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212a. TREPONEMA RUMINIS MEDIUM

Culture medium recipe

Vol.
×
Equipment needed: Hungate tubes Filter Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2 Solution A 915 ml - -
3 Solution B 30 ml - -
4 Solution C 4 ml - -
5 Solution D 4 ml - -
6 Solution E 30 ml - -
7 Solution F 1 ml - -
8 Solution G 2 ml - -
9 Solution H 10 ml - -
10 Solution I 10 ml - -
11 , might need strain adjustment
(2% w/v)
3 g/l - -
Instructions:
  1. Add and dissolve ingredients of solution A, adjust pH to 7.0 and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under the same gas atmosphere into anoxic Hungate-type tubes and autoclave. Solution B is prepared under 80% N2 and 20% CO2 gas atmosphere and autoclaved. Solutions C, D and H are autoclaved under 100% N2 gas atmosphere. Solutions E, F, G and I are prepared under 100% N2 gas and sterilized by filtration. To complete the medium appropriate amounts of solutions B to I are added to the sterile solution A in the sequence as indicated.
  2. Note: Some cultures are shipped in semi-solid medium which stimulates growth at the beginning. For agar stabs 3.00 g/l agar are added to the complete medium from a sterile anoxic stock solution (2% w/v). Upon receipt add anoxically 1 - 2 ml of the recommended freshly prepared liquid medium to the agar tube and incubate for 3 - 5 days. After incubation transfer 0.5 ml of the resulting cell suspension in the liquid phase to tubes with liquid medium.
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.50 g 0.545 4.007
2 0.33 g 0.36 1.77
3 0.40 g 0.436 7.464
4 0.40 g 0.436 8.155
5 0.05 g 0.055 0.371
6 Trace element solution SL-10 1.00 ml - -
7 Selenite-tungstate solution 1.00 ml - -
8 Clarified rumen fluid 50.00 ml - -
9
(BD BBL)
1.00 g 1.091 -
10 2.80 g 3.053 21.497
11 Vitamin K1 solution 1.00 ml - -
12 Volatile fatty acid mixture 3.10 ml - -
13
(0.1% w/v)
0.50 ml 5.5e-4 0.002
14 860.00 ml 936.159 51964.7
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 1.5 g 50 471.752
2 30.0 ml 998.207 55408.8
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.4 g 100 -
2 4.0 ml 998.207 55408.8
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.8 g 200 1110.15
2 4.0 ml 998.207 55408.8
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 Sugar mix 30 ml - -
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 Seven vitamins solution 1 ml - -
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.5 g 250 2271.82
2 2.0 ml 998.207 55408.8
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.3 g 30 170.808
2 10.0 ml 998.207 55408.8
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.4 g 40 259.318
2 10.0 ml 998.207 55408.8
Solution occurs in 161 other media (first defined in medium 385 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.5 g 0.5 12.501
2 3.0 mg 0.003 0.011
3 4.0 mg 0.004 0.012
4 1000.0 ml 998.207 55408.8
Solution occurs in 32 other media (first defined in medium 1310 )
Equipment needed: Centrifuge Filter Autoclave
Instructions:
  1. Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 minand then centrifuged at 27,000 g for 20 min The supernatant is made anoxic by sparging with 100% N2 gas for 15 min dispensed under same gas atmosphere into anoxic serum vials to 30% of volume and then stored frozen at - 20°C.
Solution occurs in 5 other media (first defined in medium 78 )
Equipment needed: Filter
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2 0.1 ml - -
3
(95 %)
20.0 ml 750.5 16291
Instructions:
  1. Dissolve 0.1 ml of vitamin K1 in 20 ml 95% ethanol and filter sterilize. Store refrigerated in a brown bottle.
Solution occurs in 4 other media (first defined in medium 330 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 548.50 ml - -
2 193.50 ml 191.565 2585.97
3 129.00 ml 123.363 1400.18
4 32.25 ml 30.315 296.822
5 32.25 ml - -
6 32.25 ml - -
7 32.25 ml - -
Solution occurs in 1 other media (first defined in medium 843 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 1.70 g 56.667 165.549
2 1.70 g 56.667 165.548
3 1.80 g 60 166.523
4 1.90 g 63.333 185.025
5 0.75 g 25 166.522
6 30.00 ml 998.207 55408.8
Solution occurs in 86 other media (first defined in medium 503 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 100 mg 0.1 0.063
2 80 mg 0.08 0.583
3 20 mg 0.02 0.082
4 200 mg 0.2 1.625
5 100 mg 0.1 0.21
6 300 mg 0.3 1.459
7 200 mg 0.2 0.536
8 1000 ml 998.207 55408.8
Solution occurs in 239 other media (first defined in medium 320 )
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2
(25%; 7.7 M)
10.0 ml 3.17 86.943
3 1.5 g 1.5 7.545
4 70.0 mg 0.07 0.514
5 100.0 mg 0.1 0.505
6 6.0 mg 0.006 0.097
7 190.0 mg 0.19 0.799
8 2.0 mg 0.002 0.012
9 24.0 mg 0.024 0.101
10 36.0 mg 0.036 0.149
11 990.0 ml 988.225 54854.8
Instructions:
  1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.0 ml.

Metadata

Media status: Official DSMZ medium
Source: DSMZ
Taxonomic range:  Bacteria
Medium type: Complex medium
Final pH: 7.0
Equipment needed: Autoclave, Centrifuge, Filter, Hungate tubes

Associated strains

All strains for this medium: 3
Strains with modifications: 3

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Distilled water 943.742 52385.6
Na2SO4 2.777 19.552
Trehalose 1.889 5.518
Maltose x H2O 1.789 4.966
Cellobiose 1.69 4.937
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Cultivation metadata from 3 strains BacDive

pH 6 - 8 1 strain
pH 8 - 10 1 strain
phyla 1 3 strains
oxygen anaerobe 2 strains
oxygen obligate anaerobe 1 strain
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