1.1.2 Culture media commonly used in Mirobiology Lab Flashcards
A general purpose, nutrient medium used for the cultivation of microbes supporting growth of a wide range of non-fastidious organisms
Nutrient Agar/Broth
What are the ingredients of Nutrient agar?
15g/L Pancreatic digest of casein
5 g/L Papaic digest of soyabean meal
5 g/L Sodium chloride
15 g/L Agar
Final pH at 25 C (7.4 +- 0.2)
What is the purpose of pancreatic digest of casein and papaic digest of soyabean meal?
Provides amino acid and long chains peptides for the growth of microorganisms
What is the purpose of sodium chloride
Maintains the osmotic balance
What is the positive control for nutrient agar?
E. coli
Pseudomonas aeruginosa
(Expected growth at 35)
A general purpose, nonselective growth medium that supports the growth of most non-fastidious gram-negative and gram positive bacteria as well as many yeast and molds
Can be use for longer storage, Maintenance purposes, Cultivation and transport
Can be use as base if you want to add blood or antibiotics
Trypticase soy agar/Broth
Tryptone soy agar
Tryptic soy agar
Soybean casein digest agar medium
What are the ingredients of TSA
300 g/L Beef infusion
17.5 g/L Casein acid hydrolysate
1.5 g/L Starch
17 g/L Agar
Final pH 7.3 +- 0.2
What is the purpose of Beef infusion and Casein acid
Main provider of nitrogenous compounds, sulfur, carbon and etc.
One of the ingredients that provides a Protective colloid against toxic substance present in the medium
Starch
What happen if there is hydrolysis on starch?
Yields a dextrose
What is the purpose of dextrose?
Serves as the source of energy
Observation for the growth of FASTIDIOUS organisms and study of hemolytic reaction is done after addition of defibrinated sheep blood
BAP
What is the procedure in creating a BAP?
- Provide blood agar base (TSA)
- Autoclave
- Cool down in 45 - 50 C
- Add defibrinated sheep blood (5%)
How to prepare defibrinated sheep blood?
100ml of TSA molten agar = 5 mL blood
Blood agar base is has similar ingredient to what agar?
TSA
Beef infusion
Casein acid hydrolysate
starch
agar
3 type of hemolytic pattern
Beta hemolysis (Complete)
Alpha hemolysis (Partial)
Gamma hemolysis (None)
What is the positive control for BAP?
Streptococcus pneumoniae (Alpha)
Streptococcus pyogenes (Beta)
Staphylococcus epidermidis (Gamma)
FASTWhat is the positive control for BAP?
Streptococcus pneumoniae (Alpha)
Streptococcus pyogenes (Beta)
Staphylococcus epidermidis (Gamma)
Observation for the growth of FASTIDIOUS organisms is done after addition of defibrinated sheep blood and heated at 80-100 C.
Chocolate agar plate
Procedure of CAP is similar to?
BAP but heated at the end (80-100 C)
What is the rationale in heating at 80 - 100 C?
To release growth factors NAD (Factor V) and Hemin (Factor X) for nutrients
What bacteria uses CAP?
Neisseria and Haemophilus
What is the Positive Controls for CAP?
Neisseria gonorrhoeae (5-10% CO2)
Haemophilus influenzae (5 -10% CO2)
In candle jar (Carbon dioxide enrichment environment)
Used as selective media for isolating gram-negative bacteria and differential media for the isolation and differentiation of lactose fermenting and non-lactose fermenting enteric bacilli
MacConkey agar
2 function of MacConkey agar
Differential (Differentiate Lactose to non Lactose) and Selective (Isolating Gram negative)
What are the ingredients of MacConkey agar?
3 g/L Peptones (Meat and casein)
17 g/L Pancreatic digest of gelatin
10 g/L Lactose monohydrate
1.5 g/L Bile salts
5.0 g/L Sodium chloride
0.001 g/L Crystal Violet
0.03 g/L Neutral red
13.5 g/L Agar
pH 25 = 7.1 +- 0.2
What is the purpose of Pancreatic digest of gelatin and peptones?
Provides the nutrient such as nitrogenous factors and vitamins
What is the purpose of Lactose monohydrate
Fermentable source of carbohydrate for the bacteria
What is the purpose of Bile salts and crystal violet
Use to inhibit most gram positive bacteria
What is the purpose of Sodium chloride
Maintains the osmotic balance
What is the purpose of Neutral red
pH indicator
What is the positive indicator for MacConkey agar?
Pink or Red = Acid (<6.8)
What is negative indicator for MacConkey agar?
Clear colonies
What is the positive control for MacConkey?
E. coli (Pink-Red with bile precipitate)
What is the control for Non-Lactose fermenter
Pseudomonas aeruginosa (Can grow in MacConkey)
What is the negative control for MacConkey?
Gram positive cocci
Example of Bacteria grows on MacConkey agar
EEK
Isolation and differentiation of lactose fermenting and non lactose fermenting enteric bacilli
Ethylene methylene blue agar (EMB)
2 functions of EMB?
Selective and Differential
What are the ingredients for EMB?
10 g/L Peptic digest of animal tissue
2 g/L Dipotassium phosphate
5 g/L Lactose
5 g/L Sucrose
0.4 g/L Eosin -Y
0.065 g/L Methylene blue
13.5 g/L Agar
pH 7.2 +- 0.2
What is the purpose of peptic digest of animal tissue
Source of nutrients such as carbon, vitamins and minerals
What is the purpose of dipotassium phosphate
Use as a buffer
What is the purpose of Lactose and Sucrose
Source of energy by being the fermentable carbohydrates
What is the purpose of Eosin-Y and Methylene blue
Use to inhibit the growth of gram positive bacteria
What is the positive control for EMB?
E.coli (Purple colonies with green metallic sheen)
Enterobacter aerogenes (Purple growth)
What is the reason of purple colonies
Lactose fermentation
What is the negative indicator for EMB?
Colorless or pink colonies
Used for selective differentiation of staphylococci
Mannitol salt agar
Mannitol salt agar is what type of agar?
Selective and differential agar
What are the ingredients of MSA?
5.0 g/L Enzymatic Digest of Casein
5.0 g/L Enzymatic Digest of animal Tissue
1.0 g/L Beef extract
10.0 g/L D-Mannitol
75.00 g/L Sodium chloride
0.025 g/L Phenol Red
15 g/L Agar
pH 7.4 +- 0.2
What is the purpose of Enzymatic digest of Casein and animal tissue, and Beef extract?
Main provider of nutrients such as carbons, nitrogen
What is the purpose of D-mannitol
Carbohydrate source (Use to differentiate mannitol fermenters)
What is the purpose of Sodium chloride
Inhibits most bacteria except Staphylococci (Reason for being Selective)
What is the purpose of Phenol red
pH indicator
What are the positive control for MSA?
S. aureus (Yellow colonies with yellow halo - Mannitol fermenter)
S epidermidis (pink Colonies with pink medium - Non-mannitol fermenter)
What is the negative control for MSA?
E. coli due to High concentration of MSA
Used as differential and selective medium for the isolation and differentiation of Salmonella and Shigella spp. from other gram-negative enteric bacilli?
Hektoen Enteric agar
What are the ingredients for HEA?
12 Protease peptone
3 Yeast extract
12 Lactose
2 Sucrose
9 Saliclin
9 Bile salt mixture
5 Sodium chloride
5 Sodium thiosulfate
1.5 Ferric ammonium citrate
0.1 Acid - fuchsin
0.065 Bromothymol blue
14 Agar
pH 7.5 +-0.2
provide Nutrients such as carbon, nitrogen, and amino acids for growth in HEA
Protease peptone
What is the Vitamin source for HEA?
Yeast extract
What is the purpose of Lactose, sucrose, Salicin
Cab source or being fermented
Inhibits gram positive and HEA
Bile salts
acid fuchsin
What is the purpose of Sodium chloride
Maintain osmotic balance
What is the purpose of Ferric ammonium citrate
Source of iron, production of hydrogen sulfide (Blackening)
What is the purpose of Bromothymol Blue
pH indicator
What is the Positive indicator for HEA?
E. coli (Partial to complete inhibition, Yellow to salmon orange colonies)
Shigella flexneri (Green colonies)
Salmonella typhimurium (Green colonies with black centers)
What is the Negative control of HEA?
Enterococcus faecalis
Selective and differential medium for the isolation of pathogenic enteric bacilli particularly Salmonella and shigella from stool, Food and Clinical Material
Salmonella shigella agar
What are the ingredients of Salmonella Shigella agar?
5.0 Beef extract
2.5 Enzymatic Digest of casein
2.5 Enzymatic digest of animal tissue
10 Lactose
8.5 Bile salts
8.5 Sodium Citrate
8.5 Sodium Thiosulfate
1.0 Ferric Citrate
0.00033 Brilliant Green
0.025 Neutral red
13.5 Agar
pH 7.0 +- 0.2
What is the source of nutrients for SSA?
Beef extract
Enzymatic digest of Casein
Enzymatic digest of animal tissue
What is the carbohydrate source of SSA?
Lactose
Inhibits gram positive and coliform bacteria, and Swarming motility
Bile salts
Sodium citrate
Brilliant green
Ingredients for H2S producers of SSA
Sodium Citrate
Sodium Thiosulfate
pH indicator of SSA
Neutral red
Unique to SSA from other media?
Does not autoclave
What is the Positive control for SSA?
Salmonella typhimurium (Colorless colonies with Black centers)
Shigella flexneri (Colorless colonies)
What is the negative control for SSA
Enterococcus faecalis
Selective differential medium for isolating and cultivating Vibrio cholerae and other Vibrio species from clinical specimens and other materials
TCBS agar
Ingredients of TCBS agar
5 Yeast extract
5 Enzymatic digest of casein
5 Enzymatic digest of animal tissue
10 Sodium Citrate
10 Sodium Thiosulfate
5 Oxbile
3 Sodium cholate
20 Sucrose
10 Sodium Chloride
1 Ferric Citrate
0.04 Bromothymol blue
0.04 Thymol blue
14 Agar
pH 8.6 +- 0.2
TCBS requires autoclave?
No
Nutrient provider for TCBS agar
5 Yeast extract
5 Enzymatic digest of casein
5 Enzymatic digest of animal tissue
Selective agents that creates alkaline pH, suppresses coliform (E.coli)
for TCBS agar
Sodium citrate
Sodium Thiosulfate
Oxbile
Sodium Cholate
Carbohydrate in TCBS
Sucrose
Act as the indicator for H2S producers in TCBS agar
Sodium Thiosulfate
ferric Citrate
pH indicators for TCBS
Bromothymol blue
Thymol blue
What is the positive Control for TCBS?
Vibrio alginolytious (Yellow, Sucrose fermenter)
Vibrio parahaemolyticus (Green, non sucrose fermenter)
