1、BSI Standards PublicationPD CEN ISO/TR 6579-3:2014Microbiology of the food chain Horizontal method for thedetection, enumeration andserotyping of SalmonellaPart 3: Guidelines for serotyping ofSalmonella spp. (ISO/TR 6579-3:2014)PD CEN ISO/TR 6579-3:2014 PUBLISHED DOCUMENTNational forewordThis Publis
2、hed Document is the UK implementation of CEN ISO/TR6579-3:2014. It supersedes BS EN ISO 6579:2002+A1:2007 which iswithdrawn.The UK participation in its preparation was entrusted to TechnicalCommittee AW/9, Microbiology.A list of organizations represented on this committee can beobtained on request t
3、o its secretary.This publication does not purport to include all the necessaryprovisions of a contract. Users are responsible for its correctapplication. The British Standards Institution 2014. Published by BSI StandardsLimited 2014ISBN 978 0 580 82085 4ICS 07.100.30Compliance with a British Standar
4、d cannot confer immunity fromlegal obligations.This Published Document was published under the authority of theStandards Policy and Strategy Committee on 31 July 2014.Amendments issued since publicationDate Text affectedTECHNICAL REPORT RAPPORT TECHNIQUE TECHNISCHER BERICHT CEN ISO/TR 6579-3 July 20
5、14 ICS 07.100.30 Supersedes EN ISO 6579:2002English Version Microbiology of the food chain - Horizontal method for the detection, enumeration and serotyping of Salmonella - Part 3: Guidelines for serotyping of Salmonella spp. (ISO/TR 6579-3:2014) Microbiologie de la chane alimentaire - Mthode horizo
6、ntale pour la recherche, le dnombrement et la srotypie des Salmonella - Partie 3: Lignes directrices pour la srotypie des Salmonella spp. (ISO/TR 6579-3:2014) Mikrobiologie von Lebensmitteln und Futtermitteln - Horizontales Verfahren zum Nachweis, zur Zhlung und zur Serotypisierung von Salmonellen -
7、 Teil 3: Leitfaden fr die Serotypisierung von Salmonella spp. (ISO/TR 6579-3:2014)This Technical Report was approved by CEN on 28 October 2013. It has been drawn up by the Technical Committee CEN/TC 275. CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Cz
8、ech 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, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom.
9、EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels 2014 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. CEN ISO/TR 6579
10、-3:2014 EPD CEN ISO/TR 6579-3:2014CEN ISO/TR 6579-3:2014 (E) 3 Foreword This document (CEN ISO/TR 6579-3:2014) 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
11、 by DIN. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. CEN and/or CENELEC shall not be held responsible for identifying any or all such patent rights. This document supersedes EN ISO 6579:2002. Endorsement notice The text of ISO
12、/TR 6579-3:2014 has been approved by CEN as CEN ISO/TR 6579-3:2014 without any modification. PD CEN ISO/TR 6579-3:2014ISO/TR 6579-3:2014(E) ISO 2014 All rights reserved iiiContents PageForeword ivIntroduction v1 Scope . 12 Normative references 13 Terms and definitions . 14 Principle 25 Culture media
13、 and sera 25.1 General . 25.2 Culture media and reagents . 25.3 Antisera . 26 Apparatus . 27 Sample . 38 Taxonomy of Salmonella . 38.1 General . 38.2 Nomenclature. 38.3 Biochemical characteristics . 48.4 Antigenic characteristics 59 Procedure for Salmonella serotyping . 79.1 General . 79.2 Example p
14、rocedure for serotyping five public health-related Salmonella serovars . 710 Quality control .1111 Reporting .11Annex A (informative) Composition and preparation of culture media and reagents 13Annex B (informative) Examples of procedures for serotyping an unknown Salmonella isolate 20Annex C (infor
15、mative) Biochemical tests 24Annex D (informative) Schematic overview for serotyping five important public-health related Salmonella serovars .26Annex E (informative) Microtitre plate method for serotyping Salmonella spp27Annex F (informative) Examples of procedures for phase inversion 29Bibliography
16、 .32PD CEN ISO/TR 6579-3:2014ISO/TR 6579-3:2014(E)ForewordISO (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
17、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
18、 Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.The procedures used to develop this document and those intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different ty
19、pes of ISO documents should be noted. This document was drafted in accordance with the editorial rules of the ISO/IEC Directives, Part 2 (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
20、 held responsible for identifying any or all such patent rights. Details of any patent rights identified during the development of the 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 informa
21、tion given for the convenience of users and does not constitute an endorsement.For an explanation on the meaning of ISO specific terms and expressions related to conformity assessment, as well as information about ISOs adherence to the WTO principles in the Technical Barriers to Trade (TBT) see the
22、following URL: Foreword - Supplementary informationThe committee responsible for this document is ISO/TC 34, Food products, Subcommittee SC 9, Microbiology. ISO 6579 consists of the following parts, under the general title Microbiology of the food chain Horizontal method for the detection, enumerati
23、on and serotyping of Salmonella: Part 1: Horizontal method for the detection of Salmonella spp.1) Part 2: Enumeration by a miniaturized most probable number technique Technical Specification2) Part 3: Guidelines for serotyping of Salmonella spp. Technical Report1) Under preparation. (Revision of ISO
24、 6579:2002)2) The main element of the series title has been changed since Part 2 was published. It is intended that upon revision, the main element of the title will be aligned with Part 3.iv ISO 2014 All rights reservedPD CEN ISO/TR 6579-3:2014ISO/TR 6579-3:2014(E)IntroductionThis part of ISO 6579
25、gives information on the taxonomy of Salmonella spp. and it gives guidance on serotyping of Salmonella serovars, based on the WhiteKauffmannLe Minor scheme (see Reference 9). ISO 2014 All rights reserved vPD CEN ISO/TR 6579-3:2014PD CEN ISO/TR 6579-3:2014Microbiology of the food chain Horizontal met
26、hod for the detection, enumeration and serotyping of Salmonella Part 3: Guidelines for serotyping of Salmonella spp.WARNING In order to safeguard the health of laboratory personnel, it is essential that tests for detecting and typing Salmonella, be undertaken only in properly equipped laboratories,
27、under the control of a skilled microbiologist, and that great care is taken in the disposal of all incubated materials.IMPORTANT Persons using this document should be familiar with normal laboratory practice. This document does not purport to address all of the safety aspects, if any, associated wit
28、h its use. It is the responsibility of the user to establish appropriate safety and health practices and to ensure compliance with any national regulatory conditions.1 ScopeThis part of ISO 6579 gives guidance on the procedure for serotyping Salmonella serovars and is applicable to the serotyping of
29、 pure cultures of Salmonella spp., independent of the source from which they are isolated.2 Normative referencesThe following documents, in whole or in part, are normatively referenced in this document and are indispensable for its application. For dated references, only the edition cited applies. F
30、or undated references, the latest edition of the referenced document (including any amendments) applies.ISO 6579-1, Microbiology of the food chain Horizontal method for the detection, enumeration and serotyping of Salmonella Part 1: Horizontal method for the detection of Salmonella spp. ISO 7218, Mi
31、crobiology of food and animal feeding stuffs General requirements and guidance for microbiological examinationsISO 11133, Microbiology of food, animal feed and water Preparation, production, storage and performance testing of culture media3 Terms and definitionsFor the purposes of this document, the
32、 following terms and definitions apply.3.1Salmonellagram-negative, oxidase-negative, facultatively anaerobic, non-spore-forming, rod-shaped bacteria which generally form colonies of 2 mm to 4 mm in diameter on solid selective media and display biochemical and serological characteristics described wh
33、en tests are carried out in accordance with this part of ISO 6579TECHNICAL REPORT ISO/TR 6579-3:2014(E) ISO 2014 All rights reserved 1PD CEN ISO/TR 6579-3:2014ISO/TR 6579-3:2014(E)3.2serotyping of Salmonelladetermination of the presence or absence of specific O-antigens, H-antigens and Vi-antigens i
34、n an isolate confirmed as Salmonella (3.1)3.3antigenic formulacombination of numbers and letters representing the O-, H-, and Vi-antigens of an isolate confirmed as Salmonella (3.1)4 PrincipleFor the serotyping of Salmonella spp. the following antigens are determined for isolates biochemically confi
35、rmed as Salmonella spp.:O-antigens, H-antigens and Vi-antigens.NOTE Alternative procedures can be used to confirm the isolate being Salmonella spp. provided the suitability of the alternative procedure is verified (see ISO 7218).5 Culture media and sera5.1 GeneralFor current laboratory practice, app
36、ly ISO 7218.For the performance testing of media, follow the recommendations of ISO 11133.5.2 Culture media and reagentsSee Annex A.5.3 AntiseraO-antisera, H-antisera and Vi-antisera are available from various commercial suppliers. Information on relevant polyvalent antisera and monovalent antisera
37、can be found in Annex B.6 ApparatusDisposable supplies are an acceptable alternative to reusable glassware if they have similar specifications.Usual microbiological laboratory equipment and, in particular, the following.6.1 Incubator, to grow Salmonella isolates, capable of operating in the range 34
38、 C to 38 C.NOTE In this part of ISO 6579, the incubation temperature is not a differential parameter. Isolates are cultured to obtain sufficient material to perform the tests on a pure culture. Therefore, culture step is performed at an optimal growth temperature. For Salmonella this is generally a
39、temperature between 34 C and 38 C.6.2 Oven (for dry sterilization) or autoclave (for wet sterilization). See ISO 7218.6.3 Refrigerator (for storage of prepared media), capable of operating at 5 C 3 C.6.4 Glass slides.6.5 Sterile inoculation instrument, e.g. needles, wires, wooden sticks, loops (e.g.
40、 of 1 l).2 ISO 2014 All rights reservedPD CEN ISO/TR 6579-3:2014ISO/TR 6579-3:2014(E)6.6 Sterile test tubes and flasks, of appropriate capacity. Flasks or bottles and test tubes with non-toxic metallic or plastic (screw) caps may be used.6.7 Sterile Petri dishes, with diameters of approximately 55 m
41、m and 90 mm.6.8 Water bath, capable of operating at 47-50 C.6.9 Water bath (or incubator), capable of operating at 50 C 2 C.7 SampleIt is important that the laboratory works with a pure culture which has been biochemically confirmed as Salmonella spp.8 Taxonomy of Salmonella8.1 GeneralApproximately
42、every 7 years, the WHO Collaborating Centre for Reference and Research on Salmonella (Institut Pasteur, Paris) publishes an update of the “Antigenic formulae of the Salmonella serovars”, which is the basis for assigning serovar names and formulas to isolates of Salmonella spp. At the time of publica
43、tion, the latest version of the WhiteKauffmannLe Minor scheme is that of 2007 (Reference 9).NOTE Supplements to the White-Kauffmann-Le Minor scheme are published in Research in Microbiology, a publication of the Institut Pasteur (formerly called Annales de lInstitut Pasteur/Microbiologie). For insta
44、nce, supplement no. 47 was published in 2010 and characterises new serovars found between 2003 and 2007 (Reference 10).This part of ISO 6579 provides guidance on the serotyping of Salmonella serovars.8.2 NomenclatureDifferent nomenclatures have been used (or are still in use) for Salmonella strains:
45、 originally, Kauffmann (Reference 12) considered each Salmonella serovar as a separate species; different type species have been used: S. enterica vs. S. choleraesuis, each having another type strain; some “important” Salmonella strains (like Salmonella Typhi and Salmonella Paratyphi) were considere
46、d to be species and not being “only” serovars of a species.The Judicial Commission of the International Committee on Systematics of Procaryotes indicated that many synonyms can be used in Salmonella nomenclature (Reference 22). In this part of ISO 6579, the widely accepted current nomenclature is us
47、ed, which is also approved by the WHO Collaborating Centre for Reference and Research on Salmonella (Reference 9), the American Society for Microbiology (Reference 20), the Centers for Disease Control and Prevention (Reference 3) and Bergeys manual (Reference 17). According to the current nomenclatu
48、re, the genus Salmonella belongs to the family of Enterobacteriaceae and consists of only two species: S. enterica and S. bongori. S. enterica is divided into six subspecies: S. enterica subsp. enterica, S. enterica subsp. salamae, S. enterica subsp. arizonae, S. enterica subsp. diarizonae, S. enter
49、ica subsp. houtenae, and S. enterica subsp. indica.Salmonella serovars belonging to S. enterica subsp. enterica are isolated most frequently (more than 99,5 % of isolated Salmonella strains) and they are designated by a name, usually related to the geographical place where the serovar was first isolated. Serovars belonging to other subspecies of S. enterica and those of S. bongori are designated by their antigenic formula.Due to combinations of subspecies and many serovars, the full names are lo
