IsEnvironmentalAntibioticResistanceaFakeIssue?

IanPepperTheUniversityofArizona

Water&EnergySustainableTechnology(WEST)Center

BiosolidsWorkshop:LinkingIndustryandResearchRMITUniversity,Australia

August2017

BACKGROUNDAntibiotics:• Naturalcompoundsproducedbysoilmicroorganismsthatkillorinhibitthegrowthofothercompetingmicroorganisms

History:• 1929AlexanderFlemingdiscoverspenicillinisolatedfromsoilfungusPenicillium

• 1943SelmanWaksmandiscoversstreptomycinisolatedfromthesoilactinomyceteStreptomyces

Zoneofinhibitionofbacterialgrowthonaspreadplate

BACKGROUND:TheConcern

• Themoreanantibioticisusedthegreaterthelikelihoodofantibioticresistantstrains

• Themoretheantibioticisusedtofightinfectiousdisease,thelesseffectiveitbecomes

• Ofparticularconcern,arebacteriaresistanttomultipleantibiotics

e.g.Methicillin-resistantStaphylococcusaureus(MRSA)

MICROBIALPRODUCTIONOFANTIBIOTICS

• Vastmajorityofantibioticssynthesizedbysoilmicroorganisms‒ PenicillineffectiveonGm+vebacteria‒ PolymixineffectiveonGm–ve‒ ChloramphenicoleffectiveagainstGm+veand-ve

• Antibioticsproducedbybacteria,actinomycetesandfungi• Utilizedasmajorformofself-defenseagainstotherindigenoussoilmicrobes

SOURCEOFANTIBIOTICSARBsandARGs

Anthropogenic:•Sewageeffluentsandbiosolids•CAFOeffluentandanimalmanures•Hospitalwastesdischargedintosewers

Natural:•Soils•Water

ENVIRONMENTALANTIBIOTICRESISTANCE

• Newtermintroducedin2013• Causedbyanthropogenicactivity•Wastewatertreatmentplantsblamedforincreasing“environmentalantibioticresistance”

Rizzoetal.,2013:“UrbanWastewaterTreatmentPlantsasHotspotsforAntibioticResistantBacteriaandGenesSpreadintotheEnvironment:AReview”

ISTHISTRUE:afreshperspective?

SOILBACTERIACulturablebacteria=108 – 1010 pergramsoilTotalbacteria=1010 – 1012 pergramsoil

Oneacrefurrowslice � 2x106 lbssoil(1acretodepthof6”)

=9x108 gramssoil

Assume109 bacteriapergramsoil

Therefore,109 x(9x108)bacteriaperacrefurrowslice� 1018

ANTIBIOTICRESISTANTBACTERIA(ARB)

Naturallyoccurringinsoilfor3billionyears(Gazeetal.,2013)

Presentinallsoilsevenpristinesoils

Foranygivenantibioticthereare106 – 107 culturableARB

Therefore,for1acrefurrowslicewehave:

9x108 x5x106 � 4.5x1015 ARB

Table1.DocumenteddetectablelevelsofARBandARGintheenvironment.

SampleType ARB(CFU/gorCFU/ml)

Ref ARG(#gene

copies/gorml)

Ref

Soil 106 – 107 Brooksetal.,2007 108 – 109* Seetext

ClassBBiosolids 104 – 109107

Muniretal.,2011McCalletal.,2016

106 - 109 Muniretal.,2011

TreatedMunicipalWastewatereffluent

101 – 103 Gaoetal.,2012 104 – 107 Muniretal.,2011

PoultryManure 105 - 108 Brooksetal.,2010 106 – 1011 Brooksetal.,2016McKinneyetal.,2010

BovineManure 105- 108 Yangetal.,2010 103 – 1010 Yangetal.,2010Muniretal.,2011

SwineManure 105 - 107 Sengelovetal.,2003 104 – 109 Brooksetal.,2014

IMPACTOFLANDAPPLICATIONOFBIOSOLIDS

Assume2metrictonsbiosolidsappliedperacre

2metrictons=2x106 gbiosolids

FromTable1,geometricmeanvalueofARB/g

biosolids=4.6x=106

Therefore,4.6x106 x 2x106 ARBadded

=9.2x1012 additionalARBapplied

%INCREASEDUETOLANDAPPLICATION

%#$%&'()' = (++,-,.$(/012),$-&,$),%).,/012)

%#$%&'()' = 9.26109:2.96109;

%#$%&'()' = 0.32%

Sample ARB(CFUperacrefurrow

slice)

ARG(genecopiesperacrefurrowslice)

Soil 4.5x1015 2.9x1017

Biosolidsapplied,2TonsPerAcre

9.2x1012 6.3x1013

1AcreFootEffluent 1.2 x1011 3.9x 1014

2TonsPerAcreSolidManure 4.3x1012 2.9x1013

Table2.ImpactoflandapplicationofClassBbiosolids,effluentandmanureonARBandARGinsoilusingTable1asareference.

MaterialApplied ARB(%increase) ARG(%increase)

Biosolids 0.32 0.02

Effluent 0.0043 0.14

Manure 0.15 0.01

Table3.HypotheticalpercentincreasesofARBandARGinsoilduetolandapplication.

Table4.Concentrationsofpathogensmeasuredinsoil,ClassBbiosolidsandtreatedwastewater.

IndigenousPathogens Ref Number(pergramsoil)

Number(peracrefurrowslice)

Bacillus anthracis Ganzetal.,2014 104 �1013

Clostridiumperfringens Konishietal.,1981 900 �109

Pathogens Number(pergram)

Number(per2tonsbiosolid)

Thermotolerant(fecal)coliforms(includingE.coli)

106 1012

FecalStreptococcus 106 9x1012

Salmonella 40 4x107

Pepperetal.,2010

A)Soil

B)ClassBBiosolids

BIGGESTENVIRONMENTALCONCERN

Animalmanuresparticularlyswinemanure

• PublicationsshowthatARGlevelsincreasefollowinglandapplications• Decreasetobackgroundlevelswithin2months

Percentantibioticresistantbacteriainlandappliedbiosolids

0

2

4

6

8

10

12

14

16

Prior to

Application

Day 0 Day 7 Day 14 Month 1 Month 2Month 3 Month 4 Month 5 Month 6

% R

esis

tanc

e/g

AmpicillinCephalothinCiprofloxacinTetracycline

IMPACTOFSEWAGEEFFLUENTSANDCLASSBBIOSOLIDSONPATHOGENIC

ARBsINSOIL• MinimalARBsaddedrelativetowhatisalreadyinsoil• Numberofentericpathogens(e.g.E.coli)introducedviaeffluentandbiosolidsislessthanpathogensindigenoustosoil(e.g.Bacillusanthracis orClostridiumperfringens)

• EntericpathogensandARBsintroducedintosoilnormallydie-offquickly

• WhenE.coli adapttosoilenvironment,pathogenicitylost

• Horizontalgenetransferinsoillimitedduetospatialseparationofcells

RISKASSESSMENTFORANTIBIOTICRESISTANTPATHOGENS:

Questions&Issues• DopathogenicARBbehavelikenon-ARBpathogens?

Wedon’tknow

• IsthedoseresponsethesameforARBandnonARBpathogens?Wedon’tknow,butsomedatasuggestARBpathogensmoreinfectiousthannon-ARB

• DoARBandnonARBpathogenssurvivesimilarlyintheenvironment?SomeARBco-evolvemobilegenetictraits(transposonsandplasmids)thatenhancemetalresistanceandotherenvironmentaltolerance.BUTplasmidsareageneticburdencausingshortersurvivalduration.

RISKASSESSMENTFORANTIBIOTICRESISTANTPATHOGENS:

Questions&Issues

• Doeshorizontalgenetransfer(HGT)betweenARBsandpathogenicbacteriainsoilpresentadanger?

HGTratesnotknown,butthoughttobelow

• DoARBsmaintainplasmid-borneresistanceinsoils?Unlikelywithoutselectivepressure

RISKASSESSMENTFORANTIBIOTICRESISTANTPATHOGENS:

Questions&Issues

• DofreshinputsofantibioticsintosoilleadtoenhancedARGs?Notnecessarilysincegeneclustersforantibioticproductionalwayspresentinsoilbutonlyexpressed

whenthereareinteractionswithothermicrobes

• Basedonissues,environmentalfateandtransportofARBsandnon-ARBsassumedtobeidentical…BUTatmanifestationofdisease,riskofmorbidityandmortalitycanbealtered

QMRA– Salmonella spp. QMRA– ARBSalmonella spp.

Assumptions

Morbidity&Mortality

U.S.Population

~314million

DiseaseOutcomes

Assumptions

Morbidity&Mortality

U.S.Population

~314million

DiseaseOutcomes

• >1milliontotal(ARBandnon-ARB)annualcases(Scallan,2011)

• 378totalannualdeaths(Scallan,2011)

• 100,000ARBannualcases(CDC,2013)

• 40ARBannualdeaths(CDC,2013)

• 2per1000riskofinfection/yrforcropingestion(Brooks,2012),thus

• ~200,000infections/yrattributedtocropingestion

• Assumeriskofinfectionequivalenttonon-ARBSalmonella

• ~1/10th oftotalcasesareARB(CDC,2013),thus

• ~19,500ARBSalm infections/yr

• Mortality~66deaths/yr• 0.036%ofinfections

• Mortality~8deaths/yr• 0.040%ofinfections

• Incidenceofinfectionssimilar• Diseaseoutcomemaybedifferentifantibioticsarerequired• MortalityratessimilarbecausemostSalmonella infectionsareself-limitinganddon’trequireantibiotics

• ForMRSAinfectionsriskofmortalityisgreater

SUMMARY• Basedon:

- IncidenceofARBsnaturallypresentinsoil- NumbersofaddedARBsaddedvialand

application- Soilmicrobialecologyprinciples

• CurrentevidencesuggestsmunicipalwasteapplicationstolanddonotcauseanylongtermincreaseinARBsorARGs

• Allsoilscurrentlycontainlarge#sofindigenousantibioticresistantpathogens– notaproblem

• ForformalriskassessmentmoreresearchneededonfateandtransportofARBs

BIOSOLIDSONTRIAL:REFLECTIONSONTHEKERNCOUNTYCASE

IanPepperTheUniversityofArizona

Water&EnergySustainableTechnology(WEST)Center

BiosolidsWorkshop:LinkingIndustryandResearchRMITUniversity,Australia

August2017

KernCountyLegalHistory• KernCountypassedordinancein1999banningClassBlandapplicationbeginningin2003

• ClassAonlyrequirementadoptedin2003withnewordinance

• VotersapprovedMeasureEin2006whichprohibitedalllandapplicationinunincorporatedpartsoftheCounty(didnotimpactClassBapplicationinincorporatedpartsofcounty;e.g.CityofBakersfield

LAv.Kern:TheRoadtoTrial

• MeasureEvoterinitiativebanningallbiosolidsinKernCountypassed2006

• Coalitionofagencies,farmersandbiosolidsmanagerssuedKernCountytooverturnMeasureE

• MeasureEneverwentintoeffect;preliminaryandpermanentinjunctionswhilecasemotionsdecidedandappealed

LAvsKern:TheRoadtoTrial• MeasureEChallengedinStateCourtJanuary2011afterreleaseoffederalcase

• PreliminaryInjunction(PI)grantedinJune2011

• PIupheldinCourtofAppeals2013

BiosolidsBanOnTrial2016

• TwoweektrialApril-May2016inTulareCountySuperiorCourt

• FourclaimsagainstMeasureE:• PreemptionofbiosolidsbanbyIWMA• Banexceedslimitsoncountypolicepowers• Baninterfereswithfederalcommerce• Baninterfereswithstatecommerce

LAvsKernTrial

• Eightdaysofevidenceonbiosolidssafetyfromeightexpertsandtwelvefactwitnesses

• KernCountydefensefocusedontraceperfluorinatedcompounds(PFCs)ingroundwater

• Plaintiffsfocusedonnoimpactsafter22yearsoflandapplication

ExpertsforKern

• Kernfocusedontraceorganiccompoundsinsoilandgroundwater(PFCs)atppblevel,potentialrisks

• ChrisHiggins(ColoradoSchoolofMines)

• GwynnJohnson(PortlandState)

• GaryHokkanen(hydrogeologist)

KeyWitnessesforPlaintiffs

LAvsKernTrial• TrialextendedovertwoweeksinApril/May2016

• PreliminarydecisionissuedlateNovember2016

• FinaldecisioninAugust2017

• RuledMeasureEisinvalidduetopre-emptionofIWMAandexceedanceofpolicepowers

• Verystrongandsupportivedecision

• SupportsresponsiblemanagementofbothClassAandB

PERFLUORINATEDCOMPOUNDS(PFCs)• Fullyfluorinatedlongchainorganiccompounds• Familyofanthropogenicchemicalsusedfordecadestomakeproductsresistanttoheat,oilstains,greaseandwater

• Perfluoroctanesulfonate(PFOS)andperfluorooctanoicacid(PFOA)producedinthelargestamountsintheU.S.

• RegardedbyEPAasan“emergingcontaminant”

TriclosanandTriclocarban• Commoningredientsineverydaycleaningproducts

- Handsoaps- Dishandlaundrysoaps- Toothpaste- Mouthwash

• Triclosanshowntobioaccumulateinfishandhumans

• Triclosanshowntobeanendocrinedisruptorinaquaticorganisms

• Foundinbiosolidsatthelowppmconcentration

• FoundinColgatetoothpasteataconcentrationof3000ppm

JudgeStrikesDownMeasureENovember2016(preliminarydecision)

• MeasureEconflictswithIntegratedWasteManagementAct’smandateforrecycling

• MeasureEexceedsKernCounty’spolicepowerbecauseofimpactsonneighbors,lackofbenefits

• CourtofAppealreachedsameconclusionsin2013whenitupheldpreliminaryinjunction

42

OpinionStressesSafetyofLandApplication

• “ThereisnobasisinfactforanydeterminationthatlandapplicationofbiosolidsposesanyrisktoKernCounty”

• “LosAngeleshasmetitsburdenofproducingevidencethatthereisnobasisinfactforMeasureE’spublicwelfareclaims”

• “THEREISNOEVIDENCEOFRISKTOHUMANHEALTH”

43

NoReasonableBasisforaBan

• “Norealandsubstantialrelationship”betweenabanandthepublicwelfare

• MeasureEdoesnotaccommodateregionalinterestsincost-efficientmanagementofbiosolids

• NocountervailinginterestsinKernservedbytheban

44

NextStepsforProtectingLandApplication

• Firsttrialonthesafetyoflandapplicationresultedinaresoundingfindingforbenefitsandlackofrisk

• KernCountyspentmillionsofdollarsonsampling,experts,andoneofcountry’slargestlawfirms

• Trialcourtandappellateopinionsshouldconvincelocalitiesthatfuturebansorregulationsclosetobanswillbestruckdownatgreatexpensetoenactingjurisdiction

LAvsKern

• SettlementreachedAugust2017(FinalDecision)

• LandapplicationatGreenAcressafeandwillcontinue

• LandapplicationelsewhereinKernCountystillindispute