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David Sanders
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1 TheCeresTrust2013GraduateStudentOrganicResearchGrant: FinalReport Project(Title: Transforming+waste:+rearing+black+soldier+flies+as+a+source+of+compost+and+ entomopathogenic+nematodes. Project(Leader:JosephS.Tourtois,GraduateStudent,MichiganStateUniversity, DepartmentofEntomology Major(Advisor:Dr.MatthewJ.Grieshop,MichiganStateUniversity,Departmentof Entomology AccomplishmentsbyObjectives: Objective1:Determine+black+soldier+fly+composting+rate+and+quality+for+plant+and+animal+ based+farm+waste+and+the+number+and+quality+of+larvae+produced+on+these+substrates.+ ( Methods(and(materials:( Wecompletedtwoexperimentaltrialstoaddressthisobjective.Onetrialtookplace atswallowtailfarmsinmason,miandthesecondtookplaceinaheatedgreenhouseatthe MSUStudentOrganicFarm.WeassignedthreeBioPodsPlus (Fig.1A)tooneoftwo treatments:(1)foodwasteonly,and(2)50:50mixtureoffoodwasteandchickenbedding (Fig.1B).WecollectedchickenbeddingfromSwallowtailFarms.Itwasstrawbasedand containedmanureandspiltchickenfeed.foodwastesconsistedofrottingfruitsand vegetablesfromthemsustudentorganicfarmandhorticulturefarm,prewconsumer wastefromamsucafeteria,andcoffeegroundsfromalocalcoffeeshop. + Trial+1+at+Swallowtail+Farm:+ WesetWuptheBioPods with5galofmaterialandapproximately700youngblack soldierflylarvae.anadditional5galofmaterialwasadded17dayslater,followedbyan additional2gal18dayslater.matureblacksoldierflylarvae(i.e.prepupae)selfwharvested bycrawlingupthebuiltwitrampofthebiopod anddroppingintothecollectionbasket, whichwasfilledwithaspenwoodchips.wecollected,counted,andweighedblacksoldier flylarvaefoundinthecollectionbasket.asampleofthedigestate remainingmaterial thatthelarvaedidnoteat fromeachbiopod wassubmittedtoa&lgreatlakes Laboratories,Inc.fornutrientanalysis. + Trial+2+in+heated+greenhouse+at+MSU+Student+Organic+Farm:+ ThroughoutthisTrialIcollaboratedwithJasonMatlock,agraduatestudentinthe MSUdepartmentofHorticulture.Feedstocksweremixedinadvancedandstoredin27gal plastictubsinawalkwinwcooleruntilneeded.biopodswereinitialstockedwith2000early instarblacksoldierfliesand5gal(7w8kg)offeedstock.additionalfeedstockinwasadded intothebiopodsonthreeadditionaldatesat1weekintervals,5galperweek.prepupae werecollected,counted,andweighedtwiceaweekfor5weeks.prepupae,digestate,and leachatewerecollectedfromeachbiopodandsubmittedfornutrientanalysisata&l GreatLakesLaboratories,Inc.

2 JasonMatlockandotherstudentsintheOrganicPestManagementclassconducteda germinationtestwithaqueousdigestateextracts.theyuseradishandlettuceseedsand fourconcentrationsoftheextractstotesttheplanttoxicityofusingblacksoldierfly digestateasasoilfertilityamendment. ( Results:( Trial+1+at+Swallowtail+Farm:+ Oneverynoticeablethingaboutwasthattheblacksoldierflylarvaedidnoteatthe strawfoundinthechickenbeddingorothersimilartoughplantstructuresthatcontained cellulose(fig.1b).eventhoughmoreblacksoldierflies(203.3±79.2)selfwharvestedfrom thefoodwastetreatmentitwasnotsignificantlymorethantheblacksoldierflies(78.0± 32.1)harvestedfromthemixtureofchickenbeddingandfoodwaste(W=8,p=0.2)(Fig. 2A).Theaveragemassoftheblacksoldierflyprepupaecollectedfromthefoodwaste (0.163±0.016g)wasnotsignificantlydifferentthanthelarvaethatfeedonfoodwasteand chickenbedding(0.169±0.004g)(w=4,p=1)(fig.2b). AsecondflyspecieswasalsocollectedfromtheBioPods.Itwasidentifiedas Ptecticus+trivittatusSay(Diptera:Stratiomyidae)whichisacloserelativeoftheblack soldierflyandcommonlyfoundinmi.ptecticus+trivittatusprepupae(0.042to0.058g)are muchsmallerthanblacksoldierflyprepuae(0.135to0.191g)andtheyhavetwocreamw coloredparallelstripesrunningthelengthoftheirbodiesbothonthedorsalandventral sides.significantlymorep.+trivittatusprepupaewereharvestedfromthebiopodsthat receivedonlyfoodwastethanthebiopodswithchickenbeddingandfoodwastemixture (138.0±86.2and5.3±0.7,respectively)(W=9,p=0.038)(Fig.3A).TheP.+trivittatus harvestedfromthechickenbeddingandfoodwastemixtureweresignificantlylargerthan theprepupaefromthetreatmentwithonlyfoodwaste(0.053±0.002gand0.045±0.001g, respectively)(w=9,p=0.038)(fig.3b). Theadditionofthechickenbeddingtothefoodwasteaffectedtheabundanceof somethenutrientsinnutrientsinthedigestate.phosophorus,calcium,zincand manganesewerehigherinthedigestatecontianingchickenbedding;whereas,potassium washigherinthedigestaewithoutchickenbedding(table1). + Trial+2+in+heated+greenhouse+at+MSU+Student+Organic+Farm:+ Oneveryapparentvisualobservationwasthatthestrawincludedinthefeedstocks wasstillintactaftertheblacksoldierflyhadfed,asobservedinthefirsttrial.wealso noticedthatseedlingsweresproutinginthebiopods,indicatingthattheblacksoldierfly didnotfeedonthem.notalloftheblacksoldierflyprepupaeselfwharvested.many remainedinthefeedingchamberofbiopod,pupated,andemergedasadults.weobserved largemitesontheadultblacksoldierfliesthatemerged.adultfliesthatemergedearlier hadahighermiteloadthanfliesthatemergedlater.miteswerenotidentified,butwe presumedthattheycamefromthechickenbeddingandwerefacultativeparasites, attachingtotheadultsfliesinhopesofdispersing.ptecticus+trivittatuswasnotobservedin thebiopodsduringthistrial. ThenumberofprepupaethatselfWharvesteddidnotdifferentbetweenthetwo treatments,225.3±39.8forfoodwasteand220.7±9.0forthechickenbeddingandfood wastemixture(w=6,p=0.7)(fig.4a).likewise,theprepupaedidnotdifferinsize,0.142 ±0.004gand0.137±0.004g,respectively(W=7,p=0.4)(Fig.4B).Theblacksoldierflies

3 reducedthemassofthefeedstockby35%inthecontroland27%inthechickenbedding treatment.certainnutrientsweremoreconcentratedinthedigestatefromthechicken beddingandfoodwastemixturethanthecontrol(table2);however,theprepupedidnot differinnutrientvalues(table3a).theywereabletosequesteronlyasmallamountof thenutrientsfromfeedstockandthisdidnotdifferbyfeedstocktreatment(table3b).a minimalamountofnutrientsleachedfromthedigestateoverthecourseoftheexperiment anddidnotdifferinnutrientconcentrationsbetweenthetwotreatments(table4). Seedlinggerminationrateswerenotaffectedbytheaqueousdigestateextracts; however,seedlinglengthwasshortestatthehighestconcentrationlevelof1:1forboth radishandlettuce(fig.5).thiswasprobablyaresultofosmoticstresscausedbythehigh concentrationofdissolvedsolids.atlowerconcentrationlevels,seedlinglengthdidnot significantlydifferfromthewatercontrolforbothradishandlettuce. Objective2:Develop+an+optimized+entomopathogenic+nematode+rearing+system+based+on+ black+soldier+fly+larvae. ( Methods(and(Materials(( Invertebrate+colonies+ Werearedlarvaeinbatchesof300individualsin946mlplasticdelicontainers coveredwithabrownpapertowelheldinplacewitharubberband.wefedlarvae10gof Gainesvillehouseflydiet(5:3:2ratioofwheatbran,alfalfamealandcornmeal)and17W20 mlofwater5daysaweekat25 C,80%relativehumidity,andinthedark. Werearedfourspeciesofentomopathogenicnematodes Heterorhabitis+ bacteriophora,steinernema+carpocapsae,s.+feltiae,ands.+riobrave+ onthehostgalleria+ mellonellaininvertedpetridishes.harvestedinfectivejuvenileswerestoredintissue cultureflasksinlaboratorydrawersatambientroomtemperature.allinfectivejuvenile usedintheexperimentsdescribedbelowwerelessthan14daysold. + Experiment+1a+ +black+soldier+fly+susceptibility+by+larval+stage+ WesetWupa5x5twoWwayfactorialexperimenttotestthesusceptibilityofblack soldierflyinstarstomultiplespeciesofentomopathogenicnematodes.thelevelsofthe firstfactorweremultipleinstars(2,4,5,6)ofblacksoldierflyplusthepositivecontrol Galleria.Thelevelsofthesecondfactorwerefourspeciesofnematodes(H.+bacteriophora, S.+carpocapsae,S.+feltiae,andS.+riobrave)plusanegativecontrol waterwithout nematodes.therewasatotalof25treatmentcombinations. Wewashedanddriedlarvaebeforedistributingthemtoinverted60mmdiameter PetridishescontainingonepieceofNo.1Whatmanfilterpaper,onelarvaperdish.Usinga micropipette,weapplied100infectivejuvenilesin500μlofdeionizedwatertothefilter paper.weprepared25replicatesforeachtreatmentandplacedinagrowthchamber (20.4±0.6 C,70.4±10.4%relativehumidity,nolight).Weassessedinsectmortalitydaily for8days.onthe8 th day,wefrozethelarvaeatw20 Ctostoreforlaterdissection.We dissectedcadaversindeionizedwaterandcountedadultnematodesunderadissecting microscope. + Experiment+1b+ +black+soldier+fly+susceptibility+at+pupal+stage+

4 Asimilar,butseparateexperimentsetWupwasusedtotestifthepupalstageofblack soldierflyweresusceptibletoentomopathogenicnematodes.a2x5factorialexperiment wassetwup.thelevelsofthefirstfactorwereblacksoldierflypupaeandthepositive control Galleria.Thelevelsofthesecondfactorwerefourspeciesofnematodes(H.+ bacteriophora,s.+carpocapsae,s.+feltiae,ands.+riobrave)plusanegativecontrol water withoutnematodes.therewasatotalof10treatmentcombinations. + Experiment+2+ +injuring+larvae+ Basedontheresultsfromthepreviousexperiment,5 th instarwereselectedforall followingexperiments.wesetwupa3x5twowwayfactorialexperimenttotestwhether injuringtheblacksoldierflylarvaeaffectsnematodeinfectivity.thelevelsofthefirst factorwhereinjuredblacksoldierflylarvae,nonwinjuredblacksoldierflylarvae,andlate instargalleria.galleriawereusedasapositivecontroltoinsurethatnematodeswere indeedinfective.thelevelsofthesecondfactorwerefourspeciesofentomopathogenic nematodes(h.+bacteriophora,s.+carpocapsae,s+feltiae,ands.+riobrave)andwatercontrol. Therewereatotal15treatmentcombinations.Weprepared40replicatesforeach treatment.theexperimenttookplaceinagrowthchamberat25 C,100%humidityand blockedbyspace. Petridishinfectionarenaswerepreparedbyinserting60mmdiameterNo.1 Whatmanfilterpapercirclesinto60mmdiameterinvertedPetridishes.Infectivejuveniles (1000)wereappliedtothefilterpaperin500μl.Controls(nonematodes)receivedjust 500μlofwater. Meanwhile,blacksoldierflylarvaewereseparatedfromfeedingmaterialand washedinatuboflukewarmtapwater.underadissectingmicroscope,weusedasize0 insectpintopuncturetwoholesinthecuticleontheventrallateraledgesofninesegments themesothorax,metathorax,andfirstsevenabdominalsegments.wepuncturedatotal of18holesintothelarva.afterthepuncturingoperation,larvaewereimmediately transferredontothepreparepetridishes.insectpinsweresterilizedbyboilinginwaterfor 1minandstoringin80%ethanol.NonWinjuredlarvaeandGalleriawereplacedinto appropriatepetridishesatthesametime.oneblockofreplicateswascompletedbefore proceedingontothenextblock. Insectmortalitywasassessedasinexperiment1.Anylarvaethatwerepreviously recordedasdeadandthenalivewererevaluateddaily.onthefifthdayofmortality assessment,wefrozealldeadindividualsplusatleastonereplicateforeachblockofthe controls.weplacedlarvaeintothefreezer(w20 C)toarrestnematodedevelopmentuntil dissectioncouldoccur. Cadaverswerewashedinwatertoremoveexteriornematodes.Cadaverswere dissectedinringerssolutionunderadissectingmicroscope.wecountedtheadult nematodes.onefifthofthenegativecontrols(nonematodes)weredissectedtoverifyno nematodecontamination.anynegativecontrolsthatdiedwerealsodissectedtoverifyno nematodecontamination. ( Results( Experiment+1a+ +black+soldier+fly+susceptibility+by+larval+stage+ WefoundthatinsectmortalitywasstageWdependent(pWvalue<0.001)andthere wasaninteractionbetweennematodespeciesandinstar(pwvalue=0.032).wefoundthat

5 mortalitywasnotsignificantlydifferentamonginstarswheninfectedwithh.+bacteriophora ors.+feltiae(fig.6).secondinstars(92%±5%)and74%±11%offourthinstarswere killedbys.+carpocapsae,whichwassignificantlymorethanfifthandsixthinstars,13%± 8%and17%±8%,respectively(Fig.6).LarvaetreatedwithS.+riobrave,71%±8%ofthe secondinstarsand69%±12%ofthefourthinstarsdied,whichwassignificantlymore thansixthinstars(8%±5%),butnotsignificantlydifferentthanfifthinstars(53%±9%) (Fig.6).SecondinstarsweremostsusceptibletoS.+carpocapsae+(92%±5%),intermediate susceptibletos.+feltiaeands.+riobrave(57%±15%and71%±8%,respectively),andless susceptibletoh.+bacteriophora(28%±10%).fourththrusixthinstarsusceptibilitydidnot varybynematodespecies. Werecoverednematodesfromonlyafewblacksoldierflycadavers(Table5).NoH.+ bacteriophoranematodeswererecoveredfromthesecondinstars.wedidfindnematodes inthefourththroughsixthinstars,butthemeanwasfourorlessnematodesfromoneor twocadavers.fors.+carpocapsae,werecoveredonaveragesixnematodesfromeight secondinstars,whichwassignificantlymorethanthetwonematodesrecoveredfromthree fourthinstarsandtwosixthinstars.nos.+carpocapsaenematodeswererecoveredfrom fifthinstars.fors.+feltiae,onlyonenematodewasrecoveredfromthreeindividualsecond instars,twonematodesfromonefourthinstar,andameanof1.7nematodesfromthree sixthinstars.nonematodeswerefoundinthefifthinstars.fors.+riobrave,werecovereda meanoffivenematodesfromthreesecondinstars,whichwasnotsignificantlydifferent fromtheonenematodeinonefourthinstarandonenematodeeachintwofifthinstars.we didnotrecoveranynematodesfromsixthinstars.heterorhabditis+bacteriophorainfected 56%oftheGalleria,S.+carpocapsaeinfected96%oftheGalleria,S.+feltiaeinfected48%of thegalleria,ands.+riobraveinfected72%ofthegalleria.wedidnotrecoverany nematodesfromtheuntreatedcontrols. + Experiment+1b+ +black+soldier+fly+susceptibility+at+pupal+stage+ Wedidnotrecoveranynematodesfromthepupae. + Experiment+2+ +injuring+larvae+ Injuringtheblacksoldierflylarvaeincreasedthemortalityratewheninfectedby Steinernema+spp.butnotH.+bacteriophora(Fig.7).ForH.+bacteriophora,theinjuredlarvae didnotdieanyfasterthanthenonwinjuredlarvae(fig.7a).onday5,alloftheinjured larvaeweredeadandallbutonenonwinjuredlarvaeweredead(fig.7a).fors.+ carpocapsae,alloftheinjuredlarvaeweredeadbyday2whereasonlyfiveofthenonw injuredlarvaeweredead(fig.7b).onday5,32ofthenonwinjuredlarvaeweredead(fig. 7B).AsimilarpatternwasobservedforS.+feltiae.Onlyoneinjuredlarvaewasstillaliveon day2whereas30nonwinjuredlarvaewerestillalive(fig.7c).allbuttwononwinjured larvaediedbyday5(fig.7c).fors.+riobrave,31injuredlarvaeand5nonwinjuredlarvae weredeadonday2and40injuredlarvaeand39nonwinjuredlarvaeweredeadonday5 (Fig.7D).OnlytwoinjuredlarvaeandnoneofthenonWinjuredlarvaediedbythefifthday (Fig.7E).AlloftheGalleriatreatedwithnematodesweredeadonday2,andonlyfive Galleriadiedintheuntreatedcontrol. InjuringblacksoldierflylarvaedidsignificantlyincreasetheinfectivityofS.+ carpocapsaeands.+feltiae,butnoth.+bacteriophoraors.+riobrave(fig.8).significantlymore S.+carpocapsaeentertheinjuredhostthanthenonWinjuredhost(Fig.9).Thenumberof

6 infectivejuvenilesthatenteredthehostwasnotaffectedbyinjuringthehostfortheother threenematodespecies(fig.9). Objective3:Demonstrate+black+soldier+fly+composting+on+organic+farms+and+deliver+project+ findings+to+organic+farmers. WepurchasedalargebioreactorandsetWupitupattheMSUStudentOrganicFarm forstudentfarmers,faculty,andfarmvisitorstoobserveblacksoldierfliesdevouring organicwastes.wearefeedingtheblacksoldierfliespostconsumerwastesfromthemsu cafeterias. JasonMatlockandIpresentedaposteratthe2014annualMOSESconferencein LaCrosse,WIdiscussingoursecondtrialwithcompostingfoodwastesandchickenbedding withblacksoldierflies. IalsoparticipatedinagrowerextensionmeetinginSaginaw,MIwhereIinstructed agroupofgrowershowtoreartheirownnematodesusingwaxworms. Conclusions:( Blacksoldierfliescanbeusedtocompostvarioussourcesoforganicmatter,except toughplantstructuresthatcontaincellulose.theyquicklybreakdownorganicmatterand greatlyreducetheamountofbiomasswithafewshortweeks.blacksoldierflycompostis safetouseassoilamendmentwhendiluted,butbecarefulaboutthepresenceofanyseeds. Wethinkthatafterblacksoldierflieshavefinishedfeeding,redwormscouldbeaddedto finishbreakingdownthecompostintoamorestablestate. Blacksoldierflylarvaearenotreadilysusceptibletoinfectionbyentomopathogenic nematodes.injuringthemdoesincreasetheirsusceptibilitytosteinernema+spp.butnoth.+ bacteriophora.

7 A B Figure1.(A)BioPodsatSwallowtailFarms,Mason,MI.(B) Feedstockfeedtotheblacksoldierflylarvae,leftischicken beddingandfoodwastemixture,rightisjustfoodwaste.

8 Figure(2.Trial1.Blacksoldierfly(Hermetia+illucens)prepupae, (A)selfWharvested,(B)mass.

9 * * Figure(3.(Trial1.Ptecticus+trivittatusprepupae,(A)selfWharvested, (B)mass.

10 Table1.NutrientanalysisofdigestatefromTrail1.MeansaregivenforeachelementandtheSEMisgiven belowinparentheses. percent'(%)' ' ppm' N P K S Mg Ca Na B Zn Mn Fe Cu Al Food 1.62' 0.41' 2.27' 0.34' 0.50' 1.35' 0.57' ' 17.7' 57.3' 269.7' ' 23.7' ' Waste' (0.10)' (0.03)' (0.20)' (0.03)' (0.01)' (0.07)' (0.23)' ' (2.0)' (4.3)' (15.2)' (351.7)' (3.3)' (265.3)' ' ' ' ' ' ' ' ' ' ' ' ' ' ' Chicken 1.37' 0.83' 1.50' 0.33' 0.48' 7.96' 0.15' ' 18.0' 305.7' 442.3' ' 32.3' ' Bedding' (0.10)' (0.06)' (0.07)' (0.01)' (0.03)' (0.34)' (0.01)' ' (0.6)' (13.1)' (21.9)' (330.2)' (0.7)' (52.1)' ' ' ' ' ' ' ' ' ' ' ' ' ' ' ' p7value*' 0.15' >0.01' 0.02' 0.77' 0.64' >0.01' 0.15' ' 0.88' >0.01' >0.01' 0.51' 0.06' 0.98' *two?tailedt?test

11 Figure'4.Trial2.Blacksoldierfly(Hermetia(illucens)prepupae (A)self8harvested,(B)mass.

12 Table2.NutrientanalysisofdigestatefromTrial2.MeansaregivenforeachelementandtheSEMisgiven belowinparentheses. percent' ' mg/kg' ' percent' Organic N P K S Mg Ca Na Fe Al Cu Mn Zn Matter OrganicC Food 4.80' 0.57' 2.82' 0.49' 0.55' 2.10' 0.30' 0.27' 0.16' ' 50.7' 219.3' 89.7' ' 74.17' 37.08' Waste (0.13)' (0.03)' (0.11)' (0.02)' (0.02)' (0.08)' (0.01)' (0.01)' (0.01)' ' (0.9)' (13.3)' (2.9)' ' (3.15)' (1.57)' ' ' Chicken ' 4.80' ' 1.01' ' 3.27' ' 0.58' ' 0.64' ' 3.35' ' 0.34' ' 0.38' ' 0.20' ' ' 58.7' ' 340.0' ' 173.3' ' ' 74.18' ' 37.09' Bedding (0.26)' (0.09)' (0.22)' (0.03)' (0.02)' (0.24)' (0.02)' (0.02)' (0.01)' ' (2.7)' (20.1)' (17.0)' ' (3.30)' (1.65)' ' ' p>value* ' 0.50' ' 0.01' ' 0.07' ' 0.06' ' 0.01' ' >0.01' ' 0.04' ' >0.01' ' 0.01' ' ' 0.02' ' >0.01' ' >0.01' ' ' 0.50' ' 0.50' *one?tailedt?test

13 Table&3A.&NutreintAnalysisofself?harvestedblacksoldierflyprepupae.Meansaregivenforeachelement andthesemisgivenbelowinparentheses. percent' ' mg/kg' ' percent' Organic N P K S Mg Ca Na Fe Al Cu Mn Zn Matter OrganicC Food 5.36' 0.57' 0.80' 0.31' 0.31' 6.33' 0.07' 0.02' 0.01' ' 9.67' 773.3' 60.3' ' 80.58' 40.29' Waste (0.13)' (0.04)' (0.02)' (0.01)' (0.02)' (0.16)' (0.00)' (0.00)' (0.00)' ' (0.67)' (32.8)' (6.3)' ' (0.47)' (0.24)' ' ' Chicken ' 5.33' ' 0.55' ' 0.81' ' 0.31' ' 0.30' ' 6.59' ' 0.07' ' 0.02' ' 0.01' ' ' 9.0' ' 824.7' ' 61.3' ' ' 80.16' ' 40.08' Bedding (0.10)' (0.01)' (0.01)' (0.00)' (0.01)' (0.24)' (0.00)' (0.00)' (0.00)' ' (0.0)' (32.1)' (1.5)' ' (0.14)' (0.07)' ' ' p>value* ' 0.85' ' 0.59' ' 0.61' ' 0.80' ' 0.67' ' 0.40' ' 1.00' ' 0.37' ' 0.37' ' ' 0.37' ' 0.33' ' 0.89' ' ' 0.44' ' 0.43' *two?tailedt?test Table&3B.Percentofnutrientsblacksoldierflysequesteredfromthefeedstock.Meansaregivenforeach elementandthesemisgivenbelowinparentheses. Organic N P K S Mg Ca Na Fe Al Cu Mn Zn Matter Food Waste 0.26' 0.26' 0.08' 0.16' 0.13' 0.82' 0.08' 0.02' 0.01' 0.04' 0.78' 0.16' 0.18' Chicken Bedding 0.25' 0.18' 0.06' 0.13' 0.16' 0.68' 0.06' 0.02' 0.02' 0.05' 0.70' 0.11' 0.16' ' ' ' ' ' ' ' ' ' ' ' ' ' p>value* 0.83' 0.13' 0.36' 0.37' 0.37' 0.48' 0.33' 0.15' 0.21' 0.22' 0.56' 0.10' 0.67' *two?tailedt?test

14 Table&4A.&NutrientAnalysisofLeachate.MeansaregivenforeachelementandtheSEMisgivenbelowin parentheses. ppm'(mg/l)' ' ' Electrical Food Waste N P K S Mg Ca Na Fe Cu Mn Zn ph Conductivity 630.0' 96.37' ' 160.0' 86.7' 159.0' 697.3' 25.48' 0.22' 1.62' 0.42' 7.70' 4.08' (132.3)' (9.72)' (497.1)' (23.0)' (13.7)' (12.5)' (76.9)' (4.91)' (0.03)' (0.31)' (0.06)' (0.10)' (0.44)' Chicken ' 473.3' ' 88.10' ' ' ' 161.0' ' 65.7' ' 120.7' ' 624.3' ' 14.33' ' 0.17' ' 1.02' ' 0.43' ' 7.77' ' 3.84' Bedding (59.3)' (4.19)' (49.5)' (10.5)' (5.5)' (9.9)' (29.8)' (1.67)' (0.02)' (0.11)' (0.07)' (0.03)' (0.08)' p>value* ' 0.34' ' 0.48' ' 0.41' ' 0.97' ' 0.23' ' 0.07' ' 0.43' ' 0.10' ' 0.25' ' 0.14' ' 0.89' ' 0.56' ' 0.62' *two?tailedt?test Table&4B.Percentnutrientsthatleachedfromthefeedstockoverthecourseofthe5?weekexperiment. N P K S Mg Ca Na Fe Cu Mn Zn Food Waste 0.21' 0.31' 0.82' 0.57' 0.26' 0.14' 5.21' 0.14' 0.06' 0.11' 0.08' Chicken Bedding 0.18' 0.24' 1.11' 0.55' 0.28' 0.10' 4.26' 0.13' 0.08' 0.07' 0.07' ' ' ' ' ' ' ' ' ' ' ' p>value* 0.64' 0.28' 0.44' 0.91' 0.74' 0.19' 0.40' 0.90' 0.45' 0.21' 0.56' *two?tailedt?test

15 Figure'5.'Germinationstudyusingaqueousdigestateextracts fromfoodwastecompostedbyblacksoldierflylarvae.

16 NS NS A A B B A A AB B Figure'6.Percentcorrectedmortalityofblacksoldierfly larvae.100juvenilesappliedperhostat20 C.Mortalityfor thesecondinstarisfromday7sincetherewashigh mortalityonday8inthecontrol.galleriamortalitywas >95%forallnematodespeciesand24±10%forthe controlonday5(datanotshown).barswithdifferent lettersindicatestatisticaldifference(anova,tukeyhsd,p <0.05).NS=nostatisticaldifference.

17 Table'5.Adultnematodesrecoveredfromfourblacksoldierflyinstars(n=25 pertreatmentcombination).100juvenilesappliedperhostat20 C.Noneof thecontrols(nonematodes)wereinfected.datanotshownforgalleria. Nematode Host instar PercentInfected ±SEM Nematodes± SEM H.#bacteriophora# 2 0.0±0.0ns 0.0 ± 0.0ns 4 8.0± ± ± ± ± ± 0.7 S.#carpocapsae# ±4.9a 6.0 ± 0.7ns ±4.9a 1.7 ± ±0.0b 0.0 ± ±4.9a 2.0 ± 0.0 S.#feltiae# ±4.9ns 1.0 ± 0.0ns 4 4.0± ± ± ± ± ± 0.2 S.#riobrave# ±8.0ns 5.0 ± 2.0ns 4 4.0± ± ± ± ± ± 0.0 *Kruskal`Wallistest,differentlettersindicatesignificant differences,ns=nosignificantdifference.

18 Alivelarvae Figure'7.Survivalcurvesforinjured(dottedline)andnon`injured(solidline) blacksoldierflylarvaeforeachnematodespecies:a)h.#bacteriophora,b)s.# carpocapsae,c)s.#feltiae,d)s.#riobrave,ande)control(nonematodes).1000 juvenilesappliedperhostat25 C.AllGalleriatreatedwithnematodesweredead onday2andonly5galleriadiedinthecontroltreatment(datanotshown).

19 * * Figure'8.Percentblacksoldierflylarvae infectedwithentomopathogenicnematodes (n=40).infectivejuveniles(1000)appliedper hostat25 C.Asterisk(*)denotessignificant difference(p<0.05).

* 37 35 40 32 39 32 39 36 Figure'9.Meannumberofentomopathogenic nematodes(firstgenerationadultsandjuveniles) recoveredfromblacksoldierflycadavers.

20 * Figure'9.Meannumberofentomopathogenic nematodes(firstgenerationadultsandjuveniles) recoveredfromblacksoldierflycadavers.1000 juvenilesappliedperhostat25 C.Numberswithin thebarsindicatenumberofinfectedlarvae(max= 40).Asterisk(*)denotessignificantdifference(P< 0.05).

2009 MN Cattle Feeder Days Jolene Kelzer University of Minnesota Beef Team

2009 MN Cattle Feeder Days Jolene Kelzer University of Minnesota Beef Team 101.8 M total US cattle and calves (July 1) Down 1% from 2008 (103.3 M) 11.6 M total US cattle on feed (July 1) Down 5% from 2008