Handbook of Microbiological Media, Fourth Edition part 105 pdf

5.0mg Preparation of Trace Elements Solution: Add components to distilled/deionized water and bring volume to 1.0L.. 0.01g Preparation of Trace Elements Solution SL-6: Add components to

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M-CP Agar Base 1035

cally add 80.0mL of sterile egg yolk emulsion, 50% Mix thoroughly

Pour into sterile Petri dishes in 20.0mL volumes

Use: For the cultivation of Clostridium botulinum.

McClung-Toabe Egg Yolk Agar, CDC Modified

(CDC Modified McClung-Toabe Egg Yolk Agar)

Compositionper liter:

Pancreatic digest of casein 40.0g

Agar 25.0g

NaHPO4 5.0g

Yeast extract 5.0g

D-Glucose 2.0g

NaCl 2.0g

Egg yolk emulsion 100.0mL

MgSO4 (5% solution) 0.2mL

pH 7.4 ± 0.2 at 25°C

Egg Yolk Emulsion:

Composition:

Chicken egg yolks 11

Whole chicken egg 1

Preparation of Egg Yolk Emulsion: Soak eggs with 1:100

dilu-tion of saturated mercuric chloride soludilu-tion for 1 min Crack eggs

Sep-arate yolks from whites for 11 eggs Mix egg yolks with 1 chicken egg

Preparation of Medium: Add components, except egg yolk

emul-sion, to distilled/deionized water and bring volume to 900.0mL Mix

thoroughly Gently heat while stirring and bring to boiling Autoclave

for 15 min at 15 psi pressure–121°C Cool to 60°C Aseptically add

100.0mL of sterile egg yolk emulsion Mix thoroughly Pour into

ster-ile Petri dishes in 20.0mL volumes

Use: For the isolation, cultivation, and differentiation of anaerobic

bacteria from foods Bacteria that produce lecithinase appear as

colo-nies surrounded by an insoluble opaque precipitate Bacteria that

pro-duce lipase activity appear as colonies with a sheen or “pearly” surface

Bacteria that possess proteolytic activity appear as colonies surrounded

by a clear zone

McClung-Toabe Egg Yolk Agar, CDC Modified

(CDC Modified McClung-Toabe Egg Yolk Agar)

Agar 25.0g

Na2HPO4 5.0g

Yeast extract 5.0g

Glucose 2.0g

NaCl 2.0g

KH2PO4 1.0g

Egg yolk emulsion, 50% 100.0mL

MgSO4·7H2O (5% solution) 0.2mL

pH 7.3 ± 0.2 at 25°C

Egg Yolk Emulsion, 50%:

Composition per 100.0mL:

Chicken egg yolks 11

Whole chicken egg 1

NaCl (0.9% solution) 50.0mL

Preparation of Egg Yolk Emulsion, 50%: Soak eggs with 1:100

dilution of saturated mercuric chloride solution for 1 min Crack eggs

and separate yolks from whites Mix egg yolks with 1 chicken egg

Beat to form emulsion Measure 50.0mL of egg yolk emulsion and add

to 50.0mL of 0.9% NaCl solution Mix thoroughly Filter sterilize Warm to 45°–50°C

emulsion, 50%—to distilled/deionized water and bring volume to 900.0mL Mix thoroughly Gently heat while stirring and bring to boil-ing Autoclave for 15 min at 15 psi pressure–121°C Cool to 50°–55°C Aseptically add 100.0mL of sterile egg yolk emulsion, 50% Mix thor-oughly Pour into sterile Petri dishes in 15.0mL volumes

Use: For the cultivation of a wide variety of anaerobic bacteria For the differentiation of anaerobic bacteria based on lecithinase production, lipase production, and proteolytic ability Bacteria that produce lecithi-nase appear as colonies surrounded by a zone of insoluble precipitate Bacteria that produce lipase appear as colonies with a pearly iridescent sheen Bacteria that produce proteolytic activity appear as colonies sur-rounded by a clear zone

McClung Toabe HiVeg Agar Base wtih Egg Yolk

Plant peptone No 3 40.0g Agar 25.0g

Na2HPO4 5.0g Glucose 2.0g NaCl 2.0g

KH2PO4 1.0g MgSO4 0.1g Egg yolk emulsion, 50% 100.0mL

pH 7.3 ± 0.2 at 25°C

Source: This medium, without egg yolk emulsion, is available as a premixed powder from HiMedia

Egg Yolk Emulsion, 50%:

Chicken egg yolks 11 Whole chicken egg 1 NaCl (0.9% solution) 50.0mL

Preparation of Egg Yolk Emulsion, 50%: Soak eggs with 1:100 dilution of saturated mercuric chloride solution for 1 min Crack eggs and separate yolks from whites Mix egg yolks with 1 chicken egg Beat to form emulsion Measure 50.0mL of egg yolk emulsion and add

to 50.0mL of 0.9% NaCl solution Mix thoroughly Filter sterilize Warm to 45°–50°C

Preparation of Medium: Add components, except egg yolk emul-sion, 50%, to distilled/deionized water and bring volume to 900.0mL Mix thoroughly Gently heat while stirring and bring to boiling Auto-clave for 20 min at 15 psi pressure–121°C Cool to 50°–55°C Asepti-cally add 100.0mL of sterile egg yolk emulsion, 50% Mix thoroughly Pour into sterile Petri dishes in 15.0mL volumes

Use: For the isolation and cultivation of Clostridium perfringens in

foods

M-CP Agar Base

Composition per liter:

Tryptose 30.0g Yeast extract 20.0g Agar 15.0g Sucrose 5.0g

L-Cysteine·HCl·H2O 1.0g MgSO4·7H2O 0.1g FeCl3·6H2O 0.09g Indoxyl β-D-glucoside 0.06g

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1036 M-CP HiVeg Agar Base with Phenolphthalein Diphosphate

Bromcresol Purple 0.04g

Selective supplement solution B 20.0mL

Selective supplement solution A 10.0mL

pH 7.6 ± 0.2 at 25°C

Source: This medium is available from HiMedia

Selective Supplement Solution A:

Compositionper 10.0mL:

D-Cycloserine 400.0mg

Polymyxin B sulfate 25.0mg

Preparation of Selective Supplement Solution A: Add

compo-nents to distilled/deionized water and bring volume to 10.0mL Mix

thoroughly Filter sterilize

Selective Supplement Solution B:

Phenolphthalein diphosphate 0.1g

Preparation of Selective Supplement Solution B: Add

phenol-phthalein diphosphate to distilled/deionized water and bring volume to

20.0mL Mix thoroughly Filter sterilize

Preparation of Medium: Add components, except selective

sup-plement solutions A and B, to distilled/deionized water and bring

vol-ume to 970.0mL Mix thoroughly Autoclave for 15 min at 15 psi

pressure–121°C Cool to 50°C Aseptically add selective supplement

solutions A and B Mix thoroughly Pour into Petri dishes or aseptically

distribute into sterile tubes

Use: Recommended by the Directive of the Council of the European

Union 98/83/EC for the isolation and enumeration of Clostridium

per-fringens from water samples using the membrane filtration technique.

M-CP HiVeg Agar Base with Phenolphthalein Diphosphate

Plant hydrolysate No 1 30.0g

Agar 15.0g

L-Cysteine·HCl 1.0g

MgSO4·7H2O 0.1g

FeCl3·6H2O 0.09

Indoxyl β-D-glucoside 0.06

Bromcresol Purple 0.04

Sucrose 5.0g

Yeast extract 20.0g

Phenolphthalein diphosphate solution 10.0mL

pH 7.2 ± 0.2 at 25°C

Source: This medium, without phenolphthalein diphosphate, is

avail-able as a premixed powder from HiMedia

Phenolphthalein Diphosphate Solution:

Preparation of Phenolphthalein Diphosphate Solution: Add

phenolphthalein diphosphate to distilled/deionized water and bring

volume to 10.0mL Mix thoroughly Filter sterilize

Preparation of Medium: Add components, except

phenolphtha-lein diphosphate solution, to distilled/deionized water and bring

vol-ume to 990.0mL Mix thoroughly Gently heat while stirring until

boiling Autoclave for 15 min at 15 psi pressure–121°C Cool to

45-50°C Aseptially add 10.0mL of sterile phenolphthalein diphosphate

solution Mix thoroughly Pour into sterile Petri dishes or distribute into

sterile tubes

Use: For the cultivation and identification of Providencia stuartii.

m-CP Medium

Tryptose 30.0g Yeast extract 20.0g Agar 15.0g Sucrose 5.0g

L-Cysteine·HCl·H2O 1.0g MgO4·7H2O 0.1g Bromcresol Purple 0.04g Phenolphthalein biphosphate tetrazolium

salt solution 10.0mL Selective supplement solution 4.0mL

Indoxyl-β-D-glucoside solution 4.0mL Ferric chloride solution 1.0mL

pH 7.6 ± 0.2 at 25°C

Selective Supplement Solution:

D-Cycloserine 0.4g Polymyxin B sulfate 25.0mg

Preparation of Selective Supplement Solution: Add compo-nents to 4.0mL of distilled/deionized water Mix thoroughly Filter ster-ilize

Phenolphthalein Biphosphate Tetrazolium Salt Solution:

Compositionper 10.0mL: Phenolphthalein biphosphate tetrazolium salt 25.0mg

Preparation of Phenolphthalein Biphosphate Tetrazolium Salt Solution: Add phenolphthalein biphosphate tetrazolium salt to 10.0mL of distilled/deionized water Mix thoroughly Filter sterilize

Indoxyl-β- D -glucoside Solution:

Indoxyl-β-D-glucoside 0.45g

Preparation of Indoxyl-β- D -glucoside Solution: Add indoxyl-β-D-glucoside to 10.0mL of distilled/deionized water Mix thoroughly Filter sterilize

Ferric Chloride Solution:

Compositionper 4.0mL: FeCl3·6H2O 30.0mg

Preparation of Ferric Chloride Solution: Add FeCl3·6H2O to 4.0mL of distilled/deionized water Mix thoroughly Filter sterilize

Preparation of Medium: Add components, except selective sup-plement solution, phenolphthalein biphosphate tetrazolium salt

solu-tion, indoxyl-β-D-glucoside solution, and ferric chloride solution, to distilled/deionized water and bring volume to 981.0mL Mix

thorough-ly Autoclave for 15 min at 15 psi pressure–121°C Cool to 45°–50°C Aseptically add 4.0mL of selective supplement solution Mix thor-oughly Aseptically add 10.0mL phenolphthalein biphosphate

tetrazo-lium salt solution, 4.0mL indoxyl-β-D-glucoside solution, and 1.0mL ferric chloride solution Mix thoroughly Pour into sterile Petri dishes or aseptically distribute into tubes

Use: A selective, chromogenic medium for the rapid identification and

enumeration of Clostridium perfringens in water samples, including

water used in food and beverage production

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MD1- Medium 1037

MCP Medium (Modified MacConkey Medium)

(MacConkey Phosphatase Medium)

Pancreatic digest of gelatin 17.0g

Agar 13.5g

Lactose 10.0g

NaCl 5.0g

Bile salts 1.5g

Peptic digest of animal tissue 1.5g

Na2HPO4 0.6g

Glucose 0.2g

Methyl Blue 0.07g

Neutral Red 0.03g

Crystal Violet 1.0mg

Phenolphthalein diphosphate solution 10.0mL

pH 7.2 ± 0.2 at 25°C

Source: This medium, without phenolphthalein diphosphate, is

avail-able as a premixed powder from HiMedia

Phenolphthalein Diphosphate Solution:

Preparation of Phenolphthalein Diphosphate Solution: Add

phenolphthalein diphosphate to distilled/deionized water and bring

volume to 10.0mL Mix thoroughly Filter sterilize

Preparation of Medium: Add components, except

phenolphtha-lein diphosphate solution, to distilled/deionized water and bring

vol-ume to 990.0mL Mix thoroughly Gently heat while stirring until

boiling Autoclave for 15 min at 15 psi pressure–121°C Cool to

45-50°C Aseptially add 10.0mL of sterile phenolphthalein diphosphate

solution Mix thoroughly Pour into sterile Petri dishes or distribute into

sterile tubes

Use: For the cultivation and identification of Providencia stuartii.

MD Medium

Agar 20.0g

L-Malic acid 20.0g

D-Glucose 5.0g

Casamino acids 3.0g

Pancreatic digest of soybean meal 1.5g

Tween™ 80 1.0g

Yeast extract 1.0g

Bromcresol Green solution 20.0mL

pH 7.0 ± 0.2 at 25°C

Bromcresol Green Solution:

Bromcresol Green 0.1g

NaOH (0.01N solution) 30.0mL

Preparation of Bromcresol Green Solution: Add Bromcresol

Green to 30.0mL of NaOH solution Mix thoroughly Filter sterilize

Preparation of Medium: Add components to distilled/deionized

water and bring volume to 1.0L Mix thoroughly Gently heat and bring

to boiling Distribute into tubes or flasks Adjust pH to 7.0 with 10N

KOH Autoclave for 15 min at 15 psi pressure–121°C Pour into sterile

Petri dishes or leave in tubes

Use: For the isolation and cultivation of Salmonella species from

foods

MD 1 Medium

Pancreatic digest of casein 3.0g MgSO4·7H2O 2.0g CaCl2 0.5g Trace elements solution 1.0mL Vitamin B12 solution 1.0mL

Trace Elements Solution:

EDTA 8.0g MnCl2·4H2O 0.1g CoCl2 0.02g KBr 0.02g ZnCl2 0.02g CuSO4 0.01g

H3BO3 0.01g NaMoO4·2H2O 0.01g BaCl2 5.0mg LiCl 5.0mg SnCl2·2H2O 5.0mg

Preparation of Trace Elements Solution: Add components to distilled/deionized water and bring volume to 1.0L Mix thoroughly

Vitamin B 12 Solution:

Vitamin B12 5.0mg

Preparation of Vitamin B 12 Solution: Add vitamin B12 to dis-tilled/deionized water and bring volume to 10.0mL Mix thoroughly

Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L Mix thoroughly Distribute into tubes

or flasks Autoclave for 15 min at 15 psi pressure–121°C

Use: For the cultivation of myxobacteria

MD1- Medium (DSMZ Medium 1118)

Casitone 3.0g MgSO4·7H2O 2.0g CaCl2·2H2O 0.7g Vitamin solution 10.0mL Trace elements solution SL-4 .1.0mL

pH 7.1 ± 0.2 at 25°C

Trace Elements Solution SL-4:

EDTA 0.5g FeSO4·7H2O 0.2g Trace elements solution SL-6 100.0mL

MnCl2·4H2O 0.5g

H3BO3 0.3g CoCl2·6H2O 0.2g ZnSO4·7H2O 0.1g

Na2MoO4·2H2O 0.03g

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1038 MDPA with Calcium Carbonate

NiCl2·6H2O 0.02g

CuCl2·2H2O 0.01g

Preparation of Trace Elements Solution SL-6: Add components

to distilled/deionized water and bring volume to 1.0L Mix thoroughly

Autoclave for 15 min at 15 psi pressure–121°C

Filter sterilize

Vitamin Solution:

Compositionper 10.0ml:

Vitamin B12 0.5mg

Preparation of Vitamin Solution: Add components to distilled/

deionized water and bring volume to 10.0mL Mix thoroughly Filter

sterilize

Preparation of Medium: Add components, except vitamin

solu-tion, to distilled/deionized water and bring volume to 990.0mL Mix

thoroughly Adjust pH to 7.1 Distribute into tubes or flasks Gently

heat while stirring and bring to boiling Mix thoroughly Autoclave for

15 min at 15 psi pressure–121°C Aseptically add 10.0mL sterile

vita-min solution Mix thorougly Aseptically distribute into sterile tubes or

flasks

Use: For the cultivation of Myxococcus xanthus.

MDPA

See: Malt Dextrose Peptone Agar

MDPA with Calcium Carbonate

Agar 25.0g

Malt extract 20.0g

Glucose 20.0g

CaCO3 5.0g

Peptone 1.0g

to boiling Distribute into tubes or flasks Autoclave for 15 min at 15

psi pressure–121°C Pour into sterile Petri dishes or leave in tubes

Use: For the cultivation of Dekkera species

MDPYA4

See: Malt 4% Dextrose Peptone Yeast Agar

MDYA4

See: Malt 4% Dextrose Yeast Agar

m-E Agar

See: E Agar

Me15% MH Agar (DSMZ Medium 582)

NaCl 121.5g

Agar 20.0g

MgSO4 14.4g

MgCl2 10.5g

Yeast extract 10.0g

Proteose peptone no 3 5.0g

KCl 3.0g

Glucose 1.0g CaCl2 0.54g NaBr 0.039g NaHCO3 solution 10.0mL

pH 7.5 ± 0.2 at 25°C

NaHCO 3 Solution:

NaHCO3 0.9g

Preparation of NaHCO 3 Solution: Add NaHCO3 to distilled/de-ionized water and bring volume to 100.0mL Mix thoroughly Sparge with 80% N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure– 121°C Cool to 50°C

Preparation of Medium: Add components, except NaHCO3 solu-tion, to distilled/deionized water and bring volume to 990.0mL Mix thoroughly Autoclave for 15 min at 15 psi pressure–121°C Cool to 50°C Aseptically add 10.0mL NaHCO3 solution Mix thoroughly Pour into Petri dishes or aseptically distribute into sterile tubes

Use: For the cultivation of Bacillus halophilus.

Me15% MH Medium (DSMZ Medium 582)

NaCl 121.5g MgSO4 14.4g MgCl2 10.5g Yeast extract 10.0g Proteose peptone no 3 5.0g KCl 3.0g Glucose 1.0g CaCl2 0.54g NaBr 0.039g NaHCO3 solution 10.0mL

pH 7.5 ± 0.2 at 25°C

NaHCO 3 Solution:

NaHCO3 0.9g

Preparation of NaHCO 3 Solution: Add NaHCO3 to distilled/de-ionized water and bring volume to 100.0mL Mix thoroughly Sparge with 80% N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure– 121°C Cool to 25°C

Preparation of Medium: Add components, except NaHCO3 solu-tion, to distilled/deionized water and bring volume to 990.0mL Mix thoroughly Autoclave for 15 min at 15 psi pressure–121°C Cool to 25°C Aseptically add 10.0mL NaHCO3 solution Mix thoroughly Aseptically distribute into sterile tubes or flasks

Use: For the cultivation of Bacillus halophilus.

MEA

See: Malt Extract Agar

Meat Extract with Peptone (Pepted Meat Broth)

NaCl 15.0g Peptic digest of animal tissue 10.0g Meat extract 3.0g

pH 7.2 ± 0.2 at 25°C

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Medium A for Producing Lysates 1039

to boiling Distribute into tubes or flasks Autoclave for 15 min at 15

psi pressure–121°C Pour into sterile Petri dishes or leave in tubes

Use: For the cultivation and maintenance of Alcaligenes species.

Meat Extract with Peptone and 1.5% Salt

NaCl 15.0g

Peptone 10.0g

Meat extract 3.0g

water and bring volume to 1.0L Mix thoroughly Distribute into tubes

Use: For the cultivation and maintenance of Alcaligenes species.

Meat Infusion Agar, HiVeg

(Standard Infusion Agar, HiVeg)

Agar 25.0g

Plant infusion 10.0g

Plant peptone 10.0g

NaCl 5.0g

pH 7.5 ± 0.2 at 25°C

Source: This medium is available as a premixed powder from

Hi-Media

Use: For the mass cultivation of organisms for vaccine or toxin

pro-duction

M-EC Test Agar

Agar 15.0g

Lactose 10.0g

NaCl 7.5g

Proteose peptone 5.0g

Dipotassium phosphate 3.3g

Yeast extract 3.0g

KH2PO4 1.0g

Sodium lauryl sulphate 0.2g

Sodium deoxycholate 0.1g

Bromcresol Purple 0.08g

Bromphenol Red 0.08g

pH 7.3 ± 0.2 at 25°C

to boiling Distribute into tubes or flasks Do not autoclave Pour into

sterile Petri dishes or leave in tubes

Use: For the detection of Escherichia coli in water samples using the

membrane filter technique

MED IIa

Tris buffer stock solution 10.0mL CaCl2 (5.0% solution) 10.0mL MgSO4·7H2O (3.33% solution) 1.0mL

pH 7.2 ± 0.2 at 25°C

Tris Buffer Stock Solution:

Tris(hydroxymethyl)aminomethane·HCl 35.01g Tris(hydroxymethyl)aminomethane 3.35g

Preparation of Tris Buffer Stock Solution: Add components to distilled/deionized water and bring volume to 500.0mL Mix

thorough-ly Adjust pH to 7.2

Use: For the cultivation and maintenance of Vampirovibrio chlorellavo-rus.

Medium A

D-Glucose 20.0g Agar 20.0g Yeast extract 10.0g Biotin 1.0mg Calcium pantothenate 1.0mg

pH 7.3 ± 0.2 at 25°C

Preparation of Medium: Add components, except biotin and cal-cium pantothenate, to distilled/deionized water and bring volume to 990.0mL Mix thoroughly Gently heat and bring to boiling Autoclave for 15 min at 15 psi pressure–121°C Cool to 45°–50°C Add biotin and calcium pantothenate to distilled/deionized water and bring volume to 10.0mL Mix thoroughly Filter sterilize Aseptically add the sterile bi-otin and calcium pantothenate solution to the cooled sterile basal me-dium Mix thoroughly Pour into sterile Petri dishes or distribute into sterile tubes

Use: For the cultivation and maintenance of Zymomonas mobilis.

Medium A for Producing Lysates

Nutrient broth 8.0g KCl 1.0g MgSO4·7H2O 0.25g MnCl2 1.25mg FeSO4 (1.0mM solution) 1.0mL

Ca(NO3)2 (1.0M solution) 1.0mL

pH 7.0–7.2 at 25°C

Preparation of Medium: Add components, except FeSO4 and Ca(NO3)2, to distilled/deionized water and bring volume to 998.0mL Mix thoroughly Adjust pH to 7.0–7.2 Autoclave for 30 min at 15 psi

pressure–115°C Cool to 45°–50°C Prepare 1.0mM FeSO4 solution

and 1.0M Ca(NO3)2 solution separately Filter sterilize both solutions Aseptically add the sterile FeSO4 solution and sterile Ca(NO3)2 solu-tion to the cooled sterile basal medium Mix thoroughly Distribute into sterile tubes or flasks

Use: For the cultivation of microorganisms to be lysed

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1040 Medium 2A

Medium 2A

Arginine 10.0g

NaCl 5.0g

Agar 4.0g

Peptone 1.0g

K2HPO4·3H2O 0.3g

Phenol Red 0.01g

pH 7.2–7.4 at 25°C

to boiling Distribute into tubes Autoclave for 15 min at 15 psi

pres-sure–121°C

Use: For the cultivation and differentiation of Pseudomonas species

based on their production of arginine dihydrolase activity

Medium AS4

Sucrose 80.0g

PPLO broth without Crystal Violet 500.0mL

Horse serum 200.0mL

Phenol Red (0.5% solution) 5.0mL

pH 7.2 ± 0.2 at 25°C

PPLO Broth without Crystal Violet:

Beef heart, solids from infusion 11.53g

Peptone 2.33g

NaCl 1.15g

Source: PPLO broth without Crystal Violet is available as a premixed

powder from BD Diagnostic Systems

components to distilled/deionized water and bring volume to 500.0mL

Mix thoroughly Beef heart for infusion may be substituted; 100.0g of

beef heart for infusion is equivalent to 500.0g of fresh heart tissue

Preparation of Medium: Add components, except horse serum, to

distilled/deionized water and bring volume to 800.0mL Mix

thorough-ly Adjust pH to 7.2 Autoclave for 10 min at 15 psi pressure–121°C

Cool to 45°–50°C Aseptically add 200.0mL of noninactivated, sterile

horse serum Mix thoroughly Aseptically distribute into sterile tubes

or flasks

Use: For the cultivation and maintenance of Spiroplasma melliferum.

Medium for Acetivibrio cellulolyticus

See: BC Medium

Medium for Aciduric, Thermophilic Bacillus Strains

Solution A 500.0mL

Solution B 500.0mL

pH 4.3 ± 0.2 at 25°C

Solution A:

KH2PO4 3.0g

Glucose 1.0g

Starch 1.0g

Tryptone 1.0g

Yeast extract 1.0g

MgSO4·7H2O 0.5g

CaCl2·2H2O 0.25g (NH4)2SO4 0.2g

Preparation of Solution A: Add components to distilled/deionized water and bring volume to 500.0mL Mix thoroughly Adjust pH to 4.3 Autoclave for 15 min at 15 psi pressure–121°C Cool to 50°–55°C

Solution B:

Agar 20.0g

Preparation of Solution B: Add agar to distilled/deionized water and bring volume to 500.0mL Mix thoroughly Autoclave for 15 min

at 15 psi pressure–121°C Cool to 50°–55°C

Preparation of Medium: Aseptically combine 500.0mL of solu-tion A and 500.0mL of solusolu-tion B Mix thoroughly Pour into sterile Petri dishes or distribute into sterile tubes

Use: For the cultivation of aciduric, thermophilic Bacillus strains.

Medium 2508-85-1 with Amino Acids

Agar 20.0g Nutrient broth 8.0g D-Glucose 5.0g Polypeptone™ 5.0g Yeast extract 5.0g L-Lysine 0.1g L-Methionine 0.05g Diaminopimelic acid 0.05g

Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L Mix thoroughly Gently heat and bring

to boiling Distribute into tubes or flasks Autoclave for 30 min at 15 psi pressure–121°C Pour into sterile Petri dishes or leave in tubes

Use: For the cultivation and maintenance of Escherichia coli.

Medium for Ammonia Oxidizers

MgSO4·7H2O 0.2g (NH4)2SO4 0.13g

K2HPO4 0.09g CaCl2·2H2O 0.02g Chelated iron 1.0mg MnCl2·4H2O 0.2mg

Na2MoO4·2H2O 0.1mg ZnSO4·7H2O 0.1mg CuSO4·5H2O 0.02mg CoCl2·6H2O 2.0μg

Use: For the isolation, cultivation, and enrichment of ammonia-oxi-dizing bacteria from soil

(NH4)2SO4 2.0g MgSO4·7H2O 0.2g CaCl2·2H2O 0.02g

K2HPO4 0.02g Chelated iron 1.0mg

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Medium for Ammonia-Oxidizing Bacteria 1041

MnCl2·4H2O 0.2mg

Na2MoO4·2H2O 0.1mg

ZnSO4·7H2O 0.1mg

CuSO4·5H2O 0.02mg

CoCl2·6H2O 2.0μg

Use: For the isolation, cultivation, and enrichment of

ammonia-oxi-dizing bacteria from soil

K2HPO4 0.5g

(NH4)2SO4 0.5g

Phenol Red 0.5g

MgSO4·7H2O 0.05g

CaCl2·2H2O 0.02g

NaCl 0.02g

Na2MoO4·2H2O 2.4μg

Metals “44” 1.0mL

Metals “44”:

ZnSO4·7H2O 1.1g

FeSO4·7H2O 0.5g

EDTA 0.25g

MnSO4·7H2O 0.154g

CuSO4·5H2O 0.04g

Co(NO3)2·6H2O 0.025g

Na2B4O7·10H2O 0.018g

Preparation of Metals “44”: Add a few drops of H2SO4 to

dis-tilled/deionized water to inhibit precipitate formation Add

compo-nents to acidified distilled/deionized water and bring volume to

100.0mL Mix thoroughly

(NH4)2SO4 0.5g

KH2PO4 0.2g

CaCl2·2H2O 0.04g

MgSO4·7H2O 0.04g

Ferric citrate 0.5mg

Phenol Red 0.5mg

Medium for Ammonia Oxidizers, Brackish

CaCO3 5.0g

NH4Cl 0.5g

K2HPO4 0.05g Seawater 400.0mL

Use: For the isolation, cultivation, and enrichment of ammonia-oxidiz-ing bacteria from brackish specimens

Medium for Ammonia Oxidizers, Marine

(NH4)2SO4 1.32g MgSO4·7H2O 0.2g Chelated iron 0.13g

K2HPO4 0.11g CaCl2·2H2O 0.02g ZnSO4·7H2O 0.1mg CuSO4·5H2O 0.02mg CoCl2·6H2O 2.0μg MnCl2·4H2O 2.0μg

Na2MoO4·2H2O 1.0μg Seawater 1.0L

Preparation of Medium: Combine components Mix thoroughly Distribute into tubes or flasks Autoclave for 15 min at 15 psi pressure– 121°C

Use: For the isolation, cultivation, and enrichment of marine ammo-nia-oxidizing bacteria

Medium for Ammonia-Oxidizing Bacteria

(NH4)2SO4 235.0mg

KH2PO4 200.0mg CaCl2·2H2O 40.0mg MgSO4·7H2O 40.0mg Iron-EDTA-Phenol Red solution 1.0mL

Na2CO3 solution variable

Na2CO3Solution:

Na2CO3 5.0g

Preparation of Na 2 CO 3 Solution: Add Na2CO3 to distilled/de-ionized water and bring volume to 100.0mL Mix thoroughly Auto-clave for 15 min at 15 psi pressure–121°C

Iron-EDTA-Phenol Red Solution:

FeSO4·7H2O 50.0mg Sodium EDTA 50.0mg Phenol Red 50.0mg

Preparation of Iron-EDTA-Phenol Red Solution: Add compo-nents to distilled/deionized water and bring volume to 100.0mL Mix thoroughly

Preparation of Medium: Add components, except Na2CO3 solu-tion, to distilled/deionized water and bring volume to 1.0L Mix thor-oughly Distribute into tubes or flasks Autoclave for 15 min at 15 psi pressure–121°C Add enough sterile Na2CO3 solution to turn the

medi-um pale pink During incubation and growth of bacteria, add additional sterile Na2CO3 solution to restore the pale pink color Growth is com-plete when no further color change is observed

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1042 Medium B for Sulfate Reducers

Use: For the cultivation of Nitrosolobus multiformis and

Nitrosomo-nas europaea.

Medium B for Sulfate Reducers

(Postgate’s Medium B for Sulfate Reducers)

Sodium lactate 3.5g

MgSO4·7H2O 2.0g

NH4Cl 1.0g

CaSO4 1.0g

Yeast extract 1.0g

KH2PO4 0.5g

FeSO4·7H2O 0.5g

Ascorbic acid 0.1g

Thioglycollic acid 0.1g

pH 7.0–7.5 at 25°C

Preparation of Medium: Add components, except ascorbic acid

and thioglycollic acid, to tap water and bring volume to 1.0L For

ma-rine bacteria, NaCl may be added or seawater used in place of tap

wa-ter Mix thoroughly Adjust pH to 7.0–7.5 Thioglycolate and ascorbate

should be added immediately prior to sterilization Distribute into

tubes or flasks Autoclave for 15 min at 15 psi pressure–121°C

Use: For the isolation, cultivation, and maintenance of Desulfovibrio

species and Desulfotomaculum species This medium turns black as a

result of H2S production due to bacterial growth

Medium for Bacillus schlegelii

Agar 15.0g

Na2HPO4·2H2O 2.9g

KH2PO4 2.3g

NH4Cl 1.0g

MgSO4·7H2O 0.5g

NaHCO3 0.5g

CaCl2·2H2O 0.01g

MnSO4·H2O 10.0mg

Ferric ammonium citrate solution 20.0mL

Trace elements solution SL-6 5.0mL

pH 6.8 ± 0.2 at 25°C

Ferric Ammonium Citrate Solution:

Ferric ammonium citrate 0.05g

Preparation of Ferric Ammonium Citrate Solution: Add

fer-ric ammonium citrate to distilled/deionized water and bring volume to

20.0mL Mix thoroughly Autoclave for 15 min at 15 psi pressure–

121°C

MnCl2·4H2O 0.5g

H3BO3 0.3g

CoCl2·6H2O 0.2g

ZnSO4·7H2O 0.1g

Na2MoO4·2H2O 0.03g

NiCl2·6H2O 0.02g

CuCl2·2H2O 0.01g

Preparation of Trace Elements Solution SL-6 : Add components

Preparation of Medium: Add components, except ferric

ammoni-um citrate solution, to distilled/deionized water and bring volammoni-ume to 980.0mL Mix thoroughly Gently heat and bring to boiling Autoclave for 15 min at 15 psi pressure–121°C Aseptically add 20.0mL of sterile ferric ammonium citrate solution Mix thoroughly Pour into sterile Pe-tri dishes or disPe-tribute into sterile tubes

Use: For the chemolithotrophic growth of Bacillus schlegelii.

Medium for Bacillus stearothermophilus

NH4Cl 1.0g

K2HPO4 0.5g Yeast extract 0.2g Casamino acids 0.1g MgSO4·7H2O 0.02g Phenol solution 100.0mL Trace elements solution SL-4 1.0mL

pH 7.4 ± 0.2 at 25°C

Phenol Solution:

Phenol 0.47g

Preparation of Phenol Solution: Add phenol to distilled/deion-ized water and bring volume to 100.0mL Mix thoroughly Filter ster-ilize

EDTA 0.5g FeSO4·7H2O 0.2g Trace elements solution SL-6 100.0mL

MnCl2·4H2O 0.5g

H3BO3 0.3g CoCl2·6H2O 0.2g ZnSO4·7H2O 0.1g

Na2MoO4·2H2O 0.03g NiCl2·6H2O 0.02g CuCl2·2H2O 0.01g

to distilled/deionized water and bring volume to 1.0L Mix thoroughly Autoclave for 15 min at 15 psi pressure–121°C

to distilled/deionized water and bring volume to 1.0L Mix thoroughly Filter sterilize

Preparation of Medium: Add components, except phenol solution and trace elements solution SL-4, to distilled/deionized water and bring volume to 900.0mL Mix thoroughly Autoclave for 15 min at 15 psi pressure–121°C Aseptically add 100.0mL of sterile phenol solu-tion and 1.0mL of sterile trace elements solusolu-tion SL-4 Mix thoroughly Aseptically distribute into sterile tubes or flasks

Use: For the cultivation of Bacillus stearothermophilus.

Medium BG11 for Cyanobacteria

See: BG11 Agar and BG11 Medium

Medium BG11 for Marine Cyanobacteria

See: BG11 Marine Agar and BG11 Marine Broth

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Medium for Carbon Monoxide Oxidizers 1043

Medium with Biphenyl (DSMZ Medium 457d)

Na2HPO4 2.44g

KH2PO4 1.52g

(NH4)2SO4 0.5g

MgSO4·7H2O 0.2g

CaCl2·2H2O 0.05g

Biphenyl solution 25.0mL

pH 6.9 ± 0.2 at 25°C

EDTA 0.5g

FeSO4·7H2O 0.2g

H3BO3 0.3g

CoCl2·6H2O 0.2g

ZnSO4·7H2O 0.1g

MnCl2·4H2O 0.03g

Na2MoO4·H2O 0.03g

NiCl2·6H2O 0.02g

CuCl2··2H2O 0.01g

Adjust pH to 3.4

Biphenyl Solution:

Biphenyl 10.0g

Preparation of Biphenyl Solution: Add biphenyl to 1.0L ethanol

Mix thoroughly Filter sterilize using a cellulose filter membrane

Preparation of Medium: Add components, except biphenyl

solu-tion, to 1.0L distilled/deionized water Adjust pH to 6.9 Autoclave for

15 min at 15 psi pressure–121°C Cool to room temperature Add an

al-iquot of the biphenyl solution to a sterile flask so that the final

concen-tration will be 0.25g/L biphenyl, and let the ethanol evaporate

Aseptically add sterile medium to the crystal-layered flask

Use: For the cultivation of biphenyl- utilizing bacteria

Medium C for Sulfate Reducers

(Postgate’s Medium C for Sulfate Reducers)

Sodium lactate 6.0g

Na2SO4 4.5g

NH4Cl 1.0g

Yeast extract 1.0g

KH2PO4 0.5g

Sodium citrate·2H2O 0.3g

CaCl2·6H2O 0.06g

MgSO4·7H2O 0.06g

FeSO4·7H2O 0.004g

pH 7.5 ± 0.2 at 25°C

water and bring volume to 1.0L For marine bacteria, NaCl may be

added or sea water used in place of distilled/deionized water Mix thor-oughly Adjust pH to 7.5 Distribute into tubes or flasks Autoclave for

15 min at 15 psi pressure–121°C

Use: For detection, culturing, and storage of Desulfovibrio species and many Desulfotomaculum species This medium should be used when a

clear culture medium is desired such as for chemostat culture This medium may be cloudy after sterilization but usually clears on cooling

It turns black as a result of H2S production due to bacterial growth

Medium for Campylobacter DSM 806

(DSMZ Medium 121)

Na-aspartate 10.0g MgSO4·7H2O 1.0g Yeast extract 0.2g CaCl2·2H2O 28.0mg Resazurin 1.0mg Cysteine phosphate solution 100.0mL Trace elements solution SL-10 1.0mL

pH 7.0 ± 0.2 at 25°C

FeCl2·4H2O 1.5g CoCl2·6H2O 190.0mg MnCl2·4H2O 100.0mg ZnCl2 70.0mg

Na2MoO4·2H2O 36.0mg NiCl2·6H2O 24.0mg

H3BO3 6.0mg CuCl2·2H2O 2.0mg HCl (25% solution) 10.0mL

Preparation of Trace Elements Solution SL-10: Add FeCl2·4H2O

to 10.0mL of HCl solution Mix thoroughly Add distilled/deionized water and bring volume to 1.0L Add remaining components Mix thor-oughly Gas under 80% N2 + 20% CO2

Cysteine Phosphate Solution:

K2HPO4 0.75g NaH2PO4 0.25g Cysteine-HCl·H2O 0.25g

Preparation of Cysteine Phosphate Solution : Add components

to 100.0mL distilled/deionized water Mix thoroughly Gas under 80%

N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure–121°C

Preparation of Medium: Add components, except cysteine phos-phate solution, to distilled/deionized water and bring volume to 900.0mL Mix thoroughly Adjust pH to 7.0 Gently heat and bring to boiling Autoclave for 15 min at 15 psi pressure–121°C Cool to 25°C Aseptically add 100.0mL of sterile cysteine phosphate solution Mix thoroughly Aseptically distribute into sterile tubes or flasks

Use: For the cultivation of Sulfurospirillum sp.

Medium for Carbon Monoxide Oxidizers

Agar 12.0g

Na2HPO4·12H2O 4.5g

NH4Cl 1.5g

KH2PO4 0.75g MgSO4·7H2O 0.2g

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1044 Medium for Carbon Monoxide Oxidizers

CaCl2·2H2O 0.03g

pH 7.0 ± 0.2 at 25°C

MnCl2·4H2O 0.5g

H3BO3 0.3g

CoCl2·6H2O 0.2g

ZnSO4·7H2O 0.1g

Na2MoO4·2H2O 0.03g

NiCl2·6H2O 0.02g

CuCl2·2H2O 0.01g

to boiling Autoclave for 15 min at 15 psi pressure–121°C Pour into

sterile Petri dishes or distribute into sterile tubes After inoculation,

in-cubate in an atmosphere of 80% CO +10% O2 + 10% N2

Use: For the chemoautotrophic cultivation and maintenance of

Alcal-igenes species, Pseudomonas carboxydohydrogena, and other

Pseudo-monas species.

Medium for Carbon Monoxide Oxidizers

Agar 12.0g

Na2HPO4·12H2O 4.5g

Sodium acetate 3.0g

NH4Cl 1.5g

KH2PO4 0.75g

MgSO4·7H2O 0.2g

CaCl2·2H2O 0.03g

pH 7.0 ± 0.2 at 25°C

MnCl2·4H2O 0.5g

H3BO3 0.3g

CoCl2·6H2O 0.2g

ZnSO4·7H2O 0.1g

Na2MoO4·2H2O 0.03g

NiCl2·6H2O 0.02g

CuCl2·2H2O 0.01g

to boiling Autoclave for 15 min at 15 psi pressure–121°C Pour into

sterile Petri dishes or distribute into sterile tubes After inoculation

in-cubate in air

Use: For the chemoorganotrophic cultivation and maintenance of

Alcaligenes species, Pseudomonas carboxydohydrogena, and other

Pseudomonas species.

Medium for Chlorate Respirers (DSMZ Medium 908)

Solution A 1.0L Solution B 10.0mL Solution C 10.0mL Vitamin solution 5.0mL Trace elements solution SL-10 1.0mL

pH 7.2 ± 0.2 at 25°C

Solution A:

NaHCO3 2.5g Na-acetate 1.36g NaClO3 1.0g NaH2PO4 0.6g

NH4Cl 0.25g KCl 0.1g

Preparation of Solution A: Add components to distilled/deionized water and bring volume to 1.0L Mix thoroughly Sparge with 80% N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure–121°C Cool to room temperature

Solution B:

MgSO4 30.0mg CaCl2·2H2O 10.0mg

Preparation of Solution B: Add components to distilled/deionized water and bring volume to 10.0mL Mix thoroughly Sparge with 80%

N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure–121°C Cool

to room temperature

Solution C:

Na2MoO4 25.0mg

Na2WO4·2H2O 25.0mg

Preparation of Solution C: Add components to distilled/deionized water and bring volume to 10.0mL Mix thoroughly Sparge with 80%

N2 + 20% CO2 Autoclave for 15 min at 15 psi pressure–121°C Cool

to room temperature

Vitamin Solution:

Vitamin B12 50.0mg Pantothenic acid 50.0mg Riboflavin 50.0mg Alpha-lipoic acid 50.0mg

p-Aminobenzoic acid 50.0mg

Thiamine-HCl·2H2O 50.0mg Nicotinic acid 25.0mg Nicotinamide 25.0mg Biotin 20.0mg Folic acid 20.0mg Pyridoxamine-HCl 10.0mg

Preparation of Vitamin Solution: Add components to distilled/ deionized water and bring volume to 1.0L Mix thoroughly Filter ster-ilize

FeCl2·4H2O 1.5g

H3BO3 300.0mg CoCl2·6H2O 190.0mg MnCl2·4H2O 100.0mg

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