ASTM_D_6734_-_01_2009.pdf

Designation:D673401(Reapproved 2009)Standard Test Method forLow Levels of Coliphages in Water1This standard is issued under the fixed designation D6734;the number immediately following the designation indicates the year oforiginal adoption or,in the case of revision,the year of last revision.A number in parentheses indicates the year of last reapproval.Asuperscript epsilon()indicates an editorial change since the last revision or reapproval.1.Scope1.1 This test method covers the determination of coliphagesinfective for E.coli C in water.The test method is simple,inexpensive,and yields an indication of water quality within6.5 h.This coliphage method can determine coliphages inwater down to 1 coliphage per volume of water sampled.1.2 The test method is applicable to natural fresh watersamples and to settled,filtered or finished water samples.1.3 The values stated in SI units are to be regarded asstandard.No other units of measurement are included in thisstandard.1.4 This standard does not purport to address all of thesafety concerns,if any,associated with its use.It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2.Referenced Documents2.1 ASTM Standards:2D1129 Terminology Relating to WaterD1193 Specification for Reagent WaterD3370 Practices for Sampling Water from Closed ConduitsD4201 Test Method for Coliphages in Water(Withdrawn2005)33.Terminology3.1 For definitions of terms used in this test method,refer toTerminology D1129.3.2 Definitions of Terms Specific to This Standard:3.2.1 bacterial lawn,nconfluent growth of bacteria cul-tured on an agar plate.3.2.2 coliphage,nbacterial virus capable of plaquing onthe wide-range E.coli host strain used in this assay.3.2.3 plaque,nthe circular zone of clearing(lysis)of thevisible growth of bacteria on a one or two layer agar plate,caused by the action of one or more bacteriophage.3.2.4 plaque forming unit(PFU),nthe term used to reportthe number of plaques formed on an agar culture platepreviously seeded with a microorganism susceptible to abacteriophage.Although theoretically,each plaque developsfrom the action of a single bacteriophage,microbiologists usethe term,PFU,to acknowledge that a plaque may have beenformed from the action of two or more bacteriophage in closeproximity,which is indistinguishable from that formed by asingle phage.4.Summary of Test Method4.1 Ameasured water sample is adjusted to pH 6.0 with HClor NaOH and filtered through a positively-charged filter.Thecoliphages trapped in the filter are eluted with Trypticase SoyBroth(TSB)at pH 8.5.The total eluate is divided between fourTubes of melted modified nutrient agar(MNA)and E.coli Chost culture is added to each tube.The contents of each-tubeare mixed and poured into a petri plate.The plates areincubated at 35C for 6 h.The coliphages present infect thehost bacteria and form plaques.The total number of plaques onthe four plates represents the number of coliphages in thevolume of water sample filtered.5.Significance and Use5.1 Coliphage organisms may serve as indicators of fecalcontamination.The presence of coliphages in water in theabsence of a disinfectant indicates the probable presence offecal contamination.The absolute relationship between thenumber of coliforms and coliphages in natural waters has notbeen conclusively demonstrated.Coliphages are generallymore resistant than coliforms to chlorination and may havesome advantage over coliforms as an indicator of treatmentefficiency in disinfected waters.The detection of coliphages ina water sample depends upon the use of a sensitive host strainin the coliphage assay.Coliphages may be detected by thisconcentration procedure in 6.5 h to provide important same-day information on the sanitary quality of water.The lowerdetection limit of this concentration procedure is 1 coliphageper volume of water sample tested.1This test method is under the jurisdiction of ASTM Committee D19 on Waterand is the direct responsibility of Subcommittee D19.24 on Water Microbiology.Current edition approved April 1,2009.Published April 2009.Originallyapproved in 2001.Last previous edition approved in 2001 as D6734 01.DOI:10.1520/D6734-01R09.2For referenced ASTM standards,visit the ASTM website,www.astm.org,orcontact ASTM Customer Service at serviceastm.org.For Annual Book of ASTMStandards volume information,refer to the standards Document Summary page onthe ASTM website.3The last approved version of this historical standard is referenced onwww.astm.org.Copyright ASTM International,100 Barr Harbor Drive,PO Box C700,West Conshohocken,PA 19428-2959.United States1 6.Interferences6.1 High salt concentrations,such as these found in saline orbrackish water,interfere with this test method.6.2 Water sample turbidity in excess of 25 NTU(nepholo-metric turbidity units using Ratio Turbidimeter)results indecreased plaque formation because bacterial viruses aretr