1、BRITISH STANDARD BS ISO 18593:2004 Microbiology of food and animal feeding stuffs Horizontal methods for sampling techniques from surfaces using contact plates and swabs ICS 07.100.30 BS ISO 18593:2004 This British Standard was published under the authority of the Standards Policy and Strategy Commi
2、ttee on 17 June 2004 BSI 17 June 2004 ISBN 0 580 43959 3 National foreword This British Standard reproduces verbatim ISO 18593:2004 and implements it as the UK national standard. The UK participation in its preparation was entrusted to Technical Committee AW/9, Microbiology, which has the responsibi
3、lity to: A list of organizations represented on this committee can be obtained on request to its secretary. Cross-references The British Standards which implement international publications referred to in this document may be found in the BSI Catalogue under the section entitled “International Stand
4、ards Correspondence Index”, 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
5、 does not of itself confer 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 the UK interests informed; monitor related international and European d
6、evelopments and promulgate them in the UK. Summary of pages This document comprises a front cover, an inside front cover, the ISO title page, pages ii to iv, pages 1 to 8, an inside back cover and a back cover. The BSI copyright notice displayed in this document indicates when the document was last
7、issued. Amendments issued since publication Amd. No. Date Comments Reference number ISO 18593:2004(E) OSI 4002INTERNATIONAL STANDARD ISO 18593 First edition 2004-06-01 Microbiology of food and animal feeding stuffs Horizontal methods for sampling techniques from surfaces using contact plates and swa
8、bs Microbiologie des aliments Mthodes horizontales pour les techniques de prlvement sur des surfaces, au moyen de botes de contact et dcouvillons BSISO18593:2004IS:39581 O4002(E) DPlcsid Fremia ihTs PDF file may ctnoian emdebt dedyfepcaes. In ccaocnadrw eith Aebods licensilop gnic,y this file mairp
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13、 74 E-mail coirypthgiso.o gr We bwww.is.o gro Pulbisdehi n Switlrez dnaii ISO 4002 Allr ithgsr esedevrBSISO18593:2004IS:39581 O4002(E) I SO 4002 All irthgs ersedevr iiiForeword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member
14、 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
15、 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 Directiv
16、es, 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
17、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 18593 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee
18、SC 9, Microbiology. BSISO18593:2004IS:39581 O4002(E) iv I SO 4002 All irthgs ersedevrIntroduction It can be important to determine the presence of, or the number of, viable microbes on the surfaces of utensils, work surfaces and other equipment in contact with food to estimate the level of contamina
19、tion during production, or the effectiveness of cleaning and disinfecting protocols. The horizontal methods described in this International Standard include a surface contact method using contact plates and a swab method. The contact plate method is only applicable to flat surfaces, whereas the swab
20、 method can be used for all types of surfaces. For the sampling of large surfaces ( 100 cm 2 ), sterile cloths or sponges can be used. This alternative method is useful for the estimation of the microbial load of surfaces. Results are often presented as hygiene scores based on the number of colony-f
21、orming units (CFU) per square centimetre present on a test surface. BSISO18593:2004INTENRATIONAL TSANDADR IS:39581 O4002(E)I SO 4002 All irthgs ersedevr 1Microbiology of food and animal feeding stuffs Horizontal methods for sampling techniques from surfaces using contact plates and swabs 1 Scope Thi
22、s International Standard specifies horizontal methods for sampling techniques using contact plates or swabs on surfaces in the food industry environment (and food processing plants), with a view of detecting or enumerating viable microorganisms. NOTE The term “environment” means any item in contact
23、with the food product or likely to represent a contamination or recontamination source, for example, material, premises, operators. 2 Normative references The following referenced documents are indispensable for the application of this document. For dated references, only the edition cited applies.
24、For undated references, the latest edition of the referenced document (including any amendments) applies. ISO 6887-1, Microbiology of food and animal feeding stuffs Preparation of test samples, initial suspension and decimal dilutions for microbiological examination Part 1: General rules for the pre
25、paration of the initial suspension and decimal dilutions ISO 7218, Microbiology of food and animal feeding stuffs General rules for microbiological examinations 3 Principle 3.1 Because these methods are not quantitatively reliable or reproducible, results should only be used in a “trend analysis”. 3
26、.2 A contact plate (or dipslide) filled with a suitable agar medium is pressed against the surface to be tested. After incubation, an estimate of the surface contamination is obtained by counting the number of developed colonies. 3.3 Using the swab method, a specified area of the surface to be exami
27、ned is marked (e.g. using a template) and then wiped. The swab sticks are broken into a tube or bottle containing a sterile dilution fluid or neutralizing fluid and mixed by hand. If the surface is wiped with a sterile (damp) cloth or sponge, the sampling device is stored in a known volume of diluti
28、on liquid (e.g. 100 ml for 100 cm 2 ). In the laboratory the initial suspension and, if necessary, further decimal dilutions are used to determine the number of microorganisms using the procedures described in the methods for the enumeration of the (groups of) microorganisms to be investigated. NOTE
29、 The incubation time and temperature depend on the type of microorganisms to be detected. BSISO18593:2004IS:39581 O4002(E) 2 I SO 4002 All irthgs ersedevr3.4 For selective media, appropriate confirmatory tests can be performed. The number of colony-forming units of specific microorganisms per square
30、 centimetre or per swab is calculated from the number of (confirmed) colonies. 3.5 After sampling, the surface is cleaned and disinfected, if necessary, to avoid traces of nutrients resulting from the sampling procedure remaining on the sampled surface. 4 Culture media and dilution fluid NOTE For fu
31、rther information, see the relevant International Standards for the target microorganisms to be detected or enumerated. 4.1 Neutralizing liquid In general, the base for neutralizing liquid is buffered peptone water, or peptone salt, or any other appropriate diluent (such as quarter-stength Ringers s
32、olution, phosphate buffer at pH 7,5, peptone solution at 1 g/l). In cases where residues of disinfectants are expected, appropriate neutralizers should be added to the dilution fluid and to the media used on the contact plates to prevent any inhibitory effect of the disinfectants on the growth of mi
33、croorganisms. 5 Apparatus and glassware For general requirements, see ISO 7218. Disposable apparatus is an acceptable alternative to reusable glassware if it has similar specifications. Usual microbiological laboratory apparatus and, in particular, the following. 5.1 Contact plate, plastic dish with
34、 diameter 65 mm, filled with a controlled volume of agar medium (chosen according to the target microrganisms), especially made for the sampling of surfaces. Dishes vary in diameter or area, according to the type of surface to be sampled. The agar shall form a convex meniscus with the dish. NOTE It
35、is also possible to use any other device (nutritive medium in a flexible or rigid container) which enables contact with the surface to be sampled, such as a dipslide (5.2). 5.2 Dipslide, synthetic slide (7 cm 2to 10 cm 2 ), one or both sides of which are covered with a layer of a solid growth medium
36、 (chosen according to the target microorganisms). NOTE Various growth media are available according to the microorganism(s) sought. 5.3 Swab, breakable stick with cotton or synthetic material (such as alginate or rayon) swab contained in a tube or envelope. The swab shall be individually wrapped and
37、 sterilized. The material used shall be documented free of inhibiting substances. NOTE For some types of surface, the cotton residues can contaminate the internal parts of these surfaces after sampling. 5.4 Cloth, damp, sterile woven material, free from antimicrobial substances, packed individually
38、in sterile plastic bags, used for the sampling of large surfaces (W 100 cm 2 ). 5.5 Sponge, damp, sterile square of flat sponge, free from antimicrobial substances, packed individually in sterile plastic bags, used for the sampling of large surfaces (W 100 cm 2 ). BSISO18593:2004IS:39581 O4002(E) I
39、SO 4002 All irthgs ersedevr 35.6 Containers, such as bottles, tubes or flasks, suitable for the sterilization and storage of culture media. 5.7 Cool box, insulated, capable of maintaining the samples at low temperature during transportation to the laboratory. 5.8 Graduated pipettes, having wide open
40、ings and a nominal capacity of 1 ml, graduated in 0,1 ml divisions, or automated pipettes delivering 100 l and 1 000 l. 5.9 Mixer, for mixing liquids in culture tubes. 5.10 Peristaltic homogenizer or homogenizer using horizontal shaking, with sterile plastic bags to prepare initial suspensions by pe
41、ristaltic movement (peristaltic homogenizer) or vibration movement (homogenizer using horizontal shaking). 5.11 Petri dishes, made of plastic, of diameter 65 mm. 5.12 pH-meter, capable of being read to the nearest 0,01 pH unit at 25 C 1 C, enabling measurements to be made which are accurate to 0,1 p
42、H unit. 5.13 Template, made of a corrosion-resistant material (e.g. a frame of stainless steel enclosing an area of 20 cm 2to 100 cm 2 ), which is easy to clean and can be sterilized. 6 Sampling techniques 6.1 General It is important that the laboratory receive a sample which is representative of th
43、e surface tested and has not been changed during transport and storage, or by residues of disinfectants. Disinfectants are generally formulated for a disinfection contact time of 5 min to 15 min. Wait for a period in accordance with the disinfectant specification before investigating the surface wit
44、h swabs or contact plates to assess the performance of the cleaning and disinfection programme (or otherwise according to the disinfectant specification). An appropriate neutralizer for all situations cannot be prescribed. Generally, sorbitan monooleate (30 g/l) and lecithin (3 g/l) are useful to ne
45、utralize residues of absorbed disinfectants (e.g. quaternary ammonium compounds, amphotericides). Sodium thiosulfate (5 g/l) is a good neutralizer for halogen-based products. In the case of peroxide-based disinfectants, catalase or peroxidase may be used as neutralizer. One unit of these enzymes cat
46、alyses the decomposition of 1 mol of hydrogen peroxide per minute at 25 C and at pH 7,0 0,2. A number of disinfectant neutralizers are recommended in EN 1276 1 , EN 1650 2 , EN 13697 3and EN 13704 4 . The components of a neutralizer which may be used in most situations is given in Table 1. Make up i
47、n a solution of peptone (1 g/l) and sodium chloride (8,5 g/l), distribute in tubes or bottles, and sterilize for 15 min at 121 C. Table 1 Neutralizer which can be used in most situations Component Conc. Sorbitan monooleate (Polysorbate 80) 30 g/l Lecithin 3 g/l Sodium thiosulfate 5 g/l L-Histidine 1
48、 g/l Saponin 30 g/l BSISO18593:2004IS:39581 O4002(E) 4 I SO 4002 All irthgs ersedevr6.2 Contact plate method After removal from the transport containers, press the agar surface of the contact plate or the dipslide firmly and without any lateral movement against the test surface. From the literature
49、it is known that optimal results for contact plates are obtained with a contact time of 10 s and a pressure obtained with a mass of 500 g. Close the contact plates or dipslides immediately after inoculation and put them back in the transport container. 6.3 Swab or cloth/sponge method 6.3.1 Swab method Remove a swab from the sterile wrapping and moisten the tip by immersing it in a tube containing dilution liquid. Press the tip
