Beauveria bassiana as an endophyte in grapevine Vitis vinifera · PDF fileBeauveria bassiana...

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Potential of the entomopathogenic fungus

Beauveria bassiana as an endophyte in

grapevine Vitis vinifera

Prof. Dr. M.-H. Poehling

Prof. Dr. A. Reineke

Yvonne Rondot and Annette Reineke

Hochschule GEISENHEIM University

Institute of Phytomedicine

IOBC-WRPS meeting "Integrated Protection

and Production in Viticulture"

Ascona,16th Oct. 2013

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• entomopathogenic fungus

• worldwide distribution, naturally occurring

in soil

• wide host range

• is able to colonize plants endophytically

• commercial products:

• Naturalis® (contains strain ATCC 74040)

• BotaniGard® 22WP (contains strain GHA)

• strains are registered on Annex 2,

Regulation (EC) No. 1107/2009

Beauveria bassiana

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Tritrophic interaction

pathogens e.g. Planococcus ficus, Otiorhynchus sulcatus,

Plasmopara viticola

plant Vitis vinifera

endophyte Beauveria bassiana

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Influence on insect pests

Endophytic establishment of B. bassiana

Potential against plant pathogens

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Influence on insect pests

• Is any effect of endophytic B. bassiana on mortality and growth of pest insects evident?

• Are the insects influenced by endophytic B. bassiana in their host choice behavior?

Endophytic establishment of B. bassiana

• Is it possible to establish B. bassiana as an endophyte within Vitis vinifera plants?

• Is any effect of endophytic B. bassiana on plant growth of the plants evident?

Influence on plant pathogens

• Is any effect of endophytic B. bassiana on the disease severity of Plasmopara viticola evident?

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Inoculation

Treatments:

• different strains (ATCC 74040 and GHA)

• different conidia concentrations

• conidia suspension vs. product Naturalis®

• inoculation conditions: wrapping the plants in plastic bags for 24 h after inoculation

all treatments were inoculated with handheld-sprayer on the upper and lower leaf surface

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Re-isolation of B. bassiana

surface sterilization

placing on BSM

visual assessment

incubation (24 C, dark)

leaf discs

2 min in 0.5 % NaOCl, 2 min in 70 % ETOH,

rinsed with sterile dest. water

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2 5 0 0 0

5 0 0 0 0

7 5 0 0 0

1 0 0 0 0 0

1 2 5 0 0 0

1 5 0 0 0 0

1 7 5 0 0 0

7 5 1 0 0 1 2 5 1 5 0 1 7 5 2 0 0 2 2 5 2 5 0

B a 0 1 _ B a _ 1 2 _ B a 1 3 _ A 6 .F 0 1 _ 1 2 0 8 2 3 1 1 8 D

S ize (nt)

Dye S

ignal

molecular detection

optaining 6 leaf discs with a cork borer

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Endophytic establishment of B. bassiana

• Is it possible to establish B. bassiana as an endophyte within Vitis vinifera plants?

• Is any effect of endophytic B. bassiana on plant growth of the plants evident?

• endophytic colonization of grapevine leaves by B. bassiana is

possible

• presence of B. bassiana in grapevine plants was confirmed up to

28 days after inoculation

• different treatments had no significant effect on the colonization

rates

• endophytic B. bassiana had no negative effect on growth of

potted grapevine plants

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Influence on insect pests

• Is any effect of endophytic B. bassiana on mortality and growth of pest insects evident?

• Are the insects influenced by endophytic B. bassiana in their host choice behavior?

Endophytic establishment of B. bassiana

• Is it possible to establish B. bassiana as an endophyte within Vitis vinifera plants?

• Is any effect of endophytic B. bassiana on plant growth of the plants evident?

Influence on plant pathogens

• Is any effect of endophytic B. bassiana on the disease severity of Plasmopara viticola evident?

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Vine mealybug Planococcus ficus

Symptoms/Damage

• phloem feeding causes vine weakening,

defoliation

• honeydew excretion covers berries and

leaves, growth of sooty mold

• transferring Grapevine leafroll-associated

virus-3 (GLRaV-3)!

Occurrence

• in viticulture mainly in South Europe and

California

Fig.: Scagliusi et al. 2002 Fig.: Bovey et al. 1980

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Detached leaf assay with P. ficus

• spray inoculation of plants with B. bassiana

(Naturalis® 3%)

• assessment of numbers of alive mealybugs and size

every week

W 0

W 1

W 2

W 3

W 4

W 5

infestation with mealybugs

spray inoculation

1st assessment

surface sterilization

2nd assessment

3rd assessment

weeks

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Influence on insect pests

• Is any effect of endophytic B. bassiana on mortality and growth of pest insects evident?

• Are the insects influenced by endophytic B. bassiana in their host choice behavior?

Endophytic establishment of B. bassiana

• Is it possible to establish B. bassiana as an endophyte within Vitis vinifera plants?

• Is any effect of endophytic B. bassiana on plant growth of the plants evident?

Influence on plant pathogens

• Is any effect of endophytic B. bassiana on the disease severity of Plasmopara viticola evident?

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Black vine weevil Otiorhynchus sulcatus

Symptoms and damage:

• maturation feeding of adults on leaves and

flowers

characteristic crescent shaped notches

• root feeding of soil-borne larvae feed

results in wilting/stunting or could be lethal

for host plants

Host plants:

• major pest in many horticultural crops

• partly in forestry

• in viticulture mainly in grapevine nurseries

photo: HS Geisenheim

Rene Eisenbart

http://blog.oregonlive.com

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Choice assay with Otiorhynchus sulcatus

• spray inoculation of plants with

B. bassiana (Naturalis® 3%)

formulation of Naturalis® without conidia

water as control

• one adult O. sulcatus per tube

• each test lasted 1 h

• pre-test for validation of the olfactometer test system

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Choice assay – validation of the test system

control ↔ empty

p ≤ 0.05 exact binominal test

0

2

4

6

8

10

12

14

16

18

20

control no decision empty

No

. of

de

cisi

on

s

N = 35

the weevils are able to select a tube with a plant over an empty tube

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• a significant effect of endophytic B. bassiana on growth and on

mortality one week after the initial settlement of vine

mealybugs was evident

• adult O. sulcatus chose significantly more often the control

plants as a host plant compared to grapevine plants with

endophytic B. bassiana

Influence on insect pests

• Is any effect of endophytic B. bassiana on mortality and growth of pest insects evident?

• Are the insects influenced by endophytic B. bassiana in their host choice behavior?

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Influence on insect pests

• Is any effect of endophytic B. bassiana on mortality and growth of pest insects evident?

• Are the insects influenced by endophytic B. bassiana in their host choice behavior?

Endophytic establishment of B. bassiana

• Is it possible to establish B. bassiana as an endophyte within Vitis vinifera plants?

• Is any effect of endophytic B. bassiana on plant growth of the plants evident?

Influence on plant pathogens

• Is a protective effect of endophytic B. bassiana on the disease severity of Plasmopara viticola evident?

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Inoculation tests with P. viticola

treatment with B. bassiana

assessment of disease severity

inoculation with P. viticola

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2

3

4

5

6

7

8

9

10

leaf level

day 0

day 7

day 14

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• a significant protective effect of endophytic B. bassiana on

disease severity of Plasmopara viticola is evident

• a systemic protection of the newley formed leaves could not

be observed

Influence on plant pathogens

• Is a protective effect of endophytic B. bassiana on the disease severity of Plasmopara viticola evident?

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Tritrophic interaction

pathogens e.g. Planococcus ficus, Otiorhynchus sulcatus,

Plasmopara viticola

plant Vitis vinifera

endophyte Beauveria bassiana

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Thanks for your attention!

Thanks to:

Deutsche Bundesstiftung Umwelt (DBU)

Prof. Dr. A. Reineke

Prof. Dr. H.-M. Poehling

Roman Peiter (M. Sc. Thesis)

Gordan Behaderovic (B. Sc. Thesis)