MICROBIAL REACTION IN CULTERED MEDIA

A. Mannitol Salt Agar (MSA)

-contains NaCl C. Eosin methylene blue agar (EMB)
-pH dye: Phenol Red -pH indicator: Eosin y & methylene blue
-isolate, identify, and differentiation of lactose and non-lactose
Results: fermenting gram-negative bacteria
Mannitol fermenter: yellow (acidic)
Non-mannitol fermenter: red Results:
Lactose fermenter: green metallic sheen
Organism: Non-lactose fermenter: no metallic sheen
-Staphylococcus cohnii
-Staphylococcus aureus Organism:
-Escheria coli -Pseudomonas putida
-Klebsiella oxytoca

B. Macckonkey Agar (MAC)

-nitrogen source: peptone D. Catalase Test (NA)
-carbon source: lactose -ability to produce catalase and breakdown H2O2 to water and
-Bile salts = reduce surface tension oxygen
-Nutrient Agar medium
Results:
Lactose fermenter: red colonies Organism:
Non-lactose fermenter: colorless or translucent -Enterobacter aerogenes

Organism:
-Escheria coli -Pseudomonas putida
- Klebsiella oxytoca

E. Starch Hydrolysis (Starch agar)

-enzyme: amylase
-w/ gram iodine solution to react w/ starch = blue black

Result:
Positive: colorless zone surrounding colonies

Organism:
-Staphylococcus cohnii -Bacillus subtilis
MICROBIAL REACTION IN CULTERED MEDIA
-Escherichia coli
-Enterobacter aerogenes

F. Carbohydrate/Sugar Fermentation Medium I. Reactions in TSIA/KIA

-Durham Tube method K:red/alkaline slant/butt
-acid production: red to yellow A:yellow/acid
-gas production: bubbles
-pH indicator: phenol red or bromcresol purple K/A (glucose only) non-lactose fermenter
A/A (glucose, lactose, sucrose) acid slant and butt
Organism: K/K (no glucose,lactose,sucrose) degrade peptone from
-Escherichia coli presend and absence of Oxygen
K/NC (no glucose, lactose, sucrose) only utilize peptone
G. Methylene Red Voges-Proskauer test (NB + 0.5% aerobically
Glucose)
-test for production acid or acetoin Black ppt = acid condition exists
-MR pH indicator: methyl red Cracking/splitting = gas produced
-VP reagent: A alpha-naphthol
40% KOH
-with butylene-glycol or butaneidol fermentation as basis Organism:
-Escherichia coli -Pseudomonas putida
Result: -Klebsiella oxytoca
Acid production: red
Negative MR: yellow

AMC production: pink to red

Negative AMC: no change in color

Organism:
-Escherichia coli -Pseudomonas putida
-Klebsiella oxytoca

J. Gelatin Hydrolysis Test

-test for proteases
-enzyme: gelatinase
-degrade to amino acids and peptides

Result:
gelatinase produced: gelatin remains fluid
H. Citrate Utilization (SCA)
-more citrate = more ammonia
Organism:
-source of nitrogen: ammonium dihydrogen phosphate
-Bacillus subtilis
-ph indicator: bromthymol blue

Results:
Citrate as source of Carbon: blue
-turns yellow when acidic

Organism:
MICROBIAL REACTION IN CULTERED MEDIA

K. Indole Production (Tryptone Broth) M. Tributyrin Agar (lipase test)

-breakdown of tryptophan and Kovacs reagent (p- -reactant: glycerol tributyrate
dimethylamniobenzaldehyde) -lipolytic degrades glycerol tributyrate
-enzyme: lipase
Result:
Indole present: red ring at surface Result:
Lipase and breakdown of tributyrin: clear zones around
Organism: colonies
-Escherichia coli -Pseudomonas putida
-Klebsiella oxytoca -Enterobacter aerogenes

L. Lysine Decarboxylation and deamination (LIA)
-pH indicator: bromcresol purple
-decarboxylation only occurs at acidic medium
-acidified by glucose fermentation
-Lysine give rise to ammonia, reacts with ferric ammonium
citrate = red color (slant)
-Iron sulfide formation = blackening of medium
Results:
purple:alkaline, red:ammonia and ferric ammonium
citrate, yellow: acid
N. Litmus Milk
-dehydrated skim milk + litmus
Results:
1.Acid pH pink (fermentation of lactose)
2. Alkaline pH purple/blue (no fermentation of lactose)
3. Reduction white (litmus as electron acceptor)
4. Acid curd precipitation of casein by acid from
lactose
5. Rennet Curd soft curd followed by peptonization
6. Peptonization dissolution of clot (digestion of
curd/milk by proteolytic enzymes)
7. Gas production bubbles

1. Negative purple/yellow
2. Lysine Deamination red/yellow
3. Lysine Decarboxylation purple/purple
4. H2S blackening

Organisms:
-Escherichia coli -Pseudomonas putida
-Klebsiella oxytoca -Seratia marcesens
MICROBIAL REACTION IN CULTERED MEDIA
Organism: -oxidative use of carbohydrates by gram-negative bacilli
-Escherichia coli -Pseudomonas putida -pH indicator: bromthymol blue
-Klebsiella oxytoca -Seratia marcesens
Result:
Positive: yellow color (acidic environment)
Negative: green or blue-green

Organism:
-Pseudomonas putida

O. Nitrate Reduction Test (Nitrate Broth)

-reagent A: sulfanillic acid
-reagent B: naphthylamine
-test for presence of unreduced nitrate: zinc powder

Result:
Negative: red color
Positive: no red color appears
=reduced to N2 or NH3

Organism:
-Escherichia coli -Pseudomonas putida
-Klebsiella oxytoca -Seratia marcesens Q. Urease Test (Stuarts Urea Broth)
-Nitrogen metabolism

Results:
Urease production: red

Organisms:
-Escherichia coli -Pseudomonas putida
-Klebsiella oxytoca -Seratia marcesens

P. Oxidative-Fermentative Test
-carbohydrate 1% and peptone 0.02%