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Material Safety Data Sheet

Required Products

Organisms

Organisms in the Clinical sector this product works with:

Dehydrated Culture Media

Sector: Clinical

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VRE BROTH BASE

Code: CM0984

Selective medias for the isolation of Vancomycin Resistant Enterococci (VRE) and High Level Aminoglycoside Resistant Enterococci (HLARE) from clinical samples.

Typical Formula*

gm/litre

Calf brain infusion solids

12.5

Beef heart infusion solids

5.0

Proteose peptone

10.0

Glucose

2.0

Sodium chloride

5.0

Disodium phosphate

2.5

pH 7.4 ± 0.2 @ 25°C

 
* Adjusted as required to meet performance standards 

VRE AGAR BASE

Code: CM0985

Typical Formula*

gm/litre

Tryptone

20.0

Yeast extract

5.0

Sodium chloride

5.0

Sodium citrate

1.0

Aesculin

1.0

Ferric ammonium citrate

0.5

Sodium azide

0.15

Agar

10.0

pH 7.0 ± 0.2 @ 25°C

 
* Adjusted as required to meet performance standards 

MEROPENEM SUPPLEMENT

Code: SR0184

Vial Contents (each vial is sufficient for 500ml of medium)
per vial
per litre

Meropenem

1.0mg

2.0mg

GENTAMICIN SUPPLEMENT

Code: SR0185

Vial Contents(Each vial is sufficient to supplement 500ml of medium)
per vial
per litre

Gentamicin

256.0mg

512.0mg

VANCOMYCIN SUPPLEMENT

Code: SR0186

Vial Contents(Each vial is sufficient to supplement 500ml of medium)
per vial
per litre

Vancomycin

3.0mg

6.0mg

Directions
VRE Broth
Suspend 37.0g of VRE Broth Base in 1 litre of distilled water. Warm to dissolve completely, sterilise by autoclaving at 121°C for 15 minutes and cool to 50°C. Supplement the medium as shown in the table below. Then mix well and distribute into final sterile containers.
VRE Agar
Suspend 42.6g of VRE Agar Base in 1 litre of distilled water. Warm to dissolve completely, sterilise by autoclaving at 121°C for 15 minutes and cool to 50°C. Supplement the medium as shown in the table below. Then Mix agar well and distribute into sterile Petri dishes.

Supplement

(mg per vial )

Reconstitution Volume

(Sterile distilled water)

VRE Agar Base

(per litre)

VRE Broth

(per litre)

VRE’s

HLARE’s

Meropenem (SR0184)
1mg

2ml

1 vial

-

2 vials

Gentamicin
(SR0185)
256mg

3ml

-

2 vials

-

Vancomycin (SR0186)
3mg

2ml

2 vials

-

-

Description
Selective media for the isolation of Vancomycin Resistant Enterococci (VRE) and High Level Aminoglycoside Resistant Enterococci (HLARE) from clinical samples. NB Enterococci containing the Van C genes will not be isolated on this medium.

The proliferation of enterococci, resistant to many commonly used antimicrobials is on the increase1. The recent emergence of VRE is of great concern as enterococci can cause bacteraemia, endocarditis and urinary tract infections. The use of VRE Broth Base and VRE Agar Base complies with recommendations from the Centre for Disease Control and Prevention (CDC) to detect VRE infection in its early stages2.
Resistant enterococci can be isolated either directly by inoculation of the clinical sample onto supplemented VRE Agar, or indirectly isolated with a selective enrichment through VRE Broth followed by inoculation onto supplemented VRE Agar. VRE Agar Base contains an indicator system to detect the growth of aesculin-hydrolysing organisms. Enterococci produce black zones around the colonies from the formation of black iron phenolic compounds derived from aesculin-hydrolyis products and ferrous iron.

Oxoid has developed three antibiotic supplements to selectively isolate antibiotic resistant populations amongst pathogenic Enterococci:
Meropenem Supplement SR0184 is used at 2mg/l in VRE Broth Base , and 1mg/l in VRE Agar Base for the suppression of contaminating flora, particularly Gram-negatives and Enterococcus gallinarum. It has been reported that some Enterococcus faecalis can be sensitive to meropenem. To isolate these strains the level of meropenem may need to be reduced, or the supplement omitted from the formulation.
Gentamicin Supplement SR0185 is used at 512mg/l4 in VRE Agar Base for the selective isolation of HLARE.
Vancomycin Supplement SR0186 is used at 6mg/l in VRE Agar Base for the selective isolation of VRE.

Technique
Broth
Add ‘pea sized’ amounts of faecal samples directly to the supplemented VRE Broth and vortex to ensure emulsification. Incubate at 37°C for a minimum of 18 hours and subculture onto VRE Agar.
Agar
Inoculate faecal samples or VRE Broth culture onto supplemented VRE Agar plates with a sterile swab and spread with a loop using the diminishing sweep technique. Incubate at 37°C for 48 hours and examine at 24 hours. Re-incubate negative plates for a further 24 hours.
Enterococci appear as round grey / pale brown colonies about 1mm in diameter surrounded by black zones.

Storage conditions and Shelf life
VRE Broth Base and VRE Agar Base should be stored tightly capped in the original container at 10-30°C. Meropenem Supplement SR0184, Gentamicin Supplement SR0185 and Vancomycin Supplement SR0186 should be stored in the dark at 2-8°C. When stored as directed, the media will remain stable until the stated expiry date. Prepared medium may be stored for up to two weeks at 2-8°C in the dark.

Appearance
Dehydrated medium: Broth: Cream coloured, free-flowing powder. Agar: Cream coloured, free-flowing powder
Prepared medium: Broth: Straw coloured solution. Agar: Straw green coloured gel with blue hue

Quality control
VRE Broth

Positive control:

 Expected results

Enterococcus faecalis NCTC 12201

Growth

Negative control:

 

Escherichia coli ATCC® 25922*

Inhibited

* This organism is available as a Culti-loop®

VRE Agar

Positive control:

 Expected results

Enterococcus faecalis NCTC 12201

Growth

Negative controls:

 
Enterococcus faecalis ATCC® 33186Inhibited

Escherichia coli ATCC® 25922*

Inhibited

* This organism is available as a Culti-loop®

HLARE Agar ( CM0895 + SR0185)

Positive control:

 Expected results

Enterococcus faecalis ATCC® 51299*

Growth

Negative control:

 

Enterococcus faecalis ATCC® 29212*

Inhibited

* This organism is available as a Culti-loop®

Precautions
VRE Broth Base, VRE Agar Base , Meropenem Supplement SR0184, Gentamicin Supplement SR0185 and Vancomycin Supplement SR0186 should only be used for in vitro diagnostic purposes. Do not use any of these beyond expiry date or if the products show any signs of deterioration.
Warning: Sodium azide is harmful if swallowed, please take all necessary precautions.

References
1. King, W. K. (1996) Bug Bytes Vol. 2 No. 19.
2. CDC Preventing the spread of vancomycin resistance: a report from the Hospital Infection Control Practices Advisory
    Committee (1994). Fed Regist. May 17.
3. Gold, H. S. & Moellering R.C. Jr. (1996) N. Engl. J. Med.; 335(19): 1445-53.
4. Weinbren, M. J., Johnson, A.P. & Woodford, N. (2000) J. Antimicrobial Chemotherapy; 45 :404-405.

 
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