Part of Thermo Fisher Scientific
Organisms this product works with:
Other products used in the isolation of Cronobacter:
ENTEROBACTER SAKAZAKII ISOLATION AGAR
CODE: CM1134
A chromogenic medium for the detection and enumeration of Cronobacter (formerly E. sakazakii) in milk and milk samples as described in ISO TS/22964:20061. To be used in conjunction with mLST/vancomycin medium (CM1133 & SR0247E)
Typical Formula* |
gm/litre |
Pancreatic digest of casein† |
7.0 |
Yeast extract |
3.0 |
Sodium chloride |
5.0 |
Sodium deoxycholate†† |
0.6 |
5-Bromo-4-chloro-3-indolyl α-D-glucopyranoside |
0.15 |
Crystal violet |
0.002 |
Agar |
12.0 |
pH 7.0 ± 0.2 @ 25°C |
Directions
Suspend 27.75g in 1 litre of distilled water. Bring to the boil to dissolve completely. Sterilize by autoclaving at 121°C for 15 minutes. Cool to approximately 50°C. Mix well and pour into sterile Petri dishes.
Description
Cronobacter (formerly Enterobacter sakazakii) are Gram-negative rod-shaped members of the Family Enterobacteriaceae that have been implicated in outbreaks of disease in premature infants (neonates), causing sepsis, meningitis and necrotising enterocolitis. Neurological damage can be permanent, and the death rate is reported to be as high as 40-80%1.
Cronobacter spp. have been isolated at low levels from powdered infant formulae, and the organisms’ high tolerance to desiccation provides a competitive advantage in the dry environments of milk powder factories, increasing the risk of product contamination2.
In response to concerns over this pathogen, a collaborative technical specification for the detection of Cronobacter in milk and milk products was jointly issued by ISO and the IDF, as ISO/TS22964:20063. This recommended pre-enrichment in BPW, selective enrichment in mLST/vancomycin broth and plating on Enterobacter sakazakii Isolation Agar. Positive colonies are confirmed on the basis of sugar utilisation, decarboxylase activity, as well as yellow pigmentation.
Unlike most other members of the Enterobacteriaceae, Cronobacter possess the enzyme α-glucosidase, which is used as a diagnostic feature in chromogenic medium. Enterobacter sakazakii Isolation Agar contains a substrate for this enzyme, 5-bromo-4-chloro-3-indolyl α-D-glucopyranoside (X-α-gluc). Cronobacter hydrolyse this colourless chromogen to produce characteristic blue-green colonies for identification on the plate. Sodium desoxycholate and crystal violet are also incorporated in the medium to inhibit the growth of Gram-positive organisms.
The European Union has directly specified limits for the levels of Cronobacter in dried infant formulae, dried dietary foods for special medical purposes and food intended for infants below six months of age, in Commission Regulation no 2073/20054. This regulation states for products placed on the market, during their shelf life, 30 samples of 10g have to be tested, and the food producer has to demonstrate absence of Cronobacter in each of these samples, using ISO/TS 22964:20063.
Technique
Refer to the relevant standard method for detailed instructions. The following method is a summary of ISO/TS 22964:20063
Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
The prepared medium is best used freshly prepared, but may be suitable for longer storage following in-house validation.
Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Purple coloured gel
Quality Control
Positive Controls |
Expected results
|
Cronobacter sakazakii ATCC® 12868 |
1-2mm blue-green colonies
|
Cronobacter sakazakii ATCC® 29004 |
1-2mm blue-green colonies |
Cronobacter muytjensii ATCC® 51329* |
1-2mm blue-green colonies |
Negative Controls |
|
Escherichia coli ATCC® 25922* |
0.25-1.5mm purple colonies |
Staphylococcus aureus ATCC® 25923* |
Inhibited or no growth |
References