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SELENITE BROTH BASE (LACTOSE)

Code: CM0395

An enriched medium for the isolation of Salmonella from faeces and food products.

Directions
Dissolve 4g of sodium biselenite LP0121 in 1 litre of distilled water and then add 19g of Selenite Broth Base. Warm to dissolve, mix well and fill out into containers. Sterilise in a boiling water bath, or in free flowing steam, for 10 minutes. DO NOT AUTOCLAVE.

To minimise any possible risk of teratogenicity to laboratory workers, the sodium biselenite must be added as a solution to this medium.
Robertson¹ reported miscarriages and possible teratogenic effects on pregnant laboratory assistants which may have been caused by ingested sodium biselenite. Oxoid therefore removed this substance from the powdered medium.

Although no further reports have been received sodium biselenite is now considered to be very toxic and should be handled with great care.

SODIUM BISELENITE (SODIUM HYDROGEN SELENITE)

Code: LP0121

Directions
Dissolve 4g in 1 litre of distilled water and use this solution to reconstitute the base medium.
Toxic by inhalation and if swallowed. Danger of cumulative effects.

Description
Klett² first demonstrated the selective inhibitory effects of selenite and Guth³ used it to isolate Salmonella typhi. It was twenty years later before Leifson⁴ fully investigated selenite and promoted wide use of the medium.

Selenium toxicity to certain micro-organisms is not fully understood but it is suggested that it reacts with sulphur and sulphydral groups in critical cell components^5,6. Liefson⁴ suggested that it is best to tube the medium to a depth of 2 inches (50mm) or more.

Proteus and Pseudomonas species appear to be resistant to its effects⁵. Lactose is added as a fermentable carbohydrate to prevent a rise in pH value during incubation because any increase in pH will reduce the selective activity of selenite. The fact that Proteus and Pseudomonas species do not ferment lactose may explain why they escape inhibition.

There have been many modifications and alterations to the original medium described by Leifson, including mannitol to replace lactose (Mannitol Selenite Broth CM0399), addition of cystine (Selenite Cystine Broth CM0699), brilliant green, sodium taurocholate, sulphapyridine and streptomycin. The performance of these modifications has been investigated but with no overall agreement⁷.

Technique
For routine purposes Selenite Broth cultures should be incubated at 35°C for 18 to 24 hours and then sub-cultured on any combination of greater and lesser inhibitory selective agars for Enterobacteriaceae. The development of Escherichia coli and Proteus species is not indefinitely retarded in selenite media. Where the initial proportion of these organisms is high, it is often advantageous to sub-culture on to the solid media after 6 hours as well as after 18 hours.

If a high proportion of debris is present, in the sample of material being examined, the selective powers of the selenite may be nullified. This is well established in the examination of faeces and egg powder. It is common practice to emulsify the specimen in sterile saline, allow the gross particles to settle, and inoculate the medium with the supernatant. An alternative method is as follows: Add 2 to 3g of solid specimen to 15ml of saline in a wide-necked 1oz. bottle, emulsify, separate the debris by slowly pressing a plug of cotton-wool down through the suspension. Withdraw approximately 1ml of the supernatant and inoculate 10 ml of Selenite Broth.

Harvey and co-workers^8,9 showed that incubation of the selenite broth at 43°C facilitated the isolation of Salmonella paratyphi B from faeces. They recommended the use of this principle for the examination of sewage and river water containing large numbers of other bacteria that preferred a lower temperature for growth. It was also suggested that the procedure was of value for all salmonellae except Salmonella typhi. For urines, the broth should be made double strength and inoculated with its own volume of the specimen.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared medium at 2-8°C away from light.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Light straw coloured solution

Quality control

* This organism is available as a Culti-Loop®

Precautions
Discard the prepared medium if large amounts of reduced selenite can be seen as a red precipitate in the bottom of the bottles.
Do not incubate longer than 24 hours because the inhibitory effect of selenite is reduced after 6-12 hours incubation ¹⁰.
Take sub-cultures of broth from the upper third of the broth column, which should be at least 5 cm in depth.

References
1. Robertson D. S. F. (1970) Lancet i. 518-519.
2. Klett A. (1900) Zeitsch. fÏr Hyg. und Infekt. 33. 137-160.
3. Guth F. (1916) Zbl. Bakt. I. Orig. 77. 487-496.
4. Leifson E. (1936) Amer. J. Hyg. 24. 423-432.
5. Weiss K. F., Ayres J. C. and Kraft A. A. (1965) J. Bact. 90. 857-862.
6. Rose M. J., Enriki N. K. and Alford J. A. (1971) J. Food Sci. 36. 590-593.
7. Fagerberg D. J. and Avens J. S. (1976) J. Milk Food Technol. 39. 628-646.
8. Harvey R. W. S. and Scott T. (1953) Mon. Bull. Min. Hlth. & PHLS. 12. 149-150.
9. Harvey R. W. S. and Price T. H. (1979) J. Appl. Bact. 46. 27-56.
10. Chattopadhyay W. and Pilford J. N. (1976) Med. Lab. Sci. 33. 191-194.