1、DD ISO/TS11059:2009ICS 67.100.01NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWDRAFT FOR DEVELOPMENTMilk and milk products Method for theenumeration ofPseudomonas spp.This Draft for Developmentwas published under theauthority of the StandardsPolicy and StrategyCommittee on 31
2、March2010 BSI 2010ISBN 978 0 580 66341 3Amendments/corrigenda issued since publicationDate CommentsDD ISO/TS 11059:2009National forewordThis Draft for Development is the UK implementation of ISO/TS11059:2009.This publication is not to be regarded as a British Standard.It is being issued in the Draft
3、 for Development series of publications andis of a provisional nature. It should be applied on this provisional basis,so that information and experience of its practical application can beobtained.Comments arising from the use of this Draft for Development arerequested so that UK experience can be r
4、eported to the internationalorganization responsible for its conversion to an international standard.A review of this publication will be initiated not later than 3 years afterits publication by the international organization so that a decision can betaken on its status. Notification of the start of
5、 the review period will bemade in an announcement in the appropriate issue of Update Standards.According to the replies received by the end of the review period,the responsible BSI Committee will decide whether to support theconversion into an international Standard, to extend the life of theTechnic
6、al Specification or to withdraw it. Comments should be sent tothe Secretary of the responsible BSI Technical Committee at BritishStandards House, 389 Chiswick High Road, London W4 4AL.The UK participation in its preparation was entrusted to TechnicalCommittee AW/5, Chemical analysis of milk and milk
7、 products.A list of organizations represented on this committee can be obtained onrequest to its secretary.This publication does not purport to include all the necessary provisionsof a contract. Users are responsible for its correct application.Compliance with a British Standard cannot confer immuni
8、tyfrom legal obligations.DD ISO/TS 11059:2009Reference numbersISO/TS 11059:2009(E)IDF/RM 225:2009(E)ISO and IDF 2009TECHNICAL SPECIFICATION ISO/TS11059IDF/RM225First edition2009-08-01Milk and milk products Method for the enumeration of Pseudomonas spp. Lait et produits laitiers Mthode de dnombrement
9、 des Pseudomonas spp. DD ISO/TS 11059:2009ISO/TS 11059:2009(E) IDF/RM 225:2009(E) PDF disclaimer This PDF file may contain embedded typefaces. In accordance with Adobes licensing policy, this file may be printed or viewed but shall not be edited unless the typefaces which are embedded are licensed t
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13、any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO or IDF at the respective address below. ISO copyright office International Dairy Federation Case postale 56 CH-1211 Geneva 20 Diamant Building Boulevard Auguste Reyers 80 B-1030 B
14、russels Tel. + 41 22 749 01 11 Tel. + 32 2 733 98 88 Fax + 41 22 749 09 47 Fax + 32 2 733 04 13 E-mail copyrightiso.org E-mail infofil-idf.org Web www.iso.org Web www.fil-idf.org Published in Switzerland ii ISO and IDF 2009 All rights reservedDD ISO/TS 11059:2009ISO/TS 11059:2009(E) IDF/RM 225:2009(
15、E) ISO and IDF 2009 All rights reserved iiiForeword 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
16、 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 Elect
17、rotechnical 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 adopte
18、d 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. In other circumstances, particularly when there is an urgent market requirement for such documents, a technic
19、al committee may decide to publish other types of document: an ISO Publicly Available Specification (ISO/PAS) represents an agreement between technical experts in an ISO working group and is accepted for publication if it is approved by more than 50 % of the members of the parent committee casting a
20、 vote; an ISO Technical Specification (ISO/TS) represents an agreement between the members of a technical committee and is accepted for publication if it is approved by 2/3 of the members of the committee casting a vote. An ISO/PAS or ISO/TS is reviewed after three years in order to decide whether i
21、t will be confirmed for a further three years, revised to become an International Standard, or withdrawn. If the ISO/PAS or ISO/TS is confirmed, it is reviewed again after a further three years, at which time it must either be transformed into an International Standard or be withdrawn. Attention is
22、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/TS 11059IDF/RM 225 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and m
23、ilk products, and the International Dairy Federation (IDF). It is being published jointly by ISO and IDF. DD ISO/TS 11059:2009ISO/TS 11059:2009(E) IDF/RM 225:2009(E) iv ISO and IDF 2009 All rights reservedForeword IDF (the International Dairy Federation) is a non-profit organization representing the
24、 dairy sector worldwide. IDF membership comprises National Committees in every member country as well as regional dairy associations having signed a formal agreement on cooperation with IDF. All members of IDF have the right to be represented on the IDF Standing Committees carrying out the technical
25、 work. IDF collaborates with ISO in the development of standard methods of analysis and sampling for milk and milk products. The main task of technical committees is to prepare International Standards. Draft International Standards adopted by the Action Teams and Standing Committees are circulated t
26、o the National Committees for voting. Publication as an International Standard requires approval by at least 50 % of the IDF National Committees casting a vote. In other circumstances, particularly when there is an urgent market requirement for such documents, a Standing Committee may decide to publ
27、ish an other type of normative document which is called by IDF: Reviewed method. Such a method represents an agreement between the members of a Standing Committee and is accepted for publication if it is approved by at least 50 % of the committee members casting a vote. A Reviewed method is equal to
28、 an ISO/PAS or ISO/TS and will, therefore, also be published jointly under ISO conditions. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. IDF shall not be held responsible for identifying any or all such patent rights. ISO/TS 110
29、59IDF/RM 225 was prepared by the International Dairy Federation (IDF) and Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and milk products. It is being published jointly by IDF and ISO. All work was carried out by the Joint ISO-IDF Action Team on Microbiological harmonization
30、of the Standing Committee on Microbiological methods of analysis under the aegis of its project leader, Mrs P. Rollier (FR). DD ISO/TS 11059:2009TECHNICAL SPECIFICATION ISO/TS 11059:2009(E)IDF/RM 225:2009(E) ISO and IDF 2009 All rights reserved 1Milk and milk products Method for the enumeration of P
31、seudomonas spp. 1 Scope This Technical Specification describes a method for the enumeration of Pseudomonas spp. in milk and milk products. The method allows the isolation of all pigmented and non-pigmented psychrophilic Pseudomonas spp. This Technical Specification is also applicable to dairy enviro
32、nmental samples. 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-
33、5, Microbiology of food and animal feeding stuffs Preparation of test samples, initial suspension and decimal dilutions for microbiological examination Part 5: Specific rules for the preparation of milk and milk products ISO 7218, Microbiology of food and animal feeding stuffs General requirements a
34、nd guidance 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/TS 11133-2, Microbiology of
35、 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 of this document, the following terms and definitions apply. 3.1 Pseudomonas spp. species of bacteri
36、a of the genus Pseudomonas which form colonies in penicillin and pimaricin agar (PPA) at 25 C displaying the biochemical characteristics described, when tested as described in this Technical Specification 4 Principle The surface of a solid selective culture medium is inoculated with a specified quan
37、tity of the test sample if the product is liquid, or with a specified quantity of the initial suspension in the case of other products. The inoculation is done, under the same conditions, using decimal dilutions of the test sample or of the initial suspension. DD ISO/TS 11059:2009ISO/TS 11059:2009(E
38、) IDF/RM 225:2009(E) 2 ISO and IDF 2009 All rights reservedThe plates are incubated aerobically at 25 C for 48 h. The number of Pseudomonas is calculated per millilitre, or per gram, of sample from the number of colonies obtained on plates at dilution levels chosen so as to give a significant result
39、, and after confirmation of the selected colonies by the oxidase test and glucose fermentation test. 5 Diluent, culture media and reagent 5.1 General For current laboratory practice, see ISO/TS 11133-1 and ISO/TS 11133-2. 5.2 Diluent See ISO 6887-5. 5.3 Penicillin and pimaricin agar (PPA) 5.3.1 Basi
40、c medium 5.3.1.1 Composition Component Amount Enzymatic digest of gelatine 16,0 g Enzymatic digest of casein 10,0 g Potassium sulfate (K2SO4) 10,0 g Magnesium chloride (MgCl2) 1,4 g Agar 12,0 g to 18,0 g aWater 1 000 ml aDepending on the gel strength of the agar. 5.3.1.2 Preparation Dissolve the com
41、ponents or the dehydrated complete medium in the water by boiling. Adjust the pH, if necessary, so that after sterilization it is 7,2 0,2 at 25 C. Dispense the basic medium into flasks or bottles of appropriate capacity. Sterilize the medium in the autoclave (6.1) at 121 C for 15 min. 5.3.2 Inhibito
42、r solutions 5.3.2.1 Penicillin solution 5.3.2.1.1 Composition Component Amount Penicillin G, potassium salt 106IU Water 10 mlDD ISO/TS 11059:2009ISO/TS 11059:2009(E) IDF/RM 225:2009(E) ISO and IDF 2009 All rights reserved 35.3.2.1.2 Preparation Dissolve the penicillin in the water and sterilize the
43、solution by filtration. The penicillin solution can be stored at 5 C 3 C for one week or frozen in aliquots at 20 C for 6 months. 5.3.2.2 Pimaricin solution 5.3.2.2.1 Composition Component Amount Pimaricin (natamycin) 0,1 g Water 10 ml5.3.2.2.2 Preparation Dissolve the pimaricin in the water. Steril
44、ize the solution in the autoclave (6.1) at 110 C for 20 min. Pimaricin in solution is not stable, and has to be protected from light. Therefore, use the pimaricin solution on the day of its preparation. Alternatively, the solution can be stored frozen in aliquots at 20 C for 6 months. 5.3.3 Complete
45、 medium 5.3.3.1 Composition Volume Final concentration Basic medium 100 ml Penicillin solution 0,1 ml 100 000 IU/l Pimaricin solution 0,1 ml 0,01 g/l 5.3.3.2 Preparation Under aseptic conditions, add the inhibitor solutions to the basic medium, melted and maintained at between 44 C and 47 C. Careful
46、ly mix the thus prepared complete medium. 5.3.4 Preparation of PPA agar plates For preparation and drying of the plates, see ISO/TS 11133-1. If prepared in advance, the agar plates can be kept in the dark at 5 C 3 C for no longer than 1 day. DD ISO/TS 11059:2009ISO/TS 11059:2009(E) IDF/RM 225:2009(E
47、) 4 ISO and IDF 2009 All rights reserved5.3.5 Performance testing for quality control 5.3.5.1 Productivity Incubation: At 25 C 1 C for 48 h 2 h Strain: Pseudomonas fluorescens ATCC 13525 or Pseudomonas aeruginosa ATCC 27853 Reference medium: Soybean-casein digest agar medium (TSA) Method of control:
48、 Quantitative Criteria: Productivity ratio, PR 0,5 and non-characteristic colonies 5.3.5.2 Selectivity Incubation: At 25 C 1 C for 48 h 2 h Strain: Escherichia coli ATCC 25922 or Staphylococcus aureus ATCC 25923 Method of control: Qualitative Criteria: Total inhibition 5.4 Nutrient agar 5.4.1 Compos
49、ition Component Amount Meat extract 3,0 g Enzymatic digest of animal tissues 5,0 g Agar 12,0 g to 18,0 gaWater 1 000 ml aDepending on the gel strength of the agar. 5.4.2 Preparation Dissolve the dehydrated components or the dehydrated complete medium in the water by boiling. Adjust the pH, if necessary, so that after sterilization it is 7,0 0,2 at 25 C. Dispense the culture medium into tubes or bottles of appropriate capacity. Sterilize the nutrient agar in the autoclave (6.1) at 121 C
