Entomo-pathogenic Fungi

An entomopathogenic fungus can act as a parasite of insects and kills or seriously disables them.Targets are distributed among 10 insect orders:
Hemiptera (59.6%), Coleoptera (40.9%), Lepidoptera (17.5%), Thysanoptera (14.6%), Orthoptera (9.4%), Diptera (7.0%), Hymenoptera (2.9%), Isoptera (2.3%), Siphonoptera (1.2%), and Blattodea(0.6%).

1. Mass production of
Entomopathogenicfungi

2. Introduction
Increasingpublicsensitivitytoenvironmentalpollution
andproblemsofpestresistancetochemicalpesticideshas
ledtoaglobalconsensustoreduceorphaseoutextremely
noxiouspesticides(e.g.methylbromide).
Thereisurgentneedtomakeefforttodevelopnew,
environmentallyfriendlycropprotectionstrategies.
Atpresent,cropprotectionistrappedbetweenthe
increasingnumberofprohibitedchemicalpesticidesand
thelackofsafe,efficientalternatives.Entomogenousfungi
offeranenvironmentfriendlyalternativetochemical
pesticides.

3. Entomopathogenic fungus (EPF)
Anentomopathogenicfungusisafungusthat
canactasaparasiteofinsectsandkillsor
seriouslydisablesthem.
Thesefungicompriseaheterogenousgroupof
over100generawithapproximately750
species.
Targets are distributed among 10 insect
orders:
Hemiptera(59.6%), Coleoptera(40.9%),
Lepidoptera (17.5%), Thysanoptera(14.6%),
Orthoptera (9.4%), Diptera (7.0%),
Hymenoptera (2.9%), Isoptera(2.3%),
Siphonoptera(1.2%), and Blattodea(0.6%).

4. History
•InFranceandItaly,wheresilkproductionwasimportantinthe16thand17
th
centuries,heavylossesoflarvalsilkwormsoccurredfrom"muscardinedisease.“
•In1835,theItalianscientistAgostinoBassideLodi(the"FatherofInsect
Pathology")showedthatthemuscardinediseaseofsilkwormswasactually
causedbyafungusthatmultipliedinandonthebodyoftheinsect.
B. Bassiana infected larva

5. Cond…
Themuscardinefunguswasthefirstmicroorganismtobe
recognizedasacontagiousagentofanimaldiseasewhich
waslateridentifiedasB.bassiana.
In1883,Metchinikoffinitiatedmassculturingoffungus
andcarriedoutthefirstexperimentwithtwobeetlepests.
Thegreenmuscardinefungus,Metarhiziumanisopliaewas
describedforthefirsttimebyMetschnikoffin1879.
Metarhiziumanisopliae(Metschnikoff)Sorokinisthesecond
mostwidelyexploitedentomogenousfungusinbiocontrol
afterB.bassiana
Verticilliumlecanii(Zimm.)popularlycalledthe“whiteholo”
isknowntocausemycosisinanumberofinsectsbelonging
totheinsectordersHomoptera,Coleopteraand
Lepidoptera.

6. Examples
Fly infected by
Entomophthora
fungus
The asexual (anamorph) phases:
Beauveriabassiana(against Colorado potato beetle,
cabbage looper, grasshoppers, silkworm,)
Paecilomycesfumosoroseus(against white flies, thripsand
aphids)
Metarhiziumspp. (against soil insects like beetles,
locusts, white grub, spider mites)
Verticilliumlecanii(against white flies, thripsand aphids)
Nomuraea
Hirsutella(against mites)
Entomophthora
The sexual (teleomorph) state:
Cordycepsspecies that infect a wide
spectrum of arthropods.

7. Life cycle
Dispersalofconidiafromalarval
cadaver,attachmenttothecuticleofa
newhost,breachingthecuticle,
proliferationintheformofbudding
hyphalbodies,germinationandgrowth
ofmycelium,formationofmycelialmass,
exitthroughthecuticleandformationof
newaerialconidia.

8. Mode of infection
Attachment:-EGFhavenoknownsexualcyclesoinsectsare
infectedbyconidia(asexualpropagules)whichattachtothehost
cuticle.
Conidiagermination:-inanenvironmentwithhighhumidity.
Penetrationandmultipication:-Thegermtubesdevelopingfrom
theconidiapenetratethehostcuticleandinvadethehaemocoel.
Myceliaproliferateandreleaseblastosporesbybudding.
Secretionofenzymesandtoxin:-fordegradationofproteins,
chitinandlipidsintheinsectintegument.
Deathofinsect:-Hostinsectsarekilledduetodepletionoftheir
haemolymphnutrientsand/orduetotoxemiacausedbyfungal
toxicmetabolites.
Undermoistconditions,thefungusemergesandproducesalayer
ofaerialconidiaonthesurfaceofhostcadavers.

10. ENVIRONMENT INFLUENCE ON
PATHOGENICITY
Ofalltheecofactorsthatinfluenceepizooticofamycopathogen,the
mostcriticalforsporulation,germinationandinvasionofthehostis
highhumidity(>90%RH).
Therapidityofmycelialdevelopmentandinfectiondependson
temperature.
Ingeneral,optimumvaluesfallbetween20°Cand30°C(forexample,
23°CforBeauveriabrongiartii,24°CforEntomophthoraobscura,25°C
forBeauveriabassianaandNomuraearileyi,20°C-25°CforVerticillium
lecaniiand27°C-28°CforMetarhiziumanisopliae).
ConidiaofM.anisopliaedonotgerminateat93%RH.

11. Microbial
control/Mycopesticides
Massmultiplicationofentomogenousfungiinvolvesinundative
augmentationmethod,whichisoneofthethreestrategiesofbiological
control.
Itisapplicationofthefungus,ofteninlargeamounts,forrapidshort-
termcontrolwithnoexpectationofsecondaryinfection.Inthisway,
thefungusisusedinasimilarwaytoachemicalinsecticide.
Theterms“mycopesticide”or“mycoinsecticide”havebeenusedto
describethisapproach.

12. Advantages over chemicals
Theygenerallydonotaffectnontargetorganismsso,extremelysafeto
use.
Eco-friendlyandnoresidualtoxicity.
Relativelyeasytomassproduce(deuteromycetesfungi).
Mostcommercialfungalproductsareformulatedasspores,whichare
easilyadaptedtoexistingapplicationtechnology,likespraying.
Havebroadhostrangesocontrolofmultiplepestswiththesame
productispossible.
Unlikeotherpotentialbiocontrolagents,fungidonothavetobe
ingestedtoinfecttheirhostsbutinvadedirectlythroughthecuticle.

13. Mass production steps
Isolation & Culture Culture (nutrient media)
blastospore/conidia
Mass production harvested at 72 hrs
centrifugation (conc.)
Formulation filtered
dried paste
Storage aeration/ventilation
adjuvant added
stored
Production of adequate quantities of a good quality inoculum is an
essential component of the biocontrol programme.

14. Requirements/ considerations
Forsuccessfulandcommercialproductoin:
1.Afungalisolatemustbeselectedwithrapidgrowth,abundant
sporulationandhighpathogenicitytothetargetpests.
2.Productioncostsmustbeminimal;mediumusedshouldbesimple,
cheap,easilyavailable,witheasyproductionprocedure.
3.Microbialproductsmustbeformulatedtocontroldifferenttarget
pests.
4.Formulatedproductsmustbesuitableforlong-termstorageunder
naturalconditionswithoutsignificantlylosingviabilityand
infectivity.
5.Shelf-lifeisconsideredanimportantfactorthatdeterminesthe
commercialsuccessofabiocontrolagentaswellasitsfieldefficacy.
6.An18-monthshelf-lifeisrecommendedfortheagriculturalmarket.

15. Isolation of pathogen
B.bassianaandP.fumosoroseusareisolatedfromthediseased
caterpillarofS.lituracollectedfromthefields.
Thediseasedlarvae(d.l)showedwhitecolourforB.bassianaand
slightreddishmycelialsurfacegrowthforP.fumosoroseus.
The(d.l)aresurfacesterilizedwith0.1%mercuricchlorideforfew
secondsthenthoroughlywashedwithsterilizeddoubledistilled
water.
Theexcesswaterisremovedbykeepingthe(d.l)onWhatmanfilter
paperNo.1.
Diseasedlarvaewerethencutintosmallpieceswiththehelpof
sterilebladeandthebitsareasepticallytransferredontothe
sabourandagarenrichedwith1%yeastextract(SMYA)slantswith
thehelpofsterileinoculationneedle.
Theslantsarekeptat25±1
o
C.

16. Culture on media
Types:
Whole grain media
Whole grains vizRice, Wheat, Raghi, Sorghum, Pearl millet
and Maize can be used for the sporulation of B. bassiana, P.
fumosoroseus and V. lecanii
Liquid media
Rice wash water, wheat wash water, coconut water and
rice cooked water are used for the growth and sporulation
of fungi.
Solid media
Non-synthetic solid media; carrot, ladies finger, jack seeds,
rice husk, and saw dust are used.

17. Suitable media
Variousagriculturalproductsandbyproductssuchasgrains,
vegetablewastes,seeds,ricehusk,sawdustandliquidmediasuch
ascoconutwater,riceandwheatwashedwaterandricecooked
waterhavebeenevaluatedformassproductionof3EPF:
Beauveriabassiana,(Bals.)Vuil.
Paecilomycesfumosoroseus(Wize)BrownandSmith
Verticilliumlecanii.(Zimm)Viegas.
Amongthegrains,wheatsupportedmaximumsporeproductionfor
B.bassianawhilesorghumrecordedmaximumsporeproductionin
P.fumosoroseusandV.lecanii.
Similarlycarrot,jackseedsandladiesfingeralsosupportedgood
growthandsporulationofallthethreetestedfungi.
Coconutwatersupportedmaximumgrowthandsporulation.

18. Production in Solid media
100 g/ml of media is taken (washed well and soaked in
water overnight for grain, for rice 2-3hr)
The excess water is drained by decanting and shade drying.
Autoclave at 15 psi for 1 h; inoculation incubated at, 28-
30
0
C for 15 days (under laminar air flow chamber)
Shaking of flasks to avoid clumping, after 7 days of
inoculation.
After incubation, 10 g/ml homogenized sample is taken for
sporulatingbottle
Transfer to 100 ml sterilized distilled water
The flasks are shaken in mechanical shaker for 10 min
The suspension is filtered through double layered muslin
cloth
Counting of spores are made after the serial dilution of the
suspension using double ruled haemocytometer

19. Production in Liquid Media
Threetypesofreactorsorfermentersarecommonlyused:
1.Thestirredtank
2.Thetowerand
3.Theloopfermenters.
1.Thestirredtank
Thestirredtankfermenterhastheverticalcylinderwiththeagitator
mechanismplacedcentrally.
Thesereactorsproduceaviolentagitationoftheculturemediumwithgood
homogenizationofthebrothandahighgastransfer,thusavoidmycelial
aggregationandsubsequentpelletformation.
Onedrawbackisdamagetothemyceliumoncontactwiththestirring
mechanism.
Stirredtankfermentershavebeenemployedfortheproductionoftheboth
myceliumandyeast-likecellsofallthecommonentomopathogenicfungi.

20. Cont…
2. The tower fermeter
The tower fermenter is a vertical cylinder with a
height/diameter ratio greater than six
Lack any mechanical agitation
Nutrient mixing is promoted by the injection of
gas at the base of the reactor
Most fungi produce mycelial aggregates in this
type of fermenter

21. Cont…
3.Theloopfermenter
Theloopfermenterisamodificationofthelatter
inwhichtheculturemediumisforcedback
downtothebottomofthereactor.
Therecyclingofthemediumisachievedbythe
incorporationofatube(internalrecirculatingor
byapipe(externalrecirculating)
Restingsporescanbeproducedinthistypeof
fermenter.

22. Fermenters for the production of entomopathogenic
fungi
Diagram. A -Stirred tank fermenter, B -Tower tank fermenter, C and D loop
fermenter with internal (C) or external (D) recirculating of medium, E -Tray
reactor, F -homogeneous solid reactor, G -rotating disk fermenter: a -motor,
b -air sparger, c -heating and cooling system, d -impeller, e -baffle, f -foam
breaker, g -rotating discs, h -granular media.

23. Types of fermentations
1. Solid substrate fermentation (SSF)
Uses solid substrate for mass multiplication.
Advantages:
1.Yieldsconidia(naturalsporulationbymycelia)
2.Conidiaobtainedisstable(notheatsensitive)
3.Moreinfectiousthanblastospores.
Disadvantages:
1.Yieldislow
2.Probleminharvesting

24. conti…
2. Submerged fermentation
It is designed to take advantage of existing industrial
fermenters which gives higher yield in liquid media.
Using a mineral medium (0.9% NaNO3, 0.25% KH2PO4,
0.075% MgSO4, 1.25% CaCl2) supplemented with 1%
sucrose, a yield of 3-5 X 10
8
conidia/ ml is obtained after
72-h fermentation in fermenters up to 63 m3 capacity, at
26-28°C, with proper aeration
Inclusion of peptone in the medium result in an average
yield of 2-3 X 10
9
spores ml/L.

25. Advantages:
1.Yieldishigh
2.Harvestingiseasy
3.Cheaper
Disadvantages:
Oftengivesrisetoblastosporesoramixtureof
blastosporesandconidia.
Blastosporesshowshorterviability(suceptibilityto
drying)
Lessinfectiousthanconidia

26. conti…
3.Diphasicfermentation
Itcombinestheadvantagesofbothsolidandliquid
media
Thefungusisallowedtogrowinfermentersuptothe
endofthelogphaseformaximalproductionof
mycelialbiomass
Subsequentlytransferredontoinertsubstratesin
traysfortheproductionofaerialconidiaintheform
ofnaturalinoculum

27. Advantage:
Itissimple
itistoincresegrowthofconidiaharvest
Disadvantages:
Consideredtobethemostexpensiveand
labour-intensive
Unsuitableforconventionalprocessingof
fungalmaterialinfermenters

28. Green muscardine fungus
ThecolonyofM.anisopliae
appearswhitewhenyoungbut
astheconidiamature,thecolour
turnstodarkgreen.
Achainofconidiaisformedon
eachconidiophore
Effectiveinthesuppressionof
soilbornepests.

Light micrographof
submerged spores of M.
anisopliae var. acridum

29. Productionprocedure
Thefunguscanbemassproducedin
conventionallaboratorymediaaswellason
crushedgrains.Thecheapestmediatilldate
knownare:
(1)Cassavachipsmixedwithricebran
supplementedwithureaorfishmealextract
(2)Coconutwaterwastedfromcopramaking
industry
(3)carrotbroth.

30. Host insets of Metarhizium anisopliae
Helicoverpa armigera
Dimond Black Moth
Brown plant hopper
Root grub
Locusta migratoria infected
with Metarhizium anisopliae

31. ThemortalityofHelicoverpaarmigeraapplied
withMetarhiziumanisopliae
1. Healthy larvae (8 days old larvae)
2. Dead larvae ( 5 days after spore application)
3. Dead larvae with thick fungal mat (7 days after spore application)
4. Dead larvae covered with green coloured conidia (10 days
after spore application)

32. Mass production
1.On coconut water(liquid media)
Coconutwater(40ml)containedin375mlliquorbottles
pluggedwithcottonplugaresterilizedinbatchesof9-10
bottlesin12litrepressurecookerfor20minutesat15
psi.
Thebottlesareinoculatedwith1mlsuspension
containingsporesofthefunguswiththehelpofasterile
injectionsyringe.
Thebottlesareinoculatedinalaminarflowchamberand
restedonflatsurfacefor2daysortillthesurfaceof
mediumisfullycoveredbytheolivegreensporulated
fungus.
Thewholecultureiscrushedthoroughlyinanordinary
mixerandusedinthefield.
Fromasingleaveragesizecoconut,5-6bottlesofcultures
canbemade.

33. 2)On rice(solid media)
Culture on
rice
Ricegrainscontinuetobethemostusedsubstratefor
conidialproduction.
Mostcompaniesusefrom300to500gofriceperplastic
bag.
Riceissoakedinwater,drainedandtransferredtoplastic
bagsfollowingsterilization
Conidialsuspensions(5mlsuspension/300gricebag)are
usedtoinoculate.
Bagsarehandmixedtodistributetheinoculumevenlyand
todisaggregateclustersofriceforadequatecolonization
andgoodsporulation.
Incubationtemperaturesrangefrom25-30
0
Cfor14
days.
Duringthedryseason,thisproceduretakes56daysfor
satisfactorydehydrationandconidiacanthenbe
harvested.

34. 3) Production by diphase system
InBrazilandRussia-
Thisinvolvessubmergedfermentationusingcornextractand
molasseswithcontinuousaeration
After40-50hrresultingmycellialmassandbiomassistransferredinto
storedpanesortrays.Thenincubatedfor6days.
Yield-5to7.5x10
12
/kgofdriedpreparation.
InAustralia–
Granularbitformulationby2phasemethod.
10,000to12000Lofliquidmedium(Agriculturalwaste,sucrose,Ca,
water)areinoculatedwithblastospore
Incubationfor5daysinafermenter
Yield4x10
12
conidia/kgbait

35. Ideal condition for mass production
Thesporecount,radialgrowth,sporulationandbiomasswere
maximumwhenM.anisopliaewereculturedbothCzapeck’sdoxagar
andpotatodextroseagarmedia
HighestsporulationofM.anisopliaeobservedwhenculturedinwheat
(8,300x10
4
).Hence,itisrecommendedasbestsolidmediaforthe
massproductionofM.anisopliae
Theoptimumtemperature-25-30°Cand
IdealpH7.0forthemassproduction
High-densitypolyethylene(HDPE)bagsareidealandalsocost
effectiveandgivegoodresultforthemassproductionofM.anisopliae

36. B. bassiana
Thefungusiscalledas
white muscardine
fungus.
Thefungussporesand
myceliaaremilkywhite
andfoundsproutingon
thebodyoflepidopterous
insectslikeHelicoverpa
armigera,Spodoptera
litura.
White muscardine fungus Conidia on conidiophores

37. Conti…
Helicoverpa armigeralarvae
infected with Beauveria bassia
B. bassianaon S. lituralarval cadavers

38. Culture and Mass production
Culture
B. bassianais easily cultured on solid and liquid media, e.g.
Saubouraudor potato dextrose agar and broth.
Streptomycin (500 mg/l) must be added to agar to obtain a
pure culture free from any bacteria.
Seed inoculumis better prepared as a submerged culture in
seed fermenteror in Erlenmeyer flasks on shaker.
Aerialconidiaareproducedonsolidmedia.
Massproduction
Atpresent,massproductionofB.bassianaisgenerallybased
ondiphasicandsubmergedfermentationtechniques.

39. Mass production
On carrot
Carrotcutintosmallpieces(40g)is
washedinpotablewaterandtransferred
toconicalflask(250ml)and15mlof
distilledwaterisadded.
Theconicalflasksarepluggedwith
cottonandautoclavedfor20minat15
psi.
Theflasksareallowedtocoolandtaken
tolaminarflowchamberforinoculation.
Fromacleanuncontaminatedmother
cultureinslantloopfulquantitiesofB.
bassianasporesaretransferred
aseptically.
Theflasksareincubatedatroom
temperature.Thesporesareobtainedin
afortnight
 Beauveriabassianacultured on solid substrates in polypropylene
bags
Beauveria bassianacultured on solid
substrates in polypropylene bags

40. On cereal
B. bassianais cultivated on different media:
wheat bran, wheat hybrid, oat semolina.
The mass of 30g of media with 45 ml water is poured
to glass bowls.
The treatment sterilized for 30 minutes at 120
o
C and
pressure 1 mPa/cm2.
The media, after cooling to approx. 35
o
C, is inoculated
by the spores from the agar (PDA) or by blastospores
and mycelium from a seed fermenter.
The culture is stored in a thermostat at 25
o
C +/-1
o
C
and 75 % R.H.

41. White halo fungus,Verticilliumlecanii
V.lecanicommonlycalledas
whitehalofungus.
Deadlarvaegenerallymummify
duetofungalinfection.The
cadaversshowanwhitemycelial
growthontheinsectsurface
(exceptontheheadcapsule)
Thefungusisknowntocause
epizootic when the
environmentalconditionsare
favourable.“
Aphid infection
Spirilling whitefly infection

42. Production procedure
Thefungusismultipliedoncheapmediaforlargescale
production.
OnSorghum:grainsdevoidofpesticideresidues(40g)iswashed
andtransferredtoconicalflask(250ml)and15mlofdistilled
waterisadded.
Theconicalflasksarepluggedwithcottonandautoclavedfor20
minat15psi.
Theflasksareallowedtocoolandtakentolaminarflowchamber
forinoculation.
Fromacleanuncontaminatedmothercultureinslantloopful
quantitiesofV.lecanisporesaretransferredaseptically.
Theflasksareincubatedatroomtemperature.Thesporesare
obtainedinafortnight.

43. Contd….
Paecilomyces farinosus
Thediseasedlarvaeshowslight
reddishmycelialsurface.
Itismassproducedonwheatbran,
sporulatedfungusisharvestedfrom
substrate,driedandappliedinsoil
granularform.
Nomuraearileyi
Forcontrolofcabbageandsoybean
looper,itisgrow onsolid
media(Agar).
Yield-6.3x10
8
conidia/cm
2
ofsurface.

44. Formulation and stability
Mycopesticidesaredefinedasproductsbasedonlivingfungal
propagulesintendedtocontrolpeststhroughinundativeor
inoculativeapplications.
Toenhancestabilityandimprovethehandlingoftheconidia,
2typesofformulationsareprepared:-
1)Theproductasapowderinheat-sealedplastic-linedfoilsachets,
theadditionofskimmedmilkpowderimproveslongevityby
absorbinghumidity,andfunctionsasastickerandsunscreenafter
application.
2)Productispreparedwithmineraloilthatcontains1.2%emulsifier.
[email protected]×10
8
conida/mloil.
Thistypeofformulationhasshownbetterstabilitybothatroom
temperatureandunderrefrigeration,whichisimportantforretail
sale.

45. Formulation types
Wettablepowder(WP):-‘‘Apowderformulationtobe
appliedasasuspensionafterdispersioninwater.’’
Granule(GR).‘‘Afree-flowingsolidformulationwith
particlesofcontrolledanduniformsizerangereadyfor
useandwiththeactiveingredientstronglyadheredtoor
incorporatedintothegranule.
Waterdispersiblegranule(WG).‘‘Aformulation
consistingofgranulestobeappliedafterdisintegration
anddispersioninwater.’’
Contactpowder(CP).‘‘Insecticidalformulationinpowder
formfordirectapplication.’’
Free-flowingpowderssuitablefordustingaretermed
dustablepowders(DP).

46. Cond…
Suspensionconcentrate(SC).‘‘Astablesuspensionofactive
ingredient(s)inwater,intendedfordilutionwithwaterbeforeuse.’’
oilmisciblesuspension.‘‘Astablesuspensionofactive
ingredient(s)inafluidintendedfordilutioninanorganicliquid
beforeuse.’’
Ultra-lowvolume(ULV)suspension(SU).‘‘Asuspensionready
forusethroughULVequipment.’
’
Oildispersion(OD).‘‘Astablesuspensionofactiveingredientina
water-immisciblefluid,whichmaycontainotherdissolvedactive
ingredient(s),intendedfordilutioninwaterbeforeuse.’’

47. Common formulation types
Technicalconcentratesintheformoffungus-colonized
substrates(26.3%),wettablepowders(20.5%),andoil
dispersions(15.2%).
Theremainingtypesincludegranules(2.9%),technical
materials(2.9%),baits(1.8%),waterdispersiblegranules
(1.8%),oilmiscibleflowableconcentrates(1.2%),ULV
suspensions(0.6%),suspensionconcentrates(0.6%),and
contactpowders(0.6%).
Oftheover750speciesoffungiknowntobepathogenicto
insects,sixhavebeencommercializedandthecosmopolitan
pathogens,suchasB.bassiana,M.anisopliaearethebest
knownsofar.

48. Doses and application
Asaninsecticide,thesporesaresprayedonaffected
cropsasanemulsifiedsuspensionorwettablepowder
TheMetarhizium-basedmycoinsecticideconidiaare
appliedbyaerialsprayingwhichshouldbemixedwith
dieseloilorkerosenebeforesprayingatarateof2.5–
5×10
12
conidia/hatocontrolthespittlebugs.
Boverinifappliedalone3to20kg/ha(uneconomic).So2
kgboverin+¼insecticides(400g)/ha.
SoilapplicationonB.brongniartiiblastospore-
2x10
13
/ha
Aerialsprayofmetaquino(M.anisoplae)@6x10
11
to
1.2x10
12
conidia/hacauses65%mortality

49. Commercial Products
OfB.bassiana
Sincethelate1980s,focushasbeenonthedevelopmentofB.
bassianaasacommercialmycoinsecticidebyMycotech.
In2000,MycotechwastakenoverbyEmeraldBioAgriculture,
whocontinuetomarketthesefungal-basedformulations.
Boverin,wasdevelopedintheUSSRduringthe1970s.Ithas
beenusedextensivelytocontroltheColoradopotatobeetle,the
larvaeofcodlingmoth
“Mycotrol”wasfullyregisteredin1999bytheEPAoftheUSA.Its
WPformulationcausehighmortalityofaphids,whiteflies,thrips
ontomatoandornamentalcrops.
“Metaquino”hasbeeninuseinBrazil.Itwasalsousedagainst
coffeeberryborerinBrazilandcoconutleafbeetleinTaiwan.

50. Others
MetarhiziumanisopliaeisregisteredintheU.S.forcontrolof
cockroachesas“GreenMuscle”.
Verticilliumlecaniisoldas‘’Vartalec’’.Itisproducedbyliquid
fermentationasblastosporeswhichareformulatedwitha
nutrientsourceinawettablepowder
HirsutellasoldasMycar.
RecentadvancesbyKoppertinformulationtechnologyhave
resultedinanadjuvantbasedonanemulsifiablevegetableoil,
called“Addit”,whichcanenhanceactivityof“Mycotrol”atlow
humidities
PaecilomycesfumosoroseusStrain97hasbeenapprovedforuse
onornamentalstomanagewhiteflies,aphids,thrips,andspider
mitesandissoldbyThermoTrilogyCorp.

51. Commercial products in India
B.bassiana–Biopower,Biogard
Metarhiziumanisopliae-Biomagic,Biomet
Verticilliumlecanii-Biovert,Biocatch
Hirsutella-Metehit,Mycohit
Paecilomycesfumosoroseus-PaciHit

52. Limitations and disadvantages
Themajorissuesinvolvedinmassproductionandutilizationof
mycopathogensareselectionofeffectivestrains,developmentofcost
effectivemethodsformassrearing,developmentofeffectivemethodsfor
storageandshipmentandcreationofeffectiveformulation.
Highconcentrationsofsporesareoftenneededtogetadequatecontrol.
Thekilltimeisrelativelylong(~1weekformostfungi).
Theirbroadhostrangecansometimesbeaproblem,forbeneficial
insects.
Environmentalfactorsliketemperature,humidityandsunlightplaya
profoundroleonthefieldpersistenceofentomopathogenicfungi.
Prolongedexposuretosunlightcanalsoinactivatespores,reducing
persistenceinthecrop.

53. Major technical hurdles
They include:
Improvements in screening and quality control to ensure
that only viable, virulent inoculum reaches the market
Development of media to increase the virulence and
ecological fitness of conidia
Improved formulations to increase both the virulence and
shelf life of the inoculum
Better application strategies to ensure that sufficient
inoculum contacts the pest because mortality is dose-
related above certain thresholds

54. Future prospects
Currently,ayieldofsubmergedconidiainmuchlower
quantitiesthanblastosporesremainsanobstacletotheuse
ofthesubmergedculturemethodforcommercialproduction
ofconidia
Attemptstousethesefungiasbioinsecticideshaveusually
failedastheyaretoodifficulttomassproduceandareoften
ineffectiveunderconditionsofmoderatehumidityinthe
field
Thus,geneticengineeringmayhelpinimprovingefficacyof
mycoinsecticidesbut,allhazardsmustbeassessed,stepby
stepconcerningallpotentialdangerstonon-target
organismsbeforereleaseintotheenvironment.

55. Conclusion
Formassproductionfungalisolatemustbeselectedwith
rapidgrowth,abundantsporulationandsufficientlyhigh
pathogenicitytothetargetpests.
productioncostsmustbeminimal;mediumshouldbe
simpleincomposition,cheapinpriceandavailableinlarge
quantitieswitheasyproduction.
Formulatedproductsmustbesuitableforlong-term
storageundernaturalornearlynaturalconditionswithout
significantlylosingtheirviabilityandinfectivity.
Shelf-lifeisconsideredapivotalfactorthatdeterminesthe
commercialsuccessofabiocontrolagentaswellasitsfield
efficacy.

56. References
Soundarapandian,P.andR.Chandra,2007.Massproductionof
endomopathogenic fungusMetarhiziumanisopliae
(Deuteromycota;Hyphomycetes)inthelaboratory.Res.J.
Microbiol.,2:690-695.
Cienc.Rural[online].1999,vol.29,n.3,pp.389-394.ISSN0103-
8478.
K.SahayarajandS.KarthickRajaNamasivayam,2008.Mass
productionofentomopathogenicfungiusingagricultural
productsandbYproductsAfricanJournalofBiotechnologyVol.7
(12),pp.1907-10
M.G.FengabandT.J.Poprawskicd,1994.Beauveriabassianafor
insectcontrol:currentstatusBiocontrolScienceandTechnology
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P.A.Shah·J.K.Pell.2003.Entomopathogenicfungiasbiological
controlagentsApplMicrobiolBiotechnol61:413–423