1、Microbiology of the food chain Horizontal method for the determination of Vibrio spp.Part 1: Detection of Vibrio parahaemolyticus and Vibrio cholerae (ISO 21872-1:2017)BS EN ISO 21872-1:2017BSI Standards PublicationWB11885_BSI_StandardCovs_2013_AW.indd 1 15/05/2013 15:06EUROPEANSTANDARDNORME EUROPEN
2、NE EUROPISCHE NORMEN ISO 21872-1 July 2017ICS 07.100.30EnglishVersionMicrobiology of the food chain - Horizontal method for thedetermination of Vibrio spp. - Part 1: Detection ofpotentially enteropathogenic Vibrio parahaemolyticus,Vibrio cholerae and Vibrio vulnificus (ISO 21872-1:2017)Microbiologie
3、 de la chane alimentaire - Mthode horizontale pour la dtermination des Vibrio spp. -Partie 1: Recherche des espcesde Vibrio parahaemolyticus, Vibrio cholerae et Vibrio vulnificuspotentiellement entropathognes (ISO 21872-1:2017)Mikrobiologie von Lebensmitteln und Futtermitteln -Horizontales Verfahren
4、 zum Nachweis von potentiellenteropathogenen Vibrio spp.- Teil 1: Nachweis von vibrio parahaemolyticus und vibrio cholerae (ISO21872-1:2017)This European Standard was approved by CEN on 14 May 2017.CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditio
5、ns for giving thisEuropean Standard the status of a national standard without any alteration.Up-to-date lists and bibliographical referencesconcerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member. This European Standard exists in th
6、ree official versions (English, French, German). A version in any other language made bytranslation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC ManagementCentre has the same status as the officialversions.CEN members are the nationalstandards bodies
7、 of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Sloven
8、ia, Spain, Sweden, Switzerland, Turkey and United Kingdom.EUROPEAN COMMITTEE FOR STANDARDIZATIONCOMIT EUROPEN DE NORMALISATIONEUROPISCHES KOMITEE FR NORMUNGCEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels 2017 CEN All rights of exploitation in any form and by any means reservedworldw
9、ide for CENnational Members.Ref. No. EN ISO 21872-1:2017 ENational forewordThis British Standard is the UK implementation of EN ISO 21872-1:2017. It is identical to ISO 21872-1:2017. It supersedes DD ISO/TS 21872-1:2007 which is withdrawn.The UK participation in its preparation was entrusted to Tech
10、nical Committee AW/9, Microbiology.A list of organizations represented on this committee can be obtained on request to its secretary.This publication does not purport to include all the necessary provisions of a contract. Users are responsible for its correct application. The British Standards Insti
11、tution 2017 Published by BSI Standards Limited 2017ISBN 978 0 580 51145 5ICS 07.100.30Compliance with a British Standard cannot confer immunity from legal obligations. This British Standard was published under the authority of the Standards Policy and Strategy Committee on 30 September 2017.Amendmen
12、ts/corrigenda issued since publicationDate Text affectedBRITISH STANDARDBS EN ISO 218721:2017EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN ISO 21872-1 July 2017 ICS 07.100.30 English Version Microbiology of the food chain - Horizontal method for the determination of Vibrio spp. - Part 1: Dete
13、ction of potentially enteropathogenic Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus (ISO 21872-1:2017)Microbiologie de la chane alimentaire - Mthode horizontale pour la dtermination des Vibrio spp. - Partie 1: Recherche des espces de Vibrio parahaemolyticus, Vibrio cholerae et Vibri
14、o vulnificus potentiellement entropathognes (ISO 21872-1:2017)Mikrobiologie von Lebensmitteln und Futtermitteln - Horizontales Verfahren zum Nachweis von potentiell enteropathogenen Vibrio spp. - Teil 1: Nachweis von vibrio parahaemolyticus und vibrio cholerae (ISO 21872-1:2017) This European Standa
15、rd was approved by CEN on 14 May 2017. 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
16、national standards may be obtained on application to the CEN-CENELEC 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 l
17、anguage and notified to the CEN-CENELEC Management Centre has the same status as the official versions. CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Gre
18、ece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR
19、 NORMUNG CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels 2017 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN ISO 21872-1:2017 EBS EN ISO 218721:2017EN ISO 21872-1:2017 (E) 3 European foreword This document (EN ISO
20、 21872-1:2017) has been prepared by Technical Committee ISO/TC 34 “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 publ
21、ication of an identical text or by endorsement, at the latest by January 2018 and conflicting national standards shall be withdrawn at the latest by January 2018. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. CEN shall not be he
22、ld responsible for identifying any or all such patent rights. This document has been prepared under a mandate given to CEN by the European Commission and the European Free Trade Association. According to the CEN-CENELEC Internal Regulations, the national standards organizations of the following coun
23、tries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Po
24、land, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. Endorsement notice The text of ISO 21872-1:2017 has been approved by CEN as EN ISO 21872-1:2017 without any modification. ISO 21872-1:2017(E)Foreword vIntroduction vi1 Scope . 12 Normative
25、 references 13 Terms and definitions . 24 Principle 24.1 General . 24.2 Primary enrichment in a liquid selective medium . 24.3 Secondary enrichment in a liquid selective medium . 34.4 Isolation and identification 34.5 Confirmation . 35 Culture media and reagents 35.1 Enrichment medium: alkaline sali
26、ne peptone water (ASPW) 45.2 Solid selective isolation media 45.2.1 First medium: thiosulphate, citrate, bile and sucrose agar medium (TCBS) . 45.2.2 Second medium. 45.3 Saline nutrient agar (SNA) 45.4 Reagent for detection of oxidase . 45.5 Biochemical tests 45.5.1 L-lysine decarboxylase saline med
27、ium (LDC) 45.5.2 Arginine dihydrolase saline medium (ADH) 45.5.3 Reagent for detection of -galactosidase 55.5.4 Saline medium for detection of indole 55.5.5 Saline peptone waters . 55.5.6 Sodium chloride solution . 55.6 PCR 55.6.1 Tris acetate EDTA buffer (TAE) (or a buffer allowing similar performa
28、nce for the purpose) 55.6.2 Mastermix 55.6.3 Primers and probes . 55.6.4 Positive control material . 55.6.5 Negative extraction control 66 Equipment and consumables 67 Sampling 68 Preparation of the test sample . 69 Procedure (See Figure A.1) 79.1 Test portion and initial suspension . 79.2 Primary s
29、elective enrichment 79.3 Secondary selective enrichment 79.4 Isolation and identification 89.5 Confirmation . 89.5.1 General 89.5.2 Selection of colonies for confirmation and preparation of pure cultures 99.5.3 Tests for presumptive identification . 99.5.4 Biochemical confirmation . 109.5.5 PCR conf
30、irmation .129.5.6 DNA extraction .129.5.7 Conventional PCR .129.5.8 Real-time PCR 1310 Expression of results .13 ISO 2017 All rights reserved iiiContents PageBS EN ISO 218721:2017ISO 21872-1:2017(E)Foreword vIntroduction vi1 Scope . 12 Normative references 13 Terms and definitions . 24 Principle 24.
31、1 General . 24.2 Primary enrichment in a liquid selective medium . 24.3 Secondary enrichment in a liquid selective medium . 34.4 Isolation and identification 34.5 Confirmation . 35 Culture media and reagents 35.1 Enrichment medium: alkaline saline peptone water (ASPW) 45.2 Solid selective isolation
32、media 45.2.1 First medium: thiosulphate, citrate, bile and sucrose agar medium (TCBS) . 45.2.2 Second medium. 45.3 Saline nutrient agar (SNA) 45.4 Reagent for detection of oxidase . 45.5 Biochemical tests 45.5.1 L-lysine decarboxylase saline medium (LDC) 45.5.2 Arginine dihydrolase saline medium (AD
33、H) 45.5.3 Reagent for detection of -galactosidase 55.5.4 Saline medium for detection of indole 55.5.5 Saline peptone waters . 55.5.6 Sodium chloride solution . 55.6 PCR 55.6.1 Tris acetate EDTA buffer (TAE) (or a buffer allowing similar performance for the purpose) 55.6.2 Mastermix 55.6.3 Primers an
34、d probes . 55.6.4 Positive control material . 55.6.5 Negative extraction control 66 Equipment and consumables 67 Sampling 68 Preparation of the test sample . 69 Procedure (See Figure A.1) 79.1 Test portion and initial suspension . 79.2 Primary selective enrichment 79.3 Secondary selective enrichment
35、 79.4 Isolation and identification 89.5 Confirmation . 89.5.1 General 89.5.2 Selection of colonies for confirmation and preparation of pure cultures 99.5.3 Tests for presumptive identification . 99.5.4 Biochemical confirmation . 109.5.5 PCR confirmation .129.5.6 DNA extraction .129.5.7 Conventional
36、PCR .129.5.8 Real-time PCR 1310 Expression of results .13 ISO 2017 All rights reserved iiiContents PageBS EN ISO 218721:2017ISO 21872-1:2017(E)11 Performance characteristics of the method 1311.1 Interlaboratory study . 1311.2 Sensitivity . 1411.3 Specificity . 1411.4 LOD50. 1412 Test report 14Annex
37、A (normative) Diagram of procedure .15Annex B (normative) Composition and preparation of the culture media and reagents .17Annex C (informative) Conventional PCR for the detection of Vibrio parahaemolyticus, thermostable direct haemolysin (tdh) and thermostable direct related haemolysin (trh) genes,
38、 Vibrio cholerae and Vibrio vulnificus .23Annex D (informative) Real-time PCR for the detection of Vibrio parahaemolyticus, thermostable direct haemolysin gene (tdh) and Vibrio vulnificus 27Annex E (informative) Results of an interlaboratory study .29Bibliography .32iv ISO 2017 All rights reservedBS
39、 EN ISO 218721:2017ISO 21872-1:2017(E)11 Performance characteristics of the method 1311.1 Interlaboratory study . 1311.2 Sensitivity . 1411.3 Specificity . 1411.4 LOD50. 1412 Test report 14Annex A (normative) Diagram of procedure .15Annex B (normative) Composition and preparation of the culture medi
40、a and reagents .17Annex C (informative) Conventional PCR for the detection of Vibrio parahaemolyticus, thermostable direct haemolysin (tdh) and thermostable direct related haemolysin (trh) genes, Vibrio cholerae and Vibrio vulnificus .23Annex D (informative) Real-time PCR for the detection of Vibrio
41、 parahaemolyticus, thermostable direct haemolysin gene (tdh) and Vibrio vulnificus 27Annex E (informative) Results of an interlaboratory study .29Bibliography .32iv ISO 2017 All rights reserved ISO 21872-1:2017(E)ForewordISO (the International Organization for Standardization) is a worldwide federat
42、ion 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 committe
43、e. 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.The procedures used to develop this document and th
44、ose intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different types of ISO documents should be noted. This document was drafted in accordance with the editorial rules of the ISO/IEC Directives, Part 2
45、(see www .iso .org/ directives).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. Details of any patent rights identified during the development of the
46、 document will be in the Introduction and/or on the ISO list of patent declarations received (see www .iso .org/ patents).Any trade name used in this document is information given for the convenience of users and does not constitute an endorsement.For an explanation on the meaning of ISO specific te
47、rms and expressions related to conformity assessment, as well as information about ISOs adherence to the World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see the following URL: www .iso .org/ iso/ foreword .html.This document was prepared by the European Committee f
48、or Standardization (CEN) Technical Committee CEN/TC 275, Food analysis Horizontal methods, in collaboration with ISO Technical Committee TC 34, Food products, Subcommittee SC 9, Microbiology, in accordance with the agreement on technical cooperation between ISO and CEN (Vienna Agreement).This first
49、edition cancels and replaces ISO/TS 21872-1:2007, which has been technically revised. It also incorporates ISO/TS 21872-1:2007/Cor.1:2008.The main changes are as follows: introduction of optional molecular identification methods for major food borne Vibrio spp. (V. parahaemolyticus, including potentially enteropathogenic strains, V. vulnificus and V. cholerae); performance characteristics of the method have been added in Annex E.A list of all parts in the ISO 21872 series can be found on the ISO website. ISO 2017 All rights reserved
