Selenite F Broth Pdf Download
Nichelle Gruger
Selenite broth is used as a selective medium for the isolation of Salmonella species.[1] Selenite broth was originated by Leifson,[2] while observing good recovery of Salmonella spp. and reduced growth of fecal coliforms. Selenite broth is used as a selective enrichment for the cultivation of Salmonellaspp. that may be present in small numbers and competing with intestinal flora. This medium must not be autoclaved. Once prepared, it is steamed at 100C for 30 minutes. There should be a very slight red precipitate. To minimize the risk of teratogenicity to workers, sodium selenite must be added separately to the medium. It has a pH of approximately 7.1. Selenite broth gives pale or colorless colonies.
Selenite F Broth is the medium used for the selective enrichment of Salmonella spp from both clinical and food samples. It is a buffered Lactose Peptone Broth to which Sodium Biselenite is added as the selective agent.
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.
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 components5,6. 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 35C 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-workers8,9 showed that incubation of the selenite broth at 43C 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.
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.
A medium for the selective enrichment of Salmonella spp from both clinical and food samples. It is a buffered Lactose Peptone Broth to which Sodium Biselenite is added as the selective agent. Subcultures should be made from the top 1/3 of the broth after not more than 24 hours incubation as after this time there is a loss of selectivity.
Description
This medium is similar to the modification of Leifson2 enrichment medium described by Hobbs & Allison3 for the isolation of Salmonella typhi and Salmonella paratyphi B. Liefson2 suggested that it is best to tube the medium to a depth of 2 inches (50mm) or more.
Hobbs & Allison3 compared two sets of selenite media, one containing lactose and the other mannitol. Of 38 positive stools Salmonella typhi. was sub-cultured from both media in 32 instances, from the mannitol selenite alone in 5 instances and from the lactose selenite alone once. Comparisons showed that the mannitol selenite broth was superior to three other liquid media in its selective value for Salmonella typhi and that it was as good as tetrathionate for the isolation of Salmonella paratyphi B.
Precautions
Observe the precautionary comments made about sodium biselenite in Selenite Broth Base CM0395.
Discard the prepared medium if large amounts of reduced selenite can be seen as a red precipitate in the bottom of the bottle.
Do not incubate longer than 24 hours because the inhibitory effect of selenite is reduced after 6-12 hours incubation.
Mannitol fermentation by salmonella helps correct the alkaline pH swing which can occur during incubation.
Take sub-cultures of broth from the upper third of the broth column, which should be at least 5 cm in depth.
Selenite Broth was devised by Leifson, who demonstrated that selenite was inhibitory for coliforms and certain other microbial species, such as fecal streptococci, present in fecal specimens and, thus, was beneficial in the recovery of Salmonella species. Selenite-F Broth is used as an enrichment medium buffered with Lactose Peptone Broth to which Sodium Biselenite is added as the selective agent for the isolation of Salmonella from feces, urine, water, foods, and other materials of sanitary importance.
Selenite Broth Base contains lactose as a source of fermentable carbohydrate which helps to prevent any rise in pH value during incubation, thereby preventing any loss of selective activity of selenite.
Klett1 first demonstrated the selective inhibitory effects of selenite, and Guth2 used it to isolate Salmonella typhi. It was 20 years later before Leifson3 fully investigated selenite and promoted wide use of the medium.
Proteus and Pseudomonas species appear to be resistant to its effects4. 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.
To minimise any possible risk of teratogenicity to laboratory workers, the sodium biselenite must be added as a solution to this medium. Robertson5 reported miscarriages and possible teratogenic effects on pregnant laboratory assistants which may have been caused by ingested sodium biselenite. This substance has, therefore, been removed from the Oxoid powdered medium.
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The relative effectiveness of Rappaport-Vassiliadis (RV) medium, selenite cystine (SC) broth, and tetrathionate (TT) broth for the recovery of Salmonella spp. from foods with a low microbial load was determined. RV medium made from its individual ingredients and incubated at 42 degrees C was compared with a commercial preparation of SC broth, incubated at 35 degrees C, and TT broth incubated at 35 and 43 degrees C, for the recovery of Salmonella spp. Twenty-one artificially contaminated food types that included dairy foods, spices, and egg products, as well as other low-microbial-load foods, were analyzed. The foods were inoculated with single Salmonella serovars at target levels ranging from 0.04 to 0.4 CFU/g. No significant differences (P< or =0.05) among the selective enrichment broths for the recovery of Salmonella spp. from 18 of the foods were observed. Significantly fewer Salmonella-positive test portions of gelatin, guar gum, and nonfat dry milk were recovered with RV medium than with SC broth incubated at 35 degrees C and TT broth incubated at 35 and 43 degrees C. TT broth incubated at 35 degrees C recovered the greatest number of Salmonella-positive test portions. For the recovery of Salmonella spp. from foods with a low microbial load, it is recommended that TT broth incubated at 35 degrees C and RV medium incubated at 42 degrees C be used.
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