Mineral Salts Peptonized Milk Agar 1185 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 and maintenance of Rhodococcus rhodo- chrous. Mineral Salts Medium with Methanol Composition per liter: NaNH 4 HPO 4 ·4H 2 O 1.74g NaH 2 PO 4 ·H 2 O 0.54g MgSO 4 ·7H 2 O 0.2g KCl 0.04g FeSO 4 ·7H 2 O 5.0mg Methanol 5.0mL Trace minerals solution 1.0mL pH 7.2 ± 0.2 at 25°C Trace Minerals Solution: Composition per liter: H 3 BO 3 2.86g MnCl 2 ·4H 2 O 1.81g ZnSO 4 ·7H 2 O 0.22g CuSO 4 ·5H 2 O 0.08g CoCl 2 ·6H 2 O 0.06g Na 2 MoO 4 ·2H 2 O 25.0mg Preparation of Trace Minerals Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components, except methanol, 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. Cool to 50°C. Filter sterilize methanol. Aseptically add sterile methanol to cooled, sterile basal medium. Use: For the cultivation and maintenance of Rhodococcus rhodo- chrous. Mineral Salts Medium with Methanol and Yeast Extract Composition per liter: NaNH 4 HPO 4 ·4H 2 O 1.74g NaH 2 PO 4 ·H 2 O 0.54g MgSO 4 ·7H 2 O 0.2g Yeast extract 0.2g KCl 0.04g FeSO 4 ·7H 2 O 5.0mg Methanol 5.0mL Trace minerals solution 1.0mL pH 7.2 ± 0.2 at 25°C Trace Minerals Solution: Composition per liter: H 3 BO 3 2.86g MnCl 2 ·4H 2 O 1.81g ZnSO 4 ·7H 2 O 0.22g CuSO 4 ·5H 2 O 0.08g CoCl 2 ·6H 2 O 0.06g Na 2 MoO 4 ·2H 2 O 25.0mg Preparation of Trace Minerals Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components, except methanol, to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 50°C. Filter sterilize methanol. Aseptically add sterile methanol to cooled, sterile basal medium. Aseptically distribute into sterile tubes or flasks. Use: For the cultivation and maintenance of Pseudomonas species. Mineral Salts Medium for Thermophiles Composition per liter: NaNO 3 0.25g NH 4 Cl 0.25g Na 2 HPO 4 210.0mg MgSO 4 ·7H 2 O 200.0mg NaH 2 PO 4 90.0mg KCl 40.0mg CaCl 2 15.0mg FeSO 4 1.0mg Trace minerals solution 10.0mL n-Heptadecane 1.0mL Trace Minerals Solution: Composition per liter: ZnSO 4 ·7H 2 O 7.0mg H 3 BO 4 1.0mg MoO 3 1.0mg CuSO 4 ·5H 2 O 500.0μg CoSO 4 ·7H 2 O 18.0μg MnSO 4 ·5H 2 O 7.0μg Preparation of Trace Minerals Solution: Add components to distilled/deionized water and bring volume to 1.0L. 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 Bacillus thermoleovorans. Mineral Salts Peptonized Milk Agar (SPMA) Composition per liter: Agar 15.0g Milk, peptonized 1.0g Mineral solution 100.0mL Mineral Solution: Composition per 100.0mL: MgSO 4 ·7H 2 O 0.5g CaCl 2 0.25g K 2 HPO 4 0.25g (NH 4 ) 2 SO 4 0.1g FeCl 3 ·6H 2 O 0.01g MnCl 2 0.1mg Preparation of Mineral Solution: Add components to distilled/ deionized water and bring volume to 100.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except mineral solu- tion, to distilled/deionized water and bring volume to 900.0mL. Mix thoroughly. Gently heat and bring to boiling. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 45°–50°C. Aseptically add 100.0mL of sterile mineral solution. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. © 2010 by Taylor and Francis Group, LLC 1186 Mineral Salts for Thermophiles Use: For the cultivation of freshwater Myxobacterium species. Mineral Salts for Thermophiles (L Salts for Thermophiles) Composition per liter: NaNO 3 0.25g NH 4 Cl 0.25g Na 2 HPO 4 0.21g MgSO 4 ·7H 2 O 0.2g NaH 2 PO 4 0.09g KCl 0.04g CaCl 2 0.02g FeSO 4 1.0mg Trace minerals solution 10.0mL n-Heptadecane 1.0mL Trace Minerals Solution: Composition per liter: ZnSO 4 ·7H 2 O 7.0mg H 3 BO 4 1.0mg MoO 3 1.0mg CuSO 4 ·5H 2 O 500.0μg CoSO 4 ·7H 2 O 18.0μg MnSO 4 ·5H 2 O 7.0μg Preparation of Trace Minerals Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components, except n-heptadecane, to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Aseptically add the n- heptadecane. Mix thoroughly. Distribute into sterile tubes or flasks. Use: For the cultivation and maintenance of Bacillus thermoleo- vorans. Minerals Modified Glutamate Agar Composition per liter: Agar 15.0g Lactose 10.0g Sodium glutamate 6.35g NH 4 Cl 2.5g K 2 HPO 4 0.9g Sodium formate 0.25g MgSO 4 ·7H 2 O 0.1g L-Aspartic acid 0.024g L-Arginine 0.02g L-Cystine 0.02g Bromcresol Purple 0.01g CaCl 2 ·2H 2 O 0.01g Ferric ammonium citrate 0.01g Nicotinic acid 1.0mg Pantothenic acid 1.0mg Thiamine 1.0mg pH 6.7 ± 0.2 at 25°C 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 10 min at 11 psi pressure–116°C. Pour into sterile Petri dishes in 20.0mL volumes. Use: For the cultivation of coliform bacteria from foods. Minerals Modified Medium Composition per liter: Lactose 20.0g Sodium glutamate 12.7g NH 4 Cl 5.0g K 2 HPO 4 1.8g Sodium formate 0.5g MgSO 4 ·7H 2 O 0.2g L-Aspartic acid 0.048g L-Cystine 0.04g L-Arginine 0.04g Ferric ammonium citrate 0.02g CaCl 2 ·2H 2 O 0.02g Bromcresol Purple 0.02g Thiamine 2.0mg Nicotinic acid 2.0mg Pantothenic acid 2.0mg pH 6.7 ± 0.2 at 25°C Source: This medium is available as a premixed powder from Oxoid Unipath. Preparation of Medium: Add NH 4 Cl to distilled/deionized water and bring volume to 800.0mL. Add remaining components and bring volume to 1.0L. Mix thoroughly. Adjust pH to 6.7. Distribute into tubes or flasks. Autoclave for 10 min at 10 psi pressure–116°C. Check pH af- ter autoclaving. This medium is double strength. Use: For the enumeration of coliform bacteria in water. Minimal Agar I Composition per liter: Agar 20.0g Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: L-Tryptophan 20.0mg Uracil 20.0mg L-Histidine·HCl 20.0mg © 2010 by Taylor and Francis Group, LLC Minimal Agar, Davis 1187 Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, 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 50°–55°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Agar II Composition per liter: Agar 20.0g Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: Adenine sulfate 20.0mg L-Arginine·HCl 20.0mg L-Histidine·HCl 20.0mg Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, 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 50°–55°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Agar III Composition per liter: Agar 20.0g Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: L-Leucine 30.0mg L-Tryptophan 20.0gm Uracil 20.0mg Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, 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 50°–55°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Agar, Davis Composition per liter: Agar 15.0g K 2 HPO 4 7.0g KH 2 PO 4 2.0g (NH 4 ) 2 SO 4 1.0g Glucose 1.0g Sodium citrate 0.5g MgSO 4 ·7H 2 O 0.1g pH 7.0 ± 0.2 at 25°C Source: This medium is available as a premixed powder from BD Di- agnostic Systems. Preparation of Medium: Add components to cold distilled/deion- ized water and bring volume to 1.0L. Mix thoroughly. Gently heat and bring 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 isolation, cultivation, and characterization of nutritional mutants of Escherichia coli. © 2010 by Taylor and Francis Group, LLC 1188 Minimal Broth I Minimal Broth I Composition per liter: Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: L-Tryptophan 20.0mg Uracil 20.0mg L-Histidine·HCl 20.0mg Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, to distilled/deionized water and bring volume to 990.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure– 121°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Aseptically distribute into sterile tubes or flasks. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Broth II Composition per liter: Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: Adenine sulfate 20.0mg L-Arginine·HCl 20.0mg L-Histidine·HCl 20.0mg Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, to distilled/deionized water and bring volume to 990.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure– 121°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Aseptically distribute into sterile tubes or flasks. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Broth III Composition per liter: Agar 20.0g Glucose 20.0g (NH 4 ) 2 SO 4 5.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.5g NaCl 0.1g CaCl 2 ·2H 2 O 0.1g Inositol 2.0mg KI 1.0mg H 3 BO 3 0.5mg ZnSO 4 ·7H 2 O 0.4mg MnSO 4 ·4H 2 O 0.4mg Thiamine·HCl 0.4mg Pyroxidine·HCl 0.4mg Niacin 0.4mg Calcium pantothenate 0.4mg p-Aminobenzoic acid 0.2mg Riboflavin 0.2mg FeCl 3 0.2mg Na 2 MoO 4 ·4H 2 O 0.2mg CuSO 4 ·5H 2 O 0.04mg Folic acid 2.0μg Biotin 2.0μg Growth supplement solution 10.0mL pH 5.5 ± 0.2 at 25°C Growth Supplement Solution: Composition per 10.0mL: L-Leucine 30.0mg L-Tryptophan 20.0mg Uracil 20.0mg Preparation of Growth Supplement Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Add components, except growth supple- ment solution, to distilled/deionized water and bring volume to © 2010 by Taylor and Francis Group, LLC Minimum Essential Medium with Bicarbonate, Serum, and Antibiotics 1189 990.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure– 121°C. Aseptically add 10.0mL of sterile growth supplement solution. Mix thoroughly. Aseptically distribute into sterile tubes or flasks. Use: For the cultivation of Saccharomyces cerevisiae. Minimal Broth, Davis Composition per liter: K 2 HPO 4 7.0g KH 2 PO 4 2.0g (NH 4 ) 2 SO 4 1.0g Glucose 1.0g Sodium citrate 0.5g MgSO 4 ·7H 2 O 0.1g pH 7.0 ± 0.2 at 25°C Source: This medium is available as a premixed powder from BD Di- agnostic Systems. Preparation of Medium: Add components to cold distilled/deion- ized 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 isolation, cultivation, and characterization of nutritional mutants of Escherichia coli. Also recommended for the isolation and characterization of nutritional mutants from wild-type strains of Bacil- lus subtilis when used in conjunction with minimal agar Davis and antibiotic medium 3. Minimal F-Top Agar Composition per liter: NaCl 8.0g Agar 4.5g K 2 HPO 4 2.1g KH 2 PO 4 .0.6g (NH 4 ) 2 SO 4 .0.3g Glucose 0.3g Sodium citrate 0.15g MgSO 4 ·7H 2 O 0.03g pH 7.0 ± 0.2 at 25°C Preparation of Minimal Agar: Add components to cold distilled/ deionized water and bring volume to 1.0L. Mix thoroughly. Gently heat and bring 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 distribution of bacteriophage or bacterial cells evenly in a thin layer over the surface of a plate. Minimal Lactate Medium See: ML Medium Minimal Medium for Denitrifying Bacteria Composition per liter: Solution A 980.0mL Solution B 10.0mL Solution C 10.0mL Solution A: Composition per 980.0mL: KNO 3 5.0g Carbon source 4.0g (NH 4 ) 2 SO 4 1.0g K 2 HPO 4 ·3H 2 O 0.87g KH 2 PO 4 0.54g Preparation of Solution A: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Solution B: Composition per 100.0mL: MgSO 4 ·7H 2 O 2.0g Preparation of Solution B: Add MgSO 4 ·7H 2 O to distilled/deion- ized water and bring volume to 100.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Solution C: Composition per 100.0mL: CaCl 2 ·2H 2 O 0.2g FeSO 4 ·7H 2 O 0.1g MnSO 4 ·H 2 O 0.05g CuSO 4 ·5H 2 O 0.01g Na 2 MoO 4 ·2H 2 O 0.01g HCl (0.1N solution) 100.0mL Preparation of Solution C: Combine components. Mix thorough- ly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Preparation of Medium: Aseptically combine 980.0mL of cooled sterile solution A, 10.0mL of cooled sterile solution B, and 10.0mL of cooled sterile solution C. Mix thoroughly. Aseptically distribute into sterile tubes or flasks. Use: For the isolation and cultivation of denitrifying bacteria. Minimal Medium for Penicillium Interspecific Hybrids Composition per liter: Glucose 40.0g NaNO 3 3.0g KH 2 PO 4 1.0g KCl 0.5g MgSO 4 ·7H 2 O 0.5g FeSO 4 ·7H 2 O 0.01g pH 6.0 ± 0.2 at 25°C Preparation of Medium: Add components to distilled/deionized wa- ter and bring volume to 1.0L. Mix thoroughly. Adjust pH to 6.0. Distribute into tubes or flasks. Autoclave for 15 min at 15 psi pressure–121°C. Use: For the cultivation of genetic variants of Penicillium species. Minimum Essential Medium with Bicarbonate, Serum, and Antibiotics Composition per 1100.0mL: Minimum essential medium 950.0mL Fetal bovine serum, heat inactivated 100.0mL NaHCO 3 solution 40.0mL Penicillin-streptomycin solution 10.0mL pH 7.2 ± 0.2 at 25°C Minimum Essential Medium (MEM): Composition per liter: Inorganic salt solution 400.0mL Other component solution 400.0mL Amino acid solution 100.0mL Vitamin solution 100.0mL © 2010 by Taylor and Francis Group, LLC 1190 Minimum Salts with HiVeg Acid Hydrolysate Inorganic Salt Solution: Composition per 400.0mL: NaCl 6.8g KCl 0.4g CaCl 2 , anhydrous 0.2g NaH 2 PO 4 ·H 2 O 0.14g MgSO 4 , anhydrous 97.67mg Preparation of Inorganic Salt Solution: Add components to dis- tilled/deionized water and bring volume to 400.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Other Component Solution: Composition per 400.0mL: D-Glucose 1.0g Phenol Red 10.0mg Preparation of Other Component Solution: Add components to distilled/deionized water and bring volume to 400.0mL. Mix thor- oughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to 25°C. Amino Acid Solution: Composition per 100.0mL: L-Glutamine 292.0mg L-Arginine·HCl 126.1mg L-Lysine·HCl 72.5mg L-Isoleucine 52.0mg L-Leucine 52.0mg L-Tyrosine, disodium salt 52.0mg L-Threonine 48.0mg L-Valine 46.0mg L-Histidine·HCl·H 2 O 42.0mg L-Phenylalanine 32.0mg L-Cysteine·2HCl 31.0mg L-Methionine 15.0mg L-Glutamic acid 14.7mg L-Aspartic acid 13.3mg L-Asparagine·H 2 O 13.2mg L-Proline 11.5mg L-Serine 10.5mg L-Tryptophan 10.0mg L-Alanine 8.9mg Glycine 7.5mg Preparation of Amino Acid Solution: Add components to dis- tilled/deionized water and bring volume to 100.0mL. Mix thoroughly. Filter sterilize. Vitamin Solution: Composition per 100.0mL: i-Inositol 2.0mg D-Ca pantothenate 1.0mg Choline chloride 1.0mg Folic acid 1.0mg Niacinamide 1.0mg Pyridoxal·HCl 1.0mg Thiamine·HCl 1.0mg Riboflavin 0.1mg Preparation of Vitamin Solution: Add components to distilled/ deionized water and bring volume to 100.0mL. Mix thoroughly. Filter sterilize. Preparation of Minimum Essential Medium (MEM): Asepti- cally combine 400.0mL of sterile inorganic salt solution, 400.0mL of sterile other component solution, 100.0mL of sterile amino acid solu- tion, and 100.0mL of sterile vitamin solution. NaHCO 3 Solution: Composition per 40.0mL: NaHCO 3 2.0g Preparation of NaHCO 3 Solution: Add NaHCO 3 to distilled/de- ionized water and bring volume to 40.0mL. Mix thoroughly. Filter ster- ilize. Penicillin-Streptomycin Solution Composition per 10.0mL: Penicillin 0.01g Streptomycin 0.01g Preparation of Penicillin-Streptomycin Solution: Add compo- nents to distilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Preparation of Medium: Aseptically combine 950.0mL of sterile minimum essential medium, 100.0mL of sterile heat inactivated fetal bovine serum, 40.0mL of sterile NaHCO 3 solution, and 10.0mL of ster- ile penicillin-streptomycin solution. Adjust pH to 7.2 with humidified 10% CO 2 in 90% air. Use: For the cultivation of Encephalitozoon cuniculi, Encephalito- zoon hellem, Encephalitozoon intestinalis, Naegleria fowleri, and Nosema corneum. Minimum Salts with HiVeg Acid Hydrolysate Composition per liter: Na 2 HPO 4 6.8g Glucose 4.0g Plant acid hydrolysate 4.0g KH 2 PO 4 3.0g NH 4 Cl 1.0g NaCl 0.5g MgSO 4 0.24g pH 6.8 ± 0.2 at 25°C Source: This medium is available as a premixed powder from Hi- Media. Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Heat gently and bring to boiling. Distribute into tubes or flasks. Autoclave for 15 min at 15 psi pressure–121°C. Use: For the cultivation of Escherichia coli strains used for genetic and molecular studies. MIO HiVeg Medium (Motility Indole Ornithine HiVeg Medium) Composition per liter: Plant hydrolysate 10.0g Plant peptone 10.0g L-Ornithine hydrochloride 5.0g Yeast extract 3.0g Agar 2.0g Glucose 1.0g Bromcresol Purple 0.02g pH 6.6 ± 0.2 at 25°C Source: This medium is available as a premixed powder from Hi- Media. Preparation of Medium: Add components to distilled/deionized water and bring to 1.0L. Mix thoroughly. Gently heat and bring to boil- © 2010 by Taylor and Francis Group, LLC Mitsuokella dentalis Medium 1191 ing. Distribute into tubes or flasks. Autoclave for 15 min at 15 psi pres- sure–121°C. Use: For the differentiation of Gram-negative enteric bacteria based on their motility, indole production, and ornithine decarboxylase activ- ity. MIO Medium See: Motility Indole Ornithine Medium Mist Agar Composition per liter: Cow manure, dry 50.0g Agar 15.0g Preparation of Medium: Add cow manure to 1.0L of tap water. Boil for 1 hr. Filter through cheesecloth. Filter through paper. Add agar to filtrate and bring volume to 1.0L with tap water. Gently heat and bring 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 Streptomyces fragmen- tosporus. Mitis Salivarius Agar Composition per liter: Sucrose 50.0g Agar 15.0g Enzymatic digest of protein 10.0g Proteose peptone 10.0g K 2 HPO 4 4.0g Dextrose 1.0g Trypan Blue 0.08g Crystal Violet 0.8mg Na 2 TeO 3 solution 1.0mL pH 7.0 ± 0.2 at 25°C Source: This medium is available as a premixed powder from BD Di- agnostic Systems. Na 2 TeO 3 Solution: Composition per 10.0mL: Na 2 TeO 3 0.1g Preparation of Na 2 TeO 3 Solution: Add Na 2 TeO 3 to 10.0mL of distilled/deionized water. Mix thoroughly. Filter sterilize. Caution: Potassium tellurite is toxic. Preparation of Medium: Add components to distilled/deionized water and bring volume to 999.0mL. Mix thoroughly. Gently heat and bring to boiling. Autoclave for 15 min at 15 psi pressure–121°C. Cool medium to 50°–55°C. Aseptically add 1.0mL of the sterile Na 2 TeO 3 solution to the cooled basal medium. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Use: For the selective isolation of Streptococcus mitis, Streptococcus salivarius, and other viridans streptococci and enterococci. Mitis Salivarius HiVeg Agar Base with Tellurite Composition per liter: Sucrose 50.0g Agar 15.0g Plant hydrolysate 15.0g Plant peptone 5.0g K 2 HPO 4 4.0g Glucose 1.0g Trypan Blue 0.075g Crystal Violet 0.8mg Na 2 TeO 3 solution 1.0mL pH 7.0 ± 0.2 at 25°C Source: This medium, without tellurite, is available as a premixed powder from HiMedia. Na 2 TeO 3 Solution: Composition per 10.0mL: Na 2 TeO 3 0.1g Preparation of Na 2 TeO 3 Solution: Add Na 2 TeO 3 to 10.0mL of distilled/deionized water. Mix thoroughly. Filter sterilize. Caution: Potassium tellurite is toxic. Preparation of Medium: Add components to distilled/deionized water and bring volume to 999.0mL. Mix thoroughly. Gently heat and bring to boiling. Autoclave for 15 min at 15 psi pressure–121°C. Cool medium to 50°–55°C. Aseptically add 1.0mL of the sterile Na 2 TeO 3 solution to the cooled basal medium. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Use: For the selective isolation of Streptococcus mitis, Streptococcus salivarius, and other viridans streptococci and enterococci. Mitsuokella dentalis Medium Composition per 1003.3mL: Yeast extract 10.0g Beef extract 5.0g Glucose 5.0g Trypticase™ 5.0g L-Cysteine·HCl 0.5g (NH 4 ) 2 SO 4 0.5g Resazurin 1.0mg Bovine serum 50.0mL Mineral solution 40.0mL Hemin solution 1.0mL Vitamin K 1 solution 0.2mL pH 6.9 ± 0.2 at 25°C Bovine Serum: Composition per 50.0mL: Bovine serum 50.0mL Preparation of Bovine Serum: Incubate 50.0mL of bovine serum in a GasPak™ container overnight to make anaerobic. Mineral Solution: Composition per liter: NaHCO 3 10.0g NaCl 2.0g K 2 HPO 4 1.0g KH 2 PO 4 1.0g MgSO 4 ·7H 2 O 0.48g CaCl 2 ·2H 2 O 0.3g Preparation of Mineral Solution: Add components to distilled/ deionized water and bring volume to 1.0L. Mix thoroughly. Hemin Solution: Composition per 100.0mL: Hemin 5.0mg NaOH (0.002% solution) 1.0mL © 2010 by Taylor and Francis Group, LLC 1192 Mixed Cereal Agar Preparation of Hemin Solution: Add hemin to 1.0mL of NaOH solution. Mix thoroughly. Vitamin K 1 Solution: Composition per 100.0mL: Vitamin K 1 1.09g Preparation of Vitamin K 1 Solution: Add vitamin K 1 to 99.0mL of absolute ethanol. Store in the dark at 4°C. Preparation of Medium: Add components, except L-cysteine·HCl and bovine serum, to distilled/deionized water and bring volume to 950.0mL. Mix thoroughly. Gently heat and bring to boiling. Continue boiling for 5 min. Cool to room temperature while sparging with 100% CO 2 . Add L-cysteine·HCl. Mix thoroughly. Adjust pH to 6.0 with 8N NaOH. After pH has been reached, change sparging gas to 100% N 2 . Anaerobically distribute into bottles under 100% N 2 . Autoclave for 15 min at 15 psi pressure–121°C. Aseptically and anaerobically add 50.0mL of bovine serum. Mix thoroughly. Use: For the cultivation and maintenance of Mitsuokella dentalis. Mixed Cereal Agar Composition per liter: Gerber™ mixed cereal (oats, wheat, corn, barley) 50.0g Agar 15.0g Sucrose 15.0g Thiamine·HCl 5.0mg 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 15 min at 15 psi pressure–121°C. Pour into sterile Petri dishes or leave in tubes. Use: For the cultivation and maintenance of Tilletia caries. Mixotrophic Nitrobacter Medium (DSMZ Medium 756a) Composition per liter: NaNO 2 2.0g Yeast extract 1.5g Peptone 1.5g Na-pyruvate 0.55g Stock solution 100.0mL Trace elements solution 1.0mL pH 7.4 ± 0.2 at 25°C Stock Solution: Composition per liter: NaCl 5.0g KH 2 PO 4 1.5g MgSO 4 ·7H 2 O 0.5g CaCO 3 0.07g Preparation of Stock Solution: Add components to distilled/deion- ized water and bring volume to 1.0L. Mix thoroughly. Trace Elements Solution: Composition per liter: FeSO 4 ·7H 2 O 97.3mg H 3 BO 3 49.4mg ZnSO 4 ·7H 2 O 43.1mg (NH 4 ) 6 Mo 7 O 24 ·4H 2 O 37.1mg MnSO 4 ·2H 2 O 33.8mg CuSO 4 ·5H 2 O 25.0mg Preparation of Trace Elements Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Adjust pH to 7.4. Dis- tribute into tubes or flasks. Autoclave for 15 min at 15 psi pressure– 121°C. Use: For the cultivation of Nitrobacter vulgaris. Mixotrophic Nitrobacter Medium (LMG Medium 246) Composition per liter: NaNO 2 2.0g Yeast extract 1.50g Peptone 1.50g Na-pyruvate 0.55g Stock solution 100.0mL Trace elements solution 1.0mL pH 7.4 ± 0.2 at 25°C Stock Solution: Composition per liter: NaCl 5.0g KH 2 PO 4 1.5g MgSO 4 ·7H 2 O 0.5g CaCO 3 0.07g Preparation of Stock Solution: Add components to distilled/de- ionized water and bring volume to 1.0L. Mix thoroughly. Trace Elements Solution: Composition per liter: (NH 4 )Mo7O 2 437.10mg FeSO 4 ·7H 2 O 97.30mg ZnSO 4 ·7H 2 O 43.10mg H 3 BO 3 39.40mg MnSO 4 ·H 2 O 33.80mg CuSO 2 ·5H 2 O 25.00mg Preparation of Trace Elements Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Adjust pH to 7.4 with NaOH. Distribute into tubes or flasks. Autoclave for 15 min at 15 psi pressure–121°C. Use: For the cultivation of mixotrophic Nitrobacter spp. Mixotrophic Nitrobacter Medium, 10% (DSMZ Medium 756b) Composition per liter: NaNO 2 2.0g Yeast extract 0.15g Peptone 0.15g Na-pyruvate 0.055g Stock solution 100.0mL Trace elements solution 1.0mL pH 8.6 ± 0.2 at 25°C Stock Solution: Composition per liter: NaCl 5.0g KH 2 PO 4 1.5g © 2010 by Taylor and Francis Group, LLC MJ Medium 1193 MgSO 4 ·7H 2 O 0.5g CaCO 3 0.07g Preparation of Stock Solution: Add components to distilled/deion- ized water and bring volume to 1.0L. Mix thoroughly. Trace Elements Solution: Composition per liter: FeSO 4 ·7H 2 O 97.3mg H 3 BO 3 49.4mg ZnSO 4 ·7H 2 O 43.1mg (NH 4 ) 6 Mo 7 O 24 ·4H 2 O 37.1mg MnSO 4 ·2H 2 O 33.8mg CuSO 4 ·5H 2 O 25.0mg Preparation of Trace Elements Solution: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Preparation of Medium: Add components to distilled/deionized water and bring volume to 1.0L. Mix thoroughly. Adjust pH to 8.6. Dis- tribute into tubes or flasks. Autoclave for 15 min at 15 psi pressure– 121°C. Use: For the cultivation of Nitrobacter hamburgensis and Nitrobacter vulgaris. MJ Medium (DSMZ Medium 1011) Composition per liter: NaCl 30.0g MgCl 2 ·6H 2 O 4.18g MgSO 4 ·7H 2 O 3.4g KCl 0.33g NH 4 Cl 0.25g KH 2 PO 4 0.14g CaCl 2 ·2H 2 O 0.014g Fe(NH 4 ) 2 (SO 4 ) 2 ·6H 2 O 0.01g NiCl 2 ·6H 2 O 0.5mg Na 2 SeO 3 ·5H 2 O 0.5mg Trace elements solution 10.0mL Vitamin solution 10.0mL Thiosulfate solution 10.0mL Bicarbonate solution 10.0mL pH 6.7 ± 0.2 at 25°C Bicarbonate Solution: Composition per 10.0mL: NaHCO 3 1.5g Preparation of Bicarbonate Solution: Add components to dis- tilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Thiosulfate Solution: Composition per 10.0mL: NaS 2 O 3 ·5H 2 O 1.5g Preparation of Thiosulfate Solution: Add components to dis- tilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temper- ature. Trace Elements Solution: Composition per liter: MgSO 4 ·7H 2 O 3.0g Nitrilotriacetic acid 1.5g NaCl 1.0g MnSO 4 ·2H 2 O 0.5g CoSO 4 ·7H 2 O 0.18g ZnSO 4 ·7H 2 O 0.18g CaCl 2 ·2H 2 O 0.1g FeSO 4 ·7H 2 O 0.1g NiCl 2 ·6H 2 O 0.025g KAl(SO 4 ) 2 ·12H 2 O 0.02g H 3 BO 3 0.01g Na 2 MoO 4 ·4H 2 O 0.01g CuSO 4 ·5H 2 O 0.01g Na 2 SeO 3 ·5H 2 O 0.3mg Preparation of Trace Elements Solution: Add nitrilotriacetic acid to 500.0mL of distilled/deionized water. Dissolve by adjusting pH to 6.5 with KOH. Add remaining components. Add distilled/deionized water to 1.0L. Mix thoroughly. Vitamin Solution: Composition per liter: Pyridoxine-HCl 10.0mg Thiamine-HCl·2H 2 O 5.0mg Riboflavin 5.0mg Nicotinic acid 5.0mg D-Ca-pantothenate 5.0mg p-Aminobenzoic acid 5.0mg Lipoic acid 5.0mg Biotin 2.0mg Folic acid 2.0mg Vitamin B 12 0.1mg Preparation of Vitamin Solution: Add components to distilled/ deionized water and bring volume to 1.0L. Mix thoroughly. Sparge with 80% H 2 + 20% CO 2 . Filter sterilize. Preparation of Medium: Add components, except thiosulfate, bi- carbonate, and vitamin solutions, to distilled/deionized water and bring volume to 970.0mL. Mix thoroughly. Gently heat and bring to boiling. Boil for 1 min. Cool to room temperature while sparging with a gas mix- ture of 80% N 2 + 20% CO 2 . Dispense under the same atmosphere into culture vessels. Fill up to a volume of 20% volume of vessel. Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temperature under an atmosphere of 0% N 2 + 20% CO 2 . Aseptically add sterile thiosulfate, bicarbonate, and vitamin solutions. Mix thoroughly. Adjust pH to 6.7. Aseptically dispense into tubes, flasks, or bottles. After inoculation pres- surize vessels to 0.5 bar overpressure with 80% N 2 + 20% CO 2 gas mix- ture. Add sterile air in an amount that is equivalent to a volume of 20% of the headspace. After inoculation reduce medium with 10–20mg so- dium dithionite per liter medium, added from a 5% solution freshly prepared under N 2 and filter sterilized. Pressurize vessels to 2 bar over- pressure with 80% H 2 and 20% CO 2 . Use: For the cultivation of Sulfurimonas autotrophica, Thiomicrospira thermophila, and Desulfothermus okinawensis. MJ Medium (DSMZ Medium 1011) Composition per liter: NaCl 30.0g MgCl 2 ·6H 2 O 4.18g MgSO 4 ·7H 2 O 3.4g KCl 0.33g NH 4 Cl 0.25g KH 2 PO 4 0.14g CaCl 2 ·2H 2 O 0.014g Fe(NH 4 ) 2 (SO 4 ) 2 ·6H 2 O 0.01g © 2010 by Taylor and Francis Group, LLC 1194 MJ Medium for Thiobacter subterraneus NiCl 2 ·6H 2 O 0.5mg Na 2 SeO 3 ·5H 2 O 0.5mg Trace elements solution 10.0mL Vitamin solution 10.0mL Thiosulfate solution 10.0mL Bicarbonate solution 10.0mL Pyruvate solution 10.0mL Yeast extract solution 10.0mL Lactate soltuion 10.0mL pH 6.7 ± 0.2 at 25°C Pyruvate Solution: Composition per 10.0mL: Sodium pyruvate 0.5g Preparation of Pyruvate Solution: Add sodium pyruvate to dis- tilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Lactate Solution: Composition per 10.0mL: Sodium lactate 0.5g Preparation of Lactate Solution: Add sodium lactate to distilled/ deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Yeast Extract Solution: Composition per 10.0mL: Yeast extract 0.1g Preparation of Yeast Extract Solution: Add yeast extract to dis- tilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Bicarbonate Solution: Composition per 10.0mL: NaHCO 3 1.5g Preparation of Bicarbonate Solution: Add NaHCO 3 to distilled/ deionized water and bring volume to 10.0mL. Mix thoroughly. Filter sterilize. Thiosulfate Solution: Composition per 10.0mL: NaS 2 O 3 ·5H 2 O 1.5g Preparation of Thiosulfate Solution: Add NaS 2 O 3 ·5H 2 O to dis- tilled/deionized water and bring volume to 10.0mL. Mix thoroughly. Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temper- ature. Trace Elements Solution: Composition per liter: MgSO 4 ·7H 2 O 3.0g Nitrilotriacetic acid 1.5g NaCl 1.0g MnSO 4 ·2H 2 O 0.5g CoSO 4 ·7H 2 O 0.18g ZnSO 4 ·7H 2 O 0.18g CaCl 2 ·2H 2 O 0.1g FeSO 4 ·7H 2 O 0.1g NiCl 2 ·6H 2 O 0.025g KAl(SO 4 ) 2 ·12H 2 O 0.02g H 3 BO 3 0.01g Na 2 MoO 4 ·4H 2 O 0.01g CuSO 4 ·5H 2 O 0.01g Na 2 SeO 3 ·5H 2 O 0.3mg Preparation of Trace Elements Solution: Add nitrilotriacetic acid to 500.0mL of distilled/deionized water. Dissolve by adjusting pH to 6.5 with KOH. Add remaining components. Add distilled/deionized water to 1.0L. Mix thoroughly. Vitamin Solution: Composition per liter: Pyridoxine-HCl 10.0mg Thiamine-HCl·2H 2 O 5.0mg Riboflavin 5.0mg Nicotinic acid 5.0mg D-Ca-pantothenate 5.0mg p-Aminobenzoic acid 5.0mg Lipoic acid 5.0mg Biotin 2.0mg Folic acid 2.0mg Vitamin B 12 0.1mg Preparation of Vitamin Solution: Add components to distilled/ deionized water and bring volume to 1.0L. Mix thoroughly. Sparge with 80% H 2 + 20% CO 2 . Filter sterilize. Preparation of Medium: Add components, except lactate, pyruvate, yeast extract, thiosulfate, bicarbonate, and vitamin solutions, to dis- tilled/deionized water and bring volume to 970.0mL. Mix thoroughly. Gently heat and bring to boiling. Boil for 1 min. Cool to room tempera- ture while sparging with a gas mixture of 80% H 2 + 20% CO 2 . Dispense under the same atmosphere into culture vessels. Fill up to a volume of 20% volume of vessel. Autoclave for 15 min at 15 psi pressure–121°C. Cool to room temperature under an atmosphere of 80% H 2 + 20% CO 2 . Aseptically add sterile lactate, pyruvate, yeast extract, thiosulfate, bicar- bonate, and vitamin solutions. Mix thoroughly. Adjust pH to 6.7. Asep- tically dispense into tubes, flasks, or bottles. After inoculation pressurize vessels to 0.5 bar overpressure with 80% H 2 + 20% CO 2 gas mixture. Add sterile air in an amount that is equivalent to a volume of 20% of the headspace. After inoculation reduce medium with 10–20 mg sodium dithionite per liter medium, added from a 5% solution freshly prepared under N 2 and filter sterilized. Pressurize vessels to 2 bar overpressure with 80% H 2 and 20% CO 2 . Use: For the cultivation of Desulfothermus okinawensis. MJ Medium for Thiobacter subterraneus (DSMZ Medium 1011a) Composition per liter: MnCl 2 ·6H 2 O 4.18g NaCl 3.0g MgSO 4 ·7H 2 O 0.34g KCl 0.33g NH 4 Cl 0.25g KH 2 PO 4 0.14g CaCl 2 ·2H 2 O 0.014g Fe(NH 4 ) 2 (SO 4 ) 2 ·6H 2 O 0.01g NiCl 2 ·6H 2 O 0.5mg Na 2 SeO 3 ·5H 2 O 0.5mg Trace elements solution 10.0mL Vitamin solution 10.0mL Thiosulfate solution 10.0mL Bicarbonate solution 10.0mL pH 6.7 ± 0.2 at 25°C Bicarbonate Solution: Composition per 10.0mL: NaHCO 3 1.5g © 2010 by Taylor and Francis Group, LLC . cultivation, and characterization of nutritional mutants of Escherichia coli. Also recommended for the isolation and characterization of nutritional mutants from wild-type strains of Bacil- lus subtilis. Asepti- cally combine 400.0mL of sterile inorganic salt solution, 400.0mL of sterile other component solution, 100.0mL of sterile amino acid solu- tion, and 100.0mL of sterile vitamin solution. NaHCO 3 . Aseptically combine 950.0mL of sterile minimum essential medium, 100.0mL of sterile heat inactivated fetal bovine serum, 40.0mL of sterile NaHCO 3 solution, and 10.0mL of ster- ile penicillin-streptomycin