FRASER BROTH

INTENDED USE Fraser Broth is used for the selective and differential enrichment of Listeria monocytogenes in milk, in dairy products, and in other food products.

HISTORY The complete medium, studied by Fraser et al. in 1988, is a modification of the formulation of Donnelly and Baigent. The composition of the base is identical to that of UVM II Broth and was modified by the addition of lithium chloride as selective agent and of ferric ammonium citrate to visualize cultures that hydrolyze esculin, by the resultant blackening of the medium.

PRINCIPLES - The very good recovery of Listeria monocytogenes is assured by the concentration differences in nalidixic acid and acriflavine between Half-Fraser and Fraser, as well as the two enrichment steps themselves. Fraser broth corresponds to the secondary enrichment step. - Polypeptone, yeast extract and meat extract furnish the nutrients required for the growth of Listeria. - The high sodium chloride content increases the selectivity of the medium. - Phosphates buffer the pH of the medium. - Esculin is hydrolyzed by Listeria to glucose and esculetin, the latter compound forming a black complex with ferric ions supplied by ferric citrate, added just before use, which also favors the growth of Listeria - Lithium chloride inhibits the growth of most enterococci which can also hydrolyze esculin. - Nalidixic acid blocks the DNA replication of bacteria sensitive to this antibacterial agent. - The growth of secondary Gram-positive microflora is inhibited by acriflavine.

PREPARATION - Dissolve 55.0 g of the dehydrated base medium (BK115) or 55.0 g or dehydrated base medium Fraser Broth II (BK133) in 1 liter of distilled or deionized water. - Stir slowly until complete dissolution. - Dispense 10 mL in tubes. - Sterilize in an autoclave at 121C for 15 minutes.

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INSTRUCTIONS FOR USE - Cool the medium to 25C. - To each tube of Fraser broth base BK115(1), aseptically add 0.1 mL of a sterile 5% solution of ferric ammonium citrate : BS059 or BS062. - Using tubes prepared as above, or the ready-to-use tubes (BM013), transfer 0.1 mL of the culture obtained from the primary enrichment broth (Half-Fraser). - Mix well. - Incubate the tubes for 24 to 48 hours at 37C.
(1)

NOTE :

When using Fraser broth base II (BK133), aseptically add 0.1 mL if reconstituted Fraser Selective Supplement (BS031) to each tube of base media.

RESULTS Blackening of the cultures is an indication of the presumptive presence of Listeria. Some microbial strains hydrolyzing esculin (enterococci) may also cause the medium to blacken. Inoculate all tubes (with or without blackening) onto a selective medium : Oxford Agar (BK110 + BS003 or BM019), PALCAM Agar (BK145 + BS004/BS049 or BM020), COMPASS Listeria Agar (BM123 or BM124). Identify suspected colonies and subject them to biochemical identification tests. NOTE : A minimum period of 24 hours is necessary to permit the visualization of the black color.

TYPICAL COMPOSITION of the complete medium (can be adjusted to obtain optimal performance) For 1 liter of medium : - Polypeptone .................................................................................10.00 g - Yeast extract ..................................................................................5.00 g - Meat extract ...................................................................................5.00 g - Sodium chloride ...........................................................................20.00 g - Disodium phosphate, anhydrous....................................................9.60 g - Monopotassium phosphate ............................................................1.35 g - Esculin ...........................................................................................1.00 g - Lithium chloride ..............................................................................3.00 g - Nalidixic acid ...............................................................................20.0 mg - Acriflavin......................................................................................25.0 mg - Ferric ammonium citrate ................................................................0.50 g pH of the ready-to-use medium at 25C : 7.2 0.2.

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QUALITY CONTROL Dehydrated media : yellowish powder, free-flowing and homogeneous. Prepared media (complete) : amber yellow solution, with bluish tints. Typical culture response after 48 hours of incubation at 37C, followed by subculture on Oxford Agar : Microorganisms Listeria monocytogenes + Enterococcus faecalis + Escherichia coli Listeria monocytogenes + Enterococcus faecalis + Escherichia coli Enterococcus faecalis Escherichia coli CIP 59.53 ATCC 29212 ATCC 25922 CIP 78.31 ATCC 29212 ATCC 8739 ATCC 29212 ATCC 25922 Growth on Oxford agar 10 characteristic colonies

10 characteristic colonies

inhibited inhibited

STORAGE / SHELF LIFE Dehydrated base media (BK115, BK133) : 2-30C, shielded from light. - The expiration dates are indicated on the labels. Prepared medium from BK115 (benchmark value*) : - Base media : 15 days at 2-8C, shielded from light. - Complete media in tubes : 8 days at 2-8C, shielded from light. Prepared medium from BK133 (benchmark value*) : - Base media : 6 months at 2-8C, shielded from light. - Complete media in tubes : 8 days at 2-8C, shielded from light. Fraser Selective Supplement, Ferric Ammonium Citrate, sterile solution 5% : - Store between 2-8C, shielded from light. - The expiration dates are indicated on the labels. Ready-to-use media in tubes, - Store between 2-14C, shielded from light. - The expiration date is indicated on the label.

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PACKAGING Ready-to-use media in tubes : - 50 x 10 mL Dehydrated Fraser Broth base (without ferric ammonium citrate) : - 500 g bottle Ferric Ammonium Citrate, sterile solution 5% : - 10 x 90 mL vials - 7 x 10 mL tubes Dehydrated Fraser Broth base II (without ferric ammonium citrate, nalidixic acid or acriflavine) : - 500 g bottle - 5 kg drum Fraser Selective Supplement : - 10 vial pack

Code

BM01308

BK115HA BS05908 BS06208

BK133HA BK133GC BS03108

PHOTO SUPPORT Product reference : [BK115HA + BS05908 or BS06208] ; [BK133HA, GC + BS03108] or BM01308 Media used for : Selective and differential enrichment of Listeria monocytogenes in foods.

Negative control

Presumptive positive sample

Listeria monocytogenes Fraser broth Ref : BM01308 Incubation : 24 hours / 37C Characteristics : blackened tube (at right) a presumptive indication of Listeria monocytogenes growth (reduction of esculin), however confirmation is still required.

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BIBLIOGRAPHY
Donnelly, C.W., and Baigent, G.J. 1986. Method for flow cytometric detection of Listeria monocytogenes in milk. App. Environ. Microbiol., 52: 689-695. Fraser, J.A., and Sperber, W.H. 1988. Rapid detection of Listeria spp. in food and environmental samples by esculin hydrolysis. J. Food. Prot., 51: 762-765. NF EN ISO 11290-1 (V 08-028-1). Fvrier 1997. Microbiologie des aliments. Mthode horizontale pour la recherche et le dnombrement de Listeria monocytogenes. Partie 1 : Mthode de recherche. XP CEN ISO/TS 11133-2 (V 08-104-2). Janvier 2004. Microbiologie des aliments. Guide pour la prparation et la production des milieux de culture. Partie 2 : Guide gnral pour les essais de performance des milieux de culture. NF EN ISO 11290-1/A1 (V 08-028-1/A1). Fvrier 2005. Microbiologie des aliments. Mthode horizontale pour la recherche et le dnombrement de Listeria monocytogenes. Partie 1 : Mthode de recherche. Amendement 1 : Modification des milieux disolement, de la recherche de lhmolyse et introduction de donnes de fidlit.

*Benchmark value refers to the expected shelf life when prepared under standard laboratory conditions following manufacturers instructions. It is provided as a guide only and no warranty, implied or otherwise is associated with this information.

The information provided on the package take precedence over the formulations or instructions described in this document. The information and specifications contained in this technical data sheet date from 2009-02-17. They are susceptible to modification at any time, without warning. Code document : BK115/A/2003-01 : 7.

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