International Standard (ISO 6461/2 INTE RNATIONAL ORGANIZATION FOR STANDARDIZATION+MEKAYHAPOAHAR OPTAHM3AUMR O CTAHAAPTM3AUWWORGANISATION INTERNATIONALE DE NORMALISATION Water quality -- Detection and enumeration of the spores of sulfite-reducing anaerobes (clostridia) Part 2: Method by membrane filtration Qualite de I'eau - Recherche et denombrement des spores de micro-organismes anaerobies sulfito-reducteurs (clostridia) - Partie 2: Méthode par filtration sur membrane First edition - 1986-02-01 (3) UDC 543.39 : 579.852.13 ISO 6461/2-1986 Ref.No.ISO 6461/2-1986 (E) Descriptors : water, quality, tests, microbiological analysis, determination, micro-organisms, sulphite reducing bacteria. Price based on 3 pages Foreword 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 body interested in a subject for which a technical committee has been established has Draft International Standards adopted by the technical committees are circulated to the member bodies for approval before their acceptance as International Standards by least 75 % approval by the member bodies voting. International IStandard ISO6461/2 was prepared byTechnical Committee ISO/TC 147, Waterquality. Users shoutd note that all International Standards undergo revision from time to time latestedition,unlessotherwisestated. @InternationalOrganizationforStandardization,1986 Printed in Switzerland ISO 6461/2-1986 (E) INTERNATIONAL STANDARD Water quality - Detection and enumeration of the spores of sulfite-reducing anaerobes (clostridia) Part 2: Method by membrane filtration 0 Introduction ISO8199, Water quality General guidance for microbiological examination by enumeration :of micro- The spores of sulfite-reducing anaerobes (clostridia) are wide organisms on culture media. 1) spread in the environment. They are present in human and animal faecal matter,in waste water and in soil. Unlike Escherichia coli and other coliform organisms, the spores sur- 4 Definition vive in water for long periods as they are more resistant than vegetative forms to the action of chemical and physical factors. For the purpose of this part of Iso 6461, the following defi- They may thus give an indication of remote or intermittent nition applies. pollution. They may even be resistant to chlorination at levels which are normally used for the treatment of water, and they clostridia: Sulfite-reducing, spore-forming, anaerobic micro arethususefulforcontrol purposes organisms which belong to the Bacillaceae family and the genus Clostridium. ISO 6461 consists of the following parts : 5 Principle Part1:Method by enrichment in a liquid medium. Part 2: Method by membrane filtration. The detection of spores of sulfite-reducing anaerobes (clostridia) in a specified volume of a water sample requires the following steps. 1 Scope 5.1 Selection of spores This part of ISo 6461 specifies a method for the detection and Selection of spores in the sampie by applying heat for a period enumeration of the sporesof sulfite-reducing anaerobes of time sufficient to destroy vegetative bacteria. (clostridia)bymembranefiltration. 5.2 Membrane filtration and culture 2Field of application Filtration of the water sample through a membrane filter having a pore size such that bacterial spores (0,2 μm) are retained in or Themethodcanbeappliedtoalf typesofwater,exceptwhena on the membrane filter. large amount of particulate materiat is liable to be retained by the membrane. Placing of the filter on a specialized selective culture medium (sulfite-iron-agar), followed by incubation at 37 ± 1 oc for 20 ± 4 h and 44 ± .4 h, and counting of any black colonies. References Culture media and reagents Iso3696,Water for laboratory use - Specifications. Iso 5667, Water quality -- Sampling 6.1 Basic materials Part2:Guidanceonsamplingtechniques. In or