SIST EN ISO 6888-3:2003

SLOVENSKI STANDARD
01-maj-2003
Mikrobiologija živil in krme - Horizontalna metoda za ugotavljanje števila
koagolaza pozitivnih stafilokokov (Staphylococcus aureus in druge vrste) - 3. del:
Ugotavljanje prisotnosti in MPN-tehnika pri majhnem številu (ISO 6888-3:2003)
Microbiology of food and animal feeding stuffs - Horizontal method for the enumeration of
coagulase-positive staphylococci (Staphylococcus aureus and other species) - Part 3:
Detection and MPN technique for low numbers (ISO 6888-3:2003)
Mikrobiologie von Lebensmitteln und Futtermitteln - Horizontales Verfahren für die
Zählung von koagulase-positiven Staphylokokken (Staphylococcus aureus und andere
Species) - Teil 3: Nachweis und MPN-Verfahren für niedrige Keimzahlen (ISO 6888-
3:2003)
Microbiologie des aliments - Méthode horizontale pour le dénombrement des
staphylocoques a coagulase positive (Staphylococcus aureus et autres especes) - Partie
3: Recherche et méthode NPP pour les faibles nombres (ISO 6888-3:2003)
Ta slovenski standard je istoveten z: EN ISO 6888-3:2003
ICS:
07.100.30 Mikrobiologija živil Food microbiology
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EUROPEAN STANDARD
EN ISO 6888-3
NORME EUROPÉENNE
EUROPÄISCHE NORM
March 2003
ICS 07.100.30
English version
Microbiology of food and animal feeding stuffs - Horizontal
method for the enumeration of coagulase-positive staphylococci
(Staphylococcus aureus and other species) - Part 3: Detection
and MPN technique for low numbers (ISO 6888-3:2003)
Microbiologie des aliments - Méthode horizontale pour le Mikrobiologie von Lebensmitteln und Futtermitteln -
dénombrement des staphylocoques à coagulase positive Horizontales Verfahren für die Zählung von koagulase-
(Staphylococcus aureus et autres espèces) - Partie 3: positiven Staphylokokken (Staphylococcus aureus und
Recherche et méthode NPP pour les faibles nombres (ISO andere Species) - Teil 3: Nachweis und MPN-Verfahren für
6888-3:2003) niedrige Keimzahlen (ISO 6888-3:2003)
This European Standard was approved by CEN on 28 February 2003.
CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European
Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such national
standards may be obtained on application to the Management Centre or to any CEN member.
This European Standard exists in three official versions (English, French, German). A version in any other language made by translation
under the responsibility of a CEN member into its own language and notified to the Management Centre has the same status as the official
versions.
CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece,
Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and United
Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: rue de Stassart, 36  B-1050 Brussels
© 2003 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 6888-3:2003 E
worldwide for CEN national Members.

CORRECTED  2003-05-14
Foreword
This document (EN ISO 6888-3:2003) has been prepared by Technical Committee ISO/TC 34
"Agricultural food products" in collaboration with Technical Committee CEN/TC 275 "Food analysis
- Horizontal methods", the secretariat of which is held by DIN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by September 2003, and conflicting national
standards shall be withdrawn at the latest by September 2003.
According to the CEN/CENELEC Internal Regulations, the national standards organizations of the
following countries are bound to implement this European Standard: Austria, Belgium, Czech
Republic, Denmark, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy,
Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and the
United Kingdom.
Endorsement notice
The text of ISO 6888-3:2003 has been approved by CEN as EN ISO 6888-3:2003 without any
modifications.
NOTE Normative references to International Standards are listed in Annex ZA (normative).
Annex ZA
(normative)
Normative references to international publications
with their relevant European publications
This European Standard incorporates by dated or undated reference, provisions from other
publications. These normative references are cited at the appropriate places in the text and the
publications are listed hereafter. For dated references, subsequent amendments to or revisions of any
of these publications apply to this European Standard only when incorporated in it by amendment or
revision. For undated references the latest edition of the publication referred to applies (including
amendments).
NOTE Where an International Publication has been modified by common modifications, indicated by
(mod.), the relevant EN/HD applies.
Publication Year Title EN Year
ISO 6887-1 1999 Microbiology of food and animal EN ISO 6887-1 1999
feeding stuffs — Preparation of test
samples, initial suspension and
decimal dilutions for microbiological
examination — Part: 1: General rules
for the preparation of the initial
suspension and decimal dilutions
ISO 6888-1 1999 Microbiology of food and animal EN ISO 6888-1 1999
feeding stuffs — Horizontal method
for the enumeration of coagulase-
positive staphylococci
(Staphylococcus aureus and other
species) — Part 1: Technique using
Baird-Parker agar medium
ISO 6888-2 1999 Microbiology of food and animal EN ISO 6888-2 1999
feeding stuffs — Horizontal method
for the enumeration of coagulase-
positive staphylococci
(Staphylococcus aureus and other
species) — Part 2: Technique using
rabbit plasma fibrinogen agar
medium
ISO 8261 2001 Milk and mild products — General EN ISO 8261 2001
guidance for the preparation of test
samples, initial suspensions and
decimal dilutions for microbiological
examination
ISO/TR 11133-1 2000 Microbiology of food and animal ENV ISO 11133-1 2000
feeding stuffs — Guidelines on
preparation and production of culture
media — Part 1: General guidelines
on quality assurance for the
preparation of culture media in the
laboratory
INTERNATIONAL ISO
STANDARD 6888-3
First edition
2003-03-15
Corrected version
2004-01-15
Microbiology of food and animal feeding
stuffs — Horizontal method for the
enumeration of coagulase-positive
staphylococci (Staphylococcus aureus
and other species) —
Part 3:
Detection and MPN technique for low
numbers
Microbiologie des aliments — Méthode horizontale pour le
dénombrement des staphylocoques à coagulase positive
(Staphylococcus aureus et autres espèces) —
Partie 3: Recherche et méthode NPP pour les faibles nombres

Reference number
ISO 6888-3:2003(E)
©
ISO 2003
ISO 6888-3:2003(E)
PDF disclaimer
This PDF file may contain embedded typefaces. In accordance with Adobe's licensing policy, this file may be printed or viewed but
shall not be edited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In
downloading this file, parties accept therein the responsibility of not infringing Adobe's licensing policy. The ISO Central Secretariat
accepts no liability in this area.
Adobe is a trademark of Adobe Systems Incorporated.
Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation
parameters were optimized for printing. Every care has been taken to ensure that the file is suitable for use by ISO member bodies. In
the unlikely event that a problem relating to it is found, please inform the Central Secretariat at the address given below.

©  ISO 2003
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or
ISO's member body in the country of the requester.
ISO copyright office
Case postale 56 • CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2003 — All rights reserved

ISO 6888-3:2003(E)
Contents Page
Foreword. iv
Introduction . v
1 Scope. 1
2 Normative references . 1
3 Terms and definitions. 2
4 Principle . 2
4.1 Detection method. 2
4.2 Enumeration method . 2
5 Diluents and culture media . 3
6 Apparatus. 6
7 Sampling . 6
8 Preparation of test sample. 6
9 Procedure. 6
9.1 Detection method. 6
9.2 Enumeration method . 7
9.3 Selection of plates and interpretation. 8
10 Expression of results. 9
10.1 Detection method. 9
10.2 Enumeration method . 9
11 Precision . 10
12 Test report. 10
Bibliography . 11

ISO 6888-3:2003(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies
(ISO member bodies). The work of preparing International Standards is normally carried out through ISO
technical committees. Each member body interested in a subject for which a technical committee has been
established 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. ISO collaborates closely with the
International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 6888-3 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 9,
Microbiology.
ISO 6888 consists of the following parts, under the general title Microbiology of food and animal feeding
stuffs — Horizontal method for the enumeration of coagulase-positive staphylococci (Staphylococcus aureus
and other species):
 Part 1: Technique using Baird-Parker agar medium
 Part 2: Technique using rabbit plasma fibrinogen agar medium
 Part 3: Detection and MPN technique for low numbers
This corrected version of ISO 6888-3:2003 incorporates the following correction:
Subclause 9.1.1
The second paragraph has been amended to resolve any ambiguity.
iv © ISO 2003 — All rights reserved

ISO 6888-3:2003(E)
Introduction
Because of the large variety of food and feed products, this horizontal method may not be appropriate in every
detail for certain products. In this case, different methods, which are specific to these products, may be used if
absolutely necessary for justified technical reasons. Nevertheless, every attempt should be made to apply this
horizontal method as far as possible.
When this part of ISO 6888 is next reviewed, account will be taken of all information then available regarding
the extent to which this horizontal method has been followed and the reasons for deviations from this method
in the case of particular products.
The harmonization of test methods cannot be immediate and, for certain groups of products, International
Standards and/or national standards may already exist that do not comply with this horizontal method. It is
hoped that when such standards are reviewed they will be changed to comply with this part of ISO 6888 so
that eventually the only remaining departures from this horizontal method will be those necessary for
well-established technical reasons.

INTERNATIONAL STANDARD ISO 6888-3:2003(E)

Microbiology of food and animal feeding stuffs — Horizontal
method for the enumeration of coagulase-positive
staphylococci (Staphylococcus aureus and other species) —
Part 3:
Detection and MPN technique for low numbers
1 Scope
This part of ISO 6888 specifies a horizontal method for the enumeration and detection of coagulase-positive
staphylococci, using the most probable number (MPN) technique. It is applicable to
 products intended for human consumption and the feeding of animals, and
 environmental samples in the area of food production and food handling.
This method is recommended for products where staphylococci are expected to be stressed and in low
numbers as, for example, in dried products. Coagulase-positive staphylococci will primarily be Staphylococcus
aureus but Staphylococcus intermedius and some strains of Staphylococcus hyicus also produce coagulase.
2 Normative references
The following referenced documents are indispensable for the application of this document. For dated
references, only the edition cited applies. For undated references, the latest edition of the referenced
document (including any amendments) applies.
ISO 6887 (all parts), Microbiology of food and animal feeding stuffs — Preparation of test samples, initial
suspension and decimal dilutions for microbiological examination
ISO 8261, Milk and milk products — General guidance for the preparation of test samples, initial suspensions
and decimal dilutions for microbiological examination
ISO 6888-1:1999, Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of
coagulase-positive staphylococci (Staphylococcus aureus and other species) — Part 1: Technique using
Baird-Parker agar medium
ISO 6888-2:1999, Microbiology of food and animal feeding stuffs – Horizontal method for the enumeration of
coagulase-positive staphylococci (Staphylococcus aureus and other species) — Part 2: Technique using
rabbit plasma fibrinogen agar medium
ISO 7218, Microbiology of food and animal feeding stuffs — General rules for microbiological examinations
ISO/TS 11133-1, Microbiology of food and animal feeding stuffs — Guidelines on preparation and production
of culture media — Part 1: General guidelines on quality assurance for the preparation of culture media in the
laboratory
ISO 6888-3:2003(E)
1)
ISO/TS 11133-2:— , Microbiology of food and animal feeding stuffs — Guidelines on preparation and
production of culture media — Part 2: Practical guidelines on performance testing culture media
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
coagulase-positive staphylococci
bacteria which form typical and/or atypical colonies on the surface of a selective culture medium and which
show a positive coagulase reaction or a specific rabbit plasma reaction on rabbit plasma fibrinogen agar
NOTE For the purpose of this part of ISO 6888, the confirmation of coagulase-positive staphylococci is based on a
strongly positive coagulase reaction, but it is recognized that some strains of coagulase-positive staphylococci give weakly
positive coagulase reactions. These latter strains can be confused with other bacteria but they can be distinguished from
such other bacteria by the use of additional tests such as the production of thermonuclease (for details, see IDF 83).
3.2
enumeration of the coagulase-positive staphylococci
determination of the number of coagulase-positive staphylococci found per millilitre or per gram of sample
when the test is carried out according to the method specified in this part of ISO 6888
4 Principle
4.1 Detection method
4.1.1 A selective culture medium is inoculated with a specified quantity of the test sample if the initial
product is liquid, or a specified quantity of an initial suspension in the case of other products.
4.1.2 The tubes are incubated at 37 °C, anaerobically, for 24 h and 48 h. The presence of presumptive
coagulase-positive staphylococci is indicated by the reduction of potassium tellurite.
NOTE In this part of ISO 6888, anaerobiosis is obtained by pouring a plug of agar or paraffin into each tube, but an
alternative procedure is to incubate the tubes in a jar or an incubator under anaerobic conditions.
4.1.3 The surface of solid selective Baird-Parker medium is inoculated from presumptive positive tubes
(4.1.2) after 24 h, and all the remaining tubes after 48 h.
4.1.4 The plates are inoculated at 37 °C for 24 h and 48 h. The presence of presumptive coagulase-positive
staphylococci is indicated by the reduction of potassium tellurite and an egg yolk reaction.
4.1.5 Typical and/or atypical colonies are confirmed by a
...