1、 Collection of SANS standards in electronic format (PDF) 1. Copyright This standard is available to staff members of companies that have subscribed to the complete collection of SANS standards in accordance with a formal copyright agreement. This document may reside on a CENTRAL FILE SERVER or INTRA
2、NET SYSTEM only. Unless specific permission has been granted, this document MAY NOT be sent or given to staff members from other companies or organizations. Doing so would constitute a VIOLATION of SABS copyright rules. 2. Indemnity The South African Bureau of Standards accepts no liability for any
3、damage whatsoever than may result from the use of this material or the information contain therein, irrespective of the cause and quantum thereof. ICS 07.100.20 ISBN 0-626-16169-X SANS 10705-4:2005Edition 1ISO 10705-4:2001Edition 1SOUTH AFRICAN NATIONAL STANDARD Water quality Detection and enumerati
4、on of bacteriophages Part 4: Enumeration of bacteriophages infecting Bacteroides fragilis This national standard is the identical implementation of ISO 10705-4:2001 and is adopted with the permission of the International Organization for Standardization. Published by Standards South Africa 1 dr late
5、gan road groenkloof private bag x191 pretoria 0001 tel: 012 428 7911 fax: 012 344 1568 international code + 27 12 www.stansa.co.za Standards South Africa SANS 10705-4:2005 Edition 1 ISO 10705-4:2001 Edition 1 Table of changes Change No. Date Scope Abstract Specifies a method for the detection and en
6、umeration of bacteriophages infecting Bacteroides fragilis by incubating the sample with an appropriate host-strain. The method is applicable to all kinds of water, sediments and sludge extracts, where necessary after dilution. In the case of low phage numbers, a pre-concentration step may be necess
7、ary. Keywords bacteria count methods, bacteriophages, Bacteroides fragilis, counting, microbiological analysis, quality, verification, water, water testing. National foreword This South African standard was approved by National Committee StanSA SC 5140.19B, Water Biological treatment and assessment
8、of water, in accordance with procedures of Standards South Africa, in compliance with annex 3 of the WTO/TBT agreement. INTERNATIONALSTANDARDISO10705-4First edition2001-12-15Reference numberISO 10705-4:2001(E) ISO 2001Water quality Detection and enumeration of bacteriophages Part 4:Enumeration of ba
9、cteriophages infecting Bacteroides fragilisQualit de leau Dtection et dnombrement des bactriophages Partie 4: Dnombrement des bactriophages infectant Bacteroides fragilisISO 10705-4:2001(E)ii ISO 2001 All rights reservedPDF disclaimerThis PDF file may contain embedded typefaces. In accordance with A
10、dobes licensing policy, this file may be printed or viewed but shall not beedited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In downloading this file,parties accept therein the responsibility of not infringing Adobes licensing policy
11、. The ISO Central Secretariat accepts no liability in this area.Adobe is a trademark of Adobe Systems Incorporated.Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation parameterswere optimized for printing. Every care h
12、as been taken to ensure that the file is suitable for use by ISO member bodies. In the unlikely eventthat a problem relating to it is found, please inform the Central Secretariat at the address given below. ISO 2001All rights reserved. Unless otherwise specified, no part of this publication may be r
13、eproduced or utilized in any form or by any means,electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISOsmember body in the country of the requester.ISO copyright officeCase postale 56 CH-1211 Geneva 20Tel. + 41 22 74
14、9 01 11Fax + 41 22 749 09 47E-mail copyrightiso.chWeb www.iso.chPrinted in SwitzerlandISO 10705-4:2001(E) ISO 2001 All rights reserved iiiContents Page1 Scope . 12 Normative references . 13 Term and definition . 14 Safety precautions 25 Principle 26 Reagents 27 Apparatus . 28 Microbiological referen
15、ce cultures 39 Sampling . 410 Preparation of test materials . 411 Procedure . 712 Expression of results 913 Test report 10AnnexesA General description of bacteriophages infecting Bacteroides fragilis 11B Culture media and diluents . 12C Alternative method for preparing inoculum cultures 17D Culturin
16、g of Bacteriophage B56-3. 18Bibliography. 19ISO 10705-4:2001(E)iv ISO 2001 All rights reservedForewordISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISOmember bodies). The work of preparing International Standards is normally carried
17、out through ISO technicalcommittees. Each member body interested in a subject for which a technical committee has been established hasthe right to be represented on that committee. International organizations, governmental and non-governmental, inliaison with ISO, also take part in the work. ISO col
18、laborates closely with the International ElectrotechnicalCommission (IEC) on all matters of electrotechnical standardization.International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 3.Draft International Standards adopted by the technical committees are
19、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.Attention is drawn to the possibility that some of the elements of this part of ISO 10705 may be the subject of patentrights. ISO shall not be he
20、ld responsible for identifying any or all such patent rights.International Standard ISO 10705-4 was prepared by Technical Committee ISO/TC 147, Water quality,Subcommittee SC 4, Microbiological methods.ISO 10705 consists of the following parts, under the general title Water quality Detection and enum
21、eration ofbacteriophages: Part 1: Enumeration of F-specific RNA bacteriophages Part 2: Enumeration of somatic coliphages Part 3: Validation of methods for concentration of bacteriophages from water Part 4: Enumeration of bacteriophages infecting Bacteroides fragilisAnnexes A, B, C and D of this part
22、 of ISO 10705 are for information only.INTERNATIONAL STANDARD ISO 10705-4:2001(E) ISO 2001 All rights reserved 1Water quality Detection and enumeration of bacteriophages Part 4:Enumeration of bacteriophages infecting Bacteroides fragilis1 ScopeThis part of ISO 10705 specifies a method for the detect
23、ion and enumeration of bacteriophages infecting Bacteroidesfragilis by incubating the sample with an appropriate host-strain. The method is applicable to all kinds of water,sediments and sludge extracts, where necessary after dilution. In the case of low phage numbers, a pre-concentration step may b
24、e necessary for which a separate International Standard has been developed. The methodis also applicable to shellfish extracts.NOTE It is desirable for International Standards to be adopted as widely as possible. This part of ISO 10705 includes referenceto alternative procedures which obviate the ne
25、ed for expensive materials or equipment which may not be readily available indeveloping countries. Use of these alternatives will not affect the performance of this method.2 Normative referencesThe following normative documents contain provisions which, through reference in this text, constitute pro
26、visions ofthis part of ISO 10705. For dated references, subsequent amendments to, or revisions of, any of these publicationsdo not apply. However, parties to agreements based on this part of ISO 10705 are encouraged to investigate thepossibility of applying the most recent editions of the normative
27、documents indicated below. For undated references,the latest edition of the normative document referred to applies. Members of ISO and IEC maintain registers ofcurrently valid International Standards.ISO 3696, Water for analytical laboratory use Specification and test methodsISO 5667-1, Water qualit
28、y Sampling Part 1: Guidance on the design of sampling programmesISO 5667-2, Water quality Sampling Part 2: Guidance on sampling techniquesISO 5667-3, Water quality Sampling Part 3: Guidance on the preservation and handling of samplesISO 6887-1, Microbiology of food and animal feeding stuffs Preparat
29、ion of test samples, initial suspension anddecimal dilutions for microbiological examination Part 1: General rules for the preparation of the initial suspensionand decimal dilutionsISO 8199, Water quality General guide to the enumeration of micro-organisms by culture3 Term and definitionFor the purp
30、oses of this part of ISO 10705, the following term and definition applies.3.1 bacteriophage infecting Bacteroides fragilisbacterial virus which is capable of infecting selected Bacteroides fragilis host strains by attachment to the bacterialcell wall as the first step of the infectious processNOTE 1
31、 Such bacteriophages produce visible plaques (clearance zones) in a confluent lawn of host bacteria grown underappropriate culture conditions.ISO 10705-4:2001(E)2 ISO 2001 All rights reservedNOTE 2 A general description of bacteriophages infecting B. fragilis is given in annex A.4 Safety precautions
32、The host strain used in this part of ISO 10705 is non-pathogenic to man and animals and should be handled inaccordance with the normal (national or international) safety procedures for bacteriological laboratories.Bacteriophages infecting Bacteroides fragilis are non-pathogenic for man and animals,
33、but some types are veryresistant to drying. Appropriate precautions shall be taken to prevent cross-contamination of test materials,particularly when examining or handling cultures of high titre or when inoculating cultures of the host strains. Suchprocedures shall be carried out in a biohazard cabi
34、net or a separate area of the laboratory. Chloroform is acarcinogenic substance. Observe relevant safety precautions or use an alternative method of equal efficacy.It is recommended that personnel using this method have or acquire some experience in handling anaerobicbacteria.5PrincipleThe sample is
35、 mixed with a small volume of semi-solid nutrient medium. A culture of host-strain is added and platedon a solid nutrient medium. After this, incubation and reading of plaques take place. The results are expressed as thenumber of plaque-forming particles (also named plaque-forming units, pfu) per un
36、it of volume (pfp/ml, pfp/l, etc.).6Reagents6.1 Basic materials.Use ingredients of uniform quality and chemicals of analytical grade for the preparation of culture media andreagents and follow the instructions given in annex B. For information on storage see ISO 8199, except whereindicated in this p
37、art of ISO 10705. Alternatively, use dehydrated complete media and follow strictly themanufacturers instructions.Other grades of chemicals may be used provided they can be shown to lead to the same results.6.2 Water, for the preparation of media, glass-distilled or deionized, free from substances wh
38、ich might inhibitbacterial growth under the conditions of the test, and complying with ISO 3696.6.3 Diluent, for making sample dilution, such as peptone-saline solution or another diluent complying withISO 6887-1.7 ApparatusApart from apparatus supplied sterile, sterilize any glassware and other equ
39、ipment in accordance with ISO 8199.Usual sterile, microbiological laboratory equipment and glassware or disposable plastics-ware in accordance withISO 8199 and including the following:7.1 Hot-air oven, for dry-heat sterilization, and an autoclave.7.2 Incubator or water bath, thermostatically maintai
40、ned at .7.3 Incubator or water bath, thermostatically maintained at with a shaking device.7.4 Water bath or heating block, thermostatically maintained at .7.5 Water bath or equivalent device, for melting of agar media.7.6 pH meter, and pH paper.(36 2)C(36 2)C(45 1)CISO 10705-4:2001(E) ISO 2001 All r
41、ights reserved 37.7 Counting apparatus, with indirect, oblique light.7.8 Deep freezer, thermostatically maintained at .7.9 Deep freezer, thermostatically maintained at or liquid nitrogen storage vessel.7.10 Spectrophotometer, equipped with a filter for the range of to with a maximum bandwidth of, ca
42、pable of holding cuvettes (7.21) having an optical path length of or Hungate glass tubes (7.20) withbutyl rubber stopper and screw cap or screw-capped glass culture tubes.7.11 Anaerobic cabinet, or jars or bags, as well as anaerobiosis generators and anaerobiosis indicators.7.12 Refrigerator, temper
43、ature set at .7.13 Petri dishes, having a diameter of , and vented.7.14 Graduated pipettes, having a capacity of , , and and Pasteur pipettes.7.15 Glass bottles, of suitable volumes.7.16 Screw-capped glass bottles, of suitable volumes.7.17 Culture tubes, with caps or suitable alternatives.7.18 Screw
44、-capped glass culture tubes.7.19 Measuring cylinders, of suitable capacity.7.20 Hungate glass tubes, with butyl rubber stopper and screw cap or screw-capped glass culture tubes whichcan fit in the spectrophotometer (see Figure 1).7.21 Cuvettes, having an optical path length of .7.22 Membrane filter
45、units, for decontamination, having a pore size of , preferably low protein-bindingmembranes, as for example, those composed of polyvinylidene difluoride.7.23 Plastics vials, lidded, having a capacity of .7.24 Glass vials, screw-capped, having a capacity of .7.25 Syringes and needles.7.26 Cotton swab
46、s.8 Microbiological reference culturesThe recommended host strain is Bacteroides fragilis RYC2056 (ATCC 700786).1Use bacteriophage B56-3 (ATCC 700786-B1) infecting Bacteroides fragilis RYC2056 for the preparation of referencematerials (11.4).NOTE The ATCC strains are available from American Type Cul
47、ture Collection (ATCC), 10801 University Boulevard, Manassas,VA 20110-2209, USA. This information is given for the convenience of users of this part of ISO 10705 and does not constitue anendorsement by ISO of the product named. Equivalent products may be used if they can be shown to lead to the same
48、 results.(20 5)C(70 10)C500 nm 650 nm 10 nm 1 cm(5 3)C9cm0,1 ml 1 ml 5 ml 10 ml1cm0,2 m3ml3mlISO 10705-4:2001(E)4 ISO 2001 All rights reserved9 SamplingTake samples and deliver them to the laboratory in accordance with ISO 8199, ISO 5667-1, ISO 5667-2 andISO 5667-3. 10 Preparation of test materials1
49、0.1 Culturing and maintenance of host strains10.1.1 GeneralThe culturing and maintenance of host strains involves several stages which are summarized in Figure 2.Bacteroides fragilis is an obligate anaerobe. However, it does not require handling under conditions of strictanaerobiosis. Incubation of cultures in solid media should be carried out in an anaerobic cabinet, or anaerobic jars orbags. When using liquid media it is sufficient to ensure that containers are completely filled a
