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Appendix G
Media Glossary
January 2006
Glossary of agars, broths and other reagents
Section O
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(PDF Version - 22 K)| Monosodium phosphate solution, 1.0 M | |
|---|---|
| Sodium phosphate monobasic (NaH 2PO 4.H 2O) | 6.9 g |
| Distilled water | 45.0 mL |
| Sodium hydroxide (NaOH 30% w/v) | 3.0 mL |
| pH 7.0 | |
| ONPG | |
| ONPG | 80.0 g |
| Distilled water | 15.0 mL |
| Monosodium phosphate solution | 5.0 mL |
Complete Medium :
Monosodium phosphate solution : Suspend sodium phosphate monobasic in distilled water, and stir to dissolve. Add NaOH and adjust pH to 7.0. Bring the volume to 50.0 mL with distilled water and refrigerate.
ONPG: Dissolve ONPG in warmed distilled water (37oC) and add the monosodium phosphate solution. The solution should be colourless. Store in the refrigerator. Warm the amount required to 37oC before use.
| Basal Medium | |
|---|---|
| Tryptone or trypticase | 10.0 g |
| Yeast extract | 3.0 g |
| Dextrose | 4.0 g |
| Dipotassium phosphate (K 2HPO 4) | 2.5 g |
| Agar | 17.0 g |
| Cysteine | 0.001 g |
| Orange serum | 200 mL |
| Distilled water | 800 mL |
| pH 5.5 ± 0.2 | |
| Orange serum | |
| Freshly extracted orange juice or reconstituted frozen orange juice concentrate | 1.0 L |
| Filter aid | 30.0 g |
Complete Medium:
Suspend basal medium ingredients in 800 mL of distilled water. Heat to boiling with constant stirring to dissolve all ingredients. Autoclave at 121oC for 15 min. Do not overheat as this can result in a softer medium.
Orange serum: Prepare orange serum by heating 1.0 L of freshly extracted orange juice or reconstituted frozen orange juice concentrate to approximately 93oC (200oF). Add 30.0 g of "filter aid" and mix thoroughly. Filter under suction through a Buchner funnel using coarse filter paper precoated with "filter aid". Discard the first few mL of filtered serum.
| Oxford Agar (OXA) | |
|---|---|
| Basal Medium | |
| Columbia blood agar base | 39.0 g |
| Esculin | 1.0 g |
| Ferric ammonium citrate | 0.5 g |
| Lithium chloride (LiCl) | 15.0 g |
| Agar | 2.0 g |
| Distilled water | 1.0 L |
| pH 7.2 ± 0.2 | |
| Supplement per 10 mL (Commercially available) | |
| Cycloheximide | 400 mg |
| Colistin sulphate | 20.0 mg |
| Acriflavine | 5.0 mg |
| Cefotetan | 2.0 mg |
| Fosfomycin | 10.0 mg |
Complete Medium:
Suspend basal medium ingredients in 1.0 L of distilled water. Heat to boiling with constant stirring to dissolve all ingredients. Autoclave at 121oC for 15 min and cool to 45-50oC. Aseptically add 10 ml of supplement and dispense. Store under refrigeration in the dark (acriflavine can photo-oxidize to form compounds inhibitory to Listeria spp.).
Supplement: Dissolve supplement ingredients in 10 mL of a 1:1 mixture of ethanol and distilled water. Filter sterilize.
| Basal Medium | |
|---|---|
| N,N,N',N'-Tetramethyl-p-phenylenediamine.2HCl | 1.0 g |
| Distilled water | 100 mL |
Complete Medium:
Reagent can be kept for up to 7 days if stored in a dark glass bottle under refrigeration, otherwise it is to be used freshly prepared.
Alternately: The oxidase test may be performed using a 1% solution of N,N-dimethyl-p-phenylenediamine hydrochloride.
| Basal Medium | |
|---|---|
| Tryptone | 2.0 g |
| Sodium chloride (NaCl) | 5.0 g |
| Potassium phosphate dibasic ( K 2 HPO 4 ) | 0.3 g |
| Bromthymol blue | 0.03 g |
| Agar | 2.0 g |
| Glucose | 10.0 g |
| Distilled water | 1.0 L |
| pH 6.8 ± 0.2 | |
Complete Medium:
Suspend basal medium ingredients in 1.0 L of distilled water. Heat to boiling with constant stirring to dissolve all ingredients. Dispense in 5 mL portion into tubes and autoclave at 121oC for 15 min.
Note : For halophilic Vibrio spp. add another 15.0 g NaCl (2% NaCl, final concentration).
Note : To use OF with sugars other than glucose, prepare the medium without glucose in 900 mL of distilled water, sterilize as above and cool to 45-50oC . Add 100 mL of a 10% solution of filter-sterilized sugar to the basal medium and dispense.
| Basal Medium | |
|---|---|
| Yeast extract | 5.0 g |
| Dextrose | 20.0 g |
| Agar | 12.0 g |
| Distilled water | 1.0 L |
| pH 7.0 ± 0.2 | |
| Oxytetracycline GYE Supplement (commercially available) | |
| Oxytetracycline | 0.1 g |
| Distilled water | 10.0 mL |
Complete Medium:
Suspend basal medium ingredients in 1.0 L of distilled water. Heat to boiling with constant stirring to dissolve all ingredients. Autoclave at 121oC for 15 min and cool to 45-50oC. Aseptically add the sterile oxytetracycline solution and mix thoroughly. Dispense into plates.
Supplement: Add oxytetracycline to distilled water and bring the volume to 10.0 mL. Mix thoroughly and filter sterilize.
