Tetrathionate Broth, Base

Tetrathionate Broth Base

EMD Cat. No. 1.05285.0500

500 g

For the selective enrichment of salmonellae from various materials.

This broth complies with the specifications given in the United States Pharmacopeia XXIII (1995) and the recommendations of the APHA (1992).

Additives

EMD Cat. No.	Product	Pack Size
1.01310.0050	Brilliant green (C.I. 42040)	50 g
B10135-30	Iodine resublimed	100 g
B10212-34	Potassium iodide	500 g

Mode of Action	Preparation
Experimental Procedure and Evaluation	Quality Control
Typical Composition (g/liter)	Literature

Mode of Action
Tetrathionate and excess thiosulfate (PALUMBO and ALFORD 1970) suppress coliform microorganisms and other accompanying bacteria, whereas all tetrathionate-reducing bacteria (e.g. salmonellae and Proteus) can multiply more or less normally in this medium. Acidic tetrathionate decomposition products are formed, which are neutralized by calcium carbonate
Bile salts largely inhibit all microorganisms, which do not normally live in the intestine. The United States Pharmacopeia recommends the addition of brilliant green, which suppresses, above all, the Gram-positive microbial flora. The resulting culture medium has a very strong inhibitory effect; it is therefore sometimes better to omit the brilliant green in order to obtain satisfactory yields of salmonellae. According to JEFFRIES (1959), Proteus can be suppressed by adding 0.04 g novobiocin/liter.

Preparation
Suspend 46 g/liter, heat briefly to the boil and cool rapidly.

Do not autoclave.
NOTE: When prepared, the medium shows a pH of around 7.0 ± 0.2 at 25°C. For testings in line with FDA-BAM please adjust pH to 8.4 ± 0.2 at 25°C.

Prior to use, add 20 ml iodine/potassium iodide solution/liter, if desired 10 ml of a 0.1 % brilliant green solution/liter and if required 0.04 g novobiocin/liter. Avoid any further heating. When dispensing the prepared medium, make sure that any precipitate formed is evenly suspended.
Preparation of the iodine/potassium iodide solution: Iodine 6 g; potassium iodide 5 g; distilled water 20 ml.

The ready-to-use broth should be prepared and used the same day.
The medium is turbid and green with white sediment (calcium carbonate).

Experimental Procedure and Evaluation
Inoculate the culture medium massively with the sample material.
Incubation: 18-24 hours at 35-37 °C or 43 °C respectively (BÄNFFER 1971).

The resulting cultures are then subjected to further tests.

Quality control

Test strains	Inoculum	Growth after 24 hours
Escherichia coli ATCC 25922	approx. 99 %	<= 5 %
Salmonella typhimurium ATCC 14028	approx. 1 %	>= 95 %

Typical Composition (g/liter)
Peptone from casein 2.5; peptone from meat 2.5; bile salt mixture 1.0; calcium carbonate 10.0; sodium thiosulfate 30.0.
Also to be added:
Potassium iodide 5.0; iodine 6.0; if required brilliant green 0.01.

Literature

American Public Health Association: Compendium of methods for the microbiological examination of foods. 3 rd ed. (1992).
BÄNFFER, J.R.: Comparison of the isolation of Salmonellae from human faeces by enrichment at 37 °C and 43 °C. - Zbl. Bakt. I. Orig., 217; 35-40 (1971).
JEFFRIES, L.: Novobiocin - tetrathionate broth: A medium of improved selectivity for the isolation of salmonellae from faeces. - J. Clin. Path., 12; 568-571 (1959).
KNOX, R., POLLOCK, M.R., a. GELL, F.G.H.: The selective action of tetrathionate in bacteriological media. - J. Hyg., 43; 147-158 (1943).
PALUMBO, S., a. ALFORD, J.: Inhibitory action of tetrathionate enrichment broth. - Appl. Microbiol., 20; 970-976 (1970).
United States Pharmacopeia XXIII, Chapter "Microbiol Limit Tests", 1995.