Diabrotica with EPN - University of Sussex with EPN Ralf-Udo Ehlers [email protected]...

Christian-Albrechts-University Kiel

Institute for PhytopathologyDept. Biotechnology & Biol. Control

Biological Control of Diabrotica with EPN

Ralf-Udo [email protected]

www.e-nema.de

Diabrotica in Europe

Salzau, March 27, 2009

Entomopathogenic Nematodes and their

Symbionts

Nematodes Bacteria

Rhabditida Enterobacteriaceae

Closely related to Caenorhabditis elegans

Steiner, G. - in 1923 described Aplectana kraussei from a massive outbreak of the spruce sawfly, Cephaleia abietis in Westfalia, Germany. Krausse, a forest entomologist from Eberswalde, had sent him an infected larva. The description was without many details as he had available only 15 female and 5 male nematodes.

Historical Review into EP Nematology

Entomopathogenic nematodes and their symbiotic bacteria

• Worms of 0.5 - 0.9 mm length• Present in all soil environments • Dauer juveniles (DJ)free-living in

the soil, reproduction only in host insects

• DJ well adapted for long-term survival in the soil (fat reserves, non-feeding, ambushing)

• Symbiotic bacteria in the intestine, symbiosis during nematode growth and development

Life cycle

Dauer juveniles enter the host

The symbiotic bacteriaare released

The insect dies,nematodes developeto adults

Offspring exit from insect cadaver and searching for new hosts

Xenorhabdus and Photorhabdus spp.• Nematode transmits bacterium into sterile environment• Bacteria contribute to kill host insect • Bacteria provide essential nutrients for nematodes• Photorhabdus spp. bioluminescent (not flourescent) • A mutualistic relation: symbiosis

Xenorhabdus is not producing light

Burnell, Maynooth, Ireland

Transport of the Bacterial Symbionts

S. feltiae

2100 600 0

Number of bacterial cells

H. megidis

How are EPN are attracted by insects?

• EPN are attracted by CO2 and vibrations• Penetration behaviour is triggered by substances on the insect cuticle• EPN react to plant SOS signals (Rasman et al. 2005, Nature)

?

ß - Caryophyllene

Diabrotica larvae

Reproduction of EPN:Without the presence of the symbiotic

bacteria they cannot reproduce



Use of EPN in biocontrolIt´s mainly a matter of application costs

Nema-green (5x109/ha)

Home Gardening Internet – > 2000 €/ha

Golf Market – 800 €/ha

Nemycel (2x106/m² =20x109/ha)

Mushroom Farm + Distrib. – 500-1000 €/ha

Nemapom (1.5x109/ha)

Distributor – 150 €/ha

Control Diabrotica virgifera v. (109/ha)

Product cost must be < 100 €/ha

Nema-green: grub control

Nemycel: sciarid control

Nemapom: cydia control

Instar susceptibility

• All instars and pupae areeffectivelykilled,

• Thus timing of sprays is lessimportant

Kurtz Hiltpold 2008 BioControl

L1 L2 L3 PUPAE

Screening in the lab

Toepfer et al. (2005). Bull Entomological Res 95

Root volatiles from maize hybrids ?

Interaction

Pest

Host Biocontrol Agent

Nematode orientation in the field

Pactol (-)

Magister (Caryophyllene +)

Rasmann et al., 2005, Nature; Hiltpold 2008 phd thesis.

• Caryophyllene is mainly important for the nematode H. megidis and less for H. bacteriophora,

Maize variety +/- caryophyllene

37

63

49

0

20

40

60

80

100

St. feltiae H. bacteriophora H. megidis

Nematode species, applied in April with sowing

Red

uctio

n D

iabr

otic

a v.

virg

ifera

(% A

bbot

t cor

rect

ed)

Application atsowing (April)

25

59 57

0

20

40

60

80

100

St. feltiae H. bacteriophora H. megidis

Nematode speciey applied as row sprays in June

Redu

ctio

n of

Dia

brot

ica

v. v

irgife

ra(%

Abb

ott c

orre

cted

)

Application in June to L3H. bacteriophora again working best



Application methods + time

Diabrotica virgifera v.

5,7 5,9

21,5

8,65,6

22,9

0

5

10

15

20

25

30

35

Fluid rowspray with

sowing

Powder rowspray with

sowing

Flat sprayon soil after

sowing

Fluid rowspray in

June

Fluid flatspray in

June

Control

Applications of H. bacteriophora

% R

oots

with

eco

nom

ic d

amag

e(>

3 in

1 to

6 Io

wa

Sca

le)

75%62%6%74%75%Reduction

IOWA Scale Reduction

0.3

1.0

0.4 0.2 0.2

1.7

0

1

2

3

Fluid rowspray with

sowing

Powder rowspray with

sowing

Flat spray onsoil aftersowing

Fluid rowspray in June

Fluid flatspray in June

Control

Applications of H. bacteriophora

Dia

brot

ica

adul

ts e

mer

ged

/10

0 eg

gs /

plan

t



Application method and time

105/m² Heterorhabditis bacteriophoraStefan Toepfer

88%88%57%41%82%Reduction

Reduction Adult Emergence

Summary: Testing application techniques

Insecticide

Step 5 Lowest effective dose

y = 14.4Ln(x) + 72.1r2 = 0.4

0

1020

3040

50

6070

8090

100

0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4Million H. bacteriophora per metre

Mea

n ne

mat

ode

effic

acy

(% R

educ

tion

of ro

ot d

amag

e m

easu

red

by 0

.00

- 3.0

0 no

de in

jury

sca

le

com

pare

d to

con

trol)

Step 5 Lowest effective doseNeeded for product price estimation

• The lowest dose that canreach similar efficacies as insectides is 0.1 Million 0.1 Million H.bactH.bact / m/ m

• (=1.3 x 10 9 / ha).



Diabrotica v. virgifera

Szatymaz

00,5

11,5

22,5

33,5

44,5

5

Control Force H.b.0.03

H.b.0.1

H.b.0.2

Poncho

Treatment

Adul

t D. v

. virg

ifera

em

erge

d / p

lant

/ 10

0 eg

gs

Toepfer, Kuhlmann, Peters, Ehlers, 2007



Diabrotica v. virgifera

Toepfer, Grabenweger, Peters, Ehlers, 2007

1,06

0,500,13

3,25

1,75

7,31

0,00

1,00

2,00

3,00

4,00

5,00

6,00

7,00

8,00

9,00

10,00

11,00

12,00

nicht infiziert UK Poncho EPNspät EPNfrüh Granulat



Diabrotica Control

Recommendation: Apply granularformulation of H. bacteriophora withsowing in April/May at 1.3 x 109 ha-1

Application costs at 150 €/ha

Further reduction of costs possible:

- Increasing production capacity

- Lower application density

- Improved application technology

- Prolonged shelf life

Thanks to Stefan Töpfer and his CABI Team in Hungary

Diabrotica with EPN - University of Sussex with EPN Ralf-Udo Ehlers [email protected]...

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