Interactions between Phoma koolunga, Didymella pinodes and Phoma medicaginis var. pinodella, causal agents of ascochyta blight on field pea in

South Australia

Jenny Davidson (SARDI, University of Adelaide)&

Marzena Krysinska-Kaczmarek (SARDI)Herdina, Alan McKay(RDTS-SARDI)

&Eileen Scott (University of Adelaide)

FUNDED BY: South Australian Grains Industry Trust (SAGIT)

Field peas and ascochyta blight2009: 285,000 ha grown in Australia (47% crop in South Australia)

Value approx $100 million farm gate

Ascochyta blight (synonym blackspot)- major disease world wide

Affects all above ground parts and crown below the ground level

Annual losses ~ 15% of Australian pea crop

Can cause total failure of individual crop

▪are independent ▪exist in the same field ▪on the same plant

4 causal agents of ascochyta blight complex on field pea

Ascochyta pisiSplash borne conidiaNo known teleomorph(minor role in Australia)

Phoma medicaginis var. pinodellasplash borne conidia

Teleomorph only found once in laboratory conditions(Bowen et al. 1997, Mycological

Research 10: 80-84).

Ascochyta pinodesConsidered the major pathogenSplash borne conidia

Teleomorph: Didymella pinodesAirborne ascospores

Recently identifiedPhoma koolungaDavidson et al. 2009, Mycologia 101:120-129.Widespread in South Australia, limited distribution in Victoria and Western Australia. Similar symptoms to D. pinodes.

• P. koolunga newly recognised as an important component of the ascochyta complex on field pea in South Australia

• Appears to fit the same niche on field pea as the other fungal species

AIM

• Investigate interactions between Phoma koolunga; D. pinodes and P. medicaginis var. pinodella

What is the role of recently identified P. koolunga in ascochyta blight complex on field pea?

Competition on Potato Dextrose AgarZones of inhibition between paired pathogens including self paired colonies of P. koolunga.

a. D. pinodes c. P.medicaginis var. pinodella e. P. koolunga

b. P. koolunga d. P. koolunga f. P. koolunga

Interactions on amended Potato Dextrose AgarBroth filtrates produced for each pathogen

D. pinodes P. koolunga2 isolates

P. medicaginis var. pinodella

Sterile brothcontrol

25ml of each broth filtrate added separately to PDA

Each pathogen plated onto every type of amended agar

Culture growth measured at day 3 and day 7

Filtrate from P. koolunga inhibits growth of D. pinodes and P. medicaginis var. pinodella.

Filtrate sourceD. pinodes P. medicaginis

var. pinodellaP. koolunga a P. koolunga b

sterile broth-control 15.0 22.3 8.0 7.3D. pinodes 14.5 20.3 9.0 6.8P. medicaginis var. pinodella 15.0 23.3 8.8 7.3P. koolunga a (isolate DAR78535) 10.3 17.5 8.3 7.0P. koolunga b (isolate FT07006) 12.0 19.0 7.8 6.3LSDinteraction P(0.05) = 2.2

Effect of P. koolunga filtrate on colony growth of D. pinodes , P. medicaginis var. pinodella and P. koolunga

Diameter of colonies after 3 days (mm)

After removing cultures from filtrate normal growth is resumed = fungistasis.

Filtrate sourceD. pinodes P. medicaginis

var. pinodellaP. koolunga a P. koolunga b

sterile broth-control 44.0 52.8 24.8 23.3D. pinodes 42.5 51.5 20.5 21.8P. medicaginis var. pinodella 43.3 52.3 23.8 22.3P. koolunga a (isolate DAR78535) 43.3 50.0 20.5 22.8P. koolunga b (isolate FT07006) 43.3 52.3 22.8 23.5

mean 43.3 51.8 22.5 22.7LSDinteractionP(0.05) = not significant

Effect of removing P. koolunga filtrate from colonies of D. pinodes , P. medicaginis var. pinodella and P. koolunga

Diameter of colonies after 7 days (mm)

Interaction of pathogens in planta

Inoculated in pairs onto same leaf in controlled conditions

Lesion size by D. pinodesgreater than lesions by the two Phoma species.

No interaction in lesion size

Interaction of pathogens in planta - DNA quantification per leaf

0

2

4

6

8st

erile

aga

r**

D.p

inod

es

P.m

edic

agin

isva

r. p

inod

ella

P. k

oolu

nga

DAR

7853

5

P. k

oolu

nga

FT07

006

ster

ile a

gar*

*

D.p

inod

es

P.m

edic

agin

isva

r. p

inod

ella

P. k

oolu

nga

DAR

7853

5

P. k

oolu

nga

FT07

006

D. pinodes* P.medicaginis var. pinodella*

Log

(fg

DN

A D

pPm

p) p

er le

af a

t 7 d

p.i.

Quantity of DNA of D. pinodes and P. medicaginis var. pinodella is reduced in presence of P. koolunga

Interaction of pathogens in planta -DNA quantification per leaf

-Quantity of DNA of P. koolunga was not reduced by co-inoculation

0

2

4

6

8ste

rile

agar

**

D.p

inod

es

P.m

edic

agin

isva

r. pi

node

llaP.

koo

lung

aD

AR78

535

P. k

oolu

nga

FT07

006

steril

e ag

ar**

D.p

inod

es

P.m

edic

agin

isva

r. pi

node

llaP.

koo

lung

aD

AR78

535

P. k

oolu

nga

FT07

006

P. koolunga DAR78535* P. koolunga FT07006*Log

(fg D

NA

P. k

oolu

nga)

per

leaf

at 7

d p

.i.

Interaction of pathogens on leaf discs -Detached leaf discs floating on sterile RO water

-Measured lesion size and quantified DNA

• no interactions for DNA

• Interactions for lesion size

*denotes isolate which caused lesion measured** denotes co-inoculated isolate or sterile water for controls.

Interaction of pathogens on leaf discs – lesion size

0

5

10

15

D. p

inod

es**

P.m

edic

agin

isva

r. pi

node

llaP.

koo

lung

aD

AR78

535

P. k

oolu

nga

FT07

006

RO w

ater

D. p

inod

esP.

med

icag

inis

var.

pino

della

P. k

oolu

nga

DAR

7853

5P.

koo

lung

aFT

0700

6RO

wat

er

D. p

inod

esP.

med

icag

inis

var.

pino

della

P. k

oolu

nga

DAR

7853

5P.

koo

lung

aFT

0700

6RO

wat

er

D. p

inod

esP.

med

icag

inis

var.

pino

della

P. k

oolu

nga

DAR

7853

5P.

koo

lung

aFT

0700

6RO

wat

er

D. pinodes* P.medicaginis var.pinodella*

P. koolungaDAR78535*

P. koolunga FT07006*

Lesi

on d

iam

eter

(mm

)

7 d p.i. on leaf discs on sterile waterLesion diameter of D. pinodes and P. medicaginis var. pinodellareduced compared to controls in presence of P. koolunga isolate DAR78535 or self

Lesions size of P. koolunga isolate DAR78535 was increased in presence of P. medicaginis var. pinodella or if self paired.

Summary

• This study demonstrated interactions between the three ascochyta blight fungal species resulting in antagonism, including self antagonism.

• P. koolunga was not self antagonistic nor affected by co-inoculation with the other species.

• P. koolunga is slower growing but had fungistatic effects on the other causal pathogens.

• Anomalies possibly due to environmental effects of leaf discs.

• Further studies are warranted into the potential of fungistatic metabolites produced by P. koolunga.