Selenite F Broth- Composition, Principle, Preparation, Results, Uses
Selenite F Broth is a type of enrichment medium that is used for the isolation and cultivation of Salmonella spp. from various samples, such as feces, urine, water, foods and other materials of sanitary importance. It is also useful for the detection of some species of Shigella and the transport of strains of Vibrio cholerae.
Selenite F 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. He found that the inhibited strains would eventually breakthrough, but if subcultures were made from the enrichment broth after 8–12 h incubation, the isolation of Salmonella was possible without overwhelming growth of many members of the intestinal flora.
Selenite F Broth is a buffered Lactose Peptone Broth to which Sodium Biselenite is added as the selective agent. The F in Selenite F Broth stands for Faeces, because it is mostly used for the isolation of Salmonella and Shigella from faecal specimens.
Selenite F Broth works by inhibiting many species of gram-positive and gram-negative bacteria, including enterococci and coliforms, with sodium selenite, while allowing the growth of Salmonella and some Shigella species. The medium also maintains a neutral pH with lactose fermentation and sodium phosphate buffering, which reduces the toxicity of selenite.
Selenite F Broth is an important tool for the diagnosis and surveillance of salmonellosis, shigellosis and cholera, which are serious infectious diseases that can cause diarrhea, fever, abdominal cramps and dehydration. These diseases can be transmitted through contaminated food or water, or through contact with infected animals or people. Therefore, it is essential to identify and isolate the causative agents as quickly and accurately as possible. Selenite F Broth can help achieve this goal by enriching the target organisms and suppressing the background flora.
Selenite F Broth is a liquid medium that contains the following ingredients per liter of distilled water:
- Casein enzymic hydrolysate: 5 g
- Lactose: 4 g
- Sodium phosphate: 10 g
- Sodium hydrogen selenite (Sodium biselenite): 4 g
The pH of the medium is adjusted to 7.1 ± 0.2 at 25°C.
The casein enzymic hydrolysate provides nitrogenous substances, carbon compounds, and vitamin sources for the growth of bacteria. The lactose is a carbohydrate source that maintains the pH of the medium and allows the differentiation of lactose-fermenting and non-fermenting bacteria. The sodium phosphate buffers the medium and also reduces the toxicity of selenite. The sodium hydrogen selenite is the selective agent that inhibits many gram-positive and gram-negative bacteria, including enterococci and coliforms, while allowing the growth of Salmonella and some Shigella species.
The medium is prepared by dissolving 4 g of sodium selenite powder in distilled water, then adding the remaining 19 g of powder and bringing the volume to 1 liter. The medium is gently heated and brought to boiling, then dispensed into sterile test tubes and sterilized in a boiling water bath or free-flowing steam for 10 minutes. The medium should not be autoclaved as this may destroy the selenite. The medium should be cooled to room temperature before use.
Sodium hydrogen selenite is a very toxic, corrosive agent and causes teratogenicity. It should be handled with great care and washed off immediately with plenty of water if it comes in contact with skin.
Selenite F Broth is based on the selective inhibition of many bacteria by sodium selenite, which is a toxic and corrosive agent. Sodium selenite inhibits the growth of gram-positive and gram-negative bacteria, including enterococci and coliforms, by interfering with their cellular metabolism. However, some bacteria, such as Salmonella and Shigella, can tolerate sodium selenite and grow in the medium.
The medium also contains casein enzymic hydrolysate, which provides nitrogenous substances, carbon compounds and vitamin sources for bacterial growth. Lactose is a carbohydrate source that maintains the pH of the medium by producing acid when fermented by some bacteria. Sodium phosphate buffers the medium and reduces the toxicity of selenite by maintaining a stable pH.
The principle of Selenite F Broth is that selenite is reduced by bacterial growth and alkali is produced, which increases the pH and lessens the toxicity of selenite. This allows the overgrowth of other bacteria that are not inhibited by selenite. However, the acid produced by lactose fermentation serves to counteract the alkali production and maintain a neutral pH. This creates a balance between the selective and differential properties of the medium.
The medium is used as an enrichment broth for the isolation of Salmonella and Shigella from fecal specimens and other materials. The broth should not be incubated longer than 24 hours as the inhibitory effect of selenite decreases after 6-12 hours of incubation. The broth should also be subcultured onto selective and non-selective plating media to increase the probability of isolating pathogens.
Selenite F Broth is a liquid medium that can be prepared from a dehydrated powder or from individual ingredients. The following steps describe how to prepare the medium from the powder:
- Weigh 23 g of the dehydrated powder and dissolve it in 1 L of distilled or deionized water. The powder contains sodium selenite, sodium phosphate, lactose, and casein enzymic hydrolysate.
- Heat the solution gently and bring it to boiling. This will help to dissolve the powder completely and sterilize the medium.
- Dispense the medium into sterile test tubes, at least 5 cm deep. The volume of each tube depends on the size of the inoculum and the incubation time. Usually, 5 to 10 mL per tube is sufficient.
- Sterilize the tubes in a boiling water bath or free-flowing steam for 10 minutes. Do not autoclave the medium, as this will destroy the selenite and reduce its inhibitory effect.
- Cool the tubes to room temperature before use. Store them in a dark place at 2 to 8°C until needed.
Alternatively, you can prepare the medium from individual ingredients as follows:
- Weigh 4 g of sodium selenite and dissolve it in 100 mL of distilled or deionized water. This is the selenite solution.
- Weigh 19 g of sodium phosphate dibasic and dissolve it in 900 mL of distilled or deionized water. This is the phosphate solution.
- Weigh 4 g of lactose and dissolve it in the phosphate solution. This is the lactose-phosphate solution.
- Weigh 10 g of casein enzymic hydrolysate and dissolve it in the lactose-phosphate solution. This is the final solution.
- Adjust the pH of the final solution to 7.1 ± 0.2 at 25°C using hydrochloric acid or sodium hydroxide if needed.
- Follow steps 2 to 5 as described above for preparing the medium from the powder.
The prepared medium should have a clear to slightly opalescent appearance and a light amber color. It should be used within one month after preparation.
After incubation, the growth of organisms is indicated by turbidity in the medium. The broth can be subcultured on a selective or differential agar, such as MacConkey Agar, to obtain isolated colonies for further identification. The appearance of the colonies on the agar can help to differentiate Salmonella and Shigella from other bacteria.
- Positive Result: Colorless, good growth. Examples: Salmonella Typhimurium, Shigella sonnei, Salmonella enteritidis, etc. These organisms are able to grow in the presence of selenite and do not ferment lactose on MacConkey Agar.
- Negative Result: Pink with bile precipitate, inhibited or no growth. Examples: Escherichia coli, Enterococcus faecalis, etc. These organisms are either inhibited by selenite or ferment lactose on MacConkey Agar.
Selenite F Broth has several applications in microbiology, especially for the detection and isolation of Salmonella and Shigella species from various sources. Some of the common uses of Selenite F Broth are:
- It is used as a selective enrichment medium for the cultivation of Salmonella spp. from clinical specimens such as feces, urine, blood, and other body fluids, as well as from food samples such as meat, poultry, eggs, dairy products, and spices. Selenite F Broth enhances the growth of Salmonella by inhibiting the competing flora and reducing the toxicity of selenite.
- It is also used as an enrichment medium for the isolation of some species of Shigella from fecal specimens. Shigella species are usually present in low numbers and may be overgrown by other bacteria in primary isolation media. Selenite F Broth allows the growth of Shigella by reducing the pH and inhibiting the coliforms and enterococci.
- The broth is also recommended for the transport of strains of Vibrio cholerae, because these organisms can survive 2 to 5 days in Sodium Selenite Broth. Vibrio cholerae is the causative agent of cholera, a severe diarrheal disease that can be fatal if untreated. Selenite F Broth preserves the viability of Vibrio cholerae during transport from remote areas to laboratories for identification and confirmation.
Selenite F Broth is a useful medium for the enrichment and isolation of Salmonella, Shigella, and Vibrio cholerae from various sources. However, it should not be used as the sole isolation medium, but rather in conjunction with selective and non-selective plating media to increase the probability of isolating pathogens.
- Sodium hydrogen selenite (Sodium biselenite) is a very toxic, corrosive agent and causes teratogenicity. It can cause severe skin burns, eye damage, respiratory irritation, and reproductive harm. Handle it with great care and wear appropriate personal protective equipment. If there is contact with skin, eyes, or clothing, wash immediately with plenty of water and seek medical attention if needed.
- Enrichment broths should not be used as the sole isolation medium. They are to be used in conjunction with selective and non-selective plating media to increase the probability of isolating pathogens, especially when they may be present in small numbers. Some strains of Salmonella and Shigella may not grow well or at all in Selenite F Broth.
- The inhibitory effect of selenite decreases after 6 – 12 hours of incubation. Therefore, do not incubate the broth longer than 24 hours as it may result in overgrowth of other bacteria and loss of selectivity. Subcultures should be made from the enrichment broth after 8–12 h incubation for optimal isolation of Salmonella and Shigella.
- Selenite F Broth may not be suitable for the isolation of some rare or atypical serotypes of Salmonella and Shigella that are more sensitive to selenite or less able to reduce it. For example, Selenite F Broth may inhibit Salmonella Paratyphi A and some strains of Shigella flexneri. Alternative or supplementary enrichment media may be required for these organisms.
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