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1、BRITISH STANDARD BS EN ISO 21871:2006 Microbiology of food and animal feeding stuffs Horizontal method for the determination of low numbers of presumptive Bacillus cereus Most probable number technique and detection method The European Standard EN ISO 21871:2006 has the status of a British Standard
2、ICS 07.100.30 ? Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI BS EN ISO 21871:2006 This British Standard was published under the authority of the Standards Policy and Strategy Committee on 1 February 2006 BSI 1 February 2006 ISBN 0 580 472
3、08 6 National foreword This British Standard is the official English language version of EN ISO 21871:2006. It is identical with ISO 21871:2006. The UK participation in its preparation was entrusted to Technical Committee AW/9, Microbiology, which has the responsibility to: A list of organizations r
4、epresented on this committee can be obtained on request to its secretary. Cross-references The British Standards which implement international or European publications referred to in this document may be found in the BSI Catalogue under the section entitled “International Standards Correspondence In
5、dex”, or by using the “Search” facility of the BSI Electronic Catalogue or of British Standards Online. This publication does not purport to include all the necessary provisions of a contract. Users are responsible for its correct application. Compliance with a British Standard does not of itself co
6、nfer immunity from legal obligations. aid enquirers to understand the text; present to the responsible international/European committee any enquiries on the interpretation, or proposals for change, and keep UK interests informed; monitor related international and European developments and promulgate
7、 them in the UK. Summary of pages This document comprises a front cover, an inside front cover, the EN ISO title page, the EN ISO foreword page, the ISO title page, pages ii to v, a blank page, pages 1 to 14, an inside back cover and a back cover. The BSI copyright notice displayed in this document
8、indicates when the document was last issued. Amendments issued since publication Amd. No. DateComments Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN ISO 21871 January 2006 ICS 07.100.30
9、English Version Microbiology of food and animal feeding stuffs - Horizontal method for the determination of low numbers of presumptive Bacillus cereus - Most probable number technique and detection method (ISO 21871:2006) Microbiologie des aliments - Mthode horizontale pour le dnombrement de Bacillu
10、s cereus prsums en petit nombre - Technique du nombre le plus probable et mthode de recherche (ISO 21871:2006) Mikrobiologie von Lebensmitteln und Futtermitteln - Horizontales Verfahren zur Bestimmung niedriger Zahlen von prsumtivem Bacillus cereus - Verfahren der wahrscheinlichsten Keimzahl (MPN) u
11、nd Nachweisverfahren (ISO 21871:2006) This European Standard was approved by CEN on 28 November 2005. 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. U
12、p-to-date lists and bibliographical references concerning such national standards may be obtained on application to the Central Secretariat 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 un
13、der the responsibility of a CEN member into its own language and notified to the Central Secretariat has the same status as the official versions. CEN members are the national standards bodies of Austria, Belgium, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, I
14、celand, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG Management Centre: rue
15、 de Stassart, 36 B-1050 Brussels 2006 CENAll rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN ISO 21871:2006: E Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI Foreword This documen
16、t (EN ISO 21871:2006) 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 sta
17、ndard, either by publication of an identical text or by endorsement, at the latest by July 2006, and conflicting national standards shall be withdrawn at the latest by July 2006. According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following countries are bo
18、und to implement this European Standard: Austria, Belgium, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerlan
19、d and United Kingdom. Endorsement notice The text of ISO 21871:2006 has been approved by CEN as EN ISO 21871:2006 without any modifications. EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI Reference number ISO 21871:2006(E)
20、 INTERNATIONAL STANDARD ISO 21871 First edition 2006-01-15 Microbiology of food and animal feeding stuffs Horizontal method for the determination of low numbers of presumptive Bacillus cereus Most probable number technique and detection method Microbiologie des aliments Mthode horizontale pour le dn
21、ombrement de Bacillus cereus prsums en petit nombre Technique du nombre le plus probable et mthode de recherche EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI ii Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08
22、:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI iii Contents Page Foreword iv Introduction v 1 Scope . 1 2 Normative references. 1 3 Terms and definitions. 1 4 Principle. 2 4.1 Enumeration method 2 4.2 Detection method 2 5 Diluent, culture media and reagents. 2 5.1 General. 2 5.2 Diluent 2 5.3 Liquid
23、 selective enrichment medium: Tryptone soya polymyxin broth (TSPB) 3 5.4 Solid selective medium: Polymyxin pyruvate egg yolk mannitol bromothymol blue agar (PEMBA). 4 5.5 Solid selective medium: Mannitol egg yolk polymyxin agar (MYP) 6 5.6 Staining solutions for microscopic identification. 7 5.7 She
24、ep blood agar 8 6 Apparatus and glassware 8 7 Sampling 9 8 Preparation of test sample. 9 9 Procedure 9 9.1 Enumeration method 9 9.2 Detection method 11 10 Calculation and expression of results 12 10.1 Enumeration method for the determination of the most probable number (MPN) 12 10.2 Detection method
25、 12 11 Test report . 12 Annex A (normative) Diagram of enumeration procedure. 13 Bibliography. 14 EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI iv Foreword ISO (the International Organization for Standardization) is a wor
26、ldwide 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
27、 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
28、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 Standar
29、d 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 21871 was prepared by T
30、echnical Committee ISO/TC 34, Food products, Subcommittee SC 9, Microbiology. EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI v Introduction Because of the large variety of food and feed products, this horizontal method may
31、 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 th
32、is International Standard 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 i
33、mmediate, 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 International Standard so that eventually the on
34、ly remaining departures from this horizontal method will be those necessary for well-established technical reasons. EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI blank Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 1
35、7 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c) BSI 1 Microbiology of food and animal feeding stuffs Horizontal method for the determination of low numbers of presumptive Bacillus cereus Most probable number technique and detection method 1 Scope This International Standard specifies a horizontal
36、method for the detection or the enumeration of low numbers of viable presumptive Bacillus cereus by means of the most probable number technique. This International Standard is applicable to products intended for human consumption and the feeding of animals, and environmental samples in the area of f
37、ood production and food handling. 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) a
38、pplies. ISO 6887 (all parts), Microbiology of food and animal feeding stuffs Preparation of test samples, initial suspension and decimal dilutions for microbiological examination ISO 7218, Microbiology of food and animal feeding stuffs General rules for microbiological examinations ISO 8261, Milk an
39、d milk products General guidance for the preparation of test samples, initial suspensions and decimal dilutions for microbiological examination ISO/TS 11133-1, Microbiology of food and animal feeding stuffs Guidelines on preparation and production of culture media Part 1: General guidelines on quali
40、ty assurance for the preparation of culture media in the laboratory 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 of culture media 3 Terms and definitions For the purposes o
41、f this document, the following terms and definitions apply. 3.1 presumptive Bacillus cereus microorganism that forms typical or atypical colonies on the surface of selective culture media and which gives positive confirmation reactions under the conditions specified in this International Standard NO
42、TE For the purpose of a practical test method, this definition of presumptive Bacillus cereus, used as a basis for the procedure, does not exclusively describe strains of Bacillus cereus. In particular, the confirmatory test is inadequate to distinguish between Bacillus cereus and other closely rela
43、ted but less commonly encountered Bacillus species such as Bacillus weihenstephanensis, Bacillus anthracis, Bacillus thuringiensis, Bacillus mycoides and Bacillus pseudomycoides. EN ISO 21871:2006 Licensed Copy: sheffieldun sheffieldun, na, Fri Nov 17 10:08:30 GMT+00:00 2006, Uncontrolled Copy, (c)
44、BSI 2 4 Principle 4.1 Enumeration method 4.1.1 Inoculation of three tubes of double-strength liquid selective enrichment medium 5.3.1.1 a) with a specified quantity of the primary dilution (initial suspension). 4.1.2 Inoculation of three tubes of single-strength liquid selective enrichment medium 5.
45、3.1.1 b) with a specified quantity of the primary dilution (initial suspension). Then, under the same conditions, inoculation of single-strength liquid selective enrichment medium 5.3.1.1 b) with specified quantities of decimal dilutions of the primary dilution (initial suspension). 4.1.3 Incubation
46、 of the tubes of double and single-strength liquid selective enrichment medium (5.3) for 48 h at 30 C. 4.1.4 Inoculation of the solid selective medium (5.4 or 5.5) from the liquid selective enrichment medium (5.3). 4.1.5 Incubation of the solid selective medium (5.4 or 5.5) for 18 h to 48 h at 37 C
47、(5.4) or 30 C (5.5) and examination of the plates to check for the presence of colonies which, from their characteristics, are considered to be presumptive Bacillus cereus. 4.1.6 Confirmation of suspected colonies by means of haemolysis (9.1.5.3) or by microscopic examination (9.1.5.4). 4.1.7 Calcul
48、ation of the most probable number of presumptive Bacillus cereus per gram or per millilitre of sample from selected dilutions by reference to most probable number tables. 4.2 Detection method 4.2.1 Inoculation of a liquid selective enrichment medium (5.3) with a specified quantity of the initial sus
49、pension of the test sample. 4.2.2 Incubation of the tube for 48 h at 30 C. 4.2.3 Inoculation of a solid selective medium (5.4 or 5.5) from the liquid selective enrichment medium (5.3). 4.2.4 Incubation of the solid selective medium (5.4 or 5.5) for 18 h to 48 h at 37 C (5.4) or 30 C (5.5) and examination of the plates to check for the presence of colonies which, from their characteristics, are considered to be presumptive Bacillus cereus. 4.2.5 Confirmat