observed with a brightfield microscope.
Aerobic gram-negative helical bacilli with polar flagella=corkscrewlike motility. campylobacter, heliobacter, vibrio

Catalase Test

1st test for aerobic gram-positive
Detects catalase(iron-containing)
Catalse hydrolyzes hydrogen peroxide to water and oxygen
Catalase-positive=gas bubbles
catalase-negative=no gas bubbles
False positive-blood agar because have erythrocytes

Oxidase Test

1st biochemical test for aerobic gram-negative
Detects cytochrome C
Oxidase + =blue color from idophenol blue
oxidase - = no color change
false-positive=old oxidase reagent

Nitrate Reduction Test

ID aerobic bacteria, particularly gram-negative bacilli in Enterobacteriaceae
+ test=nitrate is reduced to nitrite by nitrate reductase or to other nitrogenous products.
1. Nitrate reduction positive where nitrate has been reduced to nitrite=reagents A an

Nitrate Agar method

Aerobic, 37 degrees for 24 hours
1 drop of reagent A(sulfanilic acid) +
1 drop of reagent B(alpha-napthylamine) added to slant
- test=Add zinc dust to slant

Nitrate Disk method

Aerobic 37 degrees, 24 hours
1 drop of reagent A(Sulfanilic acid)+1 drop of reagent B(alpha-napthylamine) are added to paper disk
nitrate reduction - =Add zinc dust to paper plate

Kilger Iron Agar

saccharolytic facultative anaerobic bacteria ferment glucose=acid reaction in butt=black precipitate by H2S producing bacteria
Contains glucose, lactose, phenol Red PH indicator(pH 7.4)
Incubated for 24 hours at 37 degrees.

Lysine Iron Agar

Saccharolytic facultative anaerobic bacteria ferment glucose=acid reaction=black precipitate by H2S producing bacteria.
Sodium thiosulfate=sulfur source
Ferric ammonium citrate=H2S indicator

Aerogenic bacteria

Produce acid and gas from a charbohydrate substrate

Anaerogenic bacteria

Produce acid from charbohydrate substrate

Methyl Red test

.5% glucose in broth medium
24 hours at 37 degrees
Tests for acid production via a mixed acid fermentation pathway of glucose.
After 24 hours add methyl red PH indicator
red=methyl red-positive with pH less than or =4.4=short chain fatty acids produced

Voges-prokauer Test

Based on fementative degradation of glucose to acetylmethylcarbinol(acetoin)(intermediate in butanediol fermentation)
acetylmethylcarbinol oxidzed
After 24 incubation, 2 drops of alpha napthol and 2 drops of creatine+KOH.
orange=positive test=acetylmethyl

Citrate Agar

Used to determine if citrate can be used as sole carbon source and energy.
Ammonium phosphate=nitrogen source
Bromothymol blue=pH indicator
1.Blue=positive=bacterial growth on agar and production of ammonium hydroxide and alkanization of medium
2. Green=n

carbohydrate metabolism

1% concentration to nutrient broth medium
pH 7.4
phenol red-pH indicator
37 degrees for 24 hours
Glucose + =yellow with acid, sometimes gas
lactose and mannitol positive=yellow and acid
Glucose, lactose, mannitol negative =red and alkaline pH

Lysine Iron Agar

Lysine, glucose, ferric ammonium citrate(H2S indicator), sodium thiosulfate(sulfur source)
pH indicator=bromocresol (pH 7.4)
Detect lysine deaminase + and lysine decarboxylase + and H2S production
24 hours, 37 degrees, stab and streak
deaminase -, decarbo

Ornithine Broth

24 hours, 37 degrees
pH indicator-bromocresol purple
Detects ornithine decarboxylase-positive reaction with increased pH, dark purple, putrescine produced
negative reaction=yellow

Tryptophane Broth

Reagent-Kovac's reagent, 2 drops added after 24 hours
37 degrees, 24 hours
Kovac's reagent reacts with indole=red compound
indole +=tryptophane hydrolyzed, producing indole, pyruvic acid and ammonia. Occur immediately
Indole negative=no color change

Urea Agar

Phenol Red-pH indicator
Urea hydrolyzed by urease=ammonia+ carbon dioxide
ammonia +water=ammonium carbonate + alkaline
urease + =pH increased, pink or red
Urease - = no color change
Urea agar with acid pH=amber or colorless