Deoxycholate Citrate Agar is an alteration of Leifson formula that is suggested for the identification of Salmonella as well as Shigella spp. It is comparable to deoxycholate agar however it is slightly more selective for enteric pathogens due to higher levels of both deoxycholate and citrate salts. The sodium deoxycholate pH range of 7.3 to 7.5 inhibits gram-positive bacteria. Citrate salts in the amount contained into the composition, act as inhibitors to gram-positive bacteria, as well as other intestinal organisms that are normal. This makes it an effective and selective media, commonly used to isolate intestinal pathogens.
Principle of Deoxycholate Citrate Agar (DCA)
HI solids are a source of nitrogen and carbon and can inhibit the growth of bacteria known as coliforms. Proteose Peptone supplies carbon nitrogen, vitamins, and minerals. Gram-positive and gram-negative bacteria are suppressed or diminished by the sodium deoxycholate and sodium citrate ferric ammonium citrate, and sodium deoxycholate. Dipotassiumphosphate buffers the media. Lactose assists in the differentiation of enteric bacilli as fermenters of lactose produce red colonies whereas non-fermenters create non-coloured colonies. Coliform bacteria, when present create pink colonies on the medium. The degrading of lactose results in acidsification to the environment around the colonies, and the pH indicator neutral red alters its color from neutral red to red. The colonies are typically covered by a turbid area of deoxycholic acid precipitated due to the acidification within the medium.
Sodium deoxycholate is a mixture of neutral red within an acidic setting and causes the color to be removed from the solution and then formation of the deoxycholate. Reduced ferric ammonium-citrate into iron sulfide is evident by creation of black iron sulfur. Salmonella or Shigella species are not fermenting lactose, however Salmonella can produce H2S, creating colorless colonies with black centers or without. The iron and the citrate (Fe) combination produces strong hydrolyzing effects on agar once you heat the medium, resulting in an unlastic and soft agar.
Composition of Deoxycholate Citrate Agar (DCA)
|Ferric ammonium citrate||2.000|
Final pH (at 25°C) 7.5±0.2
Preparation and Method of Use of Deoxycholate Citrate Agar (DCA)
- Do not suspend 70.52 grams in 1000ml of distilled water.
- Then heat to boiling, allowing the medium until the medium is completely dissolving.
- Cool to 45-50degC. Mix well before pouring into sterilized Petri plates.
- Dry the surface of the agar before using.
- Inoculate the medium extensively using feces or rectal swabs dispersing a portion of the inoculum that was originally used to collect well-separated colonies on one portion of the plates.
- Incubate for 18-24hrs at 35°C.
- If the organisms are late-developers or if there are no non-lactose fermenters are detected, allow to allow to incubate another 24 hours.
Note: DO NOT AUTOCLAVE. Avoid overheating because it can be harmful on the material.
Result Interpretation on Deoxycholate Citrate Agar (DCA)
Lactose non-fermenters create transparent colored, lighter pink, tan or light-colored colonies that have or with black centers.
Lactose fermenters create the red colony either with or without the precipitate of bile.
|Escherichia coli||Poor growth; pink with bile precipitate negative reaction for H2S|
|Salmonella Enteritidis||Good-luxuriant growth; colorless; positive reaction for H2S, black centered colonies|
|Salmonella Typhimurium||Good-luxuriant growth; colorless; positive reaction for H2S, black centered colonies|
|Shigella flexneri||Good growth; colorless|
|Salmonella Abony||Good-luxuriant growth; colorless; positive reaction for H2S, black centered colonies|
|Shigella sonnei||Colonies are smooth and initially colorless, becoming pale pink on further incubation due to late lactose fermentation|
|Enterobacter/Klebsiella spp.||Large, pale mucoid colonies with the pink center|
Uses of Deoxycholate Citrate Agar (DCA)
- Deoxycholate Citrate Agar is utilized for the identification and differentiating Gram-negative enteric bacteria in a laboratory.
- It is especially useful to identify organisms that cause bacillary dysentery. Salmonella strains which cause food poisoning as well as Salmonella Paratyphi.
Limitations of Deoxycholate Citrate Agar (DCA)
- The pathogens that are suspected of being pathogenic should be subcultured in the less inhibiting medium prior to the identification.
- Since there are a variety of bacteria that look similar to Salmonella on DCA It is generally suggested that more specific Agars be used for detection of Salmonella specifically xylose lysine deoxycholate (XLD) the agar.
- The growth medium is sensitive to heat and must be poured and chilled as soon as it is feasible following the addition of deoxycholate. Otherwise it can become extremely delicate and difficult to work with.
- For routine testing of urine and stool samples It is recommended to use other media, such as MacConkey Agar and Bismuth Sulphite Agar and so on. should be utilized to complement this medium.
- A further biochemical identification is needed to verify the species.
- Due to the variations in nutrition Certain organisms could have a slow growth rate.