at. tsurushima, bh. nakayashiki, by. tosa and bs. …...an oxidized derivative of linoleic acid...

An Oxidized Derivative of Linoleic Acid (Mag-Toxin)Produced in Pyricularia oryzae

AT. Tsurushima, BH. Nakayashiki, BY. Tosa and BS. Mayama

AHannan University, Faculty of Business, Osaka, 580-8502, Japan.BKobe University, Faculty of Agriculture, Hyogo, 657-8501, Japan.

Active factors might relate directly or indirectly in pathogenicity of blast fungi .

Example (1)factors related with disease symptom(2)determinant of specific

pathogenicity

1. Pyrichalasin H production and pathogenicity of Digitaria-specific isolates

Mol. Plant Pathol. 6 (2005)

Mol. Plant pathol. 6 (2005)

Numbers of Production of Pathogenisity isoloates Host plant Pyrichalasin H on Digitaria

rice, Itallian ryegrass, foxtail millet, common millet, green bristle grass, finger millet, wheat, Lehman

32 livegrass, Jungle rice, rice cutgrass, buffelgrass, 0 0bamboo grass, etc

32 Digitaria sanguinalis (crabgrass) 31 311 Digitaria sumutsii (pangolagrass) 1 17 Digitaria holizontalis 6 6

Total 72 38 38

Summary (pyrichalasin H production and pathogenicity on Digitaria of Pyricularia isolates)

Result 1

1. There is a correlation between pyrichalasin H production and pathogenicitｙon Digitaria.

2. Pyrichalasin H was detected in spore germination fluid of a Digitaria isolate.3. Pretreatment of leaf sheaths of crabgrass with 3 g/ml pyrichalasin H led to the

penetration and colonization by nonhost isolate.

↓Production of pyrichalasin H might be responsible for the genus-specific pathogenicity of Digitaria isolates.

Kato et al (2001) Couch and Kohn (2002)pathogenicitymating the sequences of DNA restriction fragment three genes

length polymorphisms

P. oryzae is a new fungal species distinct from P. grisea.

This phylogenic difference between Digitaria isolates and the crop isolates could be related to the production of pyrichalasin H.

Kato et al., J. Gen. Plant Pathol. 66. 30 (2001)Couch, B. C. and Kohn, L. M., Mycologia 94. 683 (2002)

2. Necrosis-inducing Factor and Chlorosis-inducing Factor

CH3

CHO

OH

CH3

OHCHO

OH

OHOH

OH

CH3

CHO

OH

OH

OH

CH3

CHO

OH

OH

CH3

OH

O

OH

OHO

O

OH

Pyriculol Epipyriculol

Pyriculariol Pyricuol

3-(1',3'-pentadienyl)-3,4-dihydro-1H-2-benzopyran-4,8-diol

4-(1'-hydroxy-2'-butenyl9-1,4-dihydro-2,3-benzodioxocin-10-ol

Necrosis-inducing factors from P. grisea

NH

HO CH3

O

OH3C H

CH3

Tenuazonic acid

Pyrichalasin H

H 3CO

OH

NH

CH

H 3C OH

H

O OAc

H 3C

の

This is also produced inthe genera Cochliobolus and Alternaria

These were only reported by Pyricularia grisea

ーーーーーーーーーーーーーーーーーーーーーーーーー

ーー_

の4-(1’-hydroxy-2’-butenyl)-1,4dihydro-2,3-benzodioxocin-10-ol

d ip p ed in aceto n e fo r 24 h

fi lter ed th ro u gh fil te r p ap er(TO Y O, No . 2 )evapo ra ted

E tO A c-fr W a te r-fr

co ncen tr ated in vacu o

d isso lved in CH C l3

C HC l3 1% M eO H

3% M eO H

5% M eO H

10% M eO H

100% M eOH

21 3 4 5 6 7 8 9 10 11 12

A v e na iso la te

cu ltu red in o at m eal m edia for 10 d ays

ir rad ia ted w ith BL B l ig h t for 3 d ays

C ultu reS po re

Ger m in ation flu id

extracted with E tOA c ( tw ice )

N ecrosisC h lorosis

Advances in Genetics. Genomics and Control of Rice Blast Disease (2009)

Fig. 6. HPLC profile of Fr.6

Time (min)0 5 10 15 20

Ref

rect

ive

inde

x

Shim-pack PREP-ODS(H)Kit (4.6 mmI.D.X250 mmL), 60% MeOH , 0.8 ml/min. Detector:RI

Yellow peak fractions were toxic to oat leaves.

(1)

(2)

CH3

CHO

OH

OH

OH

CH3

OHCHO

OH

OH

(1) Pyriculol (2) Epipyriculol

Result 1

All isolates of Pyricularia (rice, weat, oat, finger millet, foxtail millet and common millet and crabgrass) produced pyriculol and epipyriculo as mainly necrosis-inducing principles

HPLC profile of Fr.7

Solvent: 65% acetonitrile; Flow rate: 1.4ml/minColumn: Shim-pack PREP-ODS(H)Kit

(4.6mm I.D. x 250mm)

Chlorosis induction (m/z 293)

Biosci. Biotechnol. Biochem. 74 (2010)

m/z 295

m/z 295

Chemical structures of linoleic acid (1) , 8-oxo-octadeca-9E,12Z-dienoic acid (2)and 14-oxo-octadeca-9E,12Z-dienoic acid (3)


(Reference)Mycelium of Pyricularia grisea biosynthesized 8-hydroxylinoleic acid from linoleic acid (Crista, Osboun and Oliw, 2003)

ROS generation in organelle of the cell of leaf segment treated with Mag-toxin I


Effect of light on ROS generation and induction of cell death in the leaf segment treated with Mag-toxin Biosci. Biotechnol. Biochem. 74 (2010)

Result２1. Chlorosis-inducing factor was an oxidized C18 unsaturated

fatty acid named Mag-toxin.2. Chlorosis was light-dependently induced on segments by

Mag-toxin.3. ROS generation was observed at mitochondria of cell after

30 min of the toxin treatment.4. Cell death was induced on cells of segments after 2h of the

toxin treatment5. ROS generation and cell death were light-independently

induced by the toxin. -

at. tsurushima, bh. nakayashiki, by. tosa and bs. …...an oxidized derivative of linoleic acid...

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