ISO 5552:1979

Meat and meat products — Detection and enumeration of Enterobacteriaceae (Reference methods)

Viandes et produits à base de viande — Recherche et dénombrement des Enterobacteriaceae (Méthodes de référence)

General Information

• Status: Withdrawn
• Publication Date: 31-Mar-1979
• Withdrawal Date: 31-Mar-1979
• ICS: 07.100.30 - Food microbiology
• Technical Committee: ISO/TC 34/SC 6 - Meat, poultry, fish, eggs and their products
• Drafting Committee: ISO/TC 34/SC 6 - Meat, poultry, fish, eggs and their products
• Current Stage: 9599 - Withdrawal of International Standard
• Completion Date: 04-Dec-1997

Relations

• Revised: ISO 5552:1997 - Meat and meat products — Detection and enumeration of Enterobacteriaceae without resuscitation — MPN technique and colony-count technique
• Effective Date: 15-Apr-2008

Standards Content (Sample)

INTERNATIONAL STANDARD @ ’a!!@ 5552
INTERNATIONAL ORGANIZATION FOR STANDARDIZATION*MEKJY”APO~HAfl OPrAHM3AUMfl no CTAH/lAPTM3AUMlrlYRGANlSATiON INTERNATIONALE DE NORMALISATION
-
Meat and meat products - Detection and enumeration
of Enterobacteriaceae (Reference methods)
Viandes et produits à base de viande - Recherche et dénombrement des Enterobacteriaceae (Méthodes de référence)
First edition - 1979-04-01
-
UDC 637.5 : 576.8.08 : 576.851
Ref. No. IS0 5552-1979 (E)
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2 Descriptors : meat, meat products, microbiological analysis, detection, counting, bacteria, Enterobacteriaceae.
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Price based on 6 pages

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FOREWORD
IS0 (the international Organization for Standardization) is a worldwide federation
of national standards institutes (IS0 member bodies). The work of developing
International Standards is carried out through IS0 technical committees. Every
member body interested in a subject for which a technical committee has been set
up has the right to be represented on that committee. international organizations,
governmental and non-governmental, in liaison with ISO, also take part in the work.
Draft International Standards adopted by the technical committees are circulated
to the member bodies for approval before their acceptance as International
Standards by the IS0 Council.
International Standard IS0 5552 was developed by Technical Committee
34, Agricultural food products, and was circulated to the member bodies
ISO/TC
1977.
in August
It has been approved by the member bodies of the following countries :
Canada Israel Portugal
Czechoslovakia Kenya Romania
Egypt, Arab Rep. of Malaysia South Africa, Rep. of
Ethiopia Mexico Spain
France Netherlands Thailand
Germany, F. R. New Zealand Turkey
Hungary Peru
India Ph il imines
k
i ran poiaid
The member bodies of the following countries expressed disapproval of the
document on technical grounds :
I
Austria
Bulgaria
4
O International Organization for Standardization, 1979
Printed in Switzerland

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INTERNATIONAL STA. JARD IS0 5552-1979 (E)/ERRATUM
Published 1979-06-01
c
INTERNATIONAL ORGANIZATION FOR STANDARDIZATION .MEXnYHAPODHAR OPrAHMJAUMR Il0 CTAHnAPTM3AUHM .ORGANISATION INTERNATIONALE DE NORMALISATIO
Meat and meat products - Detection and enumeration of Enterobacteriaceae (Reference methods)
ERRATUM
MODIFICATION TO FOREWORD (hide from cover)
The IS0 member body for the United Kingdom has now disapproved this International Standard. The United Kingdom
should therefore be included in the list of countries whose member bodies have disapproved the document.

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IS0 5552-1979 (E)
Meat and meat products - Detection and enumeration
of Enterobacteriaceae (Reference methods)
Incubation of the tubes at 37 "C for 24 h, followed by
1 SCOPE
streaking of the cultures onto violet red bile glucose agar.
This International Standard specifies reference methods
After incubation of the streaked agar plates at 37 "C for
for the detection and enumeration of viable Entero-
24 h, subjection of suspected colonies to biochemical
bacteriaceae in meat and meat products.
lu confirmation tests.
2 FIELD OF APPLICATION
5.3 Enumeration of Enterobacteriaceae
The methods can be applied to all kinds of meat and meat
5.3.1 Most probable number (MPN) technique - where
products but are not intended for surface count of
the number is expected to be in the range of 1 to 1 O00 per
carcasses.
gram of meat or meat product
Introduction of 1 ml of the macerate and of the dilutions
3 REFERENCE
in triplicate, into tubes containing the same
and
IS0 31 00, Meat and meat products - Sampling.
selective enrichment broth as mentioned in 5.2. Incubation
of the tubes at 37°C for 24 h. From the number of
confirmed positive tubes (see 5.2), determination of the
4 DEFINITIONS
most probable number of Enterobacteriaceae per gram
4.1 Enterobacteriaceae : Micro-organisms which ferment
of sample by using the MPN table (see annex).
glucose and show a negative oxidase reaction when the test
is carried out according to the method specified.
5.3.2 Colony count - where the number is expected to
be > 1 O00 per gram
4.2 detection of Enterobacteriaceae : Determination of
the presence or absence of Enterobacteriaceae in a
Introduction of 1 ml of the macerate and of the dilutions
particular mass when the test is carried out according to
and into empty Petri dishes,
the method specified.
followed by addition of violet red bile glucose agar and
covering of this agar by an overlay of violet red bile glucose
4.3 count of Enterobacteriaceae : The number of Entero-
agar. Incubation of the dishes at 37 "C for 24 h. From the
bacteriaceae found per gram of meat or meat product
number of confirmed (see 5.2) typical colonies per Petri
when the test is carried out according to the method
dish, calculation of the number of Enterobacteriaceae per
specified.
gram of sample.
5 PRINCIPLE
5.1 Maceration and dilution
6 CULTURE MEDIA, DILUTION FLUID AND
REAGENTS
Mincing of a test sample and then maceration of a test
portion with a sterile diluent, in a mechanical blender.
6.1 Basic materials
Preparation, from the macerate, of decimal dilutions.
6.1.1 For uniformity of results, it is recommended that
5.2 Detection of the presence or absence of Entero-
uniform dehydrated culture media components oi-
bacteriaceae in a particular mass (0,l g or 0,Ol g or 0,001 g)
dehydrated complete culture media be used.
of meat or meat product
6.1.2 The basic materials - peptone, tryptone, yeast
Introduction of 1 ml of the macerate (or the dilution 10-2
extract, ox bile, bile-salts, and watei - shall meet the
or in triplicate, into tubes containing a selective
requirements for preparations of bactei iological cultut e
enrichment broth.
media Chemicals shall be of analytical reagent grade
1

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IS0 5552-1979 (E)
Preparation of agar plates
6.2 Culture media
Transfer portions of about 10 ml of the culture medium,
Buffered brilliant green bile glucose broth
6.2.1
melted and cooled to approximately 45 OC, to Petri
dishes (7.2.3) and allow to solidify.
Composition
Immediately before use, dry the plates, preferably with
peptone
the lids off and the agar surface downwards, in an oven
glucose
or incubator (7.1.4) at 50 OC for 30 min.
disodium hydrogen phosphate
If prepared in advance, the undried plates shall not be
(Na2 HPO,)
kept longer than 4 h at room temperature or 1 day at
potassium hydrogen phosphate
O to 5 Oc.
(KH,PO,)
ox bile, desiccated
brilliant green 6.2.3 Glucose agar
water
Composition
tryptone
Preparation
yeast extract
Dissolve the components or the complete medium in
glucose
the water by boiling. The medium shall not be heated
sodium chloride
longer than 30 min. Cool the medium rapidly.
bromo-cresol purple
Adjust the pH so that after boiling it is 7,2 ? 0,l at
agar
20 Oc.
water
Transfer portions of 10 ml to sterile culture tubes.
Preparation
Sterilization of the medium is not desirable.
Dissolve the medium components or the complete
1 week at O to
The medium may be stored for up to
medium in the water by boiling.
5 Oc.
Adjust the pH so that after sterilization it is 7,O f 0,l at
45 Oc.
Transfer the culture medium in quantities of 15 ml to
6.2.2 Violet red bile glucose agar
culture tubes.
Composition
Sterilize the medium for 20 min at 121 * 1 OC.
peptone
Allow the medium to set in the tubes in a vertical
position.
yeast extract
\,rc
bile salts
These tubes may be stored for up to 1 week at O to 5 OC.
glucose
sodium chloride
6.2.4 Nutrient agar
neutral red
Cornposition
crystal violet
agar beef extract
water peptone
agar
water
Preparation
Dissolve the components or the complete medium in Preparation
the water by boiling.
Dissolve the medium components or the complete
medium in the water by boiling.
Adjust the pH so that after boiling it is 7,4 f 0.1 at
45 Oc.
Adjust the pH so that after sterilization it is 7,O f 0.2
Transfer the culture medium to sterile tubes or flasks of at 45 OC.
not more than 500 ml capacity.
Transfer the culture medium to tubes or flasks of not
more than 500 ml capacity.
Sterilization of the medium is not desirable.
Sterilize the medium for 20 min at 121 f 1 OC.
This medium shall be freshly prepared.
2

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IS0 5552-1979 (E)
7.1.3 Apparatus for sterilization of glassware, blender
Preparation of agar plates
jars, culture media, etc.
Transfer portions of about 15 ml of the culture medium,
melted and cooled to approximately 45 OC, to Petri
7.1.4 Drying cabinet, oven or incubator for drying the
dishes (7.2.3) and allow to solidify.
surfaces of agar plates, preferably at 50 ? 5 OC.
Immediately before use, dry the plates, preferably with
the lids off and the agar surface downwards, in an oven
7.1.5 Incubator for maintaining the inoculated plates
or incubator (7.1.4) at 50 OC for 30 min.
and tubes at 37 f 1 OC.
If prepared in advance, the undried plates shall not be
kept longer than 4 h at room temperature or 1 day at
7.1.6 Water bath for cooling the melted culture medium
O to 5 Oc.
to 45 Oc.
6.3 Dilution fluid
7.2 Glassware
Composition
The glassware shall be resistant to repeated sterilization.
peptone
sodium chloride
7.2.1 Culture tubes and flasks for the sterilization and
water
storage of culture media and dilution fluid.
Preparation
7.2.2 Graduated pipettes, calibrated for bacteriological
Dissolve the components in the water by boiling.
use only, with a nominal capacity of 1 ml, subdivided in
0,l ml and with an outflow opening of diameter 2 to 3 mm.
Adjust the pH so that after sterilization it is 7,O f 0,l at
20 Oc.
7.2.3 Petri dishes
Transfer part of the dilution fluid in quantities of
100 ml to 300 ml flasks for the maceration and the
Dish
remainder to tubes or small flasks in such quantities
that these contain 9.0 ml after sterilization.
90I2mm
internal diameter
less than 18 mm
external height not
Sterilize the dilution fluid for 20 min at 121 f 1 OC.
The rim shall be ground in a plane parallel to the base.
6.4 Oxidase reagent
The bottom of the dish shall be flat and parallel to the base.
Composition
Lid
N,N,N',N' - te t ra me t h y I -p- p h e n y I e ne
external diameter not more than 102 mm
diamine dihydrochloride 1,o 9
Plastics Petri dishes may also be used, even if of slightly
water 100 ml
different dimensions from those mentioned.
Preparation
Dissolve the reagent in cold water.
7.3 Sterilization of glassware, etc.
The reagent shall be prepared immediately before use.
Sterilize the glassware, etc. by one of the following
methods :
- wet sterilization at not less than 121 OC for not less
than 20 min;
7 APPARATUS AND GLASSWARE
- dry sterilization at not less than 170 OC for not less
than 1 h.
7.1 Apparatus
7.1.1 Mechanical meat mincer, laboratory size, sterile,
fitted with a plate with holes of diameter not exceeding
4 mm.
8 SAMPLING
7.1.2 Mechanical blender, operating at a rotational Proceed from a representative sample of a least 200 g. See
frequency of not less than 8 O00 min-' and not more than
3100,
45 O00 min-', with glass or metal blending jars of an
appropriate capacity, fitted with lids and resistant to the The sample may be stored at a tempera
...