15

Click here to load reader

Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

  • Upload
    dothuan

  • View
    212

  • Download
    0

Embed Size (px)

Citation preview

Page 1: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

BASE Biotechnol. Agron. Soc. Environ.201620(S1),299-313

Implementingplantbiostimulantsandbiocontrolstrategiesintheagroecologicalmanagementofcultivatedecosystems.AreviewGeraldineLeMire(1,2)*,MinhLuanNguyen(1,3)*,BereniceFassotte(1,4),PatrickduJardin(3),FrançoisVerheggen(4),PierreDelaplace(3)**,M.HaissamJijakli(2)**(1)UniversityofLiège-GemblouxAgro-BioTech.TERRA.AgricultureIsLife.PassagedesDéportés,2.BE-5030Gembloux(Belgium).E-mail:[email protected](2)UniversityofLiège-GemblouxAgro-BioTech.Agrobiochem.UrbanandIntegratedPhytopathology.PassagedesDéportés,2.BE-5030Gembloux(Belgium).(3)UniversityofLiège-GemblouxAgro-BioTech.Agrobiochem.PlantBiology.PassagedesDéportés,2.BE-5030Gembloux(Belgium).(4)UniversityofLiège-GemblouxAgro-BioTech.Agrobiochem.FunctionalandEvolutionaryEntomology.PassagedesDéportés,2.BE-5030Gembloux(Belgium).

*Theseauthorscontributedequallytothiswork.**Co-lastauthors.

ReceivedonApril1,2015;acceptedonOctober14,2015.

Introduction. In the context of sustainable agricultural production, agroecology aims at optimizing the economic andenvironmental performances of beneficial ecosystem services in order to (i) increase the productivity and resilience ofcultivatedecosystemsand(ii)preservetheirnaturalresources.Themaintenanceofsuchperformancesissupportedbyresearchviathedevelopmentofnewtoolsthatenhanceplanttolerancetonumerousbioticandabioticstresses.Literature.Biostimulantscanbeusedasa tool tocomplement theuseofchemical inputs,byinvolvingnon-living-basedproducts,orliving-basedproductscontainingbeneficialrhizospheremicrobiome,suchasplantgrowth-promotingrhizobacteria(PGPR).Pestmanagementresearchhasalsomademajoradvancesinthedevelopmentofefficientbiocontrolmethods.Elicitorsandsemiochemicalsareconsideredtobesomeofthemostpromisingtoolsforinducingplantresistancetovariousdiseasesandenhancingnaturalpredation,respectively.Severalproductsarealreadyonthemarket.Thisreviewdiscussescurrentmethodsforexploitingandapplyingbiostimulantandbiocontrolproductsincontemporaryagriculturalsystems.Futureapplicationsofthesetoolsforsustainablemanagementofcultivatedecosystemsarealsodiscussed.Conclusions.Thesetoolsarestilldifficulttousebecauseoftheirlackofreliabilityinthefieldandtheiruneasyintegrationinthecroppingsystems.FurtherstudiesareneededtobetterunderstandtheparametersinfluencingtheefficiencyofPGPR,elicitorsandsemiochemicals.Specialattentionneedstobegiventotheformulationandtheinteractionsoftheseproductswithplantphysiologyandtheenvironment.Keywords. Agroecology, biological control, plant growth stimulants, biofertilizers, biopesticides, induced resistance,rhizobacteria,semiochemicals,elicitors.

Intégrer les biostimulants et les stratégies de biocontrôle dans la gestion agroécologique des écosystèmes cultivés (synthèse bibliographique)Introduction.Danslecadred’uneproductionagricoledurable,l’agroécologieviseàoptimiserlesperformanceséconomiqueset environnementales des services écosystémiques avantageux afin d’assurer (i) une augmentation en productivité et enrésiliencedesécosystèmescultivéset(ii)unepréservationdeleursressourcesnaturelles.Lemaintiendecesperformancesestsoutenuparlarecherchevialedéveloppementdenouveauxoutilspermettantd’augmenterlatolérancedesplantesauxstressbiotiquesetabiotiques.Littérature.Lesbiostimulantspeuventêtreutiliséscommeunoutilcomplémentaireàl’utilisationd’intrantschimiquesvial’usage de formulations inertes ou vivantes contenant desmicrobiontes rhizosphériques bénéfiques tels que les bactériespromotricesdelacroissancedesplantes(PGPR).Larechercheenphytopathologieaégalementfaitdesprogrèsconséquentsdansledéveloppementdeméthodesdelutte.Leséliciteursetsémiochimiquesfontpartiedesoutilsdebiocontrôlelesplus

Page 2: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

300 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

1. INTRODUCTION

Thedeclineinnaturalresourcesandtheenvironmentaldamageinflictedbycurrentagriculturalpracticeshavebecomemajorlimitationsinconventionalagriculture.Against this background, agroecology offers animportant scientific approach that takes into accountthe current societal concerns linked to agriculture,economyand,inparticular,theenvironment.Byusingecologicalprinciples,itaimsatstudyinganddesigningagricultural systems based on the interactions oftheir main biophysical, technical and socioeconomiccomponents(EuropeanCommission,2012).Researchisnowstronglyfocusedontheuseofagroecologicalprinciples to minimize potentially harmful chemicalinputsandmanageecologicalrelationshipsandagro-biodiversity.Thepastdecadehasseentheemergenceoftechnological toolsdeveloped topromote sustainableagroecosystems.The enhancement of plant toleranceto numerous abiotic stresses is increasingly beingsupported by biostimulant products, as preferredalternatives to chemical fertilizers. Biostimulantsinclude livingmicroorganisms,namelyplantgrowth-promoting fungi (PGPF) and rhizobacteria (PGPR)(Bhattacharyyaetal.,2012).Plantgrowth-promotingrhizobacteria are currently thought tobeaneffectivetoolforthebiostimulationofplantgrowth(Calvoetal.,2014).Beneficialrhizobacteriaassociatewiththerootsystemandstimulatethegrowthofhostplants,whilebeingfed in turnbyrootexudates.Pestmanagementresearchers have also made major advances in thedevelopment of efficient biocontrol methods toprotectplantsagainstbioticstresses.Biocontrolrefersto any method, product or organism using naturalmechanismsinthecontextofintegratedcropprotectionagainst bioaggressors (Herth, 2011). Biocontrolproducts includemacro-andmicroorganisms,naturalsubstances of plant, mineral or animal origin, andchemical mediators. Elicitors (also called plantresistance inducers) are considered to be biocontrolproductsinagricultureastheyinduceplantresistancetovariousdiseases.Similarly,semiochemicalsareseenas biocontrol tools because they involve push-pullstrategiesand/ormatingdisruptions.

This paper gives an overview of the presentstatus of these tools in agricultural production, aswellasadescriptionofthemodesofactionofPGPRbiostimulants, elicitors and semiochemicals. It thenaddressesthequestionofhowandwhyfuturestrategiesshouldincreasetheuseofthesetoolsbyhighlightingboththeirlimitationsandtheirpotentialforcontributingtosustainableandagroecologicalagriculture.

2. BIOSTIMULANT AND BIOCONTROL TOOLS IN CONVENTIONAL AGRICULTURE

Thedevelopmentofnewgreentechnologieshasledtogreaterresearchfocusonbiostimulantandbiocontroltools(Bolleretal.,2009;Bhattacharyyaetal.,2012).The definition and modes of action of three tools–PGPR,elicitorsandsemiochemicals–aredescribedinthischapter.

2.1. PGPR-based biostimulants

Thesustainablemanagementofsoilfertilityisamajorconcerngiventheadverseimpactandecologicalthreatsposedby theuseofconventionalchemical fertilizers(Wezel et al., 2014). In this context, biostimulantsrepresent an interesting alternative. They consist ofvarious substances and microorganisms (microbialinoculants,humicandfulvicacids, seaweedextracts,proteinhydrolysates,aminoacids),whichareusedtoenhance plant growth (du Jardin, 2012;Calvo et al.,2014).Theycanincreasecropyieldbyatleast5-10%andimprovefertilizeruseefficiencybyatleast5-25%(EuropeanBiostimulantsIndustryConsortium,2011).In2012,theglobalmarketofbiostimulantswasmainlylocated in Europe and it is projected to increase by12%annually,reaching$2,241millionby2018(Calvoetal.,2014).

Despite their growing use, there is currentlyno accepted definition of biostimulants, neither byregulatory bodies nor by the scientific community.However,withtherevisionofcurrentEuropeanUnion(EU) legislation on fertilizers, there has been someprogress.Withinthisframework,aconsultancyreport

prometteurs en stimulant, respectivement, les défenses naturelles des plantes contre de nombreuses maladies etla prédation naturelle. Plusieurs produits sont déjà disponibles sur le marché. Cette synthèse décrit les méthodesactuelles d’emploi et d’application des produits biostimulants et de biocontrôle dans les systèmes agricolesactuels. Le futur de ces techniques dans la gestion durable des écosystèmes cultivés est également abordé.Conclusions.Cesoutilssontencoredifficilementutilisablesdufaitdeleurefficacitévariableauchampetdumanquedereculsurlamanièredelesintégrerdanslessystèmesdeculture.Desrecherchessupplémentairessontnécessairesafindemieuxcomprendrelesparamètresconditionnantl’efficacitédesPGPR,deséliciteursetdessémiochimiques.L’accentdoitêtreportésurlaformulationetlesinteractionsdecesproduitsaveclaphysiologieetl’environnementdelaplante.Mots-clés.Agroécologie,luttebiologique,stimulantdecroissancevégétale,biofertilisants,biopesticide,résistanceinduite,rhizobactérie,substancesémiochimique,éliciteur.

Page 3: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 301

defined a plant biostimulant as “any substance or microorganism, in the form in which it is supplied to the user, applied to plants, seeds or the root environment with the intention to stimulate natural processes of plants benefiting nutrient use efficiency and/or tolerance to abiotic stress, regardless of its nutrients content, or any combination of such substances and/or microorganisms intended for this use” (Traonet al., 2014). This definition clearly differentiatesbiostimulantsfrombiocontrolsubstancesoragents.

In this review, we focus on biostimulants in thecategory ofmicrobial inoculants, specificallyPGPR,whichhavebeenintensivelystudiedinrecentdecades(Calvo et al., 2014). Plant growth-promoting rhizo-bacteria-basedbiostimulantsaremajorcomponentsofbiofertilizersintendedforagriculturaluse.Theyhavelong been commercialized by many manufacturersandappliedtovariousfieldcropssuchasrice,wheat,maizeandsoybean(Köhl,2010;Pérez-Montañoetal.,2013),aswellastohorticulturalcropssuchastropi-cal, subtropical and temperate fruits and vegetables(Reddy,2014).

Plant growth-promoting rhizobacteria-basedbiostimulants are considered to be easy-to-useagroecological tools for stimulating plant growthand enhancing plant nutrient uptake and abioticstresstolerance(Walkeretal.,2012;Holetal.,2013;Pérez-Montaño et al., 2013).They can also enhancebeneficialsymbioseswiththehostplant.SomePGPR-based biostimulants also have a biocontrol activity,enablingthemtoprotectplantsagainstbioticstresses(Bhattacharyyaetal.,2012).Thisactivity,however,isnotdiscussedinthisreview.

Biostimulant products can be based on a singlePGPRstrain,aPGPRmixoramixofPGPRandPGPF.Comparedwith single strain products, consortia canreachmostoftheemptynichesbecauseoftheirincreasedgeneticdiversityandtheycolonizetherootzonemuchfasterthansinglestrains(Reddy,2014).ProductswithamixofPGPRstrainscanthereforecompetespatiallywith a broader range of potential pathogens underdifferent plant growth and environmental conditions(Reddy,2014).Inaddition,recentstudieshaveshownthat PGPR used to complementmineral fertilizationcan reduce conventional fertilizer rates (Adesemoyeet al., 2009).Adesemoye et al. (2009) showed thata combined inoculation of the two PGPR strainsBacillus amyloliquefaciens IN937a and BacilluspumilusT4withastrainofthearbuscularmycorrhizafungiGlomus intraradices reduced fertilizer use by25%. This combination was as efficient as a 100%fertilizer application in termsof plant growth, yield,andnutrientuptake.Other examplesofPGPR-basedbiostimulants that enhance crop growth and reducetheamountofneededchemicalfertilizersaregiveninappendix 1.

The capacity of PGPR to stimulate plant growthrelies mainly on their capacity to produce/degradevariousplant-growthregulators.Phytohormones(e.g.auxins, cytokinins, gibberellins, ethylene) producedby PGPR can regulate multiple plant physiologicalprocesses (root initiation and elongation, roothair formation) (Calvo et al., 2014). For example,rhizosphere bacteria such as Azospirillum, Bacillus,RhizobiumandEnterobactercanreducetheproductionoftheplantstresshormoneethyleneviathesecretionof 1-amino cyclopropane-1-carboxylate deaminase,thuspreventingplantgrowthinhibition(Bhattacharyyaetal.,2012).

Plant growth-promoting rhizobacteria can alsoincreasetheavailabilityofnutrientsforthehostplant.Themechanismsinvolvedincludenitrogen(N)fixation,nutrient solubilization, PGPR production of volatileorganiccompounds(VOCs),andironsequesteringbyPGPR-produced siderophores (Bhattacharyya et al.,2012;Calvoetal.,2014).Lessthanhalf(10-40%)oftheappliednitrogeninthefieldiseffectivelyabsorbedby plants, and 60-90% of chemical N fertilizersare generally lost by nitrate leaching or ammoniavolatilization. Nitrogen fertilizer is transformed intoammoniagases,includingnitrousoxide(N2O),amajorgreenhousegas(Adesemoyeetal.,2009).SuchN-lossprocesses can result inwater and soil pollution and/orgreenhouse-gasgeneration.Plantgrowth-promotingrhizobacteria used to complement mineral fertilizerscanhelpsolvetheseissues(Calvoetal.,2014).Finally,PGPR inoculants can also enhance crop tolerance tosalinityanddrought,notablybyreducingsoil1-aminocyclopropane-1-carboxylate deaminase (ACC)and pollutants (herbicides, pesticides, heavy metaldetoxification)(Upadhyayetal.,2015).

There is therefore growing scientific evidencesupportingtheuseofPGPRinoculantsasbiofertilizersformanyplants.Forexample,FZB24®wasshowntopromoteplantgrowthandyieldincotton,tomatoandmaize(Kilianetal.,2000;Yaoetal.,2006).Similarly,RhizoVital42®provedefficientinlettuce(Chowdhuryet al., 2013; Kröber et al., 2014), BactofilA10®in rye-grass (Tállai et al., 2012), and TwinN® insugarcane(Simwingaetal.,2010).Furtherinformationon commercialized PGPR products are given inappendix 2.

ProductscontainingexogenousPGPRcompounds(e.g. exopolysaccharides, phytohormones) have alsobeen developed to enhance the growth of specificbeneficial microbes in the soil (Marks et al., 2013).The efficiency of PGPR-based products, however,stillreliesonseveralfactors.Plantspeciesandvariety(releasing different types of root exudates), soiltype, environmental conditions, and the commercialformulation are crucial determinants of the efficientand reproducible action of inoculated PGPR (Calvo

Page 4: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

302 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

etal.,2014).ThebestPGPRproductsgenerallyconsistoflocalstrainsthatarespecifictothehostplant,showgoodcapacityforphysiologicalandgeneticadaptationandco-evolvewithothernativestrains inacommonhabitat(Reddyetal.,1999;Mäderetal.,2011;Araújoetal.,2013).

2.2. Biocontrol products and crop protection

Elicitors.Inthelate1970s,itwasdiscoveredthatplantshaveinducibledefensemechanismsthatareactivatedby infectionandcouldpotentiallyprovideprotectionagainstabroadspectrumofpathogens(Schwessingeretal.,2012).Thisresistanceis triggeredbytheplantwhenitsenses“non-self”moleculesreleasedduringtheattack,knownasgeneralelicitors.Theterm“elicitor”refers to all the signal molecules that are perceivedandthatinduceadefensivereactionintheplant.Theythereforeplayakeyroleinplant-pathogeninteractions(Valladetal.,2004).

Induced resistancehas longbeen recognized as avaluableapproachindiseasecontrolstrategiesbecauseitoffersthepromiseofdurable,broad-spectrumdiseasecontrolusingtheplant’sownresistance(Waltersetal.,2014).Theelicitorproductscurrentlyinthemarketplaceareusedmainlyinintegratedpestmanagement(IPM)strategies as complementary tools to help reducechemical inputs. Until now and depending on theirefficiency, elicitors are usually applied alone or incombination with other fungicides, once or severaltimesinacropcycle(Waltersetal.,2013).Additionalinformation on currently commercialized elicitorproductsaregiveninappendix 3.

Two well-known elicitor products are the algaeextract laminarin, and benzo-(1, 2, 3)-thiadiazole-7-carbothioic acid S-methyl ester (BTH, also knownas BION® from Syngenta Europe) (Sobhy et al.,2012).BION®,however, isachemicalelicitor (witha structural analogy to the plant hormone salicylicacid)andcanthereforebeexcludedfromthecategoryof biocontrol products, defined by Herth (2011) as“agents or products which use natural mechanisms in the frame of an integrated pest management (IPM). This includes macro-organisms (insects, nematodes), micro-organisms (bacteria, fungi, viruses), chemical mediators (pheromones, kairomones), and natural substances of plant, animal or mineral origin.”Thisreviewfollowsthisdefinition.

Someelicitorproductsarealsosuitablefororganicfarming,whichhassurgedinpopularityinrecentyears(Dayanetal.,2009).Organicpracticesprohibittheuseof synthetic chemicalproducts and thereforehave toaddressimportantdiseasepressuresbecauseofthelackof pesticide applications. For acceptance in organicagriculture, the elicitor compounds should occur innature and should not be derived from genetically

modified organisms (García-Mier et al., 2013). Forexample, laminarin is considered to be suitable fororganic farming, and plant protection products thatcontain it are registered inBelgium,Greece,France,The Netherlands, the UK and Germany (EuropeanCommissionEGTOP,2011).

Therehasbeenan intensehunt forelicitors sincethediscoveryofinterestingonesthatcouldbeusedforagriculturalpurposes.Threecategoriesofelicitorshavebeendeterminedsofar:pathogen-associatedmolecularpatterns(PAMPs)emittedspecificallyfrompathogenicorganismsorparasites;microbe-associatedmolecularpatterns (MAMPs), overall released by beneficial/non-pathogenic microorganisms such as yeasts, andplantgrowth-promotingrhizobacteriaorplantgrowth-promoting fungi; endogenous molecular patternsrelatedtoinjuredplanttissue(DAMPs,fordamage-ordanger-associatedmolecularpatterns)emittedfromtheplantitself(Henryetal.,2012).Thesedanger-signalingcompoundsareessentialcomponentsofentireclassesofmicroorganismsandhavebeenshowntoprotectavarietyofplantsagainstabroadspectrumofpathogens(Schwessinger et al., 2012). Non-microbial elicitorshave also been identified, originating from an arrayoforganicsources,suchasalgaeextracts,crustaceanshellsandminerals,aswellasfromchemicalsynthesis(Henryet al.,2012).Thevarious signalingpathwaysthese three elicitor categories trigger in plantsare described in appendix 4. Most of the organicelicitors that have been characterized include fungalchitin, bacterial flagellin, lipopolysaccharides (LPS),oligogalacturonides(OGAs),ergosterol,siderophores,surfactinandfengycincycliclipopeptides,andvolatileorganiccompounds(VOCs).

Elicitor perception activates the plant’s immunesystemwhichischaracterizedbyacascadeofeventswithacomplexspatialand temporal regulation.Thisincludes a local burst of reactive oxygen species(ROS), ion fluxes across the plasmamembrane, andthe production of phytoalexins and pathogenesis-related(PR)proteins(Henry,2013).Atthescaleofthewholeplant,elicitorperceptiontriggersspecificsignaltransduction pathways involving one or several keyregulators,andresultinginoneoftwopossibleformsofinducedresistance:systemicacquiredresistance(SAR)against biotrophic and hemibiotrophic pathogens,combined with a characteristic accumulation of PRproteins;orinducedsystemicresistance(ISR)againstnecrotrophic pathogens, chewing herbivores andphloem-feedinginsects(Henryetal.,2012;Wasternacketal.,2013).Dependingontheplantandtheelicitor,asetperiodoftimeisrequiredforsystemicresistancetobeembedded.

In addition, some elicitors can trigger a processcalledpriming,whichpreparestheplantforafasterandstrongerresistanceonlywhenasubsequentpathogen

Page 5: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 303

attackoccurs (Walters et al., 2014).Priming ismorecost-effectivethanelicitationbecausetheenergycostofinducedresistanceintheplantisoptimized(Beckerset al., 2007). Although the molecular mechanismsbehind priming remain poorly understood, somenatural and synthetic compounds have demonstratedgoodpriming-inducingactivityinlaboratoryandfieldconditions, such as the nonprotein β-aminobutyricacid (BABA) (Walterset al.,2014).Thediversityofdefensemechanismsandsignalingpathwaysinvolvedin induced resistance underline the potential use ofelicitorsinplantprotection.Sincethefirstdiscoveryofelicitorsabout20yearsago,researchonthesespecificcompoundsandtheirmodeofactionhasconsiderablyincreasedunderstandingof theplant immune systemandopensthewaytothedevelopmentofnewtoolsfordisease management strategies (Schwessinger et al.,2012).

Semiochemicals. Living organisms emit a range ofsemiochemicals,i.e.moleculesthatactasmessengersand induce behavioral or physiological responsesin other individuals. They are usually classifiedinto two groups depending on the trophic levelsconcerned:intraspecific(pheromones)orinterspecific(allelochemicals)(Vetetal.,1992).

Semiochemicals are a promising alternative toinsecticidesastheyarehighlyspecific,slightlytoxicornon-toxicmolecules,andaresafefortheenvironment.Inagroecosystems,pestcontrolwithsemiochemicalsisbasedonexploitingmethodstochemicallyincrease,conserveandenhancetheefficacyofnaturalpredation(Rodriguez-Saonaetal.,2012).Push-pullstrategiesuseacombinationofstimulithatmanipulatethebehaviorofinsectpestsand/ornaturalenemiesinordertoaltertheir abundance and distribution in agroecosystems(Cooketal.,2007).Thisapproachisbasedonrepellentor deterrent chemical stimuli that push insect pestsawayfromthecrop.Atthesametime,theyareattractedbyhighlyattractivestimulicomingfromtrapsorfromtrapcropswheretheycanbeeasilycontrolled(Cooketal.,2007).Eachagriculturalsystemneedstodevelopanappropriateanduniquepush-pullstrategybasedona good understanding of the targeted pest’s biologyanditsinteractionswithitshostsandnaturalenemies(Khanetal.,2004).Oneexampleofasuccessfulpush-pull strategy was conducted in Africa against stemborersinmaizeandsorghum(Khanetal.,2001).

Anothersuccessfulapplicationofsemiochemicalsfor direct pest control involves mating disruption,whereby sex pheromones are applied in crops,saturating theenvironment inorder toconfusemalesand interfere with mating behavior. Females fail tomateandthusthenumberoffertilizedeggsandlarvaeisreducedandthereisadecreaseinpestpopulations.This technique is being applied worldwide against

mothsinorchards,vegetablecropsandforestry(Cardé,2007).

Currently, Isomate® dispensers (Shin-EtsuChemical Co. Ltd.), which consist of polyethylene-tubesloadedwithpheromones,arethemostcommondispensersinorchards(Rodriguez-Saonaetal.,2009).In recent years, interest inmethyl salicylate (MeSA)has grown because of its capacity to attract naturalenemies in cultivated fields. This volatile compoundisproducedinseveralplantspeciesandisemittedinresponsetopestattacks(Picherskyetal.,2002).Recentresearchstudieshavedemonstratedtheattractiveeffectof MeSA on natural enemies such as coccinellids,syrphids,lacewings,predatorybugsandsomeparasiticHymenoptera (Rodriguez-Saona et al., 2011). In theUSA, a commercial product containingMeSA as anactive substance is sold as PredaLure® (AgBio Inc.,Westminster,Colorado,USA).

3. IMPLEMENTATION CHALLENGES IN AGRICULTURAL PRACTICES

The literature supports the implementation ofbiostimulant and biocontrol tools in agroecologicalpractices,withcleardemonstrationsoftheirpotentialto reduce chemical inputs, save energy and providefarmers with new opportunities for sustainablefertilizationanddiseasecontrol(Mejía-Tenienteetal.,2010;Chandleretal.,2011;Calvoetal.,2014;Wezelet al., 2014). The agroecological use of these toolswillobviouslyrequireashiftinconventionalpracticesfromtotalrelianceonpesticidesandfertilizerstotheintegratedmanagement of biotic and abiotic stresses(Vallad et al., 2004; Wezel et al., 2014). However,biostimulantandbiocontrolproductsarenotyetusedas routine tools in agriculture. In the second half ofthis review,weexplain thedrawbacks restricting thewidespreaduseofPGPR,elicitorsandsemiochemicalsinagriculture,andwhatisbeingdevelopedtoenhancetheiruse,andthusmakeanimportantcontributiontothe agroecological and sustainable management ofcultivatedecosystems.

3.1. Screening biostimulant and biocontrol products

ThescreeningofsuitablePGPRinoculants,elicitorsandsemiochemicals for specificcrops,growthconditionsand pathogens is critical if the efficacy of theseproducts in the field is to be guaranteed.A commonmethod for screeninganeffectivePGPR inoculant istoisolatestrainsfromplantgrowth-promotingsoilorfrompathogen-suppressivesoil(Mendesetal.,2011).Screening failures can occur, as some PGPR strainswhich show limited ability to promote plant growth

Page 6: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

304 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

during screening trials under controlled conditionscan be among themost effective strains in the field(Araújoetal.,2013).Suchresultscouldbeduetothedifferences between field and controlled conditions,anditappearsthatPGPR,whichincreasethenutrientuptakecapacityofrootsinalargefield,cannotexpresssuch properties easily in vitro or in small-scale pots.In addition, the screening of multiple microbes inconsortia is complicated and requires considerabletimeandresearch,aswellasknowledgeofmicrobialecology and of the interactions between the strainsof a biostimulant product, the host plant and thelocal rhizomicrobial community. Recent progress inmolecular biology and biotechnology will probablyfacilitatePGPRscreening(Bhattacharyyaetal.,2012).

Similarly,thescreeningofelicitorsisusuallydoneunder controlled conditions initially, before beingdone in thefield.Screeningprotocols are adapted toa targeted disease and to the plant to be protected.Different plant genotypes showing various levelsof susceptibility to the disease can be used, as wellas one or several infectious strains of the pathogen.The amount and positioning of the elicitors need tobe optimized, as does the mode of application, thenumberoftreatmentsandtheplantdevelopmentstage(Waltersetal.,2013).Thenext step is to investigatethe signaling pathways involved in the elicitationprocess using variousmethods, such as biochemicalstudies measuring the amount of plant defense-related compounds (plant hormones, phytoalexins,enzymaticactivity,ROS)ormolecularbiologystudiesmeasuring the expression of genes associated withplant defense mechanisms (Walters et al., 2014).The final step involves investigating the influenceof various environmental parameters (temperature,relative humidity, luminosity) for subsequent fieldtrials(Waltersetal.,2014).

3.2. Formulation and application methods

Theformulationandapplicationmethodareprobablyamong the most critical parameters determining theefficiency of biostimulant and biocontrol products.The formulation must maintain an effective plant-growthpromotionorbiocontrolcapacityandbeeasyto use (Bashan et al., 2014;Walters et al., 2014). Inthe case of PGPR inoculants, Bashan et al. (2014)summarized various formulation methods, from thechoiceofcarriers (peat,coirdust,charcoal, sawdust,clay,perlite,vermiculite,polymer-likealginate)totheformulation process. They also summarized variouspractical techniques for inoculant application andproductionachievement.Seedtreatmenthasattractedattentionasasimpleandeconomicallyviabletechnique,being convenient for both farmers and industry(Bashan et al., 2014). The seeds are usually coated

with a carrier and PGPR,with orwithout adhesives(carboxymethyl cellulose, sucrose, vegetable oil,Arabicgum).ThisiscurrentlythemethodmostoftenusedtoapplyPGPRinoculantsasitensuresanoptimalthreshold number of PGPR cells per seed needed tocover the seedling roots.Although the cell thresholddiffers among strains, the common concentration is108cells per plant (Bhattacharyya et al., 2012). Soilapplications of PGPR are performed when a largepopulationofrhizobacteriaisneededataspecificandcrucialplantgrowth stage (e.g. tilleringorfloweringstages)(Bashanetal.,2014).However,soilandopen-air conditions (humidity, temperature) can affect thesuccessofthesoilapplication.Extremetemperaturescan cause a decline in the PGPR survival rate, andsoilhumiditydeterminestheeffectivemobilityoftheinoculatedbacteria in the rhizosphere (Bashanet al.,2014).Usingenoughwater(e.g.atleast100l.ha-1)inthe mixture with liquid or powder-based inoculantsalso ensures that thebacteria arepositionednear theroot system.Additional PGPR inoculations could beneeded tomaintain aminimal bacteria population inthe case of stressful conditions such as winter anddrought(Bashanetal.,2014).

In contrast, commercialized elicitor products areusually applied as a topical spray, once or severaltimes in the season, to complement fungicidetreatments(Waltersetal.,2014).Worraletal.(2012)demonstrated that seeds are also receptive to plantelicitorssuchasjasmonicacidorβ-aminobutyricacid(BABA), thereby triggering long-lasting protectionagainstawidespectrumofpathogens.Seedtreatmentsusingelicitorsrepresentapromisingtechniqueinpestmanagement for sustainable agriculture, but moreresearchisneededtounderstandthebenefitsandcostsoftheapplicationmethods.Soildrenchapplicationsofelicitorshaverecentlybeenreportedtoachievegoodresults(Waltersetal.,2014).

Finally,semiochemicalsdohavesomelimitations,linkedtotheirvolatilenature,butsignificantadvancesandnewapplicationsareexpectedinthecomingyears.Theformulationshould thereforeensureacontrolledrelease of semiochemicals over time. Ideally, slow-releasedevices shouldmeet particular specifications,such as sufficiently high aerial concentrations fordetection, reproducibility in terms of dispenserproduction and efficiency over a given time span(Heuskin et al., 2011).Dispensers should be appliedearlyinthegrowingseason,whenpestpopulationsarequitelow,becausethereleaserateofmostdispenserstendstodecreasewithtime(Witzgall,2001).Intermsof efficacy, the optimal concentration and density ofsemiochemicalsiscrucialtotheeffectivemanipulationof biological control agents (BCAs). The attractionspecificitycanalsobeimprovedbycombiningvolatilecompoundsinblends(Rodriguez-Saonaetal.,2012).

Page 7: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 305

3.3. Farmers and the use of alternative methods

Farmersdonot alwaysgreet the suggestionof usingalternativemethodswithmuchenthusiasm,especiallythoseonsmall-scalefarmsorindevelopingcountries(Gozzo et al., 2013;Bashan et al., 2014).They tendnottoadoptbiostimulantproductsorinnovativecropprotectionstrategiesunlesstheirsuccessisguaranteed.The highest number of farmers currently using bio-based products, which include plant biostimulantsand biopesticides, is in NorthAmerica, representing40%ofthebiocontrolmarket,comparedwith25%inEurope,20%inAsia,10%inSouthAmericaand5%intherestoftheworld(Coxetal.,2013).

Themainreasonforfarmerskepticismaboutthesealternativemethodsrelatestotheirvariableefficacyinthefieldcomparedwithconventionalchemicalinputs(Aroraetal.,2010;Waltersetal.,2013).Manystudieshave shown that these products can have a variablefieldperformance,incontrasttothepromisingresultsobtainedinthelaboratoryoringreenhouseconditions(Gozzoetal.,2013).Thereareseveralreasonsforthisinconsistencyinpracticalconditions.

In the case of PGPR, bacteria concentrations incommercialized products can fall below the desiredthreshold (usual concentration: 108-1011cells.ml-1),especially under long-term or inadequate storage(Bashanetal.,2014).Alesseffective interactioncanalsooccurwhenthePGPRinoculantisnotadaptedtothe host plant or the local environmental conditions(climate, soil characteristics, agronomic practices).For example, modern rice varieties selected to useNfertilizerseffectivelyarelessinteractivewithnativeN-fixing bacteria than traditional varieties (Araújoet al., 2013). Similarly, the performance of elicitorsdepends greatly on field environmental conditions(temperature,relativehumidity,diseasepressure),cropsystems (plant genotype, nutritional requirements,physiologicalstate)andtheformulation(Waltersetal.,2014).

Farmers’ decisions on whether or not to adoptnewmethods often depend on howmuch theywantto change their agricultural practices. Total relianceon new strategies can be challenging. The benefitsof these strategies have to be clearly demonstratedthrougheducationalprogramsthatfocusonfielddata(e.g. pest/disease identification, timingof infestation,crops) (Rodriguez-Saona et al., 2009). This includesdetailed knowledge about agronomic parameters anddesigningadaptedcropmanagementtechniques,withthe appropriate biostimulant or biocontrol productappliedattherighttimeandfrequency,incombinationwithothercontrolmethodsandonresponsivecultivars(Waltersetal.,2013;Bashanetal.,2014).

Henceforth, tools need to be designed that meetfarmers’demandsbyensuring:optimalcropyieldwith

lower input costs; compatibility between the appliedproducts and soil conditions, farming machines andequipment;andgoodshelflifeandlong-termsurvivalduring storage, especially with PGPR inoculants(Bashanetal.,2014).Theintegrationofbiostimulantand biocontrol products into agricultural practicesdependson their economic relevance comparedwithconventionalpractices(Rodriguez-Saonaetal.,2012;Bashan et al., 2014;Walters et al., 2014).Currently,apart from open field applications, biocontroltechniques are widely and efficiently used in thepre- and post-harvest treatments of specific productlines, such as horticultural and ornamental crops(e.g., cucumber, lettuce, cyclamen, roses) (Darras,2012). Further research is needed on improving theunderstanding of which field conditions are mostsuitedtotheuseofaspecificbiostimulantorbiocontrolproduct (Bhattacharyya et al., 2012; Walters et al.,2013).Scientistsareawareofthestakesinvolvedhereandmanypartnershipshavebeenlaunched.InFrance,an integrated network called Elicitra was created in2011with theaimofpromotingthestrategyofplantinduced resistance by elicitors through research,training and development (Bazinet, 2012). Thisnetworkincludespartnersfrompublicresearchbodies,technical institutes, crop industries and universities.PartnersinthenetworkincludeArvalis-PlantInstituteand the French National Institute for AgriculturalResearch (INRA). The successful integration ofbiostimulants and biocontrol products into planthealth care programs is generally ensured by trialscarriedoutbysuchinstitutions.Similarly,aEuropeanstakeholders association, theEuropeanBiostimulantsIndustry Council (EBIC), was founded in 2011 andprovides technicalandpractical informationonplantbiostimulants(Traonetal.,2014).

3.4. Regulatory framework

Alargenumberofbiostimulantandbiocontrolproducts,that have long been known and have been patentedfor agricultural plant growth-promotion and/or pestmanagement, are still not available commerciallyin the EU, unlike the situation in other countries inthe world (Dayan et al., 2009). Many products thatencourage plant growth or plant protection have notbeen registered and there is a lack of fit-for-purposeregulatoryprocedures in theEUbecauseof the timeand costs of registration (Arora et al., 2010; Köhl,2010;Waltersetal.,2014).TheapprovalofanyPlantProtection Product (PPP) requires the registrationof the activematerial on a list validated by the EU(EuropeanParliament,2009b).

In the case of biostimulants, these products arestill not covered by EU regulatory procedures andneedtobeincludedintheframeworkofPPPproducts.

Page 8: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

306 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

TheFertilizersRegulation2003/2003coversonlytheplacingofinorganicfertilizers(ECfertilisers)onthemarketandhasbeenunderrevisionsince2012-2013,withtheaimofextendingitsscopetootherfertilizingandrelatedmaterials,suchasplantbiostimulantsandfertilizeradditives(Traonetal.,2014).

Thelegalaspectsrelatedtousingsemiochemicalsarealsocomplicated.Semiochemicalsareregulatedasa group of biopesticides (i.e. active substances usedfor plant protection that are of natural origin or arenature-identicalsyntheticsubstances)(Chandleretal.,2011).IntheEU,biopesticidesaresubjecttothesameregistrationprocedureassyntheticpesticides,butnewguidelines facilitating their registration process arebeing developed (Czaja et al., 2015). The status ofbiopesticideswasestablishedinRegulation1095/2007(CommissionRegulation,2007).ItallowstheinsertionintoRegulation540/2011ofactivesubstancesthatdonothaveaharmfulimpactonhumanandanimalhealth,or a negative environmental impact. Each substancehastocomplywithstrictcriteria,listedinAnnexIVofRegulation1095/2007,inordertobeconsideredsafe(CommissionImplementingRegulation,2011).

The current strategy of the EU in sustaining thedevelopment of new biostimulant and biocontrolmethods in agriculture is implemented via variouslegislative procedures. Regulation1107/2009 aimsto harmonize the overall procedures authorizingplant protection products in the EU market. It alsofacilitatesapprovalofnaturalsubstances(Article23),thereby simplifying the regulation procedures fornatural preparations with low risk. The EU hasrecently proposed granting the first approvals foragrochemicals in a new category entitled “basicsubstances”. In addition, Directive2009/128/ECregulatesthesustainableuseofpesticidesinEurope:“Member States shall take all necessary measures to promote low pesticide-input pest management, giving wherever possible priority to non-chemical methods, so that professional users of pesticides switch to practices and products with the lowest risk to human health and the environment among those available for the same pest problem”(EuropeanParliament,2009a).

In2008,FranceannounceditsEcophyto2018plan,whichaimstoreducepesticideuseby50%by2018,mainlythroughtheidentificationanddevelopmentofbioactivecompoundsabletostimulateplantimmunity(Information Réglementaire Ecophyto-2018, 2011).Thereductionofconventional inputs isalsoplannedin other European countries, including Belgium(NAPAN,2013),Germany (NationalActionPlanonSustainableUse of Plant Protection Products, 2013)andtheUK(UKNAP,2013).Thepromiseofstronggrowth in thebiocontrolmarket inanear futurehasalso ledmajor agrochemical companies to invest inthese green technologies. All stakeholders in the

agriculturalsector, includingagriculturaldistributorsand plant breeders, could play an important role inpromoting the use of biostimulant and biocontrolproducts.

4. CONCLUSIONS AND PERSPECTIVES

Strong efforts are being made to improve attitudesinthefarmingcommunity,andinsocietyingeneral,towards the use of alternative methods to chemicalinputs. It iswidely agreed thatPGPRbiostimulants,elicitors and semiochemicals should not be used asstand-alonemethods in agroecologicalmanagement,but integrated into fertilization and disease/pestcontrol strategies to complement chemical inputsandcontributetoareductionintheirdosageamountsandapplicationfrequency.Althoughthesetoolshavebeen widely endorsed for their advantages, farmersandgrowersare stillnot completelyconfidentaboutusing them,mainlybecauseof theirfluctuatingfieldperformance (Beckers et al., 2007). Farmers needmore informationonhow to use these tools in theiragricultural practices.Regulators, investors, growersand consumers also need to bewell informed abouttheadvantagesofthesealternativemethodsandtheirpotentialinpromotingsustainableagriculture.

Furtherresearchisneededtobetterunderstandtheenvironmental parameters affecting the efficiencyofthese products, particularly for field crops. Specialattentionshouldalsobegiventotheformulationandthe potential interactions of these products with theplantenvironment.Multidisciplinaryresearchgroups,such as the AgricultureIsLife platform (GemblouxAgro-BioTech,UniversitédeLiège,Belgium),shouldaddress the question of howbest to use these tools,given current practices, by studying the issues thatstillneedtobeovercome(e.g.screeningmethodology,formulation,environmentalimpact).

Many challenges remain before biostimulant andbiocontrol products can be widely and successfullyusedonacommercialbasis,buttheintensiveeffortsin research and the legislative area, as well as inenhancingsociety’sawarenessoftheseproducts,willincreasetheircredibilityandacceptance(Wezeletal.,2014).Agricultural practices using these tools needtobeadapted(e.g.usingcultivarsspecificallychosenfor theappropriate responses) (Walterset al.,2014).InEurope,thelong-termobjectivetobepesticidefreeis already leading to changes in crop managementpracticesand representsamajordriver in theuseofbiostimulant and biocontrol products. Within thecontext of climate change, increasing environmentalconcerns and population increase, these alternativemethodsofferimportantpotentialtoolsforachievingsustainablefoodproduction.

Page 9: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 307

Acknowledgements

WethanktheUniversityofLiège-GemblouxAgro-BioTechandmorespecificallytheresearchplatformAgricultureIsLifeforthefundingofthisresearchproject.

Bibliography

AdesemoyeA.,TorbertH.&KloepperJ.,2009.Plantgrowth-promoting rhizobacteria allow reduced application ratesof chemical fertilizers. Microb. Ecol., 58(4), 921-929.

AraújoA.E.D.S.etal.,2013.Responseoftraditionaluplandrice varieties to inoculationwith selected diazotrophicbacteria isolated from rice cropped at the northeastregionofBrazil.Appl. Soil Ecol.,64,49-55.

AroraN.K., KhareE. & MaheshwariD.K., 2010. Plantgrowth promoting rhizobacteria: constraints inbioformulation,commercialization,andfuturestrategies.In: MaheshwariD.K., ed. Plant Growth and Health Promoting Bacteria. Dordrecht, The Netherlands:Springer.

BashanY.etal.,2014.Advancesinplantgrowth-promotingbacterial inoculant technology: formulations andpracticalperspectives(1998-2013).Plant Soil,378(1-2),1-33.

BazinetC., 2012. ELICITRA, http://elicitra.org/, (04.08.2014).

BeckersG.J. & ConrathU., 2007. Priming for stressresistance.Curr. Opin. Plant Biol.,10(4),425-431.

BhattacharyyaP.& JhaD., 2012. Plant growth-promotingrhizobacteria (PGPR):emergence inagriculture.World J. Microbiol. Biotechnol.,28(4),1327-1350.

BollerT. & FelixG., 2009. A renaissance of elicitors:perceptionofmicrobe-associatedmolecularpatternsanddanger signals by pattern-recognition receptors.Annu. Rev. Plant Biol.,60,379-407.

ÇakmakçiR.etal.,2014.Rhizobacteriaforreducedfertilizerinputs in wheat (Triticum aestivum spp. vulgare) andbarley(Hordeum vulgare)onAridisolsinTurkey.Int. J. Plant Prod.,8(2),163-181.

CalvoP.,NelsonL.&KloepperJ.,2014.Agriculturalusesofplantbiostimulants.Plant Soil.,383(1),3-41.

CardéR., 2007. Using pheromones to disrupt mating ofmothpests.In:KoganM.&JepsonP.,eds.Perspective in ecological theory and integrated pest management.CambridgeUniversityPress,122-169.

ChandlerD.etal.,2011.Thedevelopment, regulationanduse of biopesticides for integrated pest management.Philos. Trans. R. Soc. B,366(1573),1987-1998.

ChowdhuryS.P. et al., 2013. Effects of Bacillus amyloliquefaciens FZB42 on lettuce growth andhealth under pathogen pressure and its impact on therhizospherebacterialcommunity.PLoS ONE,8(7).

Commission ImplementingRegulation,2011.Commission implementing Regulation (EU) No 540/2011 of 24 May

2011 implementing Regulation (EU) No 1107/2009 of the European Parliament and of the Council as regards the list of approved active substances, http://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2011:153:0001:0186:EN:PDF,(04.08.2014).

Commission Regulation, 2007. Commission Regulation (EC) No 1095/2007 of 20 September 2007 amending Regulation (EC) No 1490/2002 laying down further detailed rules for the implementation of the third stage of the programme of work referred to in Article 8(2) of Council Directive 91/414/EEC and Regulation (EC) No 2229/2004 laying down further detailed rules for the implementation of the fourth stage of the programme of work referred to in Article 8(2) of Council Directive 91/414/EEC, http://www.pesticides.gov.uk/Resources/CRD/Migrated-Resources/Documents/C/Commission_Regulation_1095_2007.pdf,(06.01.2015).

CookS.M., KhanZ.R. & PickettJ.A., 2007. The use ofpush-pull strategies in integrated pest management.Annu. Rev. Entomol.,52,375.

CoxM. & WongB., 2013. Biological crop chemistry primer: green shoots through green products, PiperJaffrayindustrynote,http://library.constantcontact.com/download /ge t / f i l e /1102591137375-215 /Cox+Industry+Note+-+Agriculture+08.27.13+copy.pdf,(06.01.2015).

CzajaK. et al., 2015. Biopesticides – towards increasedconsumersafetyin theEuropeanUnion.PestManage. Sci.,71,3-6.

DarrasA.I.,2012.Novelelicitorsinducedefenseresponsesin cutflowers. In:CumagunC.J., ed.Plant pathology.Rijeka,Croatia:InTech.

DayanF.E., CantrellC.L. & DukeS.O., 2009. Naturalproductsincropprotection.Bioorg. Med. Chem.,17(12),4022-4034.

du JardinP., 2012.The science of plant biostimulants – a bibliographic analysis, http://ec.europa.eu/enterprise/sectors/chemicals/files/fertilizers/final_report_bio_2012_en.pdf,(07.14.2015).

EuropeanBiostimulantsIndustryConsortium,2011.Position on the revision of Reg (EC) 2003/2003, http://www.biostimulants.eu/wp-content/uploads/2011/12/EBIC_Position_7Options_29Nov2011.pdf,(07.14.2015).

European Commission EGTOP, 2011. Final report on plant protection products, http://ec.europa.eu/agriculture/organic/eu-policy/expert-advice/documents/final-reports/final_report_egtop_on_plant_protection_products_en.pdf,(07.14.2015).

European Commission, 2012. Sustainable agriculture for the future we want, http://ec.europa.eu/agriculture/events/2012/rio-side-event/brochure_en.pdf, (06.01.2015).

EuropeanParliament,2009a.Directive 2009/128/EC of the European Parliament and of the Council of 21 October 2009 establishing a framework for Community action to achieve sustainable use of pesticides, http://eur-lex.

Page 10: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

308 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2009:309:0071:0086:en:PDF,(04.08.2014).

European Parliament, 2009b. Regulation (EC) No 1107/2009 of the European Parliament and of the Council of 21 October 2009 concerning the placing of plant protection products on the market and repealing Council Directives 79/117/EEC and 91/414/EEC, http://eur-lex.europa.eu/legal-content/EN/TXT/PDF/?uri=CELEX:02009R1107-20130701&rid=1,(04.08.2014).

García-MierL. et al., 2013. Agriculture and bioactives:achieving both crop yield and phytochemicals. Int. J. Mol. Sci.,14,4203-4222.

GozzoF. & FaoroF., 2013. Systemic acquired resistance(50years after discovery):moving from the lab to thefield.J. Agric. Food Chem.,61,12473-12491.

HenryG., 2013. Study of the induced systemic resistance of plants: molecular aspects of the interaction between plant cells and amphiphilic elicitors produced by non-pathogenic rhizobacteria. PhD thesis: Universitéde Liège - Gembloux Agro-Bio Tech, Gembloux(Belgium).

HenryG., ThonartPh. & OngenaM., 2012. PAMPs,MAMPs,DAMPsandothers:anupdateonthediversityof plant immunity elicitors. Biotechnol. Agron. Soc. Environ.,16(2),257-268.

HerthA.,2011.Le bio-contrôle pour la protection des cultures : 15 recommandations pour soutenir les technologies vertes, http://www.ladocumentationfrancaise.fr/var/storage/rapports-publics/114000224.pdf,(07.14.2015).

HeuskinS. et al., 2011. The use of semiochemical slow-releasedevicesinintegratedpestmanagementstrategies.Biotechnol. Agron. Soc. Environ.,15(3),459-470.

HolG.W.H.,BezemerM.T.&BiereA., 2013.Getting theecology into interactions between plants and the plantgrowth-promotingbacteriumPseudomonas fluorescens.Front. Plant Sci.,4.

Information Réglementaire Ecophyto 2018, 2011.Information Réglementaire Ecophyto2018 : de nouveaux textes pour sécuriser et diminuer l’utilisation de produits phytopharmaceutiques, http://agriculture.gouv.fr/telecharger/36425?token=d00ad843ed4f7e8a30af796f5d68c200forme_agrement_certificat-individuel.pdf,(16.11.2015).

IslamF.etal.,2014.InfluenceofPseudomonas aeruginosaasPGPRonoxidativestresstoleranceinwheatunderZnstress.Ecotoxicol. Environ. Saf.,104(1),285-293.

JonesJ.D.G.&DanglJ.L.,2006.Theplantimmunesystem.Nature,444(7117),323-329.

KhanZ.R.,PickettJ.A.,WadhamsL.&MuyekhoF.,2001.HabitatmanagementforthecontrolofcerealstemborersandStrigainmaizeinKenya.Insect Sci. Appl.,21,375-380.

KhanZ.R.&PickettJ.A., 2004.The “Push-Pull” strategyfor stemborermanagement: a case study in exploitingbiodiversity and chemical ecology. In: GurrG.,

WrattenS.D.&AltieriM.A.,eds.Ecological engineering for pest management: advances in habitat manipulation for arthropods.Oxfordshire:CABIPublishing,155-164.

KilianM. et al., 2000. FZB24®Bacillus subtilis – modeofactionofamicrobialagentenhancingplantvitality.Pflanzenschutz-Nachr. Bayer,53(1),72-93.

KöhlJ.,2010.Microbials:theneedforapragmaticapproach.New Ag Int.,September,34-42.

KröberM. et al., 2014. Effect of the strain Bacillus amyloliquefaciensFZB42on themicrobialcommunityin the rhizosphere of lettuce under field conditionsanalyzed by whole metagenome sequencing. Front. Microbiol.,5,252.

MäderP.etal.,2011.Inoculationofrootmicroorganismsforsustainablewheat–riceandwheat–blackgramrotationsinIndia.Soil Biol. Biochem.,43,609-619.

MarksB.B. et al., 2013. Biotechnological potential ofrhizobial metabolites to enhance the performance ofBradyrhizobium spp. and Azospirillum brasilenseinoculants with soybean and maize. AMB Express, 3,1-10.

Mejía-TenienteL. et al., 2010. Use of elicitors as anapproachforsustainableagriculture.Afr. J.Biotechnol.,9(54),9155-9162.

MendesR. et al., 2011. Deciphering the rhizospheremicrobiome for disease-suppressive bacteria. Science,332(6033),1097-1100.

NAPAN, 2014. Belgian action plan to reduce the risks and impacts linked to pesticides 2013-2017, http://documentatie.leefmilieubrussel.be/documents/plan_belgian_pesticides_en.PDF?langtype=2067, (07.15.2015).

NationalActionPlanonSustainableUseofPlantProtectionProducts, 2013. National Action Plan on Sustainable Use of Plant Protection Products, http://ec.europa.eu/food/plant/pesticides/sustainable_use_pesticides/docs/nap_germany_en.pdf,(07.15.2015).

Pérez-MontañoF. et al., 2013. Plant growth promotion incereal and leguminous agricultural important plants:from microorganism capacities to crop production.Microbiol. Res.,169,325-336.

PicherskyE. & GershenzonJ., 2002. The formation andfunction of plant volatiles: perfumes for pollinatorattraction and defense. Curr. Opin. Plant Biol., 5(3),237-243.

ReddyM.V.B. et al., 1999. Chitosan treatment of wheatseedsinducesresistancetoFusariumgraminearumandimproves seed quality. J. Agric. Food Chem., 47(3),1208-1216.

ReddyP.P.,2014.Plant growth promoting rhizobacteria for horticultural crop protection.NewDelhi:Springer.

Rodriguez-SaonaC.R. & StelinskiL.L., 2009. Behavior-modifying strategies in IPM: theory and practice.In: PeshinR. & DhawanA.K., eds. Integrated pest management: innovation-development process.TheNederlands:Springer,263-315.

Page 11: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 309

Rodriguez-SaonaC. et al., 2011. Field responses ofpredaceous arthropods to methyl salicylate: a meta-analysis and case study in cranberries. Biol. Control,59(2),294-303.

Rodriguez-SaonaC., BlaauwD. & IsaacsR., 2012.Manipulation of natural enemies in agroecosystems:habitat and semiochemicals for sustainable insect pestcontrol.In:MarceloL.L.&SoloneskiS.,eds.Integrated pest management and pest control - current and future tactics.Rijeka,Croatia:InTech,89-126.

SchwessingerB. & RonaldP.C., 2012. Plant innateimmunity:perceptionofconservedmicrobialsignatures.Annu. Rev. Plant Biol.,63,451-482.

SimwingaE.,MukukaR.&KamwaleH.,2010.TheuseofTwinN(nitrogenfixingbacteriafornonlegumes)asanalternativesourceofnitrogenforsugarcaneproduction.In: S.A.S.T.Association, ed.Proceedings of the South African Sugar Technologists’ Association annual general meeting and congress,83,221-224.

SobhyI.S.etal.,2012.Less ismore: treatmentwithBTHand laminarin reduces herbivore-induced volatileemissions inmaize but increases parasitoid attraction.J. Chem. Ecol.,38(4),348-360.

TajiniF., TrabelsiM. & DrevonJ.J., 2012. Combinedinoculation with Glomus intraradices and Rhizobium tropiciCIAT899increasesphosphorususeefficiencyforsymbioticnitrogenfixationincommonbean(Phaseolus vulgarisL.).Saudi J. Biol. Sci.,19(2),157-163.

TállaiM. et al., 2012. Comparative examination of abacteriumpreparation(Bactofil®A10)andanartificialfertilizer [Ca(NO3)2] on humic sandy soil. Ann. Univ. Oradea Fascicula Protecţia Mediului,18,81-88.

TraonD. et al., 2014. A legal framework for plant biostimulants and agronomic fertiliser additives in the EU – Report to the European Commission, Enterprise & Industry Directorate – General. Brussels: ArcadiaInternational.

UKNAP,2013.UK National action plan for the sustainable use of pesticides (plant protection products), http://c-ipm.org/fileadmin/c-ipm.org/British_NAP__in_EN_.pdf,(07.15.2015).

UpadhyayS.K.&SinghD.P., 2015.Effect of salt‐tolerantplant growth‐promoting rhizobacteria on wheat plants

and soil health in a saline environment. Plant Biol.,17(1),288-293.

ValladG.E. & GoodmanR.M., 2004. Systemic acquiredresistance and induced systemic resistance inconventionalagriculture.Crop Sci.,44(6),1920-1934.

VetL.E.M.&DickeM.,1992.Ecologyofinfochemicaluseby natural enemies in a tritrophic context.Annu. Rev. Entomol.,37,141-172.

WalkerV. et al., 2012. Variation of secondary metabolitelevels in maize seedling roots induced by inoculationwithAzospirillum,PseudomonasandGlomusconsortiumunderfieldconditions.PlantSoil,356(1-2),151-163.

WaltersD.R., RatsepJ. & HavisN.D., 2013. Controllingcrop diseases using induced resistance: challenges forthefuture.J. Exp. Bot.,64(5),1263-1280.

WaltersD.R.,NewtonA.C.&LyonG.D.,eds,2014.Induced resistance for plant defense: a sustainable approach to crop protection.2nded.Oxford,UK:Wiley-Blackwell.

WasternackC.&HauseB.,2013.Jasmonates:biosynthesis,perception,signaltransductionandactioninplantstressresponse, growth and development. An update to the2007 review in Annals of Botany. Ann. Bot., 111(6),1021-1058.

WezelA. et al., 2014. Agroecological practices forsustainableagriculture.Areview.Agron. Sustain. Dev.,34(1),1-20.

WitzgallP., ed., 2001. Pheromones and other biologicaltechniquesforinsectcontrolinorchardsandvineyards.Working group “Use of pheromones and othersemiochemicals in integrated control”, Proceedingsof the meeting in Hohenheim (Baden-Württemberg,Germany), 10-12 November, 1999. IOBC/wprs Bull.,24(2).

WorrallD.etal.,2012.Treatingseedswithactivatorsofplantdefencegenerates long‐lastingprimingof resistance topestsandpathogens.New Phytol.,193(3),770-778.

YaoA.V.etal.,2006.EffectofFZB24®Bacillus subtilisas a biofertilizer on cotton yields in field tests.Arch. Phytopathol. Plant Prot.,39(4),323-328.

(72ref.)

Page 12: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

310 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

Appendix 1.Examplesofpromisingplantgrowth-promotingrhizobacteria(PGPR)-basedbiostimulantsforthereductionof chemical fertilizers under various experimental conditions — Exemples de biostimulants prometteurs à base de rhizobactéries promotrices de la croissance des plantes (PGPR), pour une réduction de l’usage des fertilisants chimiques dans des conditions d’expérimentations variées.Biostimulant Enhancement of crop growth and

reduction of chemical fertilizer level

Crop Experimental conditions

Reference

Bacillus megaterium M3,BacillusOSU-142,Azospirillum brasilense sp.245,Paenibacillus polymyxaRC05,Bacillus megateriumRC07,Bacillus licheniformisRC08,Raoutella terrigena,Burkholderia cepaciaFSTur

Plantrootandshootweightincreaseundergreenhouseconditions.SingleandcombinationsofPGPRincreasedyieldupto40.4%forwheatand33.7%forbarleyunderfieldconditionsandincombinationwithNfertilizer

Wheat,barley Greenhouseandfield

Çakmakçietal.,2014

Pseudomonas aeruginosa ImprovedNandPuptake.IncreaseinleafchlorophyllamountsandplantbiomassunderZnstress(enhancementofantioxidativeenzymes,ascorbicacidandtotalphenolics)

Wheat Greenhouse Islametal.,2014

Arthrobactersp.andBacillus subtilis

Increasedplanttolerancetosalinity.Plantdryweightincreasedupto26%and40%under2dS.m-1and6dS.m-1salinitylevel,respectively

Wheat Greenhouse Upadhyayetal.,2015

Burkholderia vietnamiensis AR112

Increasedorequivalentweightandyieldoftraditionalricecomparedwith100%Nchemicalfertilization

Rice Field Araújoetal.,2013

Bradyrhizobiumspp.andconcentratedmetabolitesfromBradyrhizobium diazoefficiens

Increasedgrainyieldby4.8%comparedwiththeexclusiveuseofBradyrhizobiumspp.

Soybean Greenhouseandfield

Marksetal.,2013

Rhizobium tropiciCIAT899,Glomus intraradices

IncreaseinPandNamountsupto40%and42%,respectively,insoil.Nodulenumberenhancedby70%andnodulemassby43%.Plantshootdryweightincreasedbyupto24%androotgrowthbyupto48%

Bean Greenhouse Tajinietal.,2012

Pseudomonas jessenii,Pseudomonas synxantha andalocalAM

PGPRorAMFaloneincreasedyieldbyupto29%and31%,respectively.CombiningPGPRandAMFincreasedtheyieldbyupto41%

Wheat Field Mäderetal.,2011

Bacillus amyloliquefaciensIN937a,Bacillus pumilus T4,Glomus intraradices

InoculationofPGPRandAMtogetherreducedfertilizeruseby25%.Combinationwasequivalentto100%fertilizerapplicationforplantgrowth,yieldandnutrientuptake

Tomato Greenhouse Adesemoyeetal.,2009

Bacillus subtilis Plantgrowthandyieldenhancedbyupto30%comparedwithNPKfertilization

Cotton Field Yaoetal.,2006

Page 13: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 311

Appendix 2. Examples of commercializedPGPR-based products inEurope,NorthAmerica andAsia— Exemples de produits à base de rhizobactéries promotrices de la croissance des plantes, actuellement commercialisés en Europe, en Amérique du Nord et en Asie.Products Rhizobacteria Crop ManufacturerAmase® Pseudomonas azotoformans

Cucumber,lettuce,tomato,pepper

LantmannenBioagri,SwedenAmniteA100® Azotobacter,Bacillus,

Pseudomonas,Rhizobium,Chaetomium

ClevelandBiotech,UK

BactoFilA10® Azospirillum brasilense,Azotobacter vinelandii,B. megaterium,B. polymyxa,P. fluorescens

Monocotyledons(cereals)

AGRO.bioHungaryKft.,HungaryBactoFilB10® Azospirillum lipoferum,

Azotobacter vinelandii,B. megaterium,B. circulans,B. subtilis,P. fluorescens

Dicotyledons(sunflower,potato,rapeseed)

Cérès® Pseudomonas fluorescens Fieldandhorticulturalcrops Biovitis,FranceCompete®Plus B. azotofixans,B. licheniformis,

B. megaterium,B. polymyxa,B. pumilus,B. subtilis

Fieldcrops,treenurseries PlantHealthCare,USA

FZB24®fl B. amyloliquefaciens ssp.plantarum

Ornamentals,vegetablesABiTEPGmbH,Germany

Rhizovital42® B. amyloliquefaciens FieldcropsGmax®PGPR Azotobacter,Phosphobacteria,

P. fluorescensFieldcrops GreenmaxAgroTech,India

Inómix®Biostimulant B. polymyxa(IAB/BP/01),B. subtilis(IAB/BS/F1)

Cereals IAB(Iabiotec),SpainInómix®Biofertilisant B. megaterium, Saccharomyces cerevisiae, Azotobacter vinelandii, Rhizobium leguminosarum

Inómix®phosphore P. fluorescens, B. megaterium, Saccharomyces cerevisiae

MicosatF®Uno Agrobacterium radiobacterAR39,B. subtilisBA41,Streptomycesspp.SB 14

Fruits,vegetables,flowers

CCSAostaSrl,ItalyMicosatF®Cereali B. subtilisBR62,Paenibacillus durusPD76,Streptomycesspp.ST 60

Cereals,tomatoes,sunflowers,beet,soybeans

Nitroguard® Azospirillum brasilenseNAB317,Azorhizobium caulinodensNAB38,Azoarcus indigensNAB04,Bacillussp. Cereals,seedrape,sugar

beet,sugarcane,vegetablesMapletonAgriBiotecPtyLtd,AustraliaTwinN® Azospirillum brasilenseNAB317,

Azorhizobium caulinodens NAB38,Azoarcus indigensNAB04

PGA® Bacillussp. Fruits,vegetables Organicatechnologies,USARhizocell®GC B. amyloliquefacienssoucheIT45 Cereals LallemandPlantCare,

CanadaSymbion®-N Azospirillum, Rhizobium,

Acetobacter, AzotobacterFieldcrops,vegetables T.Stanes&CompanyLtd,

IndiaSymbion®-P B. megaterium var. phosphaticumSymbion®-K Frateuria aurantia

Page 14: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

312 Biotechnol. Agron. Soc. Environ. 201620(S1),299-313 LeMireG.,NguyenM.L.,FassotteB.etal.

Appendix 3.Examplesof commercializedproductswith elicitorproperties—Exemples de produits commercialisés et présentant des propriétés de stimulateurs de défenses naturelles des plantes.Products Product origin Crop Disease Target ManufacturerVacciplant® Laminarinextract

frombrownalgaeLaminaria digitata

Appleorchards,tomato,lettuce,cucumber,strawberry,grapevine

Powderymildew,downymildew

LaboratoireGoëmar,France

Actigard®/Bion®/Blockade®

Acibenzolar-S-méthyl Wheat,tomato Powderymildew,bacterialdiseases

SyngentaCropProtection,USA

Elexa®4PDB Chitosan Grapevine,tomato,potato,cucumber,fieldcrops

Botrytisgreymould(Botrytis cinerea),powderymildew,downymildew

PlantDefenseBoostersInc.,USA

Armour-Zen® Chitosan Grapevine,ornamentals

Botrytisgreymould(Botrytis cinerea),whiterot(Sclerotinia sclerotiorum)

Botry-Zen2010Ltd,NewZealand

Chitoplant® Chitosan Tomato Powderymildew ChiProGmbH,Germany

Harp-N-Tek® HarpinproteinfromthebacteriaErwinia amylovara

Appleandpearorchards,grapevine,tomato

Appleandpearscab,downymildew

PlantHealthCareInc.,USA

Milsana® EthanolicleafextractfromgiantknotweedReynoutria sachalinensis

Cucumber,strawberry,tomato,wheat

Powderymildew KHHBioScience,USA;BIOFAAG,Germany

Stifenia® FEN560(Fenugrec) Grapevine Powderymildew S.O.F.T.,France

HelenaProphyt® Potassiumphosphite Fieldcrops,vineyards,orchards

Downymildew,purpleblotch(Alternariaspp.),brownrot(Monilia fructicola)

HelenaChemicalCompany,USA

Aliette®WG Fosetyl-Al Ornamentaltreesandbushes,strawberry

Downymildew BayerCropScience,Germany

AdaptedfromWaltersetal.(2014)—adapté de Walters et al. (2014).

Page 15: Implementing plant biostimulants and biocontrol … · Implementing plant biostimulants and biocontrol strategies in the agroecological management of cultivated ecosystems. ... Le

Biostimulantsandbiocontrolinagroecologicalmanagement 313

Variouselicitorcategorieshavebeendetermined.Elicitorscanbelongtomicrobe-associatedmolecularpatterns(MAMPs)emittedfromnon-pathogenicmicroorganismssuchasplantgrowth-promotingrhizobacteriaandfungi(PGPR,PGPF)oryeasts,andtriggerinducedsystemicresistance(ISR)inthehostplantthroughjasmonicacid(JA)andethylene(ET)signalingpathways;Damage-associatedmolecularpatterns(DAMPs)resultfromplantcelldegradationafterwoundingbyinsectsorherbivores;Pathogen-associatedmolecularpatterns(PAMPs)areemittedfromvariouspathogens.ChemicalssuchasAcibenzolar-S-methyl(ASM)andProbenazole,alongwithDAMPSandPAMPs,triggersystemicacquiredresistance(SAR)intheplantthroughthesalicylicacid(SA)signalingpathway,aswellastheaccumulationofpathogen-related(PR)proteins.Thevarioussignalingpathwayscitedherearenon-exhaustiveandothertypesofresistancecanoccurintheplant.TherearecomprehensivedetailsonthisinJonesetal.(2006)—Différentes catégories d’éliciteurs ont été caractérisées. Les éliciteurs peuvent faire partie des microbe-associatedmolecularpatterns(MAMPs) émis par des micro-organismes non pathogènes tels que les rhizobactéries et champignons promoteurs de la croissance des plantes (PGPR, PGPF), ou des levures, et stimulent les systèmes de défenses de la plante sous forme d’une résistance systémique induite (ISR) via les voie de signalisation de l’acide jasmonique (JA) et de l’éthylène (ET) ; les damage-associatedmolecularpatterns (DAMPs) résultent d’une dégradation de cellules végétales suite à une blessure infligée par un insecte ou un herbivore ; les pathogen-associatedmolecularpatterns (PAMPs) sont émis par une variété d’agents pathogènes. Des substances chimiques telles que l’acibenzolar-S-methyl (ASM), le probénazole, ainsi que les DAMPS et PAMPs, stimulent une résistance systémique acquise dans la plante (SAR) via la voie de signalisation de l’acide salicylique (SA), tout en entrainant une accumulation de protéines PR (pathogen-related) dans la plante. L’ensemble des voies de signalisation citées ici n’est pas une liste exhaustive et d’autres types de résistances peuvent intervenir au sein de la plante. D’excellents détails à ce sujet sont fournis dans les travaux de Jones et al. (2006).

INDUCED SYSTEMIC RESISTANCE (ISR) SYSTEMIC ACQUIRED RESISTANCE (SAR)

JA and ET signaling pathways

SA signaling pathway

MAMPs

PAMPs

DAMPs

Chemicals

PR proteinaccumulation

Pathogenic bacteria, fungi, virus

Insects, herbivores

PGPR, PGPF, yeast, etc.

Appendix 4.Elicitorsandtheplantsignalingpathwaysforinducedresistance—Éliciteurs et voies de signalisation de la plante pour la résistance induite.