The Relationship of Phenols and Oxidative Enzymes with the Resistance of Hevea to South American Leaf Blight

Phytopath. Z., 97, 332—345 (1980)© 1980 Verlag Paul Parey, Berlin und HamburgISSN 0031-9481 / ASTM-Coden: PHYZA3

Unit of the Rubber Research Institute of Malaysiaand Crop Science Department, University of the West Indies, Trinidad

The Relationship of Phenols and Oxidative Enzymeswith the Resistance of Hevea to South American Leaf Blight

By

ISMAIL HASHIM, K. H . CHEE and L. A. WILSON

With 3 figures

Received September 18, 1978

The obligate parasite, Microcyclus ulei (P. Hcnn.) Arx infects youngdeveloping Hevea brasiliensis Muell. Arg. leaves causing one of the world'smost serious diseases, South American leaf blight (SALB), on this crop. CertainHevea clones are resistant to this disease; however, the mechanism of resistanceIS little known. BLASQUEZ and OWEN (1963) found that the fungus penetratedleaves from both resistant and susceptible clones. This was confirmed byHASHIM, CHEE and DUNCAN (1979). They further suggested that resistance wasrelated to hypersensitive host cell collapse.

From histological studies, phenols were observed to accumulate in lesionsof Hevea leaves infected with M. ulei (BLASQUEZ and OWEN 1963). FIGARI(1965) mdicated that leaf extracts from a resistant Hevea clone were moremhibitory to germination of M. ulei conidia than extracts from a susceptibleclone. Using several resistant and susceptible clones, a similar result wasobtained by HASHIM (1978). From chromatographic tests, FIGARI (1965) sug-gested that the toxic substance was a flavonol (Rf-0.80) and MARTAINS et al.(1970) identified a similar substance as kaempferol 3-rhamnodigIucoside.

The mitiation of hypersensitive cell collapse in host tissues infected withplant pathogens has been attributed to accumulation of phenols at the infec-tion site (KiRALY and FARKAS 1962) and oxidation of phenols by peroxidaseand polyphenol oxidase (SIMONS and Ross 1971, WESTSTEIJN 1976). Further-more, high activities of peroxidase (FEHRMANN and DIMOND 1967, UMAERUS1959) and polyphenol oxidase (FEHRMANN and DIMOND 1967) in healthy

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The Relationship of Phenols and Oxidative Enzymes 333

tissues and increased activities of these enzymes with infection (FEHRMANNand DIMOND 1967) as well as regulation of auxins by indole acetic acid (IAA)oxidase (SEQUEIRA 1973, SHAW and HAWKINS 1958) have also been associatedwith plant resistance. This paper reports the contents of phenols and the levelof IAA oxidase, peroxidase and polyphenol oxidase in different Hevea clonesand post infection changes in phenol contents and enzyme activities. The effectof several growth substances on lesion development was also examined.

Materials and Methods

InoculationIf not specified, Hevea leaves of the species H. brasiliensis were used. Whole leaves,

7 days old from bud burst if not otherwise specified, or leaf disks (15 mm diameter) wereplaced with their abaxial surface facing upward on moistened filter paper or floated onwater or solutions of growth substances in Petri dishes. They were sprayed with a suspensionof M.ulei conidia (2 X lOS spores/ml) and incubated at 2A°C under daylight and at nightunder four 40 W white fluorescent tubes (total 250—280 lumens) as described by CHEE(1976).

Enzyme extraction and assay

Enzyme preparations were extracted by the ac£ione powder method (KONISHI andGALSTON 1964) as previously described (HASHIM, "WILSON and CHEE 1979). Leaves whidi hasbeen floated on solutions of growth substances were thoroughly washed with distilled waterbefore being processed into acetone powder. IAA oxidase activities were assayed as describedpreviously (HASHIM, WILSON and CHEE 1979) using the modified Salskowski's reagent (GOR-DON and WEBER 1951). Assays were made at 30 "C in the presence of citrate-phosphate buffer(pH 5.0) using p-coumaric acid as the promoter. Readings were taken at 530 nm.

Peroxidase activities were determined by measuring the increases in absorbance at470 nm 1 min after the addition of 1 drop of hydrogen peroxide (10 nmole) to the reactionmixture containing 2.0 ml (20 /(mole) guaiacol, 0.5 ml citrate-phosphate buffer (pH 6.0)and 0.5 ml enzyme extract.

Polyphenol oxidase activities were assayed hy determining the absorbancy increasesat 407 nm using catediol as the substrate. Readings were taken using a Bediman DB spectro-photometer 1 min after incubation of the enzyme and substrate mixture. IAA oxidase activ-ities were expressed as umoi IAA destroyed/h//fg enzyme protein while peroxidase and poly-phenol oxidase activitie.'s were expressed as unit diange in absorbance (AA)/h/m$ protein.Protein contents were determined by the method of LOWRY et al. (1951).

Phenol extraction and assay

Phenols were extracted by boiling (30 min) 3 g of thinly sliced Hevea leaves in 20 mlwater, 3 g of cross-sections of young stems in 15 ml water in experiments to determinephenol contents in healthy tissues, and 1 g leaves in 20 ml water in experiments to determinepost infection changes in phenol contents. Total phenol was assayed by the method of STAINand HiLLis (1959) using FoUn reagent prepared as described by ROSENBLATT and PELUSO(1941), Ortho-dihydroxy phenol was assayed as described by ARNOW (1937).

Quercetin was extracted by boiling (30 min) 3 g of finely chopped leaves (7 days old)in 4 ml water, filtered and the filtrate evaporated to near dryness. To this was added 2 mlhot water and the mixture hydrolysed at 70 °C with 0.5 ml 0.1 N hydrochloric acid for4—7 h and later extracted with 1 ml ethyl acetate. One //I of the acetate fraction wasdiromatographed on Whatman No. 1 diromatography paper in butanol : acetic acid: water(4 :1 :5 Y/V). Commercial preparations of kaempferol and quercetin were run as standards.

334 HASHIM, CHEE and WILSON

Electrophoresis

Leaveswere harvested, inoculated and incubated for 72 h as previously described and1 g eadi of inoculated and uninoculated leaves were frozen and ground with a pre-diilledpestle and mortar In 3 ml 0.1 M Tris (hydroxyraethyl) aminomethane (pH 8.0) containing!15% sucrose, 0.1% ascorbic acid and 0.1% cystcine hydrodiioride. Solutions squeezedthrough two layers of cheese-cloth were centrifuged (0 °C for 10 min at 10,000 rpm) and50 ul of the supernatant was applied to the separation gel. Peroxidase isozymes were separatedm vertical acrylamide gel (7.5%) slabs by the method of REID and BIELESKI (1967) usingDAVIS (1964) buffer systems. Electrophoretic runs were at 4°C for 4 h at constant voltage(130 volts). Peroxidase isozymes were stained by immersing gels in M/I5 citrate-phosphatebuffer (pH 6.0) containing 0.02 M guaiacol and 0.03 % hydrogen peroxide.

Determination of percentage germination

Conidia were harvested from naturally field infected leaves by brushing with a wetcamel's hair brush into distilled water to give a suspension of about lOS spores/ml. One mlof the conidial suspension was mixed with 1 ml of the solutions of phenolic compounds orgrowth substances. After 12—14 h incubation at 24 °C in the dark, percentage germinationwas determined.

Results

Contents of phenolic compoundsand level of enzyme activities in healthy tissues

There was no association between the total content of phenolic com-pounds from young healthy Hevea leaves and stems with their degree ofresistance to SALB (Table 1). The content of phenolic compounds in the

Tahle 1Total phenolic compounds from leaves and stem of H. brasiliensis and H. benthamiana-^

of different susceptibilities to SALB

Clone

FX516+HHB 14+64B 721FX25F3517/1664B 238RRIM 600FI0 4/14343

18A 14/23RRIM 605C 57/10

Susceptibilityrating'^

1II12223444

•ft

55

mg dilorogenic acid equivalentper g fresh weight

leaf

14.2515.5016.2518.5019.0020.5027.0024.2517.2518.2525.2519.0020.0020.50

stem

1.642.16

3.122.762.404.043.801.922.763.202.162.162.56

*) 1, highly resistant; 2, resistant; 3, intermediate; 4, susceptible; 5, highly susceptible


moderately resistant clones varied from high to moderately high levels whilethe highly resistant clones surprisingly possessed lower contents than the sus-ceptible clones. Under ultraviolet light, the yellow spots from Hevea leafextracts corresponded to quercetin (Rf = 0.6) and not to kaempferol (Rf=0.9).The eluate of the yellow spots inhibited conidial germination of M. ulei.Among the 25 Hevea clones examined, the spots were brightest with extractsfrom the resistant clones 5/26, 5/139, 6/57, 6/212, C 7, IAN 710, IAN 6645,64B 721, 64B 850, and FB 3363, the exceptions being F 351 and C 12. Mostof the susceptible clones showed light yellow spots: RRIM 501, RRIM 605,C14, C22, 64B120, 64B305, 18A 12/11, 18A 12/21, 18A 15/20 and 7/8 with64B 553 and 285 being exceptions. A similar pattern was observed with theinfected leaves of these clones.

There was some associationship between resistance of Hevea leaves toSALB with the level of activities of IAA oxidase and peroxidase. Peroxidaseand IAA oxidase activities were significantly (p=0.05) higher in leaves fromsusceptible clones. Polyphenol oxidase activities were not correlated withresistance (Table 2).

Table 2The level of activities of IAA oxidase, peroxidase and polyphenol oxidase

of preparations from young healthy Hevea leaves

FX516HHB 14FX25F3517/1664B238F10 4/14343RRIM 6057/1064B 382

Susceptibilityrating*)

1112224

4555

IAA oxidase(^mol IAAdestroyed/h//ig protein

xio-*)

5.864.386.555.854.963.41

10.3012.8012.206.048.02

Enzyme activities

peroxidaseprotein X{A A/min///g protein

X 10-1)

2.382.822.614.943.652.363.783.789.224.613.99

polyphenoloxidase

(J A/min/ug protein' X 10-1)

1.405.302.432.753.101.432.322.324.002.433.82

Differences in enzyme activities between resistant and susceptible clones were significantat P = 0.05 for IAA oxidase and peroxidase.

*) See Table 1.

Phenol dianges with infection

Total phenolic compounds (Table 3) and ortho-dihydroxy phenol(Table 4) contents decreased with period of incubation of inoculated and


healthy leaves. There was no significant quantitative difference in total con-tents of phenolic compounds in inoculated leaves as compared with healthyleaves both from resistant and susceptible clones. Similar observations appliedto ortho-dihydroxy phenol.

Table 3Total phenolic compounds' from diseased (artificially inoculated) and healthy

resistant and susceptible Hevea leaves

Treatmentmg dilorogenic acid equivalent/g fresh leaf weight

at different times (h) after inoculation

24 48 72 96 120 144

F 351 (resistant) ControlInoculated

31.00 22.25 22.63 24.75 21.13 18.50 18.50— 20.50 22.04 20.00 17.73 20.50 19.50

F 10 3/2 (resistant) ControlInoculated

34.50 27.50 27.75 24.25— 29.50 25.50 24.00

19.00 17.50 —20.00 18.00 —

7/14 (susceptible)

C 5 (susceptible)

Control 33.50 19.00 19.00 20.00 20.50 24.00 27.00— 23.50 22.50 20.00 19.50 15.00 16.00Inoculated

Control 24.50 28.00 26.00 22.50 20.00 15.00 12.50Inoculated — 25.00 25.50 19.00 17.00 17.00 20.50

Table 4Ortho-dihydroxy phenols from diseased (artificially inoculated) and healthy Hevea leaves

Clone

F 351 (resistant)

F 10 3/2 (resistant)

7/14 (susceptible)

C 5 (susceptible)

Treatment

ControlInoculated

ControlInoculated

ControlInoculated

ControlInoculated

mg chlorogenic acid equivalent/g fresh weightat different times (h) after inoculation

0

12.75

13.00

9.00

7.00

24

9.258.55

10.0011.30

5.005.70

9.406.20

48

8.2010.20

10.359.25

5.005.10

8.706.30

71

8.257.45

9.059.70

5.405.80

6.505.90

96

9.157.35

8.507.10

5.405.80

4.704.00

120

5.607.20

6.907.30

6.204.40

4.505.00

124

8.809.40

—

7.505.00

3.806.30

Enzyme changes after infection

IAA oxidase

Enzyme activities increased with the period of incubation of healthy andinoculated leaves both from resistant and susceptible Hevea clones. Inoculatedleaves, except for clone 7/14 (susceptible), usually possessed higher enzymeactivities as compared with healthy leaves and the increases were detected 24 hafter inoculation in the other three clones (Fig. 1). The main difference in the


level of IAA oxidase activities between inoculated resistant and inoculatedsusceptible leaves was that the relative activities in inoculated leaves (aspercentages of controls i.e. healthy leaves) were greater with resistant leavesthan with susceptible leaves. Forty-eight h after inoculation, relative activitiesof IAA oxidase of inoculated leaves were 271 % and 246% of the controlsfor the two resistant clones F351 and FIO 3/2 respectively, and 59% and121 % of the controls for the susceptible clones 7/14 and C5 respectively.

0.8

0.6

•E 0.40)o

Ol 0.2

0) 0.5TJ

0.4

0.3

0.2

0.1

0.8

0.6

0.4

0.2

1 LClone F 351 (R)

1.0

0.8

0.6

0.4

0.2

Clone C 5(S)

J L

Clone F 10 3/2 I R)

- O

24 48 72 96 0 24 48

Hours after inoculation72 96

Fig. 1. The level of activities of IAA oxidase in diseased {•—•) and healthy (o- - -0) leavesfrom resistant (R) and susceptible (S) Hevea clones

Peroxidase

Peroxidase activities from the four clones tested were similar to the IAAoxidase. Peroxidase activities increased with the period of incubation (Fig. 2).Inoculation of leaves stimulated increased activities of peroxidase with a peak

Phyiopath. Z., Bd. 97, Heft 422


2.0

1.5

0.5

1.4c'vo

b)E£ 1.0<

< 0.8

0.6

0.4

Clone 7/14 (S)

2.0

1.5

Clone F 351 (R)

0.5

0.8

0.6

0.4

0.

J L

Clone C 5 (S)

Clone FtO 3/2 (R)

R

Q.. - O '

J I L

0 24 48 72 96 0 24 48 72 96Hours after inoculat ion

Fig. 2. The level of activities of peroxidase in diseased (•—•) and healthy (o- - -o) leavesfrom resistant (R) and susceptible (S) Hevea clones

at 71—9(> h after inoculation in both resistant and susceptible clones. Theincreases in enzyme activities of inoculated leaves over controls were detected24 h after inoculation of resistant leaves and 72 h after inoculation of sus-ceptible leaves. Forty-eight h after inoculation, the relative activities were130% and 233% of the controls for the two resistant clones F351 andF 10 3/2 respectively and 75 % and 93 % of controls for the susceptible clones7/14 and C5 respectively.

Electrophoresis of leaf extracts of inoculated and uninoculated leavesfrom resistant and susceptible Hevea clones indicated two closely situatedperoxidase isozyme bands. No difference was detected between inoculated andcontrol leaves in the number and position of isozymes; however, darker bandswere observed in preparations from inoculated leaves.


Polyphenol oxidase

The four clones tested differed in the enzyme activities of diseased leavesas compared with healthy leaves, with no definite differences in activitiesbetween inoculated resistant and inoculated susceptible leaves as comparedwith their respective controls. However, activities in inoculated susceptibleleaves mcreased with period of inoculation and 71 h after inoculation, activitieswere comparatively similar in inoculated and uninoculated leaves. Conversely,activities in inoculated resistant leaves decreased with increasing period ofmcubation and at 71 h after inoculation, enzyme activities in inoculated leaveswere Iower than m their respective controls (Fig. 3).

0 24 48 72 96 0 24 48 72 96Hours after inoculation

Fig. 3. The level of activities of polyphenol oxidase in diseased (•—•) and healthy (o- - -o)leaves from resistant (R) and susceptible (S) Hevea clones

340 HASHIM, CHEE and "WILSON

Effect of growth substances on lesion development

The disease lesions which developed in Hevea leaf disks floated on solu-tions of growth substances were smaller than lesions in disks floated on water(Table 5). /f-naphthoxyacetic acid and kinetin were more effective in limitinglesion development than gibberellic acid and IAA. However, the latter wasstill effective in lesion suppression at 5 ppm (Table 6). Disks enlarged con-siderably when floated on gibberelHc acid. The reduction of lesion size in disksfloated on IAA solutions was greater in disks from susceptible clones (29%)than in disks from resistant clones (14%).

Tahlc 5Lesion size on Hevea leaf disks following ioculation with M. ulei

and incubation on solutions of growth substances

Growth substances

IAAKinetinGibberellic acid/?-naphthoxy acetic acid

Lesion diameter (wm)''') of leaf diskson 10 ppm solutions of growth substances

treated

318 c484 a672 NS514 b

water

740

957985981

S.E.

0.030.010.090.04

a, b,c and NS: significant at 0.01%, 0.1 %, 1 % and 10 % respectively.••') Average of 4 replicates except for IAA which has 15 replicates; thirty lesions were

measured per replicate.Table 6

Effect of different concentrations of IAA on lesion diameter (/(m)on Hevea leaf disks inoculated with M. ulei

Clone

F35164B 1203437/1064B 837

Mean

Susceptibilityrating*)

23455

0

526829785

10051052

839

IAA solution (ppm)

5 10

441 446909 944720 607853 789691 670

723 691

100

0

689472557650

464

LSD between treatment means: LSD (5%) = 150, LSD (1%) = 210, LSD (0.1%)= 300.

*) See Table 1.

The effect of certain phenolic compoundsand growth substances on conidial germination

At 5 X 10-5 M, quercetin did not inhibit conidial germination, howeverrutin, a glucoside of quercetin, at 4 X 10"^ M inhibited conidial germination


by 50%. Due to aqueous insolubility of the compounds, the effects of quer-cetin at a concentration above 5 X 10—' M was not tested.

IAA and /i-napthoxyacetic acid at concentrations of 100 ppm and abovestrongly inhibited conidial germination (Table 7) while gibberellic acid didnot have the same effect.

Table 7Effect of growth substances on germination of conidia of M. ulei

Concentration(ppm)

1000100

105

Water

IAA

0.012.691.1

114.6100.0

Germination'^)

B-naphthoxyacetic acid

0.00.0

75.982.9

100.0

Gibberellic acid

0.0128.8107.9102.7100.0

*) As percentage of control (water). Data were average of two replicates with 200 to300 conidia counted per replicate.

Enzyme activities from leaves floated on IAA solution

The activities of IAA oxidase and peroxidase preparations of leavesfloated either on water or IAA solutions were higher than in fresh leaves(Table 8). IAA oxidase and peroxidase activities of inoculated resistant leaveswere nearly double that of uninoculated leaves, both floated on water. In thesusceptible clone, inoculated leaves floated on water possessed lower or similarenzyme activities than uninoculated leaves. Inoculation of resistant leavesfloated on IAA solution, produced preparations with lower IAA oxidase

TablesIAA oxidase and peroxidase activities of Hevea leaves floated on IAA solutions.

Average of two assays

olution

WaterWaterIAAIAA

Conditionof leaf

HealthyHealthyDiseasedHealthyDiseased

Incubationperiod (h)

048484848

IAA oxidase/imol IAA destroyed/h/

//g protein (X 10-3)

F 331(resistant)

RRIM 605(suscep-tible)

0.34 0.091.66 0.492.65 0.471.26 0.771.07 0.57

PeroxidasezlA/h//ig protein

(X 10-3)

F -ic]

(resistant)RRIM 605

(suscep-tible)

0.36 0.291.30 1.062.21 0.711.19 1.040.81 1.09


activities as compared with uninoculated leaves floated on water or IAAsolution. Peroxidase activities of leaves floated on water were similar to leavesfloated on IAA solution; however, inoculation of susceptible leaves floated onIAA solution showed higher enzyme activities than inoculated leaves floatedon water.

Discussion

Varietal resistance to several diseases of certain crop plants has beenrelated to the higher activities of peroxidase (FEHRMANN and DIMOND 1967,UMAERUS 1959) and polyphenol oxidase (FEHRMANN and DIMOND 1967).Contents of phenolic compounds have also been correlated with resistance(LEE and LETOURNEAU 1958, NEWTON and ANDERSON 1929), however, inseveral other diseases no such correlation existed (KIRALY and FARKAS 1961,SEEVERS and DALY 1970). In several plants, infection of resistant varieties wasalso accompanied by increased content of phenolic compounds (KOSUGE 1969,POLLOCK and DRYSDALE 1976), however, contents of phenolic compounds ininoculated Hevea leaves were not significantly different from contents inhealthy leaves. Though the total of phenolic compounds was not affected bydisease development, a specific phenol could still be important in the media-nism of Hevea resistance to SALB. FIGARI (1965) indicated greater inhibitionof conidial germination by leaf extracts from a resistant Hevea clone thanfrom a susceptible clone and he suggested that the toxic substance was aflavonol which was later identified as kaempferol 3-rhamnodiglucoside byMARTAINS et al. (1970). Using similar chromatographic techniques, our resultsindicated that the Rf of the toxic substance (Rf^O.6) corresponded to quer-cetin and not kaempferol. It seemed that quercetin contents were generallyhigher in resistant Hevea clones. In contrast to potato, where varieties resistantto Phytophthora infestans (Mont.) de Bary possessed higher peroxidase activi-ties than in susceptible varieties (FEHRMANN and DIMOND 1967, UMAERUS1959), Hevea clones resistant to SALB possessed significantly (p ^ 0.05) lowerperoxidase and IAA oxidase activities.

Application of growth substances increased resistance to certain diseases(DAVIS and DIMOND 1953). SHAW and HAWKINS (1958) related regulation ofauxin contents by IAA oxidase to resistance of wheat to rust. Inoculation ofleaves from resistant Hevea clones stimulated increased activities of IAAoxidase and peroxidase and the increases were greater and were detectedearlier in resistant leaves. No new peroxidase isozyme was detected, however,indicating that increased peroxidase activities were due to the increase inactivities of pre-existing isozymes. Polyphenol oxidase activities seemed to btsuppressed after inoculation of resistant Hevea leaves. MELOUK and HORNER(1972) also detected lower polyphenol oxidase activities in diseased pepper-mint infected with Phoma strasseri Moesz. They attributed the lowering poly-phenol oxidase activities to continuous contact between the enzyme and theoxidized substrate which inhibited enzyme activity.


Earlier studies (HASHIM, CHEE and DUNCAN 1978) showed that resistanceof Hevea was related to the hypersensitive collapse of host tissues. In highlyresistant leaves cell collapse was detected 24 h after inoculation and much laterin less resistant leaves. These observations coincided with the fact that inresistant leaves peroxidase activities increased over the controls 24 h afterinoculation of resistant leaves and 48 h in susceptible leaves. Since peroxidasehas been related to initiation of hypersensitive host cell collapse (SIMONS andRoss 1971, WESTSTEIJN 1976), peroxidase could be important in the mecha-nism of Hevea resistance to SALB. LOON (1973) suggested that reduction ofTMV lesion size in tobacco leaves floated on IAA solution was probably dueto fungitoxic properties of IAA; however, he also suggested that peroxidasecould be related to reduction of lesion size. Similarly, peroxidase could beinvolved in reduction of lesion size in Hevea leaf disks infected with M. ulei.Lesion size reduction could be due to peroxidase since susceptible leavesfloated on IAA possessed reasonably high peroxidase activities when infected(Table 7).

Summary

There was no direct correlation between total contents of phenolic com-pounds in healthy Hevea leaves and their degree of resistance to SouthA>merican leaf blight (Microcyclus ulei). However, quercetin seemed to behigher in leaves from resistant clones. Total phenolic compounds and ortho-dihydroxyphenol contents in diseased resistant leaves were not significantlydifferent from those in diseased susceptible leaves. IAA oxidase and peroxidaseactivities were generally higher in susceptible leaves and these enzymes in-creased with infection giving greater increases, and earlier detection, aftermfcction of resistant leaves. Presence of growth substances reduced lesion sizewhich could be related to increased peroxidase activities.

Zusammenfassung

Die Beziehung von Phenolen und oxydativen Enzymenzur Resistenz von Hevea gegenuber dem sudamerikanisdien Blattbrand

Zwischen dem Gesamtgehalt an phenolischen Verbindungen in gesundenHevea-hYdnem und ihrem Resistenzgrad gegenuber dem sudamerikanisdienBlattbrand (Microcyclus ulei) gibt es keine direkte Korrelation. Jedodi scheintder Gehalt an Quercetin in Blattern von resistenten Klonen hoher zu sein. DerGesamtgehalt an phenoiischen Verbindungen und der Gehalt an ortho-Dihydroxyphenol in erkrankten resistenten Bidttern war nidit signifikantunterschiedlidi zu dem in erkrankten anfalligen Blattern. Die Aktivitaten vonIndolessigsaureoxidase und -peroxidase waren in anfalligen Blattern gewohn-hch hoher, diese Enzyme nahmen mit der Infektion zu und zeigten gegen-uber der Infektion von resistenten Blattern hohere Zuwachsraten und konntenfruher nachgewiesen werden. Die Anwesenheit von Wachstumssubstanzen ver-


ringerte die Grofie der Lasionen, was mit dem Anstieg der Peroxidaseaktivitatin Zusammenhang gebradit werden konnte.

We gratefully acknowledge financial support from the Directorate and Board of theRubber Researdi Institute of Malaysia to one of us (I.H.).

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Authors' addresses: Dr. ISMAIL HASHIM, Crop Protection and Microbiology Division,Rubber Researdi Institute of Malaysia, P.O. Box 150, Kuala Lumpur (Malaysia). Dr. K. H.CHEE and Prof. L. A. WILSON, Unit of the Rubber Researdi Institute of Malaysia and De-partment of Crop Science respectively. University of the West Indies, St. Augustine, Trinidad(West Indies).

Documents

The Relationship of Phenols and Oxidative Enzymes with the Resistance of Hevea to South American Leaf Blight