BlackSoldierFlylarvaerearedoncontaminatedsubstratebyListeriamonocytogenesandSalmonella

IstitutoZooprofilatticoSperimentaledellaLombardiaedell’Emilia-Romagna(IZSLER)Sez.ReggioEmilia,Italy

BACKGROUNDInsectshavebeenproposedasahighquality,efficientandsustainablealternativeproteinsource.Usinginsectsasaproteinsourcecancontributetoglobalfoodsecurityviafeedorasadirectfoodsourceforhumans.In2011,theworldcompoundfeedproductionhasbeenestimatedat870milliontonesandtheturnoverofglobalcommercialfeedmanufacturinggenerated an estimated annual turnover and sales value equivalent to US$ 350 billionworldwide (http://www.ifif.org/). The UN Food and Agricultural Organization (FAO)estimates that the production of food worldwide will increase of about 70% by 2050.Concerning animal protein production, the International Feed Industry Federation (IFIF)believes that the production of meat (poultry/swine/beef) will even double. This posesseverechallengestotheglobalcapacitytoprovideenoughfeed.Insectscouldbeananswerbutthesafetyofprotein frominsectsandsubsequentlythesafetyofmeatandfish fromanimalsfedonsuchadietrequiresfurtherassessment.BlackSoldier fly (Hermetia illucensL.) larvae (BSFL,Fig.1)havebeennoted to reduce themicrobial load of substrates, decreasing bacteria concentrations in compost and fecalmaterial. The purpose of our researchwas to investigate the capability of BSF larvae toreduce the concentration of Salmonella typhimurium and Listeriamonocytogenes on anartificiallycontaminatedsubstrate.

MATERIALSANDMETHODSTwosetsofexperimentswereconductedinordertoinvestigate:1)  thereductionofpathogensincontaminatedsubstratewithoutlarvae(control);2)  thebehaviorofchallengedpathogensinlarvae,pre-pupae,pupaeandtheirsubstrates.BSFlarvae:BSFLwererearedundercontrolledconditions(RH70%,photoperiod14:10h(L:D)and temperature25°C)on two substrates (Gainesvilledietandahomemadeartificialdiet).The larval substratewas contaminatedwithS. typhimuriumand L.monocytogenes (about1x108CFUg-1).Foreachtrialsubstratesand larvaeweretestedforpathogensenumeration,pH,andaw.

RESULTSIn controls (contaminated substratewithout larvae) thepH remained stableuntil theendof experiments (6.59-7.95)while awremainedstableuntil7days(0.977)butafterthisperiodawdecreasedreaching0.650.IncontrolexperimentsthecontaminationofL.monocytogenesremainconstantwhileaslowdeclineofSalmonella(-0.0077log.conc/hD=130h)wasobserved.WhenlarvaewerepresentpHandawremainedstablethroughthedurationofthestudy.Salmonellashowedafastdeclineinthefirst8days(-0.02log.conc/hD=45h)butafterthisinitialreductionthepathogensremainedconstantinthesubstrate,showingatypicalbiphasicdecline curve.WhileL.monocytogenesshowedavery slowdecline rate (0.0042 log.conc/hD=239h)during thewholedurationoftheexperiment.In larvaeSalmonella and L.monocytogenes showed the same concentration observed in the substrate,when larvae entered inpupalstage(fig.2)theircontaminationwassignificantlylesserthanthesubstrate(-2log).

*Corresponding author: paolo.bonilauri@izsler.it – PRC2015005 - E88C16000070001

F.DefilippoA.GrisendiV.ListortiM.DottoriP.Bonilauri

References [1]Newton,L.,Sheppard,C.,Watson,D.W.,Burtle,G.,Dove,R.,2005.Usingtheblacksoldierfly,Hermetiaillucens,asavalue-addedtoolforthemanagementofswinemanure.In:ReportforMikeWilliams,DirectoroftheAnimalandPoultryWasteManagementCenter.NorthCarolinaStateUniversity[2]Newton,G.L.,Booram,C.V.,Barker,R.W.,Hale,O.M.,1977.DriedHermetiaillucenslarvaemealasasupplementforswine.J.Anim.Sci.44,395–400.[3]HarinderP.S.Makkara,GillesTranb,ValérieHeuzéb,PhilippeAnkersa.2014.State-of-the-artonuseofinsectsasanimalfeed.AnimalFeedScienceandTechnology197(1–33)

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MicrobiologicalanalysisSalmonellaTyphimuriumdetection(ISO6579-1:2017)andenumeration(ISO6579-2:2012)Sampleswasenriched(1:10)inBufferedPeptoneWaterfor18±2hat36°C±2°C,100µlofsamplewasdistributedonMSRV agar plate. Sample was incubated for 24/48 h at 41.5 °C ±1°C (Fig.3). The presence of Salmonella wasconfirmedbyusingXylose-Lysine-DesoxycholateAgarandappropriatebiochemicalminiaturizedtests. MicroMPNenumerationfollowingpart2ofISO6579:2012wasapplied.Whentheloadofpathogenswasabove3logCFU/g,10folddilutionanddirectplatingonXLDandhektoenentericagarwasapplied.Listeriamonocytogenesdetection(ISO11290-1:2017)andenumeration(ISO11290-2:2017) SampleswasenrichedinHFandplatedinALOAandOXFODagar.Enumerationwasperformedby10folddilutionanddirectplatingonALOAagar.

Fig.3–MRSVagarSalmonellapositive

Fig.4:(left)Salmonellaconcentration(logCFU/g)inlarvae,substrateandcontrol(substratewithoutlarva)

Fig.5:(right)Listeriamonocytogenesconcentration(logCFU/g)inlarvae,substrateandcontrol(substratewithoutlarva)

Poster Code: 28552

CONCLUSIONThis study confirm that larvae show the same pathogens concentration of their growing substrate and their feeding activity couldreducepathogenscontaminationinthegrowingsubstrate,buttherateofthisactionseemstobelargelyinsufficienttoreachthefoodandfeedsecurityobjective.

Fig.1BlackSoldierFly(Hermetiaillucens)larvae

Fig.2Black Soldier Fly (Hermetia illucens) pupae

Pupae Pupae