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Microbiological test with spores of Geobacillus stearothermophilus for the detection of antibiotic and sulphamide residues in meat DESCRIPTION
Antimicrobial substances are given to cattle for therapeutic treatment of infections and so they can be present in meat as residual drugs. The presence
of residual antimicrobial drugs in meat is a potential hazard for the consumers since those substances interfere with the intestinal flora, can cause
allergic reactions and can contribute to the spreading of resistant microorganisms.
The detection of inhibiting substances in meat is ruled in Europe by Regulation EEC 2377/90 and following modifications.
MeRA (Meat Rapid Antibiotic) Test is a microbiological test, set for the detection of residual antibiotics and sulphamides in meat, that uses spores of
Geobacillus stearothermophilus and a culture medium designed for that purpose.
CONTENT OF THE PACKAGE
Each package contains:
n. 40 vials containing a freeze-dried culture medium n. 40 discs containing spores of Geobacillus stearothermophilus PRINCIPLE OF THE METHOD
The test detects the presence of antibiotics and sulphamides through their inhibition action against the growth in vitro, at 64 ± 0,5 °C, of Geobacillus
stearothermophilus. The inhibition of bacterial growth is made evident by the presence of a fermentable sugar and a pH indicator in the culture medium.
PROCEDURE FOR USE
Weigh about g 50 of meat. Cut up the tissue and mince further by electric mixer.
Weigh g 2 of minced meat and introduce it in a 10 mL tube. Add mL 6 of distilled water to the tube (ratio meat:water 1:3). Make sure meat extract pH is 5.6 roughly.
Homogenize for some seconds through homogenizer set at 13000 rpm.
Centrifuge the homogenized meat by tabletop centrifuge set at 4000 rpm for 15 minutes.
Remove the cup from the vial and add 1 disc of spores and mL 2 of sterile distilled water. Shake gently to aid the dissolution of the freeze-dried medium.
Put the cup and introduce the vial in a waterbath at 64 ± 0,5 °C or in a Termoblock set at 64 ± 0,5 °C. Incubate for 20’.
Remove the vial from the and let it reach room temperature (roughly 5').
Remove the cup and introduce 1 mL of the milk under test.
Put the cup to the vial and let the antibiotic/sulphamide, if present, act at room temperature for 20’.
Reintroduce the vial in the waterbath or in the Termoblock at 64 ± 0,5 °C for the second incubation for 3h - 3h 30’.
INTERPRETATION OF THE RESULTS
At the end of the second incubation observe colour of the liquid in the vials that, if green, indicates antibiotic/sulphamide residues in the examined meat
in concentrations above the detection limits; otherwise, a yellow colour indicates the absence of antibiotic/sulphamide residues in the examined meat or
their presence in concentrations under the detection limits.
Photo 1. Results obtained by MeRA Test after 3h 30' of incubation at 64 ± 0,5 °C:
in the left side, no colour change of the pH indicator: analyte-positive meat sample containing antibiotic/sulphamide residues in concentration above the detection limits; in the right side, colour change of the pH indicator: analyte-negative meat sample containing no antibiotic/sulphamide residues or containing antibiotic/sulphamide residues in concentration under the detection limits.
USER QUALITY CONTROL
Every batch of MeRA Test is submitted to quality control. The analyte-negative control is obtained using a certified biological meat sample. The analyte-
positive controls are obtained using meat samples with the addition of antibiotics/sulphamides such as ampicillin (50 g
The product MeRA Test is not classified as dangerous under current legislation. MeRA Test is a disposable device to be utilized only for in vitro use, it
is intended for use in a professional environment and must be used in laboratory by properly trained personnel, using approved asepsis and safety
methods for handling pathogenic agents.
Store MeRA Test at 2-8 °C in its original packaging. Keep away from heat sources and avoid excessive temperature changes.
In such conditions MeRA Test will remain valid until the expiry date indicated on the label. Do not use beyond that date.
Eliminate without using it if there are deterioration signs.
After use, before disposal, vials, must be properly decontaminated with chemical substances or autoclaved.
DM 26/03/1992 GU n° 90 16/04/1992: Attuazione della decisione n. 91/180/CEE concernente la fissazione di metodi di analisi e prova relativi al latte crudo e al latte trattato termicamente.
CONALAnalisi e controllo qualità dei prodotti lattiero-caseari Il latte VIII/5, 356-357 (1983).
91/180/EEC: Commission Decision of 14 February 1991 laying down certain methods of analysis and testing of raw milk and heat-treated milk. Official Journal L 093 , 13/04/1991 P. 0001 - 0048.
Van Oss J.L., Lameris S.A. Doodewaard J., Oostendrop J.G., Diffusion Test for the determination of antibiotic residues in milk. Neth. Milk Dairy J. 29, 16 (1975).
International Dairy Federation International Standard FIL-IDF 57:1970 Detection of penicillin in milk by a disk assay technique (1970).
Deborah Currie, Lorraine Lynas, D. Glenn Kennedy and W. John McCaughey. Evaluation of a modified EC Four Plate.
Method to detect antimicrobial drugs. Food Additives and Contaminants, 1998, Vol. 15, No. 6, 651-660.
Regulation 2377/90 ff EEC and 1/12/2000 update.
Table 1: Sensitivity of MeRA Test to some antibiotics/sulphamides, commonly used in veterinary medicine and respective MRL (Maximum Residue
Limits) values valid for Europe1.
MRL for bovine muscle1
MeRA Test Sensitivity in 3h 30'
1 Regulation 2377/90 ff EEC and 1/12/2000 update.
TABLE OF SYMBOLS
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