LaboratoryMS-2ColiphageTestingoftheSteriPENPure+
LindenLaboratory
Civil,EnvironmentalandArchitecturalEngineering
UniversityofColorado,Boulder
February19th2016
[email protected]
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Introduction
TheSteriPENPure+,ahandheldwaterpurificationdevicethatusesultraviolet(UV)lighttodisinfect
water,waschallengetestedwithMS-2coliphageinthelaboratory.TheSteriPENusesalowpressureUV
bulbwhichemitslightat254nm.TheUVlightisabsorbedbyanorganism’sDNA,inactivatingthe
organismfromfurtherreplication(1).
TheSteriPENismarketedasaportablewatertreatmentdevicetobeusedintheoutdoorand
backpackingspacewherebyhalftooneliterofwatercanbebatchtreatedinatraditionalwaterbottle
byswirlingtheSteriPENinthesourcewater.Thedeviceincludesasimpleactivationusingsensor
electrodessubmergedinthewatertotriggeratreatment.ThesensoronlyallowstheUVlightto
becomeilluminatedwhenthedeviceissubmergedinwater.Submergingthedeviceinwateractivatesa
48secondtreatmenttimeforahalfliterofwater.Theuserknowsthepenisworkingbyablinking
greenlightinadditiontotheUVlampbeingilluminated.Thegreenlightswitchesfromblinkingto
continuousgreentoindicateasuccessfultreatment.Ifthetreatmentwasunsuccessfulorwasstopped
beforethefulltreatmenttime,thelightturnsred.Thepencanberemovedfromthewaterandreinsertedtoactivateasecond48secondtreatmentforoneliterofwater.Thepen’sinternalbatterycan
berechargedusingamicro-USBcableconnectedtoastandardUSBportinacomputerorwallcharger.
TestOrganism
ThechallengeorganismwastheMS-2coliphage.MS-2waschosenbecauseitisacommonlyused
indicatororganisminUVvalidationtesting.MS-2hastheadvantageofbeingeasilypropagatedathigh
concentrationsandthuscandemonstratehighlogreductions(2).Thechallengeorganismwas
enumeratedusingthebacterialhostE.coliFamp.
Methods
TheSteriPENwastestedintwotestwaters,ageneraltestwaterandachallengetestwaterwhichmet
theUnitedStatesEnvironmentalProtectionAgency’sGuideStandardandProtocolforTesting
MicrobiologicalPurifiers(seeTableA1inAppendix).Thecharacteristicsofbothwatersareshownin
Table1below.Thegeneraltestwaterwasdechlorinated,activatedcarbonfilteredtapwater.The
challengewaterwaspreparedbyaddingdilutedligninsulfonicacid(LSA)andfinetestdustinproper
proportionstoachievethedesiredtransmittance,absorbance,totalorganiccarbon(TOC),andturbidity
characteristics.TemperatureandpHwasmeasuredusinganOrion8157BNUMDprobeattachedtoa
ThermoScientificOrionStarA211;pHadjustmentwasnotnecessary.TOCwasmeasuredusingaSievers
TOCAnalyzer.TurbiditywasmeasuredwithaHach2100NTurbidimeterandUVabsorption(1cm
cuvette)wasmeasuredwithaCary100Spectrophotometer.TDSwasadjustedthroughtheadditionof
NaClandmeasuredgravimetrically.PhotosofthetestwatersandSteriPENarepresentedinFigure1.
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Table1.TestWaterCharacteristics.
Parameter
GeneralTestWater
ChlorineResidual(ppm)
<0.05
pH
7.04
TotalOrganicCarbon(mg/L)
0.7
Turbidity(NTU)
<1
o
Temperature( C)
22
TotalDissolvedSolids(mg/L)
~100
UVAbsorbance(1/cm)(filtered)
0.0004
UVTransmittance(%)(filtered)
99.9
Note:requirementsfortestwaterswaterqualityareinTableA1.(3)
ChallengeTestWater
<0.05
7.57
12.7
34.4
5
~1500
0.1574
69.6
Figure1.Challengetestwater(left)Generaltestwater(right)showninirradiationvessels,withthe
SteriPENPure+.
MS-2coliphagewasacquiredfromGAPEnviroMicrobialServicesLtd.atastockconcentrationof
approximately1012PFU/mL.TheUVdoseresponsecurveforthisMS2stockispresentedinthe
AppendixinFigureA1.Abatchoffivelitersofboththegeneralwaterandchallengewaterwerespiked
withMS-2toobtainastartingconcentrationaround107PFU/mL.Duplicate15mLsampleswere
collectedfromthegeneralandchallengewatersbeforeandattheendofconductingallSteriPEN
exposures.
ThreeSteriPENPURE+units(denotedA,B,andC)werethentestedthreetimesineachwater.Foreach
test,onehalf-literofwaterwastransferredtoa500mLPyrexbeakerandtheSteriPENwasusedtotreat
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thewater.TheUVdosewasadministeredbyplacingthedeviceinthewaterandthewaterwas
continuouslystirredwiththedeviceatamoderatespeedforthedurationofeachtest.Forthegeneral
waterone48secondcyclewascompleted,whereastwo48secondcycleswerecompletedforthe
challengewater.Duplicate15mLsamplesofeachtreatedsamplewerecollectedfollowingeachtest.
EnumerationofMS-2coliphageforeachofthestocksolutionsamplesandeachofthetreatedsamples
wasconductedusingmethodsfollowingEPAmethod1602andStandardmethods9224E(4).Each
samplewasplatedintriplicateandplaquescountedtodetermineMS-2concentration.
Results
Tables2summarizesMS-2coliphageconcentrationsandlogreductionsforthegeneraltestwater
treatedbyone48seconddoseandthechallengewatertreatedbytwo48seconddoses.Logreduction
wascalculatedusingtheequation:logreduction=log(No/N),whereNoisthecontrolvalueandNisthe
treatedvalueforeachpenforeachwater.
Table2.MS-2coliphagetestresults.
SteriPEN
Pure+
Unit
A
B
C
Average
GeneralWater
Control
Treated
Log
(PFU/mL)
(PFU/mL) Reduction
6.93x103
3.88
3
N/A
8.26x10 3.81
3
3.47x10 4.18
7
3
5.27x10 6.22x10 3.96
ChallengeWater
Control
Treated
(PFU/mL)
(PFU/mL)
2.40x103
N/A
1.38x103
2.93x103
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4.13x10 2.24x103
Log
Reduction
4.34
4.58
4.25
4.39
Thetreatedvaluesareanaverageoftriplicateindividualexperiments,withtriplicateplatingforeach
experiment.Thecontrolvaluesareaveragesofmeasurementstakenfromthegeneralandcontrol
waterbatchesbeforeandafterSteriPENtesting,withtriplicateplating.Theaveragewascalculatedfor
thetriplicateplates,thestandarddeviationcalculatedforthetriplicateexperimentsandtheerror
propagatedthroughthecalculationofthelogreductionandtheoverallaverageforeachwater.These
dataarealsoplottedwiththeirrespectiveerrorinFigure2.
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Figure2.MS-2ColiphageLogReductionforGeneralandChallengeTestWaters.
Conclusion
TheSteriPENachievedanaverage3.96logreduction(99.989%)ofMS2inthegeneraltestwaterwitha
single48seconddose,anda4.39logreduction(99.996%)ofthechallengetestwaterwithtwo48
seconddoses.ProvidingtwodosesoftheSteriPENtothechallengewaterprovedeffectiveinMS2
inactivation,evenwiththedecreasedwaterquality.Basedontheseresultsandthedose-responsedata
oftheMS2stock,thegeneralwaterwasexposedtoanapproximateUVdoseof97mJ/cm2andthe
challengetestwaterwasexposedtoadoseofabout123mJ/cm2.
ThetestingconductedonbothgeneraltestwaterandchallengetestwaterindicatesthatSteriPEN
PURE+meetstherequirementssetforthbytheU.S.EnvironmentalProtectionAgency(EPA)andis
adequatefortheinactivationofbacterial,viral,andprotozoanpathogensindrinkingwater.TheEPA
GuideStandardandProtocolforTestingMicrobiologicalWaterPurifiers(3)requiresaminimum6-log
reduction/inactivationofbacteria,4-logreduction/inactivationofviruses(rotavirusandpoliovirus),and
3-logreduction/inactivationofprotozoan(Giardia)cysts.Basedonwellestablisheddatainthepeer
reviewedliterature(5)theinactivationofMS2coliphageatthelevelsreportedhereinforbothtesting
scenariosmeetandexceedalltherequirementssetforthintheEPAGuideStandardandProtocolfor
TestingMicrobiologicalWaterPurifiers.
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References
1. UltravioletLightDisinfectionTechnologyinDrinkingWaterApplication–AnOverview.United
StatesEnvironmentalProtectionAgency,OfficeofWater.EPA811-R-96-002.September,1996.
2. UltravioletDisinfectionGuidanceManual.UnitedStatesEnvironmentalProtectionAgency,
OfficeofWater,Washington,DC.,2006
3. USEPA-TaskForceReport.GuideStandardandProtocolforTestingMicrobiologicalWater
Purifiers.UnitedStatesEnvironmentalProtectionAgency,RegistrationDivision,Officeof
PesticideProgramsandCriteriaandStandardsDivision,OfficeofDrinkingWater,Washington,
DC.,1987.
4. Method1602:Male-Specific(F+)andSomaticColiphageinWaterbySingleAgarLayer(SAL)
Procedure.UnitedStatesEnvironmentalProtectionAgencyEPA821-R-01-029,2001.
5. Hijnen,W.,Beerendonk,E.,andMedema,G.“InactivationCreditofUVRadiationforViruses,
Bacteria,andProtozoanCystsinWater”.WaterResearch,40,3-22.2006.
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Appendix
TableA1.Requiredphysicalandchemicalcharacteristicsoftestwaters
LogInactivation (LogCo/C)
MS2LPUVdoseresponse curve
5.0
4.5
4.0
3.5
3.0
2.5
2.0
1.5
1.0
0.5
0.0
0
20
40
60
80
100
120
140
160
UVDose(mJ/cm2)
FigureA1.UVdoseresponsecurvefortheMS2coliphagestockusedinthisstudy.
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