• TABLE OF CONTENTS
HIDE
 Front Cover
 Front Matter
 Abstracts of papers presented at...
 Fungicidal activity of crude plant...
 Rice ragged stunt disease in the...
 Mungbean viruses in the Philippines....
 Control of nematodes with temik...
 Studies on the white rust disease...
 Resistance in soybeans [glycine...
 Transmission of rice grassy stunt...
 Population dynamics of plant parasitic...
 Back Cover














Group Title: Journal of Tropical Plant Pathology
Title: Journal of tropical plant pathology
ALL VOLUMES CITATION THUMBNAILS PAGE IMAGE ZOOMABLE
Full Citation
STANDARD VIEW MARC VIEW
Permanent Link: http://ufdc.ufl.edu/UF00090520/00020
 Material Information
Title: Journal of tropical plant pathology
Series Title: Journal of tropical plant pathology.
Alternate Title: Journal of Philippine phytopathology
Philippine phytopathology
Physical Description: v. : ill. (some col.) ; 26 cm.
Language: English
Creator: Philippine Phytopathological Society
Publisher: Philippine Phytopathological Society
Place of Publication: Philippines
College Laguna
Publication Date: January-June 1978
Frequency: semiannual
regular
 Subjects
Subject: Plant diseases -- Periodicals -- Philippines   ( lcsh )
Plants, Protection of -- Periodicals -- Philippines   ( lcsh )
Genre: periodical   ( marcgt )
 Notes
Dates or Sequential Designation: v. 1, no. 1 (January 1965)-
General Note: Title from cover.
General Note: "Official publication of the Tropical Plant Pathology."
 Record Information
Bibliographic ID: UF00090520
Volume ID: VID00020
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: oclc - 54382605
issn - 0115-0804

Table of Contents
    Front Cover
        Front Cover 1
        Front Cover 2
    Front Matter
        Front Matter 1
        Front Matter 2
    Abstracts of papers presented at the fifteenth annual meeting of the Philippine phytopathological society inc., Metro Manila, 3-6 May, 1978
        Page 1
        Page 2
        Page 3
        Page 4
        Page 5
        Page 6
        Page 7
        Page 8
        Page 9
        Page 10
        Page 11
        Page 12
        Page 13
        Page 14
        Page 15
        Page 16
        Page 17
        Page 18
        Page 19
        Page 20
        Page 21
        Page 22
    Fungicidal activity of crude plant extracts against helminthosporium oryzae
        Page 23
        Page 24
        Page 25
        Page 26
        Page 27
        Page 28
        Page 29
        Page 30
        Page 31
        Page 32
        Page 33
        Page 34
        Page 35
        Page 36
        Page 37
    Rice ragged stunt disease in the Philippines
        Page 38
        Page 39
        Page 40
        Page 41
        Page 42
        Page 43
        Page 44
        Page 45
        Page 46
        Page 47
        Page 48
        Page 49
        Page 50
        Page 51
        Page 52
        Page 53
        Page 54
        Page 55
        Page 56
        Page 57
    Mungbean viruses in the Philippines. I. Identity of a virus causing mottle in Mungbean
        Page 58
        Page 59
        Page 60
        Page 61
        Page 62
    Control of nematodes with temik 15 G aldicarb and its residue in the fruit of giant cavendish banana
        Page 63
        Page 64
        Page 65
        Page 66
        Page 67
        Page 68
    Studies on the white rust disease of kangkong
        Page 69
        Page 70
        Page 71
        Page 72
        Page 73
        Page 74
        Page 75
        Page 76
        Page 77
    Resistance in soybeans [glycine max (L.) Merr.] to root-knot nematodes and statistical analysis of correlations of assesment parameters
        Page 78
        Page 79
        Page 80
        Page 81
        Page 82
        Page 83
        Page 84
        Page 85
        Page 86
        Page 87
        Page 88
    Transmission of rice grassy stunt by three biotypes of nilaparvata lugens
        Page 89
        Page 90
        Page 91
        Page 92
    Population dynamics of plant parasitic nematodes. II. Rotylenchulus reniform on monocultures bush sitao in nemacur-treates and non-treated farmer's field
        Page 93
        Page 94
        Page 95
        Page 96
        Page 97
        Page 98
    Back Cover
        Page 99
        Page 100
Full Text





au 0 l &


LdINs dflU C. H .. . . . . .


and P. Q. i


;es in the Philippine
ns............


us Causing Moti
...........


transmission of Rice Grassy Stunt by Three Biotypes of Nilaparvata lugens -
Aguiero and K. C. Ling. ................... ..................

population Dynamics of Plant Parasitic Nematodes. II. Rotylenchulus reniform
Monocultured Bush Sitao in Nemacur-Treated and Non-Treated Farmer's Fi
R. H. Calinga and M. B. Castillo .................. ..............










3Li~tW~~fl A YEJEiII Al C#%F lI'


hst Philippines, Inc., Corner Pioneer & Reliance Sts., Mandaluyong, Metrc


I .III.PIIn. H sU os u Is...


Shell Chemical Company (Philippines), I
Metro Manila


Tarlac Development Corporation, Haciei


Twin Rivers Research Center, Twin Rive
del Norte


11111id11, L LULU I ,ILy


., Shell House, 1330 Roxas Blvd.,



I Luisita, San Miguel, Tarlac


Plantation, Inc., Madaum, Tagum, Davao


wa as









Philippine

Phytopathology


icial Organ of the Philippine Phytopathological Society, Inc.



EDITORIAL BOARD
CITA H. QUIMIO, Editor-in-Chief, Department of Plant Pathology
UPLB, College, Laguna
.DALMACIO, Associate Editor, UPLB, College, Laguna
1. CASTILLO, Associate Editor, UPLB, College, Laguna

BUSINESS MANAGEMENT

R. E. CORTEZ, Business Manager, BPI, Manila













ABSTRACTS OF PAPERS PRESENTED AT THE FIFTEENTH
ANNUAL MEETING OF THE PHILIPPINE PHYTOPATHOLOGICAL
SOCIETY INC., METRO MANILA, 3-6 MAY, 1978


Rice ragged stunt disease in the Philip- Philippines. II. Transmission K. C.
es. I. Symptomatology K. C Ling, Ling, E. R. Tiongco, V. M. Aguiero, and
R. Tiongco, V. M. Aguiero, and P. Q. P. Q. Cabauatan, IRRI.
)auatan, IRRI. Transmission of ragged stunt disease
Rice ragged stunt disease occurred by various methods was studied. The
radically in the Philippines. The transmission of the disease by mechanical
nptoms of the disease are quite com- means was unsuccessful. The disease was
x, and vary according to the stage not transmitted through soil even in the
plant growth. The diseased plants presence of stubble of diseased plants.
stunted to various degrees at all No evidence was obtained to demonstrate
wth stages. Other symptoms are clear the transmission of the disease through
en they are closely examined. At early rice seed.
wth, the symptoms are: appearance The disease is transmitted by the
ragged leaves, twisted leaves and brown planthopper Nilaparvata lugens
n-swellings. The ragged leaves refer (Stal). About 40% of the tested brown
leaves having a portion of irregular planthoppers, including nymphs, female
Ye that consists of breaking and and male adults, brachypterous and
orotic edge in between or near the macropterous forms, were active trans-
aakings. Twisting of leaves is due to mitters. The latent period ranged from
rate uneven growth of the leaf 2 to 33 days. The retention period ranged
de on bbth sides of the midrib. The from 3 to 35 days. The disease-transmit-
n-swellings (technical term is his- ting days from the time that the insect
d enation or virocecidium) are due to became infective to its death varied
liberationn of phloem cells in vascular from 3 to 100%. The virus passage was
ndles. transstadial but not transovarial. Con-
At booting or later stages, flag leaves sequently, it is categorized in the per-
the diseased plants are short and often sistent group without transovarial passage.
isted, malformed, or ragged. Diseased The brown planthoppers in Japan are
nts often flower late. Panicle emergence capable of transmitting the disease. There
often incomplete. Tillers often generate were no striking difference in percentage


tertiary tillers produced at upper nodes 1
a tiller. The nodal branches often
ar panicles. Therefore, the diseased
mts have more panicles and spikelets,
t the number of filled grains is greatly
luced.
The ragged stunt disease is systemic.
te diseased plants can survive for
reral months after flowering. Experi-
:ntal results did not yield suppression i
vxrmntnme hu tptrarvrlinp trpatmepnt


ween the brown planthoppers in
ian and in the Philippines. However,
former infected more rice seedlings,
e a higher percentage of disease-trans-
:ting days, and had a longer retention
iod. It could be due to difference in
editions during the transmission test.
rhree brown planthopper biotypes in
Philippines did not differ significantly
percentage of active transmitters nor
latent period, infected seedlings,








2


transmitting days. Hence, they we
similar in ability to transmit the disease
Rice ragged stunt disease in tV
Philippines. III. Causal agent, host rang
and dual infection K. C. Ling, E. 1
Tiongco, V. M. Aguiero and P. Q. Cabau
tan, IRRI.
The nature of the causal agent <
ragged stunt disease was studied in coll
boration with Dr. E. Shikata and h
associates in the University of Hokkaidi
Japan. Electron micrographs showe
polyhedral particles of 50-7
nm not only in dip preparations but als
in ultrathin sections of rice plants natural
ly and artificially infected with th
disease in the Philippines. The particle
appeared abundantly in phloem cel
and in cells of vein-swellings. The infei
tivity of rice ragged stunt virus we
determined by the injection method
Two weeks after injection with th
supemate of macerated diseased plant
in a buffer solution, the brown plan
hoppers became infective, and cause
the inoculated seedlings to develop
the disease symptoms. These are ev.
dence that the causal agent of the ragge
stunt disease is a virus.
In addition to Oryza sativa, 0. latifoli
and 0. nivara were also infected witl
ragged stunt disease by showing similar
symptoms that occur on 0. sativa infec
ted with the disease. The brown plant
hoppers recovered the virus from the
diseased plants and transmitted it success
fully to rice plants. Consequently, O
sativa is not the only host of the virus
Rice plants were dually infected witl
both ragged stunt and grassy stunt, ane
ragged stunt and tungro by inoculation
with respective viruliferous insects. Th(
absence of cross protection indicates thai
the causal agent of ragged stunt is noi


Kice raggea stunt disease m mt
Philippines. IV. Discussion K. C Ling
E. R. Tiongco, V. M. Aguiero, and P. Q
Cabauatan, IRRI.
Of the other 14 known virus ani
virus-like diseases of the rice plant in th
world, only two, black-streaked dwar
and grassy stunt, have some unique
similarities with ragged stunt disease
The black-streaked dwarf also causes rio
plants to produce vein-swellings. The vein
swellings of black-streaked dwarf oi
culms can be grouped as tumors that ar
more irregular in shape and wider ii
width, and protrude much more thai
ragged stunt vein-swellings on culms
Furthermore, the black streaked dwarf i
transmitted by Laodelphax striatellus
and has not been successfully transmitted
by Nilaparvata lugens, the vector o:
ragged stunt disease.

Both grassy stunt and ragged stuni
are transmitted by the same insect
vector, in the same persistent manner, and
without transovarial passage, but the)
differ strikingly in their symptoms
Furthermore they do not cross protect
each other. Consequently, the ragged
stunt is a new virus disease of the rice
plant.
The origin of the ragged stunt disease
remains obscure. It is suspected that the
disease has long been present at a low
1 ^ - .^ A -^ - -


grassy stunt or tungro. D. Daguioag and K. C Ling, IRRI.


Phytopathology











The reactions of 2,190 entries of 359
t varieties and lines to rice ragged stunt
disease by natural infection at the IRRI
um were scored on a 0 to 9 scale which ]
idicates percentages of infection or rice
Uis.
Of the rices scored, about 33% gave
susceptible reaction (7 to 9 on the
ale). However, the following rices
towed low reactions: IR32, IR36, IR38,
t2307-247-2-2-3, IR3351-38-3-1, IR-
464-75-1-1, IR4219-35-3-3, IR4427-
8-3-2, IR4417-179-5-2, IR4432-28-5,
t4432-52-64, IR4432103-6-4, and IR-
353-118-5. The reason for the low
action of these rice remains to be
vestigated. It could be due to low
disease pressure, resistance to the insect
ectorNilaparvata lugens, resistance to the
rus, or their combinations.
A rice may show different reactions in
plicated tests in the field because the
itensity of disease pressure of ragged
unt varies within a field and from time
Time. For example, the reaction of
R26 in 76 replicated tests ranged from
to 9, averaging 4.8. The reaction
F IR36 in 84 replicated tests ranged
om 1 to 7, averaging 1.9. Consequently,
136 was more resistant to the disease
i the field than IR26.
Dosage rates of Ridomil 25 W. P.
iba-Geigy) as seed treatment fungi-
ide against Philippine corn downy
mildew O. R. Exconde and A. B.
folina, Jr., UPLB.
Earlier results on the evaluation of
idomil 25 WP(25% acylalalanine)showed
iat 2, 4, 6 and 8 g ai./kg seed gave
complete protection from downy mildew
ifection from emergence until harvest
under moderately low and high inoculum
ensities. Lower rates were further
I-' *' ir.... !..1...


nsity, treated seeds of PH 801 and
CA Var. with rates of 2.0, 1.5, 1.0
d 0.5 g ai/kg seed, were complete-
free from downy mildew infection
im seedling emergence to 42 days
er emergence, compared to 64.2
d 46.3% infection on the untreated
ds of PH 801 and UPCA Var. 3,
pectively. Eight rates, namely; 4.0, 3.5,
), 2.5, 2.0, 1.5, 1.0 and 0.5 g a. i./kg
d plus untreated check were also
luated under high inoculum density,
ng UPCA Var. 1, a susceptible variety
d DMR Comp. 2, a resistant variety.
eated seeds of DMR Comp. 2 at any of
e rates tested had no infection of
wny mildew while the untreated seeds
d 63.2% infection at 42 days after
iergence. On UPCA Var. 3, the untreated
eck showed 99.5% infection as early
28 days after emergence, while no
Section was observed on treated seeds
cept at the rate of 0.5 g a.i./kg seed.
: 0.5 g ai./kg seed 2.0% infection was
'served 28 days after emergence and
is infection increased to 3.1% 42 days
ter emergence until harvest.
Potato viruses in the Philippines: I.
'entity and distribution L. T. Talens,
'B.
A survey of indigenous viruses in
)tato seed stocks collected in Mountain
ovince was conducted in 1977. The
rvey included two selected farms in
tok and Madaymen situated at different
evation. They represented two of the
major growing areas along the Mountain
rail from La Trinidad, Benguet to
antoc.
The detection and diagnosis of these
ruses were performed using micro-
recipitin test with reference antiserum
mples. The following viruses, in single
:mixed infection and their corresponding
:rcentages of distribution, were detected:


__ __










yuJfl vruuo a anilu u, alu AI 7..) .io.
Only 13.5% of the tuber samples tested
were apparently not infected with potato
viruses X, S, and Y. Potato virus Y wa
not detected in single infection but wa
found to be present in combination witi
potato viruses X, or S, or both X and S
The incidence of potato virus X (74%
and potato virus S (69%) in single o
mixed infections was high confirming
their highly infectious nature.
These data strongly support the viev
that potato viruses such as potato viru
X, potato virus S, and potato virus .
contributed to the rapid degeneration ii
the state of health of seed stocks result
ing in the decline in productivity o
potatoes in the Philippines. It is suggest(
that potato viruses X and S contribute<
significantly to the low yield of potatoes
Potato viruses in the Philippines I
Identification of a ringspot: strain o,
potato virus X. L. T. Talens, IPB.
A ringspot strain of potato virus )
(PVX) was isolated and identified front
seed stocks used for commercial an(
seed potato production in Mountair
Province. The virus isolate produced ring
spots, systemic line pattern, and necrotic
streaks on tobacco; mosaic symptoms or
Capsium annum, C. frutescens, Datun
stramonium, Lycopersicon esculentum
and N. rustic; necrotic local lesion!
on Comphrena globosa; chlorotic loca
lesions on Chenopodium amaranticolo,
and C. album. Distinct, non-systemic
lesions with chlorotic ring were produced
on C. album.
Serodiagnosis using microprecipitir
test and neutralization of infectivity or
Comphrena globosa against reference po.
tato virus antiserum, and electron
microscopic demonstration of the pre.
sence of flexuous rod-shaped virions,
15 x 550 nm in dimension, in partially


ptlu1 VU VAlUO FJicpI LIiui5 uillWillIl
the identity of the virus.
This investigation describes the firs
isolation and identification of a ringspc
strain of PVX in the Philippines. Th
implications of this finding are discussed
Properties of a ringspot strain o
potato virus X. I. A simple method tA
purify PVX for the production of anti
serum Adelaida Lacuata, Angelit/
Dolores, and L. T. Talens.
Potato virus X was purified by home
genizing infected tobacco leaves in 0.;
M. borate buffer, pH 8.2 containing
1% mercaptoethanol and clarifying lea
extract with 1/10 volume of chloroform
with stirring for 15-20 min at 25C. Th
virus was concentrated by precipitation
with 6% polyethylene glycol 6000 an
4% NaCl followed by differential (5,00(
g for 15 min and 100,000 g for 90 min
centrifugation.
Virus purity was analyzed by density
gradient centrifugation at 25,000 rpm ii
a linear sucrose gradient consisting o
10-50% sucrose (w/w) solution in 0.0!
M borate buffer, pH 8.3 using a Spinc(
SW 27 swinging rotor at 4,C. Viru
banded as a narrow zone 2.5-3.0 cn
below the meniscus of the gradien
tube.
Ultraviolet absorption spectrophoto
metric measurements revealed that th(
purified preparation contained nucleopro
tein with typical maximum and minimurr
absorption at 260 nm and 240 nm
respectively. The minimum/maximufr
absorbance ratio (240 nm/260 nm,
equaled 0.89: the 260 nm ratio was 1.18

Electron microscopic examinations oi
:urified virus preparations negatively
trained with 2% phosphotungstate
showed flexuous rod-shaped particles
with an average size of 15 x 550 nm.


---I--~-------~r











arrains of me sugarcane mosaic virus A. gossypai, Longuinguis succnuar anu
n the Philippines D. A. Benigno and Rhopalopsiphum maidis were non-vectors.
Z. D. Dosayla, UPLB.
The mosaic diseases of maize in the
Three strains of SCMV were isolated The mosaic diseases of ize in the
Philippines, their causal viruses and
rom a collection of mosaicked sugar- vectors uong Hoai Xuan. Venus J.
;ane plants from the selected sugarcane- Calilung and D. A. Benign, UPLB.
producing provinces of the country such
is Batangas, Laguna, Leyte, Negros The mosaic-type disease of corn in
accidental Pampanga and Tarlac. On the the Philippines was found to be caused
oases of symptomatology, serological by two distinct plant viruses. One
action, morphology, host ranges and was a flexible rod-like transmitted by
physical properties, these strains were five species of aphids in a non-persistent
identified as A, D and H. manner, the other was a bullet or bacilli-
form shaped virus transmitted by a
Strain A and D seem to be the most
planthopper.
commonn strains. Strain D produced the planthopper.
nost severe symptom among three iso- The physical properties of the aphid-
ated strains. borne virus, its host ranges, symptomato-
logy, and morphology resembles the
Only 4 of the 10 grass species, and 5 Dwarf Mosaic Virus reported in the
Maize Dwarf Mosaic Virus reported in the
orghum varieties and 4 corn varieties United States. Serological tests revealed
United States. Serological tests revealed
vere susceptible to the disease.Rottboella that the one present in the Philippines is
that the one present in the Philippines is
xaltata produced mottling symptom the strain A of MDMV. On the other
vhile the other grasses exhibited ordinary hand, the planthopper-bome virus is
mosaic symptom, similar, if not identical to the Maize
Corn cultivars used in the test failed Mosaic (MMV) based on morphology,
o differentiate the isolated virus strains vector-type and symptomatology.
>ut exhibited varying degrees of infection. Both viruses have limited host ranges,
The lowest percent infection was 14.35%
infecting mostly grasses belonging to
recorded by Composite 1 and the highest Poaceae and one species in Musaceae.
as 44. 93% for DMR-Composite 2. Rottboellia exaltata L. and Euchlaena
No mosaicked sugarcane plants were mexicana Schrad. were found to be
observedd in Iloilo, Davao Sur, Cebu and common hosts of both viruses.
mariness Sur at the time of the survey. Of the five aphid vectors and MDMV,
Aphid vectors of soybean mosaic Schizaphis graminum (Rond.) Longuin-
irus in the Philippines D. A. Benigno guis sacchari (Zehntner), and Myzus per-
rnd S. Boonmarkka, UPLB. sicae (Zulzer) are established for the
Five of the 8 aphid species tested as first time in the Philippines.
'ectors in this study transmitted SMV. Cowpea little leaf disease in the
kmong 5 vectors, Aphis glycines and Philippines: Possible viral etiology as
lyzus persicae were the most efficient detected by immunodiffusion technique -
Ind considered of economic importance L. T. Talens, IPB.
n the Philippines. Based on double immunodiffusion
A. cracicivora, Hysteroneura .setai- tests in agar plates, clarified leaf extracts
ae and Melanopsis indosacchari were of cowpea showing little leaf syndrome


J


----UI-- --












antisera made against bean rugose mosaic loped local and systemic chlorotic lesions
virus (BRMV), bean pod mottle virus on Chenopodium quinoa L. Insect trans-
(BPMV), blackgram mottle virus (BGMV) mission test using Aphis craccivora Koch.
broad bean wilt virus-Type I (BBWV), suggested that MMV was carried non-
and cowpea strain of southern bean mosaic persistently from infected to uninfected
virus (SBMV-cs). BRMV, BPMV, and mungbean plants.
BGMV showed precipitin lines which This study describes the first report
fused together suggesting serological on the occurrence of mungbean mottle
identity. Isometric particles measuring ,,_. +.. i::-- .... -..


ucroscope
and v
ssibility
:d virus(
cowpea.
viruses ii
a virus


ine presei
is-like pa
it at leas
caused


the Philij
using moi


lu "* prop
ice of differ
ticles relat
Stwo virus
little
P4
solar
pines weea
tie in Chap


in host
ship fr

ilation I
earum it
economicc
d A. J. (


:action and
n, black-g

savior of I
rhizosphere
plant speca
timio, UPL,


ucing a mottle reaction and use'
.6 __ _. ___p Pwim


(Vigna m
. Based oi
and sym]
rties, ai
mungbeal
: antiger
ttle virus


rta) was is(
iermal stal
matology,
morphol
irus isolat
ally similh
3MV).


me electric
viral anti
raised the
spherical-s]
leaf disease
Mungbe
I Identity
mungbean

A virus
stunted gr<
of mungbe
and identify
host reacti
logical pr
features, tl
found to
blackgram
The vin
*,.,o (\f\


vity retained at 10'6 dil
mungbean tissue have
after mechanical inocul
studies indicated that I
ted to, and even within, t
The virus reacted si
terlony double immu
with reference Bgl
Isometric particles whicl


ion) in int
ed 10-14
ion. Host
dV was n
:Legumin
ongly in (
)diffusion
V antisi
measured '


I soils p
,common
ivation ir
eed or as
were ju
n), goose
n purslai



mted to :
y associated
the Philip
a rotation
,gle rice (
rass (Eleu
e (Portulai


yza sanva), con


initial p
the inf
test ph
t from
and no
vere sar
ial decl
n urn i


ulation of
ed soils bef
s was not
ich other. I
hizosphere
led at week
in popular
__._ j I_


L ueC cAepuun ol


ctor-viru
I motth

domona,
some oj
- H. H


ium wa:
Langes o:
natural)
ral plan
ith toma
-s, either
)p; these
rinochloa
indica),
leracea),
pinosus),
a mays)
).
olanacea-
planting
lificantly
:n rhizo-
Is of the
intervals,
n of the
types of
rlaca ole-


i mrcas ui r. sounacearum population
microscopic examinations of partially in the rhizosphere soil. The rate of
purified virus preparations stained with decline of population of the bacterium
2% neutral phosphotungstate. was lowest with P. oleracea, E. echinoch-
It differed somewhat from the type loa and E. indica, while rice and corn


rce
duration at 90-95 C) and and
ih concentration (infecti-


sA 1 ,








Abstracts of Papers


showed relatively greater suppression of
the pathogen. Latent infection by P.
solanacearum was found, through isola-
tions, in P. oleracea but not in the other
test plants.
The population of P. solanacearum
was also higher in the rhizosphere soil
than in the non-rhizosphere soils of the
weed plants. On the other hand both
corn and rice had relatively higher popu-
lation of the bacterium in non-rhizo-
sphere than rhizosphere soil.
Bioassay of the soil with wilt suscepti-
ble "Yellow Plum" tomato seedlings
gave varying results, depending on which
plant had been growing in the soil. Soil
with common purslane growing in it had
the highest level of wilting while those
soils planted to corn and rice had the
lowest incidence of wilting. In compari-
son to the bare fallow soil, soils planted
to goosegrass, jungle rice and purslane
showed non-significant differences, but
significant differences were detected for
corn, rice and.spiny amaranth.
The results of the study indicate the
possibility that common weeds found in
tomato growing areas contribute to the
survival of the bacterial wilt organism in
soil, even though host plants are absent.
These weed plants could be indigenous
reservoirs of inoculum in virgin soils or
in field soils in between planting of
susceptible crops. Control of common
purslane, goosegrass and jungle rice in
tomato areas in the Philippines could
possibly lower bacterial wilt incidence,
while the rotation of crops with either
rice or corn would aid considerably in
the reduction of the disease.
Characteristics of Pseudomonas sola-
nacearum E. F. Smith infecting banana
in the Philippines A. R. Rillo and
A. J. Quimio, UPLB
Banana isolates of P. solanacearum


from Davao del Norte and Davao City
were studied to determine their 1) host
range and pathogenicity by artificial and
natural inoculations, 2) colony morpho-
logy, 3) tyrosinase activity, 4) biovar,
5) hypersensitive reaction on tobacco
leaves, and 6) susceptibility to bacterio-
phases.
In artificial inoculations, the isolates
were virulent to commercial banana cv.
Giant Cavendish (AAA), Lakatan (AAA),
and Latundan (AAB), abaca cv. Tango-
ngon, and Heliconia latispatha Benth.;
moderately virulent to virulent to white
potato; weakly virulent to virulent to
diploid banana cv. Butuhan (BB), egg-
plant, tomato, tobacco, and castor
bean; and avirulent to ginger, peanut,
cowpea, and pepper.
In natural inoculations, the isolates
were virulent to Giant Cavendish and
heliconia but avirulent to Latundan,
Butuhan, ginger, white potato, castor
bean, peanut, eggplant, tobacco, and
pepper; some isolates were weakly to
moderately virulent to Lakatan, abaca
and tomato.
Healthy Giant Cavendish seedlings
wilted when inoculated with the isolates
through the leaf by leaf pruning; the
bacteria also invaded the fruit peduncle
when the bract scars or cushions of the
banana inflorescence were spray-inocula-
ted flowers to the rhizome causing
infection and severe wilting of the suckers.
None of the isolates caused hypersensitive
reaction on tobacco leaves.
The isolates exhibited 3 morphotypes
on Kelman's medium, the characteristics
of biovar 1, and slight tyrosinase activity
on a tyrosine containing-medium. Four
distinct phase isolates from P. solana-
cearum infecting ginger and tomato
failed to lyse the isolates.
It was concluded that the Philippine


Jan. & June 1978











P. solanacearum banana isolates, now grew normally; this was noted also in
designated as Phil strain, belong to race wet season, but, only under low inoculum
2. Its pathogenic behavior and colony concentration. For the susceptible
morphology resemble closely those of the varieties, in both seasons high wilt indices
insect-transmitted Peruvian A strain, meant high percent mortality because
There are indications based on symptoms most infected plant did not recover from
of naturally infected plants and results of the disease.
banana leaf pruning transmission and Wilt incidence was lower in the dry
Wilt incidence was lower in the dry
inflorescence spray inoculation studies than wet season. In both seasons, how-
thattha n we t season. In both seasons, how-
that it maybe spread mechanically be ..


included that
St plants. Disease severity also increased
g in the com-
of Dao del with increasing levels of inoculum.
of Davao del
serious threat .
mmnI~ nft*l'- m o-I n * fl~fl nt/T r f4-if4-s


Moko disease now existing
mercial banana plantation:
Norte and Davao City is a
to the flourishing banana i
Philippines. i
Resistance of some egg
to Pseudomonas solanac
Smith A. J. Quimio a
Tribo, UPLB.
Seven eggplant cultivars
malai Brinjai (AB), Pusa
(PPC), CA-Clustered (C/
Multiple Purple (DMP), Du
Purple (DLP), Nurumo Loi
piro were evaluated for
during the dry and wet seax
injury and stem punctur


In dry season, CA-C was rated 1
most resistant followed closely by P
and AB in both inoculation technique
In wet season, AB, CA-C and PPC has 1


inoculation; in stem punctu
AR was the most resistant


SPhilippine soils H.
. Quimio, UPLB.


H. Chan


ly directly


cearum in several naturally infested soils
lamely Anna- in the Philippines, comparing several
urple Cluster media reported to be selective for P.
C), Dingras, solanacearum. Found to be most effec-


d by Nesmith and Jenkins in
)ntaining peptone 10g, dextrose
n hydrolysate 1 g, yeast extract
to-agar 18g, K2HPO4 1.18g,


'aS rnl2ru4 u.'t+g, (rNr4)2
MgSO4 .7H20 0.2g2' MnS(
7cIr nAAAI ii r ,


V. A \,
n ~nn


aeiomsec water I liter, tetrazolium chlo-
-A cn ___ -- -1-t


appm, penicllnn g Ippm, tyroin-
ppm, vancomycin 10 ppm,
50 ppm, benomyl 500 ppm,
) ppm, pamaricin 20 ppm, poly-


PPC were as resistant as AB only at low mixm 1 1lU ppm. When pure culture
inoculum levels. DMP, DLP, Nurumo suspensions of P. solanacearum were
Long, and Sinampiro were all rated surface-plated on this medium and in-
cubated at 30C for 72 hr very fluidal
susceptible in both seasons
susceptible in both seasons distinctive colonies with bright red-
In dry season, resistant plants showed diffused formazan centers and narrow
high initial wilt symptoms as revealed by white margins were produced. No sionifi-








A ~ ...'. ~


iery of the bacterium when compared was also not significant. This seems to
those on casein-glucose-peptone or indicate that the resistance is different
razolium chloride media. The reco- but it is not in a ranking sequence. The
y of P. solanacearum on Nesmith and difference of isolates in a group was
nkins medium was found to be around significant and such difference is likely
% when a known population of the associated with the aggressiveness of the
cterium was added to field soil. With isolates in each group.
s medium, populations of P. solana- When variety IR1698 was used in the
aum were easily detected from study to evaluate group I isolates, one
turally infested field soils from Laguna, portion produced susceptible lesion and
vao del Norte, Misamis Oriental and the other resistant to moderately resis-
nguet provinces. Inhibition, by the me- tant lesions. It is likely that group I iso-
Im of other soil microorganisms ranged lates were mixtures of two virulence
>m 83% to 97%. It was concluded that groups based on the infection of IR1698.
: medium of Nesmith and Jenkins When mixed inoculum of any two iso-
uld be used in population or ecolo- lates from the same virulence group was
:al studies of P. solanacearum and tested against the differentials, no reduc-
abably for routine bacterial wilt tion in lesion size was noted. When iso-
rveillance in the field. lates from different groups were mixed
for inoculum, the lesions were signifi-
Continuous variation in virulence of cantly reduced.
inthomonas oryzae Muhammad Mach-
id and T. W Mew, IRRI. Resitance to bacterial blight in rice
T. W. Mew, R. C. Reyes, & C. M. Vera
A vertical relationship was demons- Cuz, IRRI.
Lted between the isolates of X. oryzae
the Philippines and the rice differen- Five hundred rice accessions were
ils carrying various genes for bacterial tested in the field for resistance to bac-
ight resistance. The isolates were trial blight during the dry and wet
ossified into four virulence groups, season in the last two years against
Lcept group 0 which apparently has PX061, an isolate which exemplifies a
st the virulence, specificity in infection major virulence group of Xanthomonas
different rice was distinguished. oryzae in 'the Philippines. Inoculation
The virulence or the relative ability of was carried out artificially at the maxi-
The virulence or the relative ability of
e isolates to cause blight lesions within mum tillering and at booting stages of the
e isolates to cause blight lesions within
e virulence groups I and II, the two plant.
ajor groups of the bacterium occurring "Resistance of the rice was classified
this country at present, were further into five distinct groups. Group I rice
aluated. The variation in virulence was was resistant at both stages. Reaction of
und to be continuous and the host- some rice in the group varied according
irasite relationship was horizontal, to leaf positions. Young leaves were
within each group, the virulence could be more susceptible than the older ones.
nked from high to low over the dif- About 38% of rice in the group was re-
rentials. Statistically the interaction sistant to another isolate (IRN167)
__- L__. .... t .... .... A4 hl inlnn tn o -liff-r-nt virlll nlP O%11mll










at Nueva Ecija. Groups II & III were kresek. This information seems to indi-
rated moderately resistant and moderate- cate that resistance to kresek is expressed
ly susceptible, respectively at both not only between varieties but also within
stages. Only 5 out of 63 varieties in varieties at different ages.
Group II and 4 out of 36 in Group III When IR8, IR20, IR1545 and DV85
maintained the same rating when tested of varying resistance or susceptibility to
against IRN167. Four of the Group III bacterial blight were tested by the root-
rice became resistant and moderately dip method of insulation for kresek
resistant to isolate IRN167. In Group IV infection, DV85 showed lesser kresek
rice, the rating for resistance was shown than the other three. It is also apparent
with considerable variation at the two that the former three varieties also dif-
stages. Some rice in the group was sus- ferred in their response to kresek at
ceptible at the maximum tillering stage different inoculum concentrations of
while resistant at the booting stage. isolate PXO82.
A major gene Xa6 for resistance has been
identified in few of the rice, e.g. Senith, Fourteen varieties were then evaluated
Malagkit Sungsong. The resistance seems in the greenhouse and in the field for
to resemble adult plant resistance and their reaction to kresek against two
the reaction at this stage was stable. isolates, PX061 and PX082. All other
There were other rice in Group IV varieties except IR8, India Dular, Ketan
that was resistant at the maximum Lumbu, and Lakshmi Digha, varied in
tillering stage and susceptible at the reaction to the two isolates. India Dular
booting stage, but the reaction varies in remained resistant and IR8 susceptible.
accordance to Incatinn Reaction of these varieties to the three


Varietal reaction to kresek infection -
T. W. Mew, C. M. Vera Cuz, and R. (
Reyes, IRRI.
The kresek symptoms of the bacteria
blight syndrome is normally observed i
nature from 1 to 4 weeks after tran=
planting. Under experimental condition:
young seedlings produce more krese
plants than older ones.
IR8 which has no gene for bacteria
blight resistance and IR1545 which ha
a recessive gene (xa5) were selected fc
further evaluation of response to krese.
infection at different ages. The metho
of inoculation was by root-dip in ba<
trial suspension for 5 min. The result
suggested that IR8 was susceptible a
all ages ranging from 9 to 32 days afte
seeding (DAS). At 9 and 16 DAS
IR1545 was suscpetible as IR8, bu
at 23 and 32 DAS, 54% and 8% re!
pectively of the plants were infected wit,


r lulppuint ulainntiut r ups ul A uryu,
for leaf blight indicated that the reaction:
to kresek among these varieties was no
correlated with the reaction to leaf blighi
Ranking of these varieties based oi
their over-all reaction to the two isolate
in the greenhouse suggested that Indi
Dular was the most resistant, while th
susceptible check IR8 ranked 13th o
the 14 varieties. Ranking of these varic
ties in the field test by the same method
of inoculation showed India Dula
as the most resistant while IR8 the mos
susceptible. In the greenhouse the scoring
was done 3 weeks after insulation i
contrast to 6 weeks in the field.
A scoring system based on the num
ber of plants and time of scoring
expressed as days after inoculation wa
formulated. For practical purposes, 6(
to 80 plants scored from 21 to 24 day
after inoculation seems adequate fo








A ha.4-f.a nf Pano


snhouse test to show varietal differ- I
es.
Control ofnematodes with Temik 15G s
ticarb and its residue in the fruit of 1
nt Cavendish Banana M. O. San
n and Lorna D. Lozano, Twin Rivers
search Center. 1
Temik 15G Aldicarb, a systemic pesti-
e for the control of insects, mites, and
natode was experimentally tested for
natode control on Giant Cavendish
iana plants. Results showed that
rates of 10,20 and 45 grams of the
sticide provided control of plant pa-
itic nematodes particularly the Rado-
)lus similis, Meloidogyne incognita,


)ur and Kentucky 16 for burley; and
ka and Samsoum for Turkish. Results
wed that the combined effects of
olanacearum and M. incognita were
:h more serious than either alone.
number of wilted (dead) plants due
simultaneous inoculations of the
togens and inoculation with the
latode first then followed by the
terium or vice versa one week later
e higher by 10% to 70%, depending
the cultivar, than due to the bac-
im alone. In all cultivars, the rate
bacterial wilt development in plants
;ulated with the two pathogens was
consistently faster and significantly


ticularly noticeable in cultivars which
The rate of 10 grams (1.5 gm. ai.) left showed an appreciable degree of resis-
idue in the fruit pulp ranging from tance to P. solanacearum. None of the
0.02 to 0.04 ppm; 20 grams (3 gms. plants artificially inoculated or grown
) with < 0.02 to 0.08 ppm; and 45 in soil naturally infested with the nema-
ms (6.75 gms. ai.) with residues tode alone died of wilt.
Iging from < 0.02 to 0.35 ppm. From
=se and other data submitted, the Bacterial wilt root-knot interaction
vironmental Protection Agency set a complex in wilt-resistant Philippine to-
erance residue of 0.3 ppm for the matoes C. M. Napiere and A. J. Quimio,
licarb (2-methyl-2-(methythio) propio- UPLB.
ldehyde) (methylcarbamoyl) oxyme.
Increased bunch weight, as expected The effects of Pseudomonas solana-


L oeen notea wimm a year aiter me
t nematicide application.
Root-knot effects on bacterial wilt of
ilippine grown tobacco A. J. Quimio
SMarcelinaR. Villacorta, UPLB.
The effects of root knot nematodes,
loidogyne incognita, on the develop-
:nt of bacterial wilt caused by Pseudo-
nas solanacearum, on tobacco were
died using artificial and natural ino-
lation methods. The following tobacco
Itivars were used: Dixie Bright 101,
ant rneldcr and NCRY fnr Virginia:


1 their interaction on the develop-
nt and severity of bacterial wilt on
nato were studied using six resistant
tivars namely 1169, Pope, Marikit,
48-1, VC 11-1, and Venus, and
o wilt-susceptible cultivars 2029 and
Ilow plum.
On natural inoculation experiment,
t-susceptible tomato CV 2029 plants
wn in both soils infested with P.
inacearum and P. solanacearum-M.
ognita combination started to die of
t one week after transolantine: all


4 4











wilt-resistant cultivars. 1169. Pope,
48-1 and Marikit, wilted (dead) plants
soils infested with both the bacteria
and the nematode occurred 1, 1, 2, an
weeks respectively, earlier than th
plants grown in the bacterium infes
soil alone. At the termination of
experiment (after the last harvesting
fruits) per cent mortality of plants gro
in soils infested with both pathog
was higher than those plants grown
soils infested with the bacterium al
by 42.9, 75.0, 73.4, and 72.7% for (


inoculated with the bacterium alo
The percentage mortality also increg
S with increasing nematode population
the bacterium-infested soil.
It was concluded that the presence
M. incognita enhanced the develop
and severity of bacterial wilt and lowe
the yield significantly of wilt-resist
tomato cultivars.
Pre-harvest anthracnose control
mango N. S. Mendoza, Union Carb
Phils.


grown m sous Inmestea wtn oacterium- oua ireaK at truit setting and U1 da)
nematode combination was lower than before harvest gave good pre-harve:
those plants grown in soils infested with control of anthracnose (Colletotrichw
\the bacterium alone by 23.0, 20.0, 15.0 gloeosporioides) in mango. During tt
and 13.3% respectively, rainy months, more fruits developed pi
inflorescence with fungicide treatment
Similar results were obtained in ex- Application rate at 2.64 g/liter of Dacon
periments involving artificial inoculations 75WP, Dithane M45 and Manzate 8
with P. solanacearum and M. incognita 10 days before harvest gave good ai
alone or in combination. Depending on thracnose control, eliminating laboriot
the variety, disease development was postharvest treatment.


Ltea wnn me Dacterium ana me nema-
tho Philinninov di A i;


UaVt iui.U11 iul. lII WIILI-VaI BLillL LUILIVi
per cent mortality in plants inoculat,
with the bacterium-nematode combir
tion was higher than those inoculate
with the bacterium alone by 50 to 85'
The combined effects of the two path
gens lowered the yields by 16.5 to 23.
compared to those inoculated with tl
bacterium alone. None of the plan
inoculated or grown in naturally
_tr,;~n;l., ;nn^ r l~ nrr;! ...:J. -


virus diseases or me potato caus
huge economic losses and remain a there
to the potato industry of the Philippini
They are the major causes of the pi
vailing low yields and the rapid deger
ration of seed pieces. Field infectii
Ranges from 30 to 60 per cent depend
upon the virus strains and variety.
Among the most prevalent vir
diseases observed in the field are the le


Various heat treatments for aflatoxin
Results of experiment using varying decontamination in mungbean (Vigna
levels of inoculum in both artificial and radiata (L) Wilczek) Rosalinda A. Perez
natural inoculations showed that devel- and Lina L. Ilag, UPLB.
opment of bacterial wilt was consistently Various heat treatments such as








A 1-4- P- ,-


eating were tested to determine their 3 days incubation in mungbean seed
effects on reducing or eliminating the decoction agar decoctionn from 80 g
flatoxin content of contaminated mung- dried mungbean seeds; agar, 17 g; distilled
ean seeds. water, to make 1 liter) and in oatmeal-
High-temperature treatments greatly mungbean leaf extract agar (extract from
educed the aflatoxin content of mung- 37 g homogenized mungbean leaves;
ean seeds that were previously inoculated oatmeal, 70 g; agar, 17 g; water, to make
.n 1 liter).
rith an aflatoxin-forming strain of liter).
spergillus flavus. As the temperatures
nd heating periods were increased, Cercospora sp. from cowpea spo-
orresponding reductions in aflatoxin related best after 6 days of incubation
content were observed, in V-8 juice agar (V-8 juice, 200 ml;
CaCO3, 3 g; agar, 17 g; water to make 1
Boiling substantially reduced the afla- liter) and in cowpea leaf extract-oatmeal
oxin content and totally eliminated the
oxin when the boiling period was exten-
cowpea leaves; agar, 17 g; oatmeal, 70 g;
ed to 3 hours. Autoclaving in combina- water to make liter).
on with boiling was a more effective
econtamination procedure than mere Preliminary studies on sunflower
oiling. Oven-dry heat treatments also diseases in Central Luzon G. G. Nueda,
educed or eliminated the toxin. The CLSU.
extent of decontamination depended Results of the three-cropping season
n the particular combination of tempe- (October, 1976 to February, 1978)
iture and length of heat treatments to survey on the diseases affecting sunflower
whichh the beans were exposed. Moist plants in Central Luzon State University
eat also appeared to be more effective farms showed that the diseases included
ian dry heat. sclerotium wilt (Sclerotium rolfsii Sacc.).
Complete elimination of aflatoxin was seedling damping-off (Pythium debarya-
bserved in the following treatments: num Hesse), bacterial wilt (Pseudomonas
i) boiling (100 C) for 3 hours, (b) solanacearum E. F. Sm), root and stem
utoclaving 121 C for 15 min then boiling rot WOonilia fructigena Sacc), head rot
or 2 hours, (c) oven-heating at 200 C (Rhizopus arrhizus Fisher), shoot blight
or 3 hours, and (d) at 220 and 250 C in of seedlings [Pseudomonas helianthi
ie oven for 30 minutes. Samples which (Kaw) Say. ], bacterial leaf blight [Pseu-
rere heated over 200 C were scorched domonas helianthi (Kaw) Say. ], rust
nd considered unfit for consumption. (Puccinia helianthi Schw.), leaf spot
ICurvularia lunata (Wakker) Rnediin Car-


n








14 rnulppmie rnyopamnouogy vol. 14

only observed in November, 1977 to Field evaluation of some fungicides for
January, 1978 plantings and was never the control of sheath blight of rice B.
observed during the previous seasons. A. Estrada and L. M. Sanchez, IRRI.
Partial control on sclerotium wilt was The efficacy of some selected fungi.
attained using Brassicol (PCNB, 70%) at cides for the control of sheath blight ol
2-5 tbsp. per 5 gallons of water as seed rice was studied in two separate trials
treatment and head rot with SN 66 752 during the wet season of 1977 on a sus.
at 0.075% concentration injected on the eptible line IR1317-392-1-2. The
back of sunflower head where infection chemicals tested were Iprodione 50 WF
starts. The effect of SN 66 752 onRhizo- [1 isopropylcarbamoy] -3 (3.5 dichlo.
pus arrhizus was fungistatic. ropheynyl) hydantoin; Iprodione SC
Selection of mycorrhizal strains for FLOW [1-isopropylcarbamoy] -3 -
biological control of Fusarim solani f sp. (3.5 dichlorophenyl) hydantoin]
phaseoli (Burkh.) Snyder and Hansen on Chlorothalonil (tetrachloroisophthaloni-
cowpea [Vigna sinensis (Torner) Savi] trile, 54%); Polyoxine (Polyoxine com-
Severina N. Baradas and P. M. Halos, plex-B, 80%; Bihypyzi [bis (1-hydroxy-2
UPLB (IH) pyredidinethionate) zinc,
48% ); Bendiphos (0, 0-Diisopropyl-
Strains of mycorrhizal fungi were S-benzylthiophosphate, 48% ); Mebeni]
assayed for stimulatory influence on (0-methyl-benzanilide 0 toluic anilide,
growth and protective effect against 75%); Validamycin A., (3%) and EDPD
Fusarium dry root rot of cowpea. These Thalide (0-Ethyl-S, S-diphenyl phospho-
strains were collected .from fifty differ- rodithiolate (15% ) + 4, 5, 6, 7 -
ent roots and rhizosphere sofls oflegumi- Tetrachlorophthalide, 20% ) at the rates
nous, solanaceous, orchard and planta- of 3.0 kg, 3.0 1, 1.7 kg, 1.0, 1.04 1,
tion crops, and pasture grasses. The 1.25, 1.0 kg, 1.5 1 and 1.0 kg per hec-
effect on growth rates and incidence of tare, respectively.
infection were analyzed using the two- ,,, ,,,, ha +, l+


variance. It was difficult to select si
ficantly promising strains due to the ef
of soil and strain interclose interac
on growth increment and infect
However, on close analysis of the res
using Duncan's multiple range
revealed the eleven promising my
rhizal strains, namely, those der
from tomato (Lycopersicon esculenta
sweet pepper (Capsicum annum),
(Oryza sativa), soybean (Glycine mn
pigeon pea (Cajanus cajani), co
(Coffea robusta), ginger (Zingiber
ficinale) guinea grass (Panicum m


UUUIf.1 OLa5. at uivi w%,V IILL IVLu wi
one spray before and three sprays aft
inoculation.
In the first trial, the two formulation
of Iprodione wettablee and flowab
cream) excelled all other chemicals
terms of effectiveness, with respect,
disease reading of 2.6 and 3.2 cor
pared to the control with 7.7 (rate
on a scale of 0-9; 0 = no disease observe
9 = all leaves dead and a correspondii
increase in yield of 28.2 and 33.1%.

The same degree of control w;


lus), lima bean (Phaseolus lunatus) and Iprodione formulations in the second tria
mungbean (Phaseolus radiatus). with corresponding disease readings o











* Ualfu .'.i t4JwIiip.af. LuJ i .S Svti tZL
ntrol. Increase in yield was approxi- I
itely 32.3 and 32.0 per cent over the
treated plots.
Multiline trials for rice blast control -
H. Ou, F. L. Nuque and J. M. Bandong,
RL.
The use of multiline as a method of
ist control was undertaken to investi-
te its effect. Two trials consisting
5 blast resistant nearly-isogenic lines
ch, respectively, with IR8 as common
ckground were planted separately and
combination of equal proportion in
x 8 m and 2 x 2 m upland plot using
Itivar IR8 as check to compare the in-
:tion rate (r) and total disease deve-
pment. They were inoculated at the 1
nter of each plot using an isolate in-
;ting all the lines and IR8 and were
vered with plastic sheets to maintain
editions favorable for disease develop-
ent. 1
The disease incidence (percentage leaf
,a infected) was obtained by counting
ions of 10 seedlings randomly selected
)m each row at 3-4 days intervals. The
sults showed that in the first trial
3273(E) IR3259(C), IR9559(D) had
die disease. In the second trial no
;ease or negligible disease develop-
:nts were likewise observed on IR4547
), IR9559(F), IR3273(G) and IR3259
). Lines IR9669(B), IR87/Dawn(A) for
e first trial and IR9637(A) for the i
:ond trial and considerable had disease
velopments. The multilines had consi- 1
rably reduced the amount of terminal
;ease as exhibited by the disease index
blues for IR8 (44.1%) and for multilines
3CDEF (14.4%) and ABCDE (9.8%) in
e first trial. Similar results were ob-
ned in the second trial for IR8 (54.4%) 1
d for multilines ABDEFG (9.4%) and
I W1 (< OQcY- Th rpnilte nhtainprl in


ies the percentage leaf area infected
is transformed into logits and the
te of infection was computed using
gression coefficient. The theoretical
lues of terminal disease development
Iculated by mathematical addition of
rcentage leaf area infected and divided
the number of lines involved did not
ow any considerably beneficial effect
multilines over the single lines in both
als. The results in both experiments
owed no marked differences in rate
infection among isolines and multi-
es indicating that there was no re-
irkable relationship between infection
te (r) and percentage susceptible
sues. Furthermore, the results suggest
at multilines did not reduce consider-
ly the rate of total disease develop-
ent.
The data on the yield of the single
es, multilines and IR8 of the first
al were statistically analyzed and
pificant differences between treat-
ents and the control (IR8) were found.
nong single lines results obtained are
Ohly significant. However, among single
es Vs multilines and among multilines
: results are not significant indicating
it no advantageous effect was de-
)nstrated by the multilines over single
es under the test conditions.

Perfect stage of Rhynchosporium
yzae causing leaf scald disease of rice -
H. Ou, F. L. Nuque T. I. Vergel de
os and S. P. Ebron, IRRI.

The perfect stage of Rhynchosporium
yzae Hashioka and Ikegami, the patho-
n of rice leaf scald disease, was com-
only found on diseased leaves in the
lilippines and it was also recently
served on specimens from Columbia
Ad Costa Rica. The perithecia were em-


CYDYIP~YO VI Ut~~


L U








10l~y '' .J Y~Y"'6


were cylindrical to clubshaped, ofte
slightly curved. The ascospores wei
obliquely uniseriate, partly biseriatb
elliptical or fusoid, colorless, 3-septal
occasionally with 4 septa, size rang(
from 9-27 x 2.5 um. These character.
tics agree with those of Metasphaeri
albescens von Thuemen reported in 188
and redescribed by Wei in 1934. Th
perfect stage with identical morpholog
and cultural characteristics has also bee
found by several workers in Japan an
Korea. The perfect stage had been give
different generic names by variot
workers thus creating taxonomical coi
fusion. Since the first description of th
perfect stage was done by von Thueme
and the characteristics described hei
and elsewhere agreed very closely wit
the first report, it is proposed, therefore
that the pathogen, be called M. albe
cens von Thuemen with a conidial stall
R. oryzae.
The pathogen caused varied sympton
that led to the description of foi
diseases in Japan. The diseases brow
leaf blight, leaf scald, leaf tip blight, an
leaf sheath browning are caused by ti
same pathogen leaf tip drying disea,
caused by M. albescens and similar to lei
tip of Japan was described earlier i
China. Since the term "leaf scald" has bee
internationally used, it is suggested tt
same common name be used, to encon
pass all the different common nami
mentioned above.
Antagonism of some bacterial isolat
to the sclerotia of Rhizoctonia sola
causing sheath blight of rice S. H. C
and T. I Vergel de Dis, IRRI.
The primary sources of inoculu
of sheath blight disease of rice are scler
tia left in the field from the previol
crop. Proper management through tl


sclerotia.
In an attempt to study the harnessin
of microorganisms against sclerotia, dil
ferent bacteria were isolated from wate
and sclerotia collected from the field
Most bacteria isolated from the scleroti
had shown stronger antagonistic activity:
against the fungus than those isolate
from field water.
To test the effect of specific bacteria;
isolates on sclerotial survival, the lab(
ratory-cultured sclerotia were introduce
into tubes of peptone-sucrose brot
and in sterile water containing there
isolates with the strongly antagoni
tic activity and kept under room ren
perature. The survival was tested b
removing the sclerotia from the bacteriL
suspension and culturing them on PD,
plates after 3, 10 and 17 days. Th
results showed that sclerotial survive;
tests from the sterile water were erratic
However, results from the bacteria;
suspension in peptone-sucrsoe brot
showed that the sclerotia grew slowly
after 10 days and were no longer viabi
after 17 days in all the three bacteria
isolates.
To increase bacterial population an
their antagonistic activity, a preliminary
test of adding rice straw to field watt
was made in the laboratory. The result
demonstrated that bacterial population
increased 100 fold a few days afti
adding rice straw to field water, suggest
ing that rice straw favorably influent
bacterial growth.
The preliminary results seem to t
encouraging, but practical use in tt
field requires much more work.
Field and storage molds of rice -
H. Ou and T. I. Vergel de Dios, IRl
S Six hundred samples were analyze


10O













-- -- 1
tant or
e rice
qlleefpn-


uIV uly
y each
dry and
Sdetert
irious ft
vet seas'
nths sto
i rice (0
of field
ot pene
is, abou
out glul
harvest.
outside
oconis


. I In resuuis Mcumu tible var
field molds are more nrndned


and Fanny
pt to a few


tillus and Penicillium spp. re- ferent levels of horizontal resistance. We,
Mdly increased during storage as however, prefer to interpret this result
n by noate tests. This indicates .. . .^ +... .. ....-,








. iiulJ.ppiuc 1 1 Arvya..Y .uvlun j


4 alllrG ~Jl rlI U 1 I I l Jl U ; 1y i L
inoculation. The present experiment I
sures the resistance in disease deve
ment in multicycles in the field.
The same 8 cultivars were plante.
an upland plot at IRRI. Three isol
with different degree of virulence \
separately inoculated to each block
sisting of 9 plots. Each isolate was
culated to a row of seedlings in a l
Each plot was covered with plastic s
every night to insure infection.
number of lesions were counted f
plants selected randomly. Counting
lesions was done every 4 days and
apparent infection rate (r) on
cultivar was calculated.
In the resistant group (Tetep, Di
Sensho), the disease developed
slowly and there were much less dis
60 days after sowing. The interme
group (NP-125 and Dular) had also
rate of development. The suscep
group had high rate of disease deve
ment.
Studies on the expression of resist
in an infection cycle showed that T
has the smallest size of lesion with s
number of spores. Dular and NP-125
have relatively small lesions with
smallest number of spores while'.
has the biggest lesions with large am<
of spores produced. Lesions on
enlarge faster compared with the o
varieties. The slow rate of disease di
opment might be due to accumul
effect of small number of lesions, s
size of lesions and small amount of sp
produced per unit lesion area. T
components of resistance are b


a- duced from abroad through the Geni
p- Resource Center, UPLB-CA were eva]
ted for their resistance to 3 foliar
in seases, namely: rust, Helminthospori
es and Phyllachora leaf spots. Screer
re was conducted either in small gar
n- plots or in the field or both. Artifi
o- inoculation was used for Helminti
it. porium leaf spot while screening
et rust and Phyllachora depended on nati
e inoculum in the field.
m Of the total 343 lines and varie
of evaluated for resistance to rust, ab
ie 148 showed resistance. On the ol
:h hand, 13 of 119 materials showed
distance ot Helminthosporium leaf sI
n, However, none of the above matei
S showed any resistance to Phyllack
se leaf spot.
t It is apparent from the result of
w study that while resistance to rust
l Helminthosporium leaf spot is rea
p- available in sorghum germplasm, re
tance to Phyllachora is difficult to f
:e Possibly, resistance to the latter dis
sp is quantitatively manifested and has
ll been detected by qualitative mea:
so such as lesion type.
ie Chemical control of neck blast
ta Cercospora leaf spot diseases of ric
at F. L. Nuque and J. M. Bandong, II
ta
er Three conventional fungicides name
1l- EDPD Thalide (0-Ethyl-S, S-diphe
:d phosporo dithiolate (15%) + 4, 5,
ll 7-Tetrachlorophthalide, 20%), chloro
es lonil (Tetrachloroisophthalonitrile, 5,
se mancozeb (a coordination product
g zinc ion and manganese ethylene bi


--v









Abstracts of Paners


It


eight or volume of the product in the upper surface of the leaves.
sprayed twice one at early flower- Several lesions may coalesce in the later
stage and the other, one week stages of the disease forming a thin
after. The percentage of neck blast stripe. Symptoms were observed both on















ly miecrea plants aexonatea earlier (] to
Studies were conducted to assess 10 days ahead than the control) and
quantitatively the effect of different caused pod abortion, reduced seed size,
levels of rust infection in the different reduced seed number per pod and re-
growth stages of soybean plants: 1) duced overall seed weight.
onset of flowering, 2) early pod form- It could be concluded from the
ation and 3) pod filling. studies that the two trials conducted
Inoculum with different concentra- showed similar results in that photo-
tions were inoculated in the soybean synthetic activity of soybean was ag-
plants when flower buds started to gravated by minimum rust infection
develop. Three harvests for dry matter at any stage of growth and as the degree


re were


that maximized dry matter production. Field
On the other hand increase in dry weight the contr
of the rust infected plants were governed pachyrhiz
to a certain extent by the degree of in- irrr


nation of five fungicides on
if soybean rust (Phakopsora
yd.) Cecilia V. Bautista,


fiction, the total dry matter production a AV uuku V I vlnIu DUC,
ec te e Dithane M45, Manzate, Parzate and
Fermate) applied at standard recom-
The net assimilation rate (NAR) mended rates were evaluated for the
which was employed for the direct control ofPhakopsorapachyrhiziSyd on
measurement of the changes in plant soybeans.
growth was significantly affected by Results showed that all fungicides
rust population. Generally, lesser rust tested significantly reduced rust infesta-
pustules give higher NAR value and tion. Vitigram Blue (50% copper oxy-
heavier rust pustules resulted to lesser chloride) was the most effective fnllnwar


- owth parai
rate (CGR)
rate (RGR)


AuMiuu v--ar ko w/O UUIUU
:t of zinc ion and manganese
is-dithiocarbamate), Manzatt
nese ethylene bis-dithiocarbai
e (65% zine fthvltnP hi.e-


ne mgner me degree or mtection at the caroamare) ana rermate


*J-1J^.. *-..



























































"Yellow Dwarf" and This study was aimed to fii







22 Philippine Phytopathology Vol. 14

to control fruit rots in transit and storage. fruits after 12 days of storage in both
"Carabao" mangoes were used in experi- perforated plastic bags and corrugated
ments conducted in Bulacan and at U. P. cartons. The use of perforated plastic
at Los Bafios. container was noted to serve as a ripening
Results showed that mature fruits retardant thus prolonging the shelf life of
that have been clip-harvested, washed or fruit in storage.
clorox swabbed and placed in either per- Retaining fruit pedicels clipped to
forated plastic bags or corrugated cartons about 1 cm to their bases appears to be a
were less affected by rots. promising complementary measure to
Clipping the pedicels of fruits about chemical treatment in reducing stem-end
1 11 4- +, +h;. ...ono f-.. fnA +,1 ha rot incidence.


Fruits placed in perforated plastic Two sets of experiments were conduc-
bags were wrinkle-free and of better ted to assess the protective effect of
market quality, different non-mercurial fungicides against
Postharvest treatments of mango pineapple disease of sugarcane. Phil. 58260
fruit to control rots in transit and storage cane points were treated in various levels
- A. N. Pordesimo and J. M. Dangan, of the fungicides at different dipping
UPLB. times, air-dried and then inoculated with
A study on postharvest control of seven-day old agar culture of Ceratocystis
mango fruit rots in transit and storage paradoxa for 15 minutes.
showed that anthracnose caused by Hoe 17411 50 WP at 15 g/10 li water
Colletotrichum gloeosporioides Penz., used as dip treatment for 5, 10 and 20
and stem-end rot caused by Diplodia minutes was the most effective. The cane
natalensis Pole-Evans were the more points treated with the fungicide have
common fruit rots observed. Significant higher percent germination compared
reduction in fruit rots was noted in with the two other formulations of Pya-
fruits dipped in TBZ at 135 g per 100 carbolid.
liters for 5 min.
In the second set of experiment,
TBZ was superior to Maneb + Be- Labilite and Daconil at 40-55 g/5 gal
nomyl, Mancozeb, and Cupravit in water and 65 g/5 gal water, respectively,
controlling storage rots as reflected in can control the disease.
more number of clean and marketable









Received for publication: 22 February, 1978




FUNGICIDAL ACTIVITY OF CRUDE PLANT EXTRACTS
AGAINST HELMINTHOSPORIUM OR YZAE

D. B. LAPIS and EDNA E. DUMANCAS

Associate Professor, Department of Plant Pathology, College of Agriculture, U. P.
at Los Banos and former graduate student, now Instructor at Siliman University, Duma-
guete City.
Portion of thesis of the junior author presented for an MS degree in Plant Pathology,
College of Agriculture, U.P. at Los Banos.

ABSTRACT

With water as the solvent, extracts from 93 plants were screened for
activity against Helminthosporium oryzae, the causal organism of brown spot
of rice. Of these, 42 inhibited the growth of the test fungus.
Of the 42 extracts from the different plants, nine were further assayed
to test the activity of the different dilutions, the supernatant and the preci-
pitate, and the effect of time. In most cases, the extracts were active at 1:10
aqueous solutions except for Euphorbia pulcherrima which was active only
in its concentrated form. The precipitate was significantly more active than
the supernatant with the exception of Pseudocalyma alliaceum and Allium
sativum wherein their supernatant liquid were significantly more active than
their precipitates. Generally, the activity of the different extracts decreased
with time.
Impatients balsamina, P. allieaceum, A. sativum and Tagetes erecta ex-
tracts were further evaluated in vivo, to assay their activity as protectant,
therapeutant, or both, and the effect of time intervals of spraying on their
activity.
I. balsamina was found active as a therapeutant and protectant while
the other three test plants were found to be effective as therapeutant only.

The acceptance of fungicides as an ag- Until recently, the organic and inor-
ricultural input is greatly affected by ganic fungicides are in wide application.
several factors. Foremost is the cost/bene- These fungicides which are generally
fit factor.. The cost of these chemicals used by crop producers often leave toxic
has tremendously increased to become residues to the soil, water, atmosphere
prohibitive to small farmers; such farmers and most especially on plants and plant
can not gamble on chemicals unless it products eaten by animals and men.
is guaranteed that such venture is profi- This is pollution and positively endangers
table, the lives of mankind.
The availability of recommended fun-
gicides is another factor to be consi- The cost, availability, environmental
dered. Due to the increase in demand pollution and hazard to human and
for these chemicals, availability is some- animal health posed by the aforemen-
what affected. In some instances they tioned group of fungicides are good
are not readily available in the market. bases for an investigation of other



23








Philippine Phytopathology


groups of fungicides. A search for fungi-
cides that are relatively cheaper,
readily available and will not cause
pollution is necessary.
The presence of fungicidal substances
within the host plants has been suggested
from time to time as a basis for resistance
against invading pathogens (Pordesimo
and Ilag, 1976). There are two groups
of fungitoxic substances present in
plants. One is the so-called phytoalexin
which are produced as a defense me-
chanism only upon the invasion of
pathogens. The other group is composed
of substances that are inherent and/or
present in the plant with or without
pathogen. The main focus of the present
study is on the search for such naturally
occurring anti-fungal substances from
higher plants on the latter group.
This study was conducted to survey,
collect and evaluate the fungicidal activi-
ty of some plant extracts against Helmin-
thosporium oryzae Breda de Haan
through in vitro and in vivo test.

MATERIALS AND METHODS
Survey and selection of plant materials
Survey was made through personal
interviews and literature review. The
selection of the plants was based on
reports and scientific findings with
special reference to local species.
In vitro screening of plant materials
Extracts from 93 plants were screened
for their activity against H. oryzae (Table
1).
Preparation of the crude extract. Plant
materials collected the day before were in-
dividually placed in plastic bags and
stored in the refrigerator. In most cases
only the leaves were utilized. However,
some plant materials included flowers
and stems. In the case of garlic, gardenia


and ginger, cloves, flowers and rhizomes
were used, respectively.
Twenty grams of the plant materials
were used in preparing the crude extract.
The weighed plant materials were washed
with 10% chlorox after which they were
rinsed three times with sterile distilled
water. The washed plant materials were
placed in a sterilized Waring blendor and
40 ml of sterile distilled water was added.
They were homogenized until homoge-
nous mixture was obtained. The homo-
genous mixture was filtered through a
double layer of sterilized gauge using
sterilized 250 ml erlenmeyer flask as
receiver. The filtrate served as the crude
concentrated extracts.
Assay of effectivity. The effectivity
of the extracts from each plant material
was assayed using the procedure described
by Sharvelle (1961). Bio-assay disc im-
pregnated with sterile distilled water
served as control.
The plates were incubated at room
temperature and zone of inhibition was
measured 2, 4 and 8 days after weeding.
Dilution assay. Crude extracts found
promising from the above experiment
were assayed for their effectivity at dif-
ferent dilutions. Ratios of 1:10, 1:100
and 1:1000 dilutions were prepared from
the concentrated crude extracts. All
concentrations including the concentrated
crude extracts were assayed separately
following the same procedure in the
screening of plant materials.
Further assay of promising extracts.
Further assay of the promising extracts
was conducted simultaneously with the
assay of the different dilitions. The crude
sap was centrifuged for 10-15 minutes at
1400 to 1500 rpm. The supernatant
liquid was separated from the precipitate
by decantation. The precipitate was dilu-
ted with 2-5 ml of distilled water. The


Vol. 14











Table Plant materials screened for fungicidal activity.


Family Scientific name Common ni

Acanthacea Asystasia gangetica (L.) T. Anders asistacia
Barleria cristata L. violeta
Gmptophyllum pictum (L.) Griff. barabas or
atay-atay

Alliaceae Allium cepa L. onion
A. sativum L. garlic
Amaranthaceae Amaranthus spinosus L. uray
Aerva lanata (L.) Juss. apug-apugan
Anacardiaceae Mangifera indica L. mango
Annonaceae Anona squamosa L. atis
Apocyanaceae Catharanthus roseus (L.) Den. sitsirika
Nerium indicum Mill adelpha
Plumiera acuminata kalachuchi
Tabernaemontana pandacaqui pandakaki
Araceae Amorphophallus campanulatus Roxb. pungapong
Balsaminaceae Impatiens balsamina L. kamantigue
Balsellaceae Basella rubra L. alugbati
Begoniaceae Begonia sp. begonia
Bignoniaceae Crescentia cujete L. calabah
Pseudocalymna alliaceum (Lam.) garlic vine
Sandwitz
Tecoma stands (L.) HB K yellow alder
Bombacaceae Cieba Pentandra (L.) Gaerta. kapok
Boraginaceae Symphytum officinale L. comfrey
Caesalpinaceae Cassia alta L. acapulco
Piliostigma malabaricum (Roxb.)
Benth alibangbang
Caricaceae Carica papaya L. papaya
Commelinaceae Zebrina pendula Schmizi wandering d4
Compositae Ageratum conzoides L. bulak-manol
(Asteraceae) Artemisia vulgaria L. damong-mar
Blumea balsamifera D.C. Prodr. sambong
Tagetes erecta L. marigold
Wedelia tribobata (L.) Hitche. wedelia
Concalvulaceae Ipomoea aquatica Forks. kangkong
I. batatas (L.) Lamk. camote
Grassulaceae Bryophyllum pinnatum Kurz. kataka-taka
Cruciferae Brassica integrifolia O.E. Schulz. mustard
(Brassicaceae) B. oleracea var. capitata L. cabbage
Raphanus sativus L. radish








26 Philippine Phytopathology Vol. 14




Cucurbitaceae Cynnopetalus chinenses (Lour.) Merr. tamleng
Memordica charantia L. ampalaya
Elaeocarpaceae Muntingis calabura L. datiles
Erythroxylaceae Erythroxylum coca Lam. cocaine
Euphorbiaceae Antidesma pentandrum Blanco Merr. binayoyong pugo
Euphorbia hirta L. golindrina
E. pulcherrima Willd. poinsettia
J. podagrica Hook gout plant
Jatopha gossypifolia L. tua-tua or digo
Manihot esculenta Crantz. kamoteng-kahoy
Mallotus philippinensis (Lam.) Muel Arg. banato
Pedilanthus tithymaloides var. variegatus
(L.) Poit lady's slipper
Graminae Cymbopogon citratus (D.C. ex nees.) Stapt. tanglad
(Poaceae)
Labiatae Coleus blumei Benth. mayana
(Lamiaceae) Mentha arvensis L. herba buena
Pogostemon cablin (Blco.) Benth kablin
Lythraceae Lagertreoemia speciosa (L.) Pers. banaba
Malvaceae Hibiscus esculentus L. okra
H. rosa-sinensis L. gumamela
Mimosaceae Samanea saman (Jacq.) Merr. akasia
Musaceae Musa x paradisiaca L. (ripe) latundan
Musa x paradisiaca L. (unripe) lat-ndan
Myrtaceae Psidium guajava L. guava
Myotaginaceae Mirabilisjalapa L. alas cuatro
Oleaceae Jasminum multiflorum L. sampaguitang-
sungsong
Oxalidaceae Averrhoea bilimbi L. kamias
Pandanaceae Pandanus odoraticsimus L. f. pandan
Papilionaceae Derris elliptica (Roxb.) Benth, tubli
Dolichus lablab L. batao
Erythrina variegata L. dap-dap
Piper lalot D. C. lalot
Plumbaginaceae Plumbago indica L. laurel
P. auriculata Lam. Plumbagin-
bughaw
Portulaceae Portulaca oleracea L. kulasiman
Talinum triagulare (Jacq.) Gaertn. talinum
Rubiceae Gardenis jasminoides Ellis rosal
Ixora chinensis Lam. santan-tsina
I. coccinea L. santan-pula
Rutaceae Citrus madurensis Leur. calamansi








Fungicidal Activity of Crude Plant Extracts


Sapotaceae
Solanaceae


Sterculiaceae
Tiliaceae
Urticaceae
Verbenaceae







Zingiberaceae


Chrysophyllum oliviforme L.
Cestrum nocturum L.
Datum metel L.
Lycopersicum esculentum Mill.
Sterculia foetida L.
Corchorus olitorius L.
Leucosyke capitellata (Poir) Wedd.
Callicarpa candicans (Burm) Hochr.
Clerodendrum intermed jum Cham.
C. cuadricularse (Blco.) Merr.
C. siphonathus R. Br.
Lantana camera L.
Premna odorata Blco.
Stachytarpheta jamaicensis (L.) Vahl.
Vitex negundo L.
Zingiber officinale Roxc.


bubble gum tree
dama de noche
talong-punai
tomato
calumpang
saluyot
alagasi
tigan
kasopangil
bagawak
music bush
wild lantana
alagao
candi-candilaan
lagundi
ginger


amount of water added was determined
by the amount of the precipitate collected
to make a 1:1 concentration.
The supernatant liquid and the diluted
precipitate were assayed separately for
effectivity following the same procedure
as in the above.
In-vivo Bioassay in the Greenhouse
Preparation of the test plants. Test
plants were prepared by pre-germinating
seeds of variety IR-8 for at least one
week after which 20-30 seeds were sown
into earthen pots filled with lowland soil.
One pot represented one replication.
All treatments were completely rando-
mized with three replicates. Three trials
were conducted.
Spraying of the test plants with the
crude extracts. A modified Hudson
Lektic sprayer attached to pressure/va-
cuum pump with a constant pressure of
5 psi was used in spraying. Ten milliliters
of the concentrated crude extracts was
sprayed per treatment.
For the test for therapeutic effect, the
crude extracts were sprayed 1, 2, and 8


days after inoculation. Pots sprayed with
distilled water served as the control. For
protectant effect test, the plants were
sprayed with the extracts, 1, 2, 4, and 8
days before inoculation. In both cases,
spraying was done when the seedlings
were 2 weeks old.
Inoculation. Spore suspension from
an 8-day old culture calibrated.at 5000-
6000 spores per ml were used in inocula-
ting 2-week old rice seedlings by spraying
with the use of Hudson Lektric sprayer.
After inoculation the plants were placed
in a moist chamber for 24 hours and
transferred to a shaded area for further
incubation.

Gathering of data. Data were collected
during the necrotic stage of the disease.
In the protective test, 10 plants were
collected at random per treatment, one
week after inoculation. In the therapeu-
tic effect test, the same number of sam-
ples were taken at random 1 week after
spraying the crude extracts.


RESULTS


C__


Jan. & June 1978








Philippine Phytopathology


Screening of plant materials


The preliminary assay of 93 plant
materials resulted in 42 (45%) plant
extracts found active against H. oryzae
(Table 2).
Dilution and longevity assay of the
plant extracts
Nine of the 42 plant extracts found
active against H. oryzae were further
assayed for their activities at different
dilutions.
The results of the dilution assay


against the growth of H. oryzae are
shown in table 3. T. erecta, I. balsamina,
M. jalapa and A. lanata showed activity
up to 1:100 dilution; S. officinale and
A. sativum were active up to 1:10 dilu-
tion; while E. pulcherrima and A. bilimbi
were active in its concentrated form only.
All the nine plant extracts did not show
activity against the test organism at
1:1000 dilution.
The concentrate extracts and 1:10
dilution of T. erecta and S. officinale
have the same activity against the growth
of H. oryzae. The activity at 1:10 and


Table 2. Plant materials showing activity against Helminthosporium oryzae, based on
zone of inhbition taken 4 days after seeding


Diameter of Diameter of
Plant Inhibition Plant Inhibition
Materials Zone (in mm) Materials Zone (in mm)

A. sativum 45.001 C. papaya 22.66
B. rubra 31.00 C. olitorus 22.66
I. balsamina 29.00 C. jasminoides 22.23
A. bilimbi 28.66 A. lanata 22.17
N. indicum 28.33 S. saman 22.00
E. pulcherrima 28.33 M. arvensis 21.66
J. podagrica 28.17 A. campanulatus 20.66
M. jalapa 26.00 T. erecta 20.58
I. aquatic 25.83 J. gossypifolia 19.16
P. auriculata 25.75 A. spinosus 19.00
H. esculentum 25.00 S. foetida 18.91
C. capitellata 24.50 B. balsamifer 18.53
Z. pendula 24.50 P. oleracea 18.41
P. indica 24.41 S. jamaicensis 18.33
S. officinale 24.33 1. batatas 18.33
P. alliaceum 24.17 P. eleptica 17.83
C. candicans 24.00 C. roseus 17.75
E. hirta 23.83 C. oliviforme 16.75
P. guajava 23.50 Bigonia sp. 15.16
I. coccinea 23.33 B. intefrifolia 13.17
Z. officinale 23.00 A. pentandrum 13.30

1Average of three replicates, each replicate with four discs or an average of
12 discs.


Vol. 14











3. Average zones of inhibition of Helminthosporium oryzae showing
dal property of the plant materials at different dilutions (in mm)1


Plant Materials
Cone.

T. erecta 18.42ab 1
I. balsamina 20.22ab 1,
M.Fjalapa 17.28abd 1
S. officinale 18.16abc I
P. alliaceum 21.56a 1(
A. lanata 13.44d
E. pulcherrima 18.91ab
A. sativum 14.22cd
A. bilimbi 8.09e (


SEach figure is an average of thre4
together 3 trials with 3 replicates. With
ferent letters are significantly different,
connected by lines are significantly differ



1:100 dilutions of M. jalapa and A. lanat,
were not significantly different fror
each other.

The inhibitory activities of the con
centrated extracts from P. alliaceum
I. balsamina, E. pulcherrima, T. erect
and S. officinale were not significantly
different from each other. However, th
activities of the extracts from the las
four test plants were also not signify
cantly different with M. jalapa, S. off
cinale M. jalapa, and A. sativum. A
balimbi showed the least activity against
the test organism and significantly differ
ent with the rest of the plant material,
At 1:10 dilution T. erecta, I. balsa
mina, M. jalapa and S. officinale wer
most active in suppressing the growth c
H. oryzae. I. balsamina, M. jalapa an
P. alliaceum showed no significant
difference in their activity against h


:10 1:100 1:1000 Coni

14a 10.15a 0.00a 0.(
ab 7.72ab 0.00a .(
37ab 8.64a 0.00a 0.
98a 0.00c 0.00a 0.(
76bc 0.00c 0.00a 0.(
02c 4.22 0.00a 0.
00Dd 0.00 0.00a 0.(
03d 0.00c 0.00a 0.(
DO 0.00c 0.00a 0.(


readings (2, 4, 8 days after seeding) lumpec
Sa vertical column, figures followed by dif
while figures within the horizontal line no
nt at 5%level with DMRT.



oryzae, while P. alliaceum and A. lana
showed the same activity. E. pulchen
ma, A. sativum and A. bilimbi were n
active against H. oryzae at 1:10 dilutio
T. erecta, I. balsamina and M. jalaj
showed the same activity against tl
growth of H. oryzae at 1:100 dilutio
The activities of I. balsamina and
lanata were not significantly differer
while S. officinale, P. alliaceum, E. pi
cherrima, A. sativum and A. bilim
were not active against the test organist
at 1:100 dilution.
The test on the longevity of t
extracts in inhibiting the growth of ,
oryzae was evaluated at three differe
times, 2, 4 and 8 days of incubati,
after seeding. In table 4, only T. erec
showed a significantly decreasing zo
of inhibition with an increase in tin
L balsamina and A. sativum exhibited








T7-l 1 A


Table 4. Average zones of inhibition of I
dal property of plant materials m



Plant Materials
2


T. erecta 19.81a
I. balsamina 15.70b
M. jalapa 8 84cde
S. officinale 11.64bc
P. alliaceum 13.06
A. lanata 7.03de
E. pulcherrima 5.82ef
A. sativum 7.84cde
A. bilimbi 2.17f


Each figure is an average of three i
together 3 trials with 3 replicates. Within
ferent letters are significantly different, i
connected by lines are significantly differed



significant decrease in inhibition zone
by a significant increase in inhibition
zone on the fourth day of incubation
but did not show a significant change
on the eight day. Time was significant
with the rest of the plant materials.
When measurements of inhibition zone
were done on the second day, T. erecta
differed significantly from the rest of
the plant materials. I. balsamina ranked
second to T. erecta but its activity
against the growth of H. oryzae was
significantly different from S. officinale
and P. alliaceum. M. jalapa, A. sativum,
and S. offcinale showed the same activity
against the test organism. A. lanata and E.
pulcherrima together with M. jalapa
and A. sativum did not differ significantly
in their activity against H. oryzae. A.
bilimbi exhibited the least zone of in-
hibition but was not significantly different
from E. pulcherrima.


elminthosporium oryzae showing the fungici
measured at different days (in mm)


Days

4 8


15.15a 8.74cd
13.27ab 10.96abc
14.94a 14.51a
11.46a 11.03ab
9.86bcd 9.40bcd
9.21bcd 8.43cd
6.56cde 6.52e
5.54de 3.07ef
3.85e 2.08


eadings (2, 4, 8 days after seeding) lumped
a vertical column, figures followed by dif-
rhile figures within the horizontal line not
t at 5% level with DMRT.



When measurements were taken on the
4th day, the suppresive activities of
extracts from T. erecta, I. balsamina, M.
jalapa and S. officinale on the growth of
H. oryzae were not significantly different.
However, the two plant materials with
lower values, I. balsamina and S. offici-
nale, showed the same activity as with
P. alliaceum and A. lanata. The activity of
P. alliaceum and A. lanata on the other
hand were not significantly different
from that of E. pulcherrima and A.
sativum. A. bilimbi showed the least
inhibition zone but was not significantly
different from A. sativum and E. pulcher-
rima.
On the 8th day, extracts from I.
balsamina, M. jalapa and S. officinale
showed the largest zone of inhibition
and were not significantly different
from each other. P. alliaceum showed a
smaller zone of inhibition but somehow


_____ _









Il Activity of Urude I


was not lignuncanuy uiiierent iroUi
I. balsamina and S. officinale of higher
values and with A. lanata, T. erecta and
E. pulcherrima of the lower values. E.
pulcherrima and A. sativum were not
significantly different from each other
and A. sativum alone did not differ
significantly from A. bilimbi which
exhibited the lowest zone of inhibition.
Assay of supernatant liquid and
precipitates of plant extracts
A further assay of the promising ex-
tracts was conducted simultaneously with
the assay of the different dilutions. The
crude extract was centrifuged to separate
the supernatant liquid from the precipi-
tate. The two components were assayed
separately. This assay was made to
further evaluate the property of the
active compound.


Table. Average zones of inhibition o
supernatant liquid and the preci



Plant Materials
Supernatant
liquid


S. officinale 18.52a
I. balsamina 16.12ab
P. alliaceum 17.62ab
T. erecta 11.60c
A. sativum 13.90bc
A. lanata 4.97d
E. pulcherrima 0.00e
M. jalapa 0.00e
A. bilimbi 0.00a


SEach figure is an average of three
together 3 trials with 3 replicates. Within
ferent letters are significantly different,
connected by lines are significantly differs


1me isuIn u uile assay using ule super-
natant liquid and the precipitate of the
plant extracts against the growth of H.
oryzae is summarized in table 5. The
activities of the supernatant liquid and
precipitate of S. officinale, I. balsamina
and A. sativum did not differ signifi-
cantly. The supernatant liquid of P. allia,
ceum was significantly more active than
the precipitate. The rest of the plant
materials exhibited a significantly more
active precipitate against H. oryzae than
its supernatant liquid.

The activities of the supernatant
liquid of S. officinale, I. balsamina and
P. alliaceum were not significantly differ-
ent. However, those from A. sativum
showed the same activity as I. balsamina
and P. alliaceum. The activities of ex-
tracts from T. erecta, and A. sativum


Helminthosporium oryzae treated with ti
itate of the plant extracts (in mm)'


Treatments


Precipitate Contr


20.26a 0.008
16.77ab 0.00'
13.45bc 0.00O
16.83ab 0.008
12.44c 0.008
14.02b 0.008
15.88be 0.00'
13.77"b 0.00a
4.05d 0.008


eadings (2, 4, 8 days after seeding) lumped
a vertical column, figures followed by dif-
'hile figures within the horizontal line not
t at 5% level with DMRT.


dan. & June 19wt








Philippine Phytopatholog


were not significantly different. Thi
activity of the supernatant liquid of A
lanata against H. oryzae differed signi
ficantly from the rest of plant materials
The supernatant liquid ofE. pulcherrima
M. jalapa and A. bilimbi were not able tc
inhibit the growth ofH. oryzae.
The precipitate of S. officinale, I
balsamina and T erect exhibited th<
same activity against H. oryzae. However
the activities of balsamina and T. erect
did not differ significantly from P. allia
ceum, A. lanata, E. pulcherrima anc
M. jalapa. On the other hand the activi
ties of the precipitate from P. alliaceum,
A. sativum, A. lanata, E. pulcherrima and
M. jalapa were not significantly different.
A. bilimbi showed significantly low acti.
vity against H. oryzae, as compared with
the other plant materials.
The activity of the supernatant liquid
and precipitate of the plant extracts

Table 6. Average zones of inhibition c
supernatant liquid and the p
different days (in mm) '


Plant Materials ---
2

S. officinale 17.83a
I. balsamina 17.87a
P. alliaceum 19.22a
T. erecta 18.64a
A. sativum 16.43a
A. lanata 7.90b
E. pulcherrima 7.23bc
M. jalapa 6.25be
A. bilimbi 2.72e

Each figure is an average of three
together 3 trials with 3 replicates. Withii
ferent letters are significantly different,
connected by lines are significantly differe


against H. oryzae was not general]
affected by time (Table 6). Only P. allia
ceum, T erecta and A. sativum showed
significant decreases in activities against
H. oryzae with time. P. alliaceum de
creased its activity on the fourth dal
while that of T erecta and A. sativun
showed significant decreases on the eigh
day.
The activities of extracts from S. offi
cinale, I. balsamina, P. aliaceum, T. erect
and A. sativum on the second day against
H. oryzae were the same. Those from A.
lanata, E. pulcherrima and M. jalapl
showed the same activity against H
oryzae but E. pulcherrima and M. jalap&
did not differ significantly from A.
bilimbi.
On the 4th day, S. officinale, I.
balsamina, and T. erecta showed the
same effects which were not significantly
different from the results on the 2nd day

Helminthosporium oryzae treated with tj
ecipitate of the plant extracts measured



Days

4 8

19.99a 20.35a
16.58ab 14,9b
13.93c 13.46b
15.71ab 8.31d
14.02bC 9.07cd
10.30cd 10.28be
8.01d 8.59d
7.48de 6.93d
2.86e 0.49

readings (2, 4, 8 days after seeding) lumped
a vertical column, figures followed by dif-
hile figures within the horizontal line not
t at 5% level with DMRT.


Vol. 1<








Activity of Crude PI:


I. balsamina, P. alliaceum, T. erecta and
A. sativum did not show significant
differences but the activities of P. allia-
ceum and A. sativum against H. ory-
zae were not different from that of
A. lanata. A. lanata, on the other hand,
did not show a significant difference
from E. pulcherrima, and M. jalapa. The
activity ofM. jalapa did not significantly
differ with the activity ofA. bilimbi.
On the 8th day, S. officinale exhibited
a significantly larger zone of inhibition
compared with those of the other plant
extracts. I. balsamina, P. alliaceum and
A. lanata showed the same activity but
P. alliaceum and A. lanata did not differ
significantly with A. sativum. The acti-
vities of T. erecta, A. sativum, A. lanata,
E. pulcherrima and M. jalapa were not
significantly different. A. bilimbi was
significantly different from the other
plant extracts.
In-vivo bioassay in the greenhouse
The results of the in-vivo assay of the
crude extracts for their protective effect
against H. oryzae is summarized in table
7. Only I. balsamina gave a significant



'able 7. Average number of Helminthosp
crude extracts before inoculation'


Da
Treatments
1 2

balsamina 11.00 15.94
.alliaceum 28.51 23.47
sativum 44.30 37.79
.erecta 25.72 32.68
controll 36.47 38.11

Ave. 29.20 29.60

Average of 3 trials with 3 renlicates ne


effect as a protectant when compared
with the control. Spray schedule did not
show any significant difference.
Table 8 shows the therapeutic effect
of the crude extracts against H. oryzae.
All of the plant materials showed a
significant effect when compared with
the control. As a therapeutant, I. balsa-
mina ranked first followed byP. alliaceum
with an average lesion count of 11.06
and 12.72, respectively. The two plant
materials did not differ significantly in
their therapeutic effects against H.
oryzae. I. balsamina and P. alliaceum
were followed by A. sativum and T.
erecta with 15.40 and 15.04 average lesion
count respectively with no significant
difference between the two treatments.
The intervals of spray schedule for the-
rapeutic effect did not show any sig-
nificant differences.


Qualitatively, I. balsamina showed
phytotoxic effect 24 hours after spraying.
The phytotoxicity seemed temporary,
for the rice plants recovered after 3 days
and had grown normally thereafter.



orium lesions on rice plants treated with



ys Before Inoculation

4 48 Ave.

24.48 15.53 16.74
35.70 21.31 27.25
27.02 16.46 31.39
41.82 26.52 31.68
33.66 22.52 32.69

32.54 20.47

r trial and 10 nlants ner replicate.


Jan. & June 1978











Table 8. Average number ofHelminthos
extracts after inoculation'


D
Treatments
1 2

I. balsamina 11.33 10.9
P. alliaceum 12.44 11.1
T. erecta 12.83 13.3
A. sativum 16.16 15.9
Control 17.20 20.2

Ave. 13.99 14.3

'Average of 3 trials with 3 replicate
not connected by lines are significantly dif




A closer view of the leaves sprayed
with the crude extracts from I. balsamin,
showed blighted portions which eventual
ly made the whole leaf necrotic. The
young leaves that came out later wer
the ones that supported the good stan(
of the rice plant.
The appearance of the rice leaves 2Y
hours after spraying the crude extracts o:
T. erecta and P. alliaceum showed depo
sits of dark green materials which ad
hered to the leaf surfaces for 3 days. The
spray residues were still clearly visible or
the foliage even 3 days after application.


DISCUSSION

The results of screening plant materials
for fungicidal property indicated the
occurrence of active substances which
are common among higher plants. Of the
42 plant materials that showed inhibitory
activity against H. oryzae, 12 species
have been reported to possess anti-bac-
terial and/or anti-finnal nmnnrrtipe Thp


orium lesions on rice plants treated with cru,



ys After Inoculation

4 8 Av

10.29 11.71 11.1
14.51 12.79 12.'
16.39 17.73 15.(
15.97 13.55 15.,
13.50 28.05 22.!

16.13 16.77

Super trial and 10 plants per replicate. Figur
erent at 5% level with DMRT.




campanulatus, roots and leaves of C
papaya, flowers of T. erecta, leaves, root
and stem of Psidium guaijava, H. esculen
turn and roots, stem and leaves of 1
batatas, oil extracts from B. integrifolit
(Walker et al. 1937, Pryor et al. 1940)
cloves ofA. sativum (Ark and Thompson
1959; Tansey and Appleton, 1975)
extracts from I. balsamina; extracts o
M. arvensis and P. olearacea (Bishop an(
Macdonald, 1951); and leaves, stem am
flowers of G. jaminoides (Gilliver, 1947).
So far, very few active compound:
have been isolated from the test plants
These active compounds were "Plumba
gin" from Plumbago (Saint-Rat anc
Luteraan, 1943 as cited by Korzybsk
et al. 1940); and "Allicin" from A.
sativum (Cavallito et al. 1945).
The significant decreases in the zone!
of inhibition exerted T. erecta, P. allia.
ceum and A. sativum against tH. oryzae
with time seem to suggest that the active
principle of the extracts was volatile and
that it nnetihlv rliaeinatpl with time T


_ _ _


__








Activity of Crude P


not absorbed by the medium to give a
lasting effect to the test organism or
did not diffuse in the medium. Another
possible reason is the degradation of the
active compound due to exposure to the
atmosphere and light. It can also be
attributed to the transformation of the
active compound to inactive form by re-
action with the atmosphere.
The precipitates of the plant extracts
were significantly more active than their
corresponding supernatant liquids, except
the extracts from P. alliaceum, A. bilimbi
and A. sativum which exhibited a non-
active supernatant liquid. The result
might had been influenced to a great
extent by the solubility of the active
compound in the extracting solvent used.
It is therefore likely that the active com-
pounds of the above plant materials were
soluble in water while those of S. offici-
nale, I. balsamina, T. erecta, A. lanata,
E. pulcherrima and M. jalapa were of low
solubility in water.
The effect of I. balsamina extracts
that showed protective and therapeutic
properties can be attributed to the exis-
tence of 2-methoxy-1, 4-napthoquinone
in the plant cells. The compound was
found highly fungicidal to a number of
phytopathogenic fungi such as Monilinia
fruticola, Colletotrichum lindemuthia-
num, Aspergillus niger, Penicillium nota-
tur and Pythium debaryanum (Little,
et al, 1948 as cited by Korzyski, et al,
1967).
The phytoxicity of the crude extracts
of I. balsamina on the rice plants is likely
to be a contributory factor to its effect
as a protectant and therapeutant. The
blighted rice plants recovered fully after
8 days. The young leaves which came out
later were the ones that supported the
good stand of the rice plants. It is
possible that when the rice plants were
enro ud it th+ ,-lupIc P'vtriOter nf I


balsamina, the pathogens together with
plant leaves were killed, giving rise to
pathogen-free leaves. It is also possible
that the compound is systemic, that it
was absorbed by the leaf tissue and was
translocated to the meristematic tissue
of the rice plant. These possibilities re-
duced the necrotic lesions of brown spot
diseases of rice to a significant level when
compared with the control.
The active principle of P. alliaceum
has not yet been isolated. However, the
very large zone of inhibition exhibited
by its activity in in-vitro assay suggested
that the active principle was perhaps
present in the leaves in a large amount.
The odor of the crushed leaves is similar
to the crushed leaves of A. sativum. The
analysis and isolation of the active agent
of this plant require another study. P.
alliaceum was comparable to balsamina
as a therapeutant to H. oryzae.
The in-vivo assay of extracts from A.
sativum against H. oryzae revealed
their therapeutic property. The inhibi-
tory activity of A. sativum has been
attributed to allyl sulfide (Uemori, 1930),
alkyl polysulfides (Kitagawa and Amano,
1935), and "allicin" (Cavallito and
Bailey, 1944). Cavallito and Bailey
(1944) have shown that diallyl sulfide
and diallyl polysulfides have no anti-
bacterial action. Furthermore, they have
isolated from a steam distillate of garlic
cloves an antibiotic substance which
they originally named "allicin". The
name "allicin" has now been dropped to
avoid confusion with certain well estab-
lished medicinal products, (Cavallito et al,
1944). The authors believed these sub-
stances to be the active principle in A.
sativum.
Cavallito et al. (1944) have also de-
monstrated that the active principle of
A. sativum was not present as such in the
nlants hut nemlrs in the form nf a ther-


.Ian R, hlna 1 Q7(








Philippine Phytopathology


mostable precursor which when acted
upon by an enzyme, yields the anti-
biotic principle. The precursor and the
enzyme were present in different cells
of the garlic cloves and the active agent
was produced only when the cells were
crushed and their contents mixed.
Allicin contains approximately 40% sul-
ful, soluble in water to the extent of
approximately 2.5% at 10 C. The pure
product is irritating to the skin and the
odor is much more characteristically that
of garlic than that of the various allyl
sulfides.
The supporting discussions above,
strongly suggested that the antifungal
effect of A. sativum extract as a thera-
peutant is the sulfur component of the
active principle. Sulfur has been esta-
blished as a fungicide commendable
against rust and mildew diseases of plants.
The significant performance ofA. sativum
as a protectant may be, due to the high
volatility of its active component.
T. erecta has been established as a
nematicide. It has been suggested that its
roots contain chemicals toxic to nema-
todes, thus, inhibiting the growth of
plant parasitic nematodes. From the data
presented, T. erecta showed a therapeutic
effect as a fungicide. It ranked third as
a therapeutant against H. oryzae.
The presence of active principle in
higher plant is influenced by several
factors such as age of the plant, plant
parts, age of the organism, method of
extraction and time of harvesting the
plant materials.
The antimicrobial principle of plants
maybe present in large amount in


younger plants or plant parts and de-
creases with age and maybe present in
one or more or in all parts of the plant.
Work done by Wiltshire and Johnstone
(as cited by Gilliver 1947) showed that
age of the plant and foliage and the nitro-
gen content of the plant affected the in-
hibitory activity of the extract. Walker
et al. (1925) said that the reduction in
the availability and vitality in spores as the
culture aged makes the organism generally
more sensitive with age.
Different plants require extracting
solvents. Active substances may occur in
plants in inactive form and become
active only if the plant is properly pro-
cessed.
It will be noted that some plant ex-
tracts showed a larger zone of inhibi-
tion during the preliminary assay than
that of the result in dilution and further
assay of the supernatant liquid and the
precipitate. This is likely to be the
effect of variation in time of the year
when the plant materials were assayed.
The detection, isolation, characteriza-
tion, further evaluation of the active
agent and determination of plant age
when the active principle is at its maxi-
mum require the coordinated efforts of
plant pathologists, plant physiologists and
chemists.
Further studies on the extraction,
synthesis, mode of action and longevity
of the synthesized compound, economics
and dosages of the extracts are suggested
to be able to recommend the possibility
of successful large-scale production of
these extracts for crop protection.


Vn- 1 A









d Activity of Crude I


LITER)

ARK, P. A. and J. P. THOMPSON. 1!
antibiotic from garlic. (Allium sativ
BISHOP, C. J. and R. E. MACDONALD
bial substances. Canad. J. Bot. 99:S
CAVALLITO, C. J. and J. H. BAILEY.
sativum. I. Isolation, physical pro
Soc.
CAVALLITO, C. J., H. BAILEY and
Allium sativism. III. Its precursor i
67:1032.
GILLIVER, K. 1947. The effect of p
Venturia inequalis. Ann. Biol. 34:1
KITAGAWA, M. and A. AMANO. 193
Terkultura, Kjusu Imp. Univ. 6:30.
KORSZYBSKI, T., Z. KOWSZYK GIND
Origin, Nature and Properties, Vol.
PORDESIMO, A. N. and L. L. ILAG. 1
organisms. Kalikasan, Philipp. J. Bi
PRYOR, D. E., J. C. WALKER and M.
cyanate vapor to certain fungi. Am
SHARVELLE, E. C. 1961. The nature a
Co., Minnesota. 308 pp.
TANSEY, R. and J. A. APPLETON. 19
Mycologia 67:409-413.
UEMORI, T. 1929. Pharmacological in,
Jap. 9(1):21-26 Chem. Abstr. 24(1
WALKER, J. C. and K. P. LINK. 1935.
bulb parasites. Bot. Gaz. 96:468-4
WALKER, J. C., C. C. LINDERGREN
the toxicity of juice from onion sc
WALKER, J. C., S. MORRELL and H.
related sulfur compounds to certain


URE CITED

9. Control of certain diseases of plants w
% L.) Plant Dis. Reptr. 43:276-282.
L951. A survey of higher plants for antimic
)-269.
44. Allicin, the antibacterial principle of Allil
rties and antibacterial action J. Amer. Che

H. BUCK. 1945. The antibacterial principle
I "Essential oil of garlic." J. Amer. Chem. S

it extracts on germination of the conidia
1-143.
Antiseptic action of garlic. Bull. Sci. Faci
Chem: Abstr. 30(1936):3091.
ER and W. KURYLOWICZ. 1967. Antibiot
Warszawa: Pergamon Press, 1651 p.
6. Toxicity of garlic juice to plant pathoge
.5:251-258.
STAHMANN. 1940. Toxicity of allyl isoti
SJ. Bot. 27:30-38.
I uses of modern fungicides. Burgess Publish

. Inhibition of fungal growth by garlic extra;

litigation of Allium sativum. Folia pharmacy
0): 2191.
ixicity of phenolic compounds to certain on

d F. M. BACHMAN. 1925. Further studies
es. J. Agr. Res. 30:175-187.
FOSTER. 1937. Toxicity of mustard oils
Fungi. Amer. J. Bot 24:536.541.


Jan. & June LI














RICE RAGGED STUNT I

K. C. LING, E. R. TIONGCO, V.

Plant Pathologist, Senior Research
the International Rice Research Institute,
Sincere thanks are due to Mr. Mari
Cotabato, who drew the authors' attent
Philippine Bureau of Plant Industry, and
at Los Banos, for sharing their opinions
NFAC-USAID, for providing diseased ph
Laboratory, IRRI, for plant analysis; to
three biotypes of the brown planthopp
Sapporo, Japan, for providing the oxyte
Hoechst Philippines Inc., for providing tel
Awarded the best paper in Plant Pi
logical Society at the 9th Annual Nation,
Philippines held at Silahis International H

AE

Rice ragged stunt, a new virui
ippines in 1977. The symptoms ar
and twisted leaves, vein-swellings,
branches, incomplete emergence of
filled grains. The disease is caused
transmitted by the brown plantho
vector interaction belongs to the pe
Not only Oryza sativa but also 0
virus. Rice plants can be dually in
or with ragged stunt and tungro. E:
response of ragged stunt disease ph
virus is likely a member of acanthov


The first available information of the
ragged stunt disease was a telegram ad-
dressed to the Director of the Inter-
national Rice Research Institute (IRRI)
from Mr. Mariano G. Tirona, Provincial
Agriculturist of North Cotabato, Minda-
nao, Philippines, dated 14 January 1977,
which requested for assistance to conduct
immediate investigation on a new rice
malady which may endanger M-99 Rice
program phase 8 in North Cotabato. The
message also indicated that affected
plants produce 0 to 50 cavans depending
on the seriousness of the disease.


EASE IN THE PHILIPPINES

AGUIERO, and P. Q. CABAUATAN

sistant, and Research Assistants, respectively
>s Banos, Laguna.
o G. Tirona, Provincial Agriculturist of Nor
i to the disease; to Mr.. Hilario T. Bergoni
Delfin B. Lapis, University of the Philippin
)ut the disease; to Dr. Reeshon Feuer, IRB
From North Cotabato; to Analytical Servi
ntomology Department, IRRI, for providii
to Dr. E. Shikata, University of Hokkaid
icycline compound; and to Mr. J. Caldero
:ycline hydrochloride.
ology in 1978 by the Philippine Phytopath
Conference of the Pest Control Council of tl
!1, Manila, 6 May 1978.

RACT

disease, occurred sporadically in the Phil-
;unting of plants, appearance of ragged
lay in flowering, production of nodal
nicles, and panicles bearing mostly un-
a polyhedral virus of 50-70 nm, and is
er Nilaparvata lugens (Stal). The virus-
tent group without transovarial passage.
itifolia and 0. nivara are hosts of the
ted with ragged stunt and grassy stunt
rimental results did not yield a positive
s to tetracyclines. The rice ragged stunt
ses.


Similar telegrams were sent to the
Bureau of Agricultural Extension and to
the National Food and Agriculture Coun-
cil of the Republic of the Philippines
(NFAC). In response, rice pathologists of
the Philippine Bureau of Plant Industry
and the University of the Philippines at
Los Baflos went to North Cotabato to
observe the disease. The authors were
unable to make the trip but specimens of
diseased plants were received from North
Cotabato through the courtesy of Dr.
Reeshon Feuer, Crop Production Specia-
1;t n ~.osi TDo TAM ATT TC A l


QW7








tagged Stunt in the I


Rice Program.

The diseased plants from North Cota
bato were at heading stage and showed
abnormalities such as short and twisted
flag leaves, incomplete emergence o
panicles, excessive nodal branches an(
nodal panicles, and panicles bearing most
ly unfilled grains. Although the disease
was then not familiar, it could be defi
nitely said that it was due to neither the
late infection of rice tungro disease no
the delayed expression of tungro infec
tion because its symptoms differed dis
tinctly from that of rice tungro disease
and the disease could not be transmitted
to rice seedlings by using the tungrn
vector Nephotettix virescens.
In February 1977, the disease in ric
fields was observed at IRRI and later ii
farmers' fields in Luzon, Negros Occider
tal, and Mindanao. In most cases, their
were only a few diseased plants scattered,
over rice fields but in severe cases, mor
than 90% of rice hills in a field wer
diseased, and the yield was totally losi
Because the disease is definitely no
the late infection of rice tungro an
symptoms of the disease are stunting c
rice plants and appearance of ragge
leaves, the authors named the disease
"rice ragged stunt" on 6 April 1971


Table 1. Growth of healthy and ragged stl

Tillers (no.)

Plant 0a 1a 2a

Healthy 1.0 1.9 8.8
Diseased 1.0 3.2 8.1
LSD (5%) 0.7 2.3

aMonths after inoculation.


The disease was also being called "intec-
tious gall" (de la Cruz, 1977) and "kerdil
hampa" (Hibino et al., 1977) which
literally means stunt empty.
This paper reports the symptomatolo-
gy, chemical analysis, causal agent, trans-
mission, host range, dual infection, and
antibiotic treatment of the ragged stunt
disease from studies of materials either
directly collected from rice fields or
artificially inoculated in the greenhouse
made either at the IRRI or at the Uni-
versity of Hokkaido, Sapporo, Japan.

SYMPTOMATOLOGY

The symptoms of the ragged stuni
disease are quite complex, and vary
according to the stage of plant growth
They include stunting of plants, appear
ance of ragged and twisted leaves, and
vein-swellings, delay in flowering, pro
duction of nodal branches, incomplete
emergence of panicles, and panicle:
bearing mostly unfilled grains. How
ever, some features of the disease
plants do not differ strikingly front
those of the healthy ones. For instance
the color of diseased plants does no
deviate markedly from that of health
ones. The difference in number of tiller
between healthy and diseased plants i:
not consistent (Table 1), but at the tim<


it diseased plants of Taichung Native 1 in poi

Plant height (cm)
3a 0a 1a 2a 3

8.3 11.8 64.5 75.4 79
6.6 11.5 38.8 47.0 52
2.3 1.6 11.2 13.9 12


Jan. & June 197








l uiiliuRJIy x y tupjubluugy


ui 1i4ullULy u ul UllcU lrIily plants, [le
diseased plants have more dimunitive
tillers than the healthy plants in pot
experiments (Table 2).
Stunting

The diseased plants are stunted to
various degrees, at all growth stages (Fig.


Table 2. Comparison of healthy and ragged
pots at maturity


Nod
Tillers brand
Plant (no.) (no,

healthy 4 0
)iseased 11 33
SD (1%) 5 7



IA). This is the only symptom that is
observable from a distance before the
heading stage. In pot experiments, artifi-
cially infected plants of Taichung
Native 1 (TNI) were only 60 to 65% as
tall as the healthy plants during all
growth stages (Table 1). The other symp-
toms are clear only when diseased plants
are closely examined.
Ragged leaf
At early growth, the predominant
symptom is the appearance of "ragged
leaves" which refer to leaves with irre.
gularly edged portions (Fig. IB and IC).
The irregular edge consists of 1 to 17 or
more breaking and the edge between or
near the breaking. The breaking appear
as notches or indentations, and can be
observed even before the leaf blades
unroll. The depth of the breaking varies
but never crosses the midrib; the dis-
tance between two breaking varies from
D.2 to 3 cm. Occasionally, the breaking
are at a similar distance, resulting in a


OavIaRLU ,Iai. AJwL IsL1iUl U IdJLaUU VUSU
varies from 0.1 to about 20 cm, or from
less than 1% to about 70% of the length
of the leaf blade, including the chlorotic
edge beyond the extreme notches at both
ends. The ragged portion is generally
chlorotic, which may later become
yellowish brown. The chlorotic edge,


'unt diseased plants of Taichung Native 1 in



Filled Unfilled
es Panicles grains grains
(no.) (no.) (no.)

3 223 54
18 52 474
4 154 114



particularly between two notches, may
eventually disintegrate.
The ragged edges occur more on leaf
blades and less on leaf sheaths (Table 3).
The ragged edges that appear on the leaf
blades are either on one side or on both
sides of the leaf blades, and at different
portions of the leaf blades. All of the
infected rice varieties and lines examined
had ragged leaves. The ragged leaves
range from 0 to 34 per hill, or from 0 to
5 per tiller, and gradually become fewer
as the plants grow older.

Twisted leaf
Another symptom of the disease is
twisting of leaves (Fig. 1B and ID). The
twisting often occurs at the top portion
of leaf blades. It may wind one or more
turns, resulting in a spiral shape. The
twisting is caused by the alternate uneven
growth of the leaf blade on both sides
of the midrib. Almost all varieties and
lines examined had twisted leaves. About


, s







Rice Ragged Stunt in the Philippines


A "C














B D


Fig. 1. Symptoms of rice ragged stunt disease. A Healthy (left) and infected (right)
plants. B. Close-up of a diseased plant showing ragged and twisted leaves..C.
Ragged leaves. D. Twisted leaves.


Jan. & June 1978








Philippine Phytopathology


Table 3. Aspects of ragged edges on leaves and ragged leaves on rice plants infected with
ragged stunt disease


No. examined


Designation


Ragged edge
Number of breakings/edge (range
Distance between two breaking (range in cm)
Length (range in cm)
As the length of leaf blade (%)
Ragged edges on
Leaf blades (%)
Leaf sheaths (%)
Ragged edges on
One side of leaf blades (%)
Both sides of leaf blades (%)
Ragged edges at
Upper portion of leaf blades (%)
Middle portion of leaf blades (%)
Lower portion of leaf blades (%)
Ragged leaves
Number/hill (range)
Number/tiller (range)
Having ragged leaves
Rice varieties and lines (%)
Rice hills (%)
Tillers within infected hills (%)


30% of tillers within infected hills had
twisted leaves, which varied from 0 to 10
per hill, or from 0 to 3 per tiller.
Vein-swelling
Another symptom of the disease is
an outward growth of vein, resulting in
swollen veins. The technical term for
such malformation is histoid enation
(Bos, 1970) or virocecidium (Blattny,
1961), but we prefer to call it "vein-
swelling" because the term is specific,
descriptive, and understandable. The
vein-swellings (Fig. 2A) are due to the
proliferation (hyperplasia) of phloem cells
in vascular bundles (Fig. 2B).
The vein-swellings vary from about
1 mm to more than 10 cm in length; from
0.2 to about 1 mm in width; and from


0.1 to about 1 mm in height. The num-
ber of vein-swellings varies among tillers
and varieties, and ranges from 0 to 74
per tiller, or from 0 to 42 per leaf, in-
cluding both on the sheath and on the
blade. The color of vein-swellings varies,
about 82% of vein-swellings are pale
yellow or white; 2% light brown; 6% dark
brown; and 10% a combination of such
colors (a portion being pale yellow; the
other, brown). The color seems to remain
unchanged until the leaf dies (Table 4).
The vein-swellings occur on the outer
surface of leaf sheaths or the lower sur-
face of leaf blades. None have been ob-
served on the opposite sides, possibly
because the vein-swellings result from the
proliferation of phloem cells which are


Quantity


1-17
0.2-3
0.1-20
< 1-70


1359
179
439
439

513
513

495
495

932
932
932

47
640

243
1486
640


0-34
0-5


_t_


~I ___~____~


Vol. 14





Rice Ragged Stunt in the Philippines


A
I(











Fig. 2. Symptoms of rice ragged stunt disease. A. Vein-swellings. B.A cross section of a
swollen vein showing proliferation of phoem cells. C. Distortion of flag leaves
and incomplete emergence of panicles. D. Nodal branches and nodal panicles.


Jan. & June 1978








Philippine Phytopathology


Table 4. Aspects of vein-swellings on leaves and culms of rice plants infected with ragged
stunt disease


No. examined


Designation


Size of vein-swellings
Length (range in mm)
Width (range in mm)
S. Height (range in mm)
Color of vein-swellings
Pale yellow or white (%)
Light brown (%)
Dark brown (%)
Combination (%)
Vein-swellings
Number/tiller (range)
Number/leaf (range)
Vein-swelllings on
Leaf blades (%)
Leaf sheaths (%)
Culms (%)
Vein-swellings at
Upper half of leaf blades (%)
Lower half of leaf blades (%)
Vein-swellings at
Upper half of leaf sheaths (%)
Lower half of leaf sheaths (%)
Having vein-swellings
Rice varieties and lines (%)
Rice hills (%)
Tillers within infected hills (%)


at an abaxial position to the rice leaf.
The vein-swellings appear on different
parts of rice plants, but more on leaf
sheaths (Table 4). About 90% of those on
leaf sheaths are on the upper half. Most
appear near the collar. All naturally
infected rice varieties and lines examined
had vein-swellings during late tillering
and after. The percentage of tillers with
vein-swellings seems to increase as the
plants grow older.
Delayed flowering, nodal branches,
incomplete panicle emergence, and
unfilled grains


At booting stage, flag leaves of the
diseased plants are short and often
twisted, malformed, or ragged (Fig. 2C).
Diseased plants often flower late (Fig.
1A). The panicle emergence is often in-
complete (Fig. 2C). Tillers often gene-
rate nodall branches" which are second-
ary or tertiary tillers produced at upper
nodes of a tiller (Fig. 2D). All the 411
rice varieties and lines examined developed
nodal branches. The number of nodal
branches varied from 0 to 11 per tiller.
It increased as the plants grew older.
The nodal branches often bear small


Quantity


646
646
646

1099
1099
1099
1099

952
419

1006
1006
1006

37
37

56
56

66
279
952


1-> 100
0.2-1
0.1-1

82
2
6
10

0-74
0-42

21
72
7

40
60

90
10

100
96
67


-- --


Vol. 14








Rice Ragged Stunt in the Philippines


panicles nodall panicles" (Fig. 2D).
Thus, the diseased plants have more
panicles and spikelets than the healthy
ones (Table 2). The spikelets are often
not discolored. The number of filled
grains of the diseased plants is greatly
reduced (Table 2). In the field, usually
none or a few filled grains can be harves-
ted from a diseased hill.
Systemic infection
The ragged stunt disease is systemic.
When a diseased plant is ratooned, vein-
swellings are the only symptom that can
be observed in the early regenerated
growth; other symptoms appear at boot-
ing stage of the plant. The diseased plants
can survive long after flowering more
than 6 months in the IRRI greenhouse.
Inoculated seedlings
When young rice seedlings are inocu-
lated, the first noticeable symptom is
stunting, followed by the appearance of
ragged leaves and then the other symp-
toms. The vein-swellings appear on both
leaf sheaths and leaf blades of artificially
infected plants about 3 weeks after
inoculation. The vein-swellings are often
brown in color.

CHEMICAL ANALYSIS

Leaves of both healthy and ragged
stunt diseased plants of three rice lines -
IR2061-465-1-5-5, IR3839-1, and
IR4427-115-3-3-3-3 collected from
fields were chemically analyzed for
contents of N, P, K, Ca, Mg, Fe, Cu,
Mn, and Zn. The analysis revealed that
the element contents of both healthy and
diseased plants were between the critical
levels of deficiency and toxicity as given
by Tanaka and Yoshida (1970). Differen-
ces in analyzed elements between the
healthy and diseased plants were not
statistically significant.


CAUSAL AGENT

The nature of the causal agent of the
ragged stunt disease was studied with
an electron microscope and a bioassay
method at the University of Hokkaido
in collaboration with Dr. E. Shikata and
his associates.

Electron micrographs (Fig. 3) showed
polyhedral particles of 50-70 nm not only
in dip preparations but also in ultrathin
sections of rice plants naturally and
artificially infected in the Philippines.
The particles were abundant in phloem
cells and in cells of the vein-swellings
(Shikata et at, 1977).
The infectivity of rice ragged stunt
virus was determined by the injection
method. Fresh leaves of diseased plants
of IR3839-1, infected in the Philippines,
were macerated in a buffer solution and
centrifuged. The supernate was injected
through a fine glass capillary into the
body of the brown planthopper Nila-
parvata lugens (Stal), the colony of which
had been reared for 4 years in Sapporo,
Japan, where ragged stunt disease has
not been observed to occur naturally.
Twenty-one of 40 insects that received
the superate injection survived for 2
weeks afterward. They were used to ino-
culate seedlings of the rice variety Miho-
nishiki at 3 insects/seedling for an inocu-
lation access time of 3 days. The same
insects were then transferred to inoculate
another set of seedlings for another 3
days. All 14 inoculated seedlings devel-
oped symptoms of the disease, indicating
that the injected insects became infective
(Senboku et aL, 1978a).

TRANSMISSION

Experiments conducted for investiga-
ting the manner of transmission of the


Jan. & June 1978








Phuvnnathnlniv


Fig. 3. -Electron micrographs show virus
phloem cell of an ultrathin sectic
stunt disease (from Shikata et al, I


ragged stunt disease at IRRI were a
follows:
Mechanical transmission
Rice seedlings of IR3839-1, a linm
susceptible to the disease, were manually
inoculated by rubbing leaf blades witl
the sap of diseased plants, using carbo
rundum as an abrasive. Other seedling:
were inoculated by the pin-prick method
None of the 96 inoculated seedling:
developed disease symptoms, indication
that transmission of the disease b)
mechanical means was unsuccessful.
Soil transmission
To determine transmission of th(
disease through soil (actually, by soil
inhabiting microorganisms), soil nea:
diseased plants and stubble of disease
plants were collected from paddy fields
and placed in trays. Healthy rice seed
lings were immediately grown in the
trays, both with and without stubble bo


-BIT







particles in a dip preparation (left) and in
i (right) of a rice plant infected with ragg(
177).


trays. The stubble was free from t
brown planthoppers. None of the 24
seedlings became infected until mal
rity although disease symptoms dev
oped on the regenerated growth of t
stubble in the center of the trays. Thl
the disease was not transmitted throu
soil even in the presence of stubble
disease plants.
Seed transmission
Mature seeds were collected frc
diseased plants and germinated. No
of the 633 seedlings showed symptom
of the disease. Some of the seedlings we
transplanted and grown in screen caE
until maturity. They remained health
Hence, no evidence was obtained
demonstrate the transmission of ragg
stunt disease through rice seed.
Transmission by the brown planthoppei
Insects with sucking mouth parts th
appeared on diseased plants in padi


J J1








Rice Ragged Stunt in the


late rice seedlings in the greenhouse.
nly some seedlings inoculated by the
rown planthopper (Fig. 4) became
ifected. Hence, the brown planthopper
the only known vector of the disease.
The ability of the brown planthopper
> transmit rice ragged stunt disease to
eedlings of IR3839-1 was tested by daily
erial transmission after acquisition access
mes of 2, 3, 4, or more days until the
death of the insect. The tests were
:peated, and involved 1,625 insects and
8,753 inoculated seedlings, including
,906 seedlings that died before symptoms
developed.
About 40% of the tested brown plant-
oppers transmitted the disease (Table
). In a single test with a small number of
sects, from 14 to 76% of the brown
lanthoppers were active transmitters.
loth nymphs and adults transmitted
























ig. 4. Brown planthoppers, Niaparata ug

ig. 4. Brown planthoppers, Nilaparvata luge


Le disease. No striking ditterences were
)served in percentage of active
ansmitters between female and male
lults (46 versus 42% ), and between
rachypterous and macropterous forms
.2 versus 48% ). A few insects became
fective immediately after an acquisti-
on feeding, while others remained
tent for as long as 33 days. The average
tent period was 8.6 days. The insects
mained infective after molting, indicat-
g that the virus passage is transstadial.
he highest number of seedlings that an
sect infected during its life span
as 22. However, 33% of the infective
fective insects infected only one seedling
ring their life spans. On the average,
i infective insect infected about four
edlings. The highest percentage of
fective insects transmitted the disease
days after acquisition feeding. The
lily transmission pattern was generally
termittent. The retention period ranged

























s (Stal), the vector of rice ragged stunt


47








? Phytopathology


Table 5. Transmission data of rice ragged


Insects
tested (no.) Desigi

1625 Active transmitters
256 Nymphs
1369 Adults
714 Females
655 Males
940 Brachypterous fi
429 Macropterous fo
500 Latent period (days
500 As of life span (,
655 Infected seedlings/i
48 Nymphs
607 Adults
330 Females
277 Males
399 Brachypterous fi
208 Macropterous fo
500 Retention period (c
655 Disease-transmittinl
655 Nontransmitting pe
655 As of life span (1
165 Congenitally infecti




from 3 to 35 days after acqusitic
feeding, averaging 15 days. The disease
transmitting days varied from 3 1
100% (averaging 41%) of the period froi
the time that an insect became infectii
to its death. As the infective insec
became old, they often failed to infe(
seedlings. The period between the la:
successful transmission and the insect
death ranged from 0 to 30 days, averaging
7 days.
The transovarial passage of the virt
was determined by examining infectivit
of the progeny of virulifeorus insect
The progeny were obtained by rearin
the viruliferous brown planthoppers o
Monochoria vaginalis, a plant specie
that is assumed a nonhost of ragge
stunt virus but is a temporary host c


stunt disease by Nilaparvata lugens



tion Range Iv

V) 14-76
0-100
14-82
15-100
6-91
m 16-100
a 0-100
2-33
6-96
'ective insect (no.) 1-22
1-8
1-22
1-21
1-22
m 1-22
a 1-17
Vs) 3-35
lays (%) 3-100
ad (days) 0-30
0-83
Insects (no.) 0-0




the brown planthopper (Ling and Aguiei
1970). The newly hatched nymphs we
used to inoculate rice seedlings of TP
at 1 insect/seedling/day for 20 co
secutive days except when insects dii
earlier. None of 2,733 seedlings inoc
lated by 165 progeny insects becar
infected, indicating the absence
congenitally infective insects.

Consequently, the interaction ofragg(
stunt virus and the brown planthopp
classifies the virus in the persistent grot
without transovarial passage.
Transmission by the brown planthopper
in Japan
The transmission of rice ragged stui
disease by a brown planthopper color
that had been reared on rice nlants in


48








Rice Ragged Stunt in


greenhouse m sapporo, Japan tor 4
years was studied in collaboration with
Japanese scientists at the University of
Hokkaido. After 1-day acquisition access
time on diseased plants of the rice line
IR3839-1, which were infected in the
Philippines, the infectivity of 50 brown
planthoppers was tested by daily serial
transmission to 1,270 seedlings of the
variety Hihonishiki for 40 consecutive
days except when insects died earlier.
The experiment was carried out in
a temperature-controlled greenhouse
(25-28 C).

Twenty-eight percent of the test
brown planthoppers, including both fe-
male and male adults, and both brachyp-
terous and macropterous forms, were
active transmitters. The latent period
ranged from 5 to 11 days after acquisi-
tion feeding, averaging 8.6 days. The
infective insect infected from 1 to 31 or
more seedlings during their life spans. The
retention period ranged from 9 to 41 or
more days after acquisition feeding
(some insects did not die on the last
day of the transmission test, 41 days
after acquisition feeding). The disease-
transmitting days from the time that the
insect became infective until the last
day of the transmission test ranged from
33 to 100%, averaging 81% (Senboku et
al., 1978b).
The brown planthopper in Japan trans-
mits rice ragged stunt disease. This con-
firms that the brown planthopper is the
vector of the disease because the positive
transmission in Japan, where there is no
natural occurrence of the disease, could
not have been due to contamination.
Although the percentage of active
transmitters and the latent period of
the brown planthopper in Japan and
those in the Philippines were similar,
the brown planthopper in Japan infected


more rice seedlings during their life
spans, gave a higher percentage of disease-
transmitting days, and had a longer
retention period than the brown plant
hoppers tested at IRRI. The reason could
be the differences in either the ecotypes
of the insect or the conditions for the
transmission test, or both. However,
we speculate that difference in trans-
missive ability between the brown plant-
hopper in Japan and those in the Philip-
pines was primarily due to the difference
in the conditions during the transmission
test. At the IRRI, the transmission tests
were done in a greenhouse where the
temperature was much higher than that in
Japan. The effect of temperature was
substantiated by the fact that the average
life span of the brown planthoppers used
for the transmission tests in the Philip-
pines was about 22 days whereas the
average life span of the 50 brown plant-
hoppers used for the transmission test
in Japan was longer than 32 days. The
insects with longer life span should have
a greater chance to infect more rice
seedlings and a longer time for retaining
the virus. The effect of temperature on
the life span of insect is not uncommon.
At least, it is the case for N. virescens
(Ling and Tiongco, 1975).
Transmission by biotypes of the brown
planthopper
The ability of different biotypes of
the brown planthopper to transmit rice
ragged stunt disease was studied by
daily serial transmission to IR3839-1
seedlings after an acquisition access
time of 2 or 4 days until the death of
the insects. The study included three
biotypes from the Department of Ento-
mology, IRRI, that differ in the ability
to attack rice varieties carrying different
resistance genes to the brown plant
hopper (IRRI, 1976), and a colony of the
brown planthopper that had been reared


49








! PhvtoDathologv


on TN1 in the greenhouse for severe
years but whose ability to attack differ
ent rice varieties had not been detei
mined. The test involved 985 insect
and 18,477 inoculated seedlings, incluc
ing 1,382 seedlings that died before
symptoms developed.
The brown planthopper biotypes di
not differ significantly in percentage c
active transmitters, latent period, number
of infected seedlings per insect, retentio
period, or percentage of disease-transmi
ting days from the time that an insec
became infective to its death (Table 6,


Table 6. Comparison of transmission of
Nilaparvata lugens



Active I
transmitters p
Biotype (%) (

1 41
2 34
3 41
Undetermined 37
Test of significance n.s.

n.s. = no significant difference amc



All tested biotypes showed transstadi:
passage and were similar in daily seri
transmission (Fig. 5). Consequently.
the tested biotypes have similar abilil
to transmit rice ragged stunt disease
Source for transmission
Diseased rice plants of differed
varieties, from different fields and diffe
ent localities were collected for tl
transmission study. The transmissic
results were similar. There was also r


difference between naturally and artifi
cially infected plants.

HOST RANGE

Plants of several species were inocu
lated by viruliferous brown planthopper
of rice ragged stunt disease in the greer
house. Among Oryza species, 0. sativa
0. latifolia, and 0. nivara were infected,
with ragged stunt. The infected plants a
0. latifolia (Fig. 6A and 6B) and C
nivara (Fig. 6C) showed symptoms c
stunting and appearance of ragged leave:


ce ragged stunt disease by different biotypes



Infected
tent seedlings Retention Disease-
riod (no.) period transmitti
ays) (insect) (days) (%)

9 5 16 37
11 4 16 36
8 4 14 39
8 5 14 45
i.s. n.s. n.s. n.s.

.g biotypes.



vein-swellings, and nodal branches, simili
to those on 0. sativa infected with th
disease. Plants of 0. latifolia were als
infected naturally in the field.
Brown planthoppers recovered th
virus from the diseased plants of (
latifolia and 0. nivara, and transmitted
ragged stunt successfully to rice seedling
Consequently, 0. sativa is not the on]
host of the rice ragged stunt virus.


fin











Rice Ragged Stunt in the Philippines


Fig. 5. Daily serial transmission of
lugens biotypes.


20
Days
rice ragged


stunt disease by infective Nilaparvata


Jan. & June 1978








Philippine Phytopathology


W i































Fig. 6. Oryza spp. infected with ragged
stunt disease. A. An infected O.
latifolia plant. B. ragged leaves
of 0. latifolia C. Healthy (left)
and infected (right) plants of O.
nivara.
8


Vol. 14








Rice Ragged Stunt in the


DUAL INFECTION

In the Philippines, besides rice ragged
unt disease, there are four known
rus and virus-like diseases of rice plants
grassy stunt (Rivera, et al., 1966, Ling,
at, 1970), orange leaf (Rivera, et al,
ange leaf (Rivera, et al. 1963), tungro
ivera and Ou, .1965), and yellow
varf are of minor importance (Ling,
>72). Rice plants can be dually in-
cted with both tungro and grassy
mnt diseases (IRRI, 1972). Therefore,
tal infection of ragged stunt and grassy
unt or of ragged stunt and tungro was
died by inoculating seedlings of TNI
ith respective viruliferous insects in the
eenhouse.
Rice plants were dually infected with
)th ragged stunt and grassy stunt (Fig.
and 7B). The symptoms of the dually
fected plants included both symptoms
ragged stunt (ragged leaves and twisted
lives) and those of grassy stunt disease
were stunting, excessive tillering, and
rrow leaf blades). Plants were much
hinted, particularly at the late growth
ige, when dually infected than when
fected with one of the two diseases.
Rice plants were dually infected with
,th ragged stunt and tungro (Fig. 7C).
ie infected plants showed symptoms of
hunting; appearance of ragged leaves, a
mptom of ragged stunt; and yellowing
'leaves, a symptom of tungro.
The dual infection indicates the ab-
nce of cross protection either between
gged stunt and grassy stunt or between
gged stunt and tungro. Hence, the
usal agent of ragged stunt is not closely
lated to the causal agent of grassy stunt
tungro.

ANTIBIOTIC TREATMENT

etracyclines reportedly suppress


rmptoms of plant diseases caused by
lycoplasmas (Whitcomb and Davis,
970), but they have no effect on disease
used by viruses.
Rice plants of IR29 and IR30,
naturally infected with ragged stunt,
ere transplanted into nine groups of
ots at three hills per group and treated
ith two tetracycline compounds, oxy-
tracycline and tetracycline hydrochlo-
de. Two concentrations of each com-
ound 15 and 60 ppm of oxytetracy-
ine, and 50 and 200 ppm of tetracycline
ydrochloride were sprayed on the
iseased plants either once a week for
1 weeks or three times a week for 10
eeks.
At the end of the treatment and for
veral weeks thereafter, the treated
plants of both varieties did not differ
om the untreated controls in plant
eight, number of tillers, ragged leaves,
ein-swellings, nodal branches, panicles,
id grains. Hence, neither of the two
impounds suppressed the symptoms of
ie diseased plants.

DISCUSSION

rigin of the disease
The origin of the ragged stunt disease
mains obscure although there are only
iree possibilities preexistence, mu-
ition, and immigration. However, we
Aspect that the disease has long been
resent at a low level in rice fields. It
lay have been unnoticed because its
symptoms are not obvious at certain
ce growth stage, particularly when it
appears together with rice grassy stunt,
r when the disease is observed from a
instance.
The disease is not limited only in the
hilippines, Indonesia (Hibino et al.,
977; Palmer and Soepriaman, 1977),


53


































A














C


B





Fig. 7. Dual infection. A. A rice
infected with ragged stunt
grassy stunt. B. Close-up ol
infected plant showing ra
leaves. C. A rice plant infh
with ragged stunt and tungro.


V


_ __








Kice Kagged stunt in tt


na lnailana (weerapat ana rongprasert,
978), because the occurrence of
ice plants with similar disease
symptoms of ragged stunt in India and
ri Lanka has been verbally reported
ly several scientists. In almost all cases,
he existence of the ragged stunt disease
n a country was first discovered by the
scientists with knowledge of symptoms
if the disease. Hence, it appears that the
disease was there.
L new rice virus disease
Besides rice ragged stunt, 14 rice
irus and virus-like diseases are known in
he world: black-streaked dwarf, dwarf,
;iallume, grassy stunt, hoja blanca,
lecrosis mosaic, orange leaf, stripe,
ransitory yellowing, tungro (including
he tungro-like diseases), waika, yellow
Irawrf, yellow mottle, and yellow stunt
Ling, 1962; Nishi, et al., 1975; Plant
protectionn Institute, 1974; Yokoyama, et
1., 1974). They differ from the ragged
tunt disease, but two of them, black-
treaked dwarf and grassy stunt, have
ome unique similarities to ragged stunt
disease.
Besides stunting and leaf twisting
Shikata and Kitagawa, 1977), the
ilack-streaked dwarf also causes rice
plantss to produce vein-swellings (Kuri-
ayashi and Shinkai, 1952; Shinkai,
.966), but they are darker than those
)f ragged stunt. The black-streaked dwarf
ein-swellings that appear on culms can
,e grouped as tumors, which are more
regular in shape and wider, and pro-
rude much more than ragged stunt
rein-swellings on culms. Also, rice plants
infected by black-streaked dwarf produce
nore tillers and do not exhibit ragged
eaves and nodal branches. Furthermore,
hey differ in species of insect vector.
'he black-streaked dwarf is transmitted
y the small brown planthopper Lao-


relphax striatellus (Fallen), and has not
ieen successfully transmitted by the
rown planthopper N. lugens (Shinkai,
1966) the vector of ragged stunt disease.
Although both grassy stunt and ragged
tunt are transmitted by the same vector
species N lugens, in the same persistent
manner, and without transovarial passage,
hey differ in symptoms such as degree
f stunting, tillering, width of leaf blades,
nd appearance of ragged and twisted
eaves, vein-swellings, and nodal branches.
furthermore they do not cross protect
against each other.
Consequently, ragged stunt is a new
ice virus disease.
i member of acanthoviruses
The ragged stunt virus may be a new
member of plant reovirus group (Shikata
t al., 1978). It appears to fall into
ie subgroup acanthovirus, a term pro-
osed by Milne and Lovisolo (1977),
rho listed the following criteria for re-
ognizing viruses of the subgroup:
1. Graminae only are infected, with
symptoms of darker green color (except
angola stunt virus), dwarfing, enation
reduction, increased tillering, and
oppression of flowering on at least some
osts;
2. Plant hoppers (Delphacidae) propa-
atively transmit the viruses;
3. Plant or hopper material, homo-
enized in small amounts of a saline,
larified by low-speed centrifugation, and
ejected into the vectors, should render
iem inoculative in 1-2 weeks. (Myco-
lasmas would fail to be transmitted in
iese ways);
4. Thin sections of leaf nations and
f infective hoppers reveal in the cyto-
lasm, spherical virus particles about
0 nm diameter containing dense cores


55








hvtnnatholofv


Se


of 50 nm diam. Viroplasms and tubular
structures also occur in infected cells.
The viroplasms appear in the light micro-
scope as X-bodies. In plants, the viruses
are phloem associated.
5. Simple dip or crush preparation!
of nations or swollen veins can be nega
tively stained for electron microscopy
Uranyl acetate reveals spherical particle!
of 65 nm diam. with double capsids anc
also spiked core's about 50 nm in diam.



LITERA'

BLATTNY, C. 1961. Virocecidien and B
media (L.) Vill. Preslia 33:206-208.
BOS, L. 1970. Symptoms of virus disease
DE LA CRUZ, C. R. 1977. Infectious gal
7(9):17-18.
HIBINO, H., M. ROECHAN, S. SUDAR]
disease of rice (kerdil hampa) tra
lugens, in Indonesia. Contr. Centr. R
IRRI (Int. Rice Res. Inst.). 1972 Annual
IRRI (Int. Rice Res. Inst.). 1976. Annual i
KURIBAYASHI, K. and A. SHINKAI. 1
dwarf (in Japanese). Ann. Phytopatl
LING, K. C. 1972. Rice virus diseases. Int.
LING, K. C. and V. M. AGUIERO. 1970. 1
Phytopathol. 6:5-6. (Abstr.)
LING, K. C., V. M. AGUIERO, and S.
testing resistance to grassy stunt dise
LING, K. C. and E. R. TIONGCO. 1975
rice tungro virus by NEPHOTETTIX
MILNE, R. G. and O. LOVISOLO. 1977
Virus Res. 21:267-341.
NISHI, Y., T. KIMURA and I. MAEJIMA.
plants in Japan (in Japanese, Eng
41:223-227.
PALMER, L. T. and Y. SOEPRIAMAN.
Int. Rice Res. Newsl. 2(5):5-6.
PALOMAR' M. K. and C. T. RIVERA.
Philipp. Phytopathol. 3:27-34.
PLANT PROTECTION INSTITUTE Of
CHEKIANG PROVINCE. 1974. R
China. 104 p.


Phosphotungstate reveals smooth sphe-
rical cores about 55 nm in diam. though
very occasionally the outer capsid may
be retained.
The available information on rice rag.
ged stunt disease seems to fit most of the
above criteria. Consequently, the ragged
stunt virus may be a member of acantho
viruses or fijiviruses (Milne, 1978 persona
communication).




JRE CITED

nerkungen uber die Taxonomie bei Stellar

in plants. 2nd ed., Pudoc, Wageningen. 206
Disease: new threat to rice crops. Greenfiell

MAN, and D. M. TANTERA. 1977. A virl
smitted by brown planthopper, Nilaparva
s. Inst. Agric. Bogor 35:1-15.
port for 1971. Los Banos, Philippines. 238
port for 1975. Los Banos, Philippines. 418 i
52. On the new disease of rice blackstreake
1. Soc. Japan 16:41. (Abstr.)
Lice Res. Inst., Los Banos, Philippines, 142
ist range of the rice grassy stunt virus. Philip

:. LEE. 1970. A mass screening method f
;e of rice. Plant Dis. Reptr. 54:565-569.
Effect of temperature on the transmission
irescens. Philipp. Phytopathol. 11:46-57.
Maize rough drawrf and related viruses. Ad

1975. Causal agent of "waika" disease of ri
h summary). Ann. Phytopathol. Soc. Jap

977. Rice ragged stunt disease in Indones

967. Yellow dwarf of rice in the Philippini

THE ACADEMY OF AGRICULTURE (
e viruses diseases (in Chinese). Agric. Pul









Rice Ragged Stunt in the Philippine


RIVERA, C. T. and S. H. OU. 1965. Leafhopper transmission of "tungro" disease of rice.
Plant Dis. Reptr. 49:127-131.
RIVERA, C. T., S.H.OU, and T. T. IIDA. 1966. Grassy stunt disease of rice and its
transmission by the planthopper Nilaparvata lugens Stal. Plant Dis. Reptr. 50:453-
456.
RIVERA, C. T., S. H. OU, ar.d M. D. PATHAK. 1963. Transmission studies of the orange-
leaf disease of rice. Plant Dis. Reptr. 47:1045-1048.
SENBOKU, T. E. SHIKATA, E. R. TIONGCO, and K. C. LING. 1978b. Transmission
of rice ragged stunt disease by Nilaparvata lugens in Japan. Int. Rice Res. Newsl.
3(2):8.
SHIKATA, E. and Y. KITAGAWA. 1977. Rice black-streaked dwarf virus: its properties,
morphology and intracellular localization. Virology 77:826-842.
SHIKATA, E., K. LEELAPANANG, E. R. TIONGCO, and K. C. LING. 1977. Study
indicates viral nature of rice ragged stunt disease. Int. Rice Res. Newsl. 2(5):7.
SHIKATA, E., T. SENBOKU, E. R. TIONGCO, and K. C. LING. 1978. Rice ragged
stunt virus, a new member of plant reovirus group. Submitted for presentation
at the 4th International Congress for Virology, The Hague, Nertherlands, August
30 September 6.
SHINKAI, A. 1966.. Studies on insect transmissions of rice virus diseases in Japan (in
English summary). Nat. Inst. Agric. Sci., Bull. Ser. C, 14:1-112.
TANAKA, A. and S. YOSHIDA. 1970. Nutritional disorders of the rice plant in Asia. Int.
Rice Res. Inst. Tech. Bull. 10:1-51.
WEERAPAT, P. and S. PONGPRASERT. 1978. Ragged stunt disease in Thailand. Int.
Rice Res. Newsl. 3(1):11-12.
WHITCOMB, R. F. and R. E. DAVIS. 1970. Mycoplasma and phytarboviruses as plant
pathogen persistently transmitted by insects. Ann. Rev. Entomol. 15:405-464.
YOKOYAMA, S., H. SAKAI, S. IMABAYASHI, T. TAKASAKI, N. FUJIYOSHI, and J.
HAGA. 1974. Studies on the "waisei" disease of rice plants. I. Symptoms and the
occurrence (in Japanese). Proc. Asso. Plant Prot. Kyushu 20:117-119.


Jan. & June 1978








Philipp. Phytopathol. 14:58-62
Received for publication: 8 May, 1978




MUNGBEAN VIRUSES IN THE PHILIPPINES
I. IDENTITY OF A VIRUS CAUSING
MOTTLE IN MUNGBEAN

L. T. TALENS

Research Virologist, Institute of Plant Breeding, U. P. at Los Banios, College Laguna,
Philippines.
Grateful appreciation is expressed to Ms. Adelaida L. Lacuata for technical assis-
tance.
This investigation was supported by research grant provided by the Philippine
Government to the Institute of Plant Breeding, U. P. at Los Banios.

ABSTRACT

A virus inducing a mottle reaction and stunted growth in field-grown
green-gram or mungbean [Vigna radiata (L.) Wilczek] was isolated and iden-
tified. Based on thermal stability, serological property, and morphological
features, the mungbean virus isolate was found to possess physical and anti-
genic properties similar to black gram mottle virus (BgMV).
The virus was stable at high temperature (10 min duration at 90-95 C)
and occurred in high concentration (infectivity retained at 10-6 dilution) in
infected mungbean tissues harvested 10-14 days after mechanical inoculation.
The virus-reacted strongly in Ouchterlony double immunodiffusion
test with reference BgMV antiserum. Isometric particles which measured
28-30 nm in diameter were seen in electron microscopic examinations of
partially purified preparations stained with 2% neutral phosphotungstate.
This study describes the first report on the occurrence of mungbean
mottle virus in the Philippines with antigenic properties similar to black
gram mottle virus.


Under natural condition, field-grown
mungbean plants succumb to infection
by several viruses. These viruses, namely:
mungbean yellow mosaic (Nariani, 1960),
mungbean mosaic (Kaiser et al., 1968),
bean common mosaic (Kaiser and Mossa-
hebi, 1974), southern bean mosaic (Zau-
meyer and Harter, 1963), and others
(Thorberry, 1966), are oftentimes found
in mixed infections.
Although the number of viruses that
can occur in one disease complex is unde-
termined, mungbean plants with viral
infections showed symptoms consisting
of mosaic, mottle, yellows, puckering,
leaf curl, bronzing, and little leaf.


In a recent trip to Davao and Cotabato,
southern Philippines in December, 1977,
an experimental plot of mungbean was
observed to be infected with viruses
whereby their identities by field diagno-
sis were difficult to establish. Observa-
tions of insect pests suggested that there
were high incidence and vigorous activi-
ties of Aphis craccivora Koch.
The reduction in yield due to virus
infections seemed high due to severe
stunted plant growth, marked chlorosis
of the foliage and, production of few
flowers and pods. Thus, it was of interest
to investigate the causative viruses so that








eean Viruses in the Pt


methods of control can be determined.
This study reports on the isolation
and identification of a virus infecting
mungbean with serological properties
similar to black gram mottle virus, herein.
after, called mungbean mottle virus.

MATERIALS AND METHODS

The virus described here was isolated
from naturally-infected mungbean plants
collected in an experimental field of Dr
J. A. Soria of the Department of Plani
Pathology, University of Southern Min.
danao, Kabacan, Cotabato. The virus wa,
transmitted mechanically by rubbing dilu
tions of phosphate-buffered homogenate
of infected leaf tissues which contained
1% Celite. Inoculations were done or
primary leaves of 10 to 14-day old mung
bean seedlings with the aid of cheese clott
pad soaked in the virus inoculum.
Thermal inactivation point and dilu
tion end-point analysis of the virus wa!
determined according to procedures des
cribed previously (Bos et al., 1960)
Briefly, extracted sap of leaves frorr
infected mungbean plants with the
mottle virus was heated between 55"C
and 100 C. After heat treatment, the
sap was cooled immediately and rubber
onto the primary leaves of young mung
bean seedlings.
Eight 10-fold dilutions of crude viru:
sap was made in distilled water for the dilu
tion end-point analysis. Ten to 14-day olc
mungbean seedlings grown in 10 to 1!
cm clay pots containing Dowfume
sterilized sand-soil mixture were used fo:
each treatment. Inoculated plants wen
rinsed thoroughly with water and kept it
the greenhouse where temperature flue
tuated between 25-31C. Prompt viral in
fections became visible 10 to 14 day
after mechanical inoculation. Howeve
infected plants were kept under observa


tions for a period of 3 to 4 weeks.
Ouchterlony double immunodiffusioi
test (Ouchterlony, 1962) in 1.0% aga
dissolve in 0.01 M phosphate buffer, pH 7.:
containing 0.85% NaCI and 0.02% NaN:
was used for virus identification. Refei
ence antisera against several isometri
viruses were kindly supplied by Dr. J. F
Fulton of the Department of Plant Pathc
logy, University of Arkansas at Fayette
ville, Arkansas, U. S. A.
Particle morphology and size wer
determined using a JEOL-100U typ
electron microscope. Specimens consistin
of partially purified virus preparation
deposited on Formvar-coated carbon film
reinforced copper grids were stained witl
2% neutral phosphotungstate prior t4
examination in the electron microscope
A drop of 0.01% bovine serum albumii
was routinely added to the virus suspend
sion to enhance homogenous spreading
of the virus particles.
Measurements of the particle size wer
made from electron microscope negative
taken at 20,000 and 50,000 times magni
fiction. Size was measured with a 7:
Bausch and Lomb ocular lens provide<
with a calibrated micrometer scale.

RESULTS AND DISCUSSION

Symptomatology
Field-infected plants showed varyin
degrees of foliar chlorosis, yellow spol
ting of different sizes and with irregula
border, vein-banding, and overall stunte'
growth. Figure 1 shows the trifoliat
leaves of mungbean plant infected witl
the mottle virus. Symptoms of viral ir
fection at an advanced stage consisted c
irregular-sized yellow spots which tende
to coalesce giving the green-yellow mosai
pattern.
Physical properties


ian. & dune l vi








Vnl 14


Fig. 1. Symptoms of mottle v
mungbean.


The in vitro properties of the virus
was: dilution end-point, infection at 10`7
to 10'6 but not at 10"8; thermal inacti-
vation (10 min duration), infection at
90-95C but not at 100C. Longevity in
vitro studies indicated that the virus
was stable after storage for 3 to 4 weeks
at 25C.
The physical properties of MMV
agreed well with those reported by


us infection in trifoliate leaves of


when the virus antigen preparation was
tested against BgMV antiserum and not
against antisera to other isometric viruses
(Fig. 2).

Electron microscopy
Examination of partially purified
virus preparations negatively-stained with
phosphotungstate showed uniform sphe-


D~;1:~~:~n P1..,nnn)knlnav








Jan. & June 1978 Mungbean Viruses in the Philippines I. 61

















7





-6





Fig. 2. Agar gel diffusion pattern of concentrated sap of
mungbean leaf tissues infected with mottle virus.
Center well (A) contained virus antigen. Surrounding
wells contained antisera against (1) bean pod mottle
virus (21 hran ruirnaf manic virus: (M) black


(5) cucumber mosaic
bean mosaic virus,
wilt virus. Type I; ai
strain.


the virus is mechanically transmissible to
a limited number of virus indicator plants
mostly within the Leguminoseae. The
response of selected indicator plants
indicates that MMV, while possessing
antigenic similarities with BgMV, differed
from BgMV (Pathak, 1974).

Mungbean mottle virus appears to be
economically important because of the


irus, cowpea strain;(6) southern
owpea strain; (7) broad bean
cowpea mosaic virus, Arkansas



severe reaction incited by the virus in
mungbean plants. Virus infection of
young, highly susceptible plants could
result in crop losses due to the chlorotic
reaction and stunted growth of the afflic-
ted plant with the concomittant reduc-
tion in flowering, and reduction in size
of pods and seeds.









Philippine Phytopathology


Fig. 3. Electron micrograph of mi
purified virus suspension sl
tungstate. Note the present
particles which measured 28

LITERAT

BOS, L., D. J. HAGEDORN, and L. QUAI
tional identification of legume viruses
FULTON, J. P. and H. A. SCOTT. 1977. Be
transmission by beetles. Fitopatologia
KAISER, W. J. and G. H. MOSSAHEBI. 1
common mosaic virus. Phytopatholog.
KAISER, W. J., D. DANESH, M. OKHOI
pulse crops (edible legumes) in Iran. P
NARIANI. T. K. 1960. Yellow mosaic of
pathol. 13:24-29.
3UCHTERLONY, 0.1962. Diffusion-in-1
Progr. Allergy 6:20-54.
?ATHAK, N. C. 1974. Seed-borne plant v
health testing. Copenhagen. 155 pp.
rHORNBERRY, H. H. 1966. Plant Pests o
Index of Plant Virus Diseases. Agricu
Service, U. S. Department of Agriculti
7.ATTMRVRI W .17 ,A T. T. ATTAPIMT 1


bean mottle virus in partially
ned with neutral 2% phospho-
of empty (b) and full (a) virus
) nm in diameter.

IE CITED

Z. 1960. Suggested procedures for interna-
. P1. Ziekten, 66:328-343.
i rugose mosaic and related viruses and their
rasileira. 2:8-16.
'4. Natural infection of mungbean by bean
o4:1209-1214.
T and H. MOSSAHEBI. 1968. Disease of
it Dis. Reptr. 52:687-691.
iung (Phaseolus aureus L.). Indian Phyto-

methods for immunological analysis. II.

ises identification and diagnosis in seed

importance to North American Agriculture.
ire Handbook N. 307. Agriculture Research
!. 466 pp.


I-1 I .









Philipp. Phytopathol. 14:63-68
Received for publication: 10 May, 1978



CONTROL OF NEMATODES WITH TEMIK 15G ALDICARB
AND ITS RESIDUE IN THE FRUIT OF
GIANT CAVENDISH BANANA

M. O. SAN JUAN and LORNA D. LOZANO

Plant Pathologist and Nematologist, respectively, Twin Rivers Research Center,
Twin Rivers Plantation, Inc. Tagum, Davao del Norte.
Paper presented during the Ninth Annual National Pest Control Conference at the
Philippine International Convention Center, Metro Manila on May 3 6, 1978.
The authors wish to thank Union Carbide Philippines, Inc. for extending some
financial support, aside from providing the experimental nematicide and handling the
residue analysis for the experiment. The services rendered by the Statistical Division of
the Twin Rivers Research Center in analyzing the data are hereby acknowledged.
This article was published with approval of the Research Technology Committee
of TRRC.

ABSTRACT

Temik 15G Aldicarb, a systemic pesticide for the control of insects,
mites and nematodes was experimentally tested for nematode control on
Giant Cavendish banana plants. Results showed that the rates of 10, 20, and
45 g of Temik 15G provided control of plant parasitic nematodes, particular-
ly Radopholus similis, Meloidogyne incognita, Helicotylenchus spp., Praty-
lenchus spp. and Rotylenchus sp.
Increased bunch weight was somewhat apparent within 2 years after
the last nematicide application. Tipping over was less frequent among the
treated plants.
The rate of 10 g (1.5 g ai.) left residue in the fruit pulp ranging from
< 0.02 to 0.04 ppm; 20 g (3 g a.i.) with < 0.02 to 0.08 ppm; and 45 g (6.75
g a.i.) with residues ranging from < 0.02 to 0.35 ppm. From these and other
data submitted, the Environmental Protection Agency set a tolerance residue
of 0.3 ppm for the Aldicarb (2-methyl-2 (methylthio) propionaldehyde
o-(methyl-carbomoyl) oxime.


Several species of plant parasitic nema-
todes parasitize Cavendish bananas. The
most prevalent species are the burrowing,
spiral, root lesion, and root-knot nema-
todes. These species heavily infest areas
planted to abaca, banana, ramie and other
crops.
In bananas, these nematodes are the
most common that reduce fruit yelds
(Feakin, 1972). The economic loss that
results from their infestations is unknown.
However, some estimates range from 20


to 30 percent reduction in yield. Guerent
(1972) found that the growth of plants
growing in poor soils was affected when
the burrowing nematode (Radopholus
similis) population reached 1,000 per 100
g of infested roots. Other growers initiate
chemical control measures when the po-
pulation levels, regardless of soil ferti-
lity, reach 10,000 per 100 g of roots.
Since this population level is an arbitrary
standard, it is not definitely known whe-
ther this is the threshold level most



























of either Temik 10G (Aldicarb), Mocap plots were laid out according to the
10G (Ethoprop) or Nemagon 10G (DBCP) design in which each constituted at least
in 3.78 liters of water. However, soil 64 plants or hills and leaving two rows
treatments at 1 g per 20 cm pot of of buffer plants around each plot or
soil was effective only when the quanti- roughly 1,000 sq. m. per plot.
ty of the nematicide was increased to 3 The plots were applied with Temik
g each pot. 15G at dosages of 10, 20, and 45 g per
Davide and Eloja (1973) conducted hill, and check plots without any chemical
field studies on nematode species parasi- treatment. A randomized complete block
tizing abaca with the use of Temik 10G design with three replications for each of
at 67 kg/ha, and noted improvement of the four treatments were provided. The
growth, particularly in increased number nematicide was applied 1 m around each
of suckers. However, data on fiber yields mat, buried to about 5 cm deep and later
from the treated plots, which could have covered with soil after each application.
served as the ultimate basis for effective The second application of nematicide was
nematode control in bananas are reflec- done in the same manner 6 months after
ted by fruit production with due con- the first.
sideration on the chemical residue result- Monitoring of nematode populations
ing from the nematicide application. was done at monthly intervals following
In fully developed banana plantations, the first sampling of soil and root samples
the root systems of the plants are well 30 days after the application of the nema-
established. Nematode control then be- ticides. Ten samples of 200 to 300 g each
comes laborious and expensive. In view of soil and roots were randomly taken at
of this, a nematicide particularly new depths of 20 to 30 cm from each experi-
for commercial use, has to be experi- mental plot and placed in separate plastic
mentally tested. The present experiment bags. A total of 30 samples each of soil
was designed to determine the rates of and roots were then taken from each
applying Temik 15G that will provide treatment. The soil and roots samples
optimum control of nematodes. In were separately composite from which


Vol. 14


Philippine Phytopathology


addition, the residue levels in the fruit
were also determined.

MATERIALS AND METHODS

The experiment was conducted in the
Ponce area of the Twin Rivers Plantation,
Inc., Tagum, Davao del Norte from April
1975 to October 1976. An area perma-
nently planted to Cavendish banana was
surveyed through soil sampling to deter-
mine the initial population levels of plant
parasitic nematodes. Results of the survey
revealed high nematode populations in
l- . .. rl'1 . .. . . I- 1 I t1 .


economical to control.


Keetch, etal, (1976) reported a 3-fold
increase in banana yield when the plants
were given pre- and post-plant treatments
with DBCP and Nemacur. They found
that post-plant treatments alone were not
as effective. Pre-plant treatment can
either be done by dipping the planting
materials in the chemical solution or
placing the nematicide at the bottom of
the hole prior to planting.
Davide (1973) reported that Meloido-








Control of Nematodes in Banana


uu g son anra uu g root samples were iNemaroae populauonswere conssU1eI-
et aside for processing. ly higher in the non-treated plots than in
Soil samples were then processed fol- the nematicide-treated plots during all
)wing the Cobb sifting and gravity me- sampling periods (Table 1). It can be
hod using 53, 100 and 326-mesh sieves noted that there was practically an
combined with the Baermann-funnel appreciable decrease in nematode popu-
iethod (Thome, 1961). The root samples lation from the soil samples except for
rere cut into pieces, 10 to 20 mm long, the 45 dosage wherein the difference
laced in the waring blender, covered with was not as great as the rest of the treat-
laced in the waring blendor, covered with
ments. The decrease in nematode popula-
istilled water and processed until all the men. The deceased in nematode pop
tion in the 10 and 20 g Temik treatments,
oots have been reduced to small frag-
nts. The residueswere thn wasd when statistically analyzed, were found
rents. The residues were then washed
i. the i ee en to be highly significant over the non-
hrough the different screens and the
treated control, and the 20 g dosage was
rashings from the finest screen were sificanly better than the 45 g treat-
significantly better than the 45 g treat-
llowed to stand in the Baermann funnels meant. While it was expected for the
ment. While it was expected for the
r to o n d i i t highest dosage to be more effective than
ematodes under a dissecting microscope. h o ,h 1u otn w to
the others, the result obtained was to
Fruit samples were also taken from the contrary. The high populations could
nature bunches of Temik 15G-treated probably be attributed to the initially
plants by taking at least 2 kg of fruits high populations in the plots assigned
rom the first, middle and bottom hands for the particular treatment, although


separate polyethylene bags in carton
loxes for shipment to Manila, frozen,
nd later air-freighted to the Morse Lab-
iratories in Sacramento, California,
J.S.A. for residue analysis.
Production records were also taken
rom each treatment by taking the data
>n the hand class, harvesting calibration
ind bunch weights. Records were like-
vise taken on the number of plants that
iave fruited but toppled over and taking
tote of their causes for tipping over.

RESULTS AND DISCUSSION

Parasitic nematodes infesting the
>anana plants in the experimental area
vere predominantly composed of R.
imilis followed by Pratylenchus coffeae,
felicotylenchus multicinctus and M.
ncognita.


Nematode counts obtained from the
00 g root samples did not show signifi-
mt differences among the treatments.
However, the treated plots had only half
ie total number of nematodes as the
on-treated plot. The two nematicide
applications were conceivably inadequate
o bring down drastically the nematode
population in the roots. M. incognita
'as found to complete its life cycle on
celery in less than two months (Perez
nd Castillo, 1973). While the same
species infests the roots of bananas, it
i suggestive that sustained nematicide
applications should be of shorter inter-
als until such time that a negligible
population is attained thereby improve
ie root system.
Plants with poor root systems and yet
ome to fruiting seldom reach up to


in. & June 1978








. slliJuV c ly X IIY I IUIUvol. 14


Table 1. Effect of varying dosages of Temik 15G on nematode control and on fruit
production of Cavendish bananas.


Nematode Count
Bunch Weight
400 g Soil 100 g Roots kg
Dosage
Total Mean Total Mean Total Mean

10 g 9712 511.16 27600 1452.63 521.05 27.4S
20 g 7087 373.00 27588 1452.00 298.19 26.22
45 g 15301 805.32 27420 1443.16 456.15 24.01
0 19479 1025.21 42977 2261.95 507.40 26.70



these plants over specially when they are production can possibly be attained
not properly propped. In most instances, much later from the succeeding followers
however, fruit losses observed in the which benefited most from the treatment.
experiments particularly from the un- The nematicide treatments left aldi-
treated plots were due mainly to very carb residues in the fruit. Fruit samples
nnnr rnnt cretpm n Tinnino nuar tha ...


do not come up to the required matu-
rity.

The bunches harvested from plots
treated with 10 g of Temik 15G were
heavier than those harvested from other
plots (Table 1). Significant differences
in bunch weights between the untreated
plants and those treated with 20 and
45 g were obtained. It was not
ascertained, however, whether these
heavier bunch weights will be maintained
or surpassed by the other treatments as
more plant followers are produced. It
is expected that fruit production per
unit area will be much higher in the treated
plots due to increased nutrient uptake
through an improved root system. While
there were reductions in the nematode
populations in the treated plots, fruit
production has not correspondingly in-
creased within the 2-year period. Increased


IyLru WJLuwVu vaiying wu rcLuIillaLuuulln ui
the active chemical (Table 2). It can be
noted that there was no appreciable
difference in the residue between the
1.5 g and 3 g treatments except in those
fruits with the higher concentration 90
days after the second treatment. How-
ever, the residue levels decreased there-
after.
Higher concentrations of Aldicarb in
the fruit were detected from those plants
given the 6.75 g active material per plant.
An increasing trend was obtained up to
60 days after each nematicide treatment.
The residue levels decreased gradually
after that period as the fruits reached
maturity.
The tolerance as requested for com-
bined residue of the insecticide Aldi-
carb (2 methyl-2-methylthio-propional-
dehyde 0-methylcarbamoyl) oxime and
its cholisterenase-inhibiting metabolites


Vol. 14








Control of Nematodes in banana


able 2. Aldicarb residues in the fruit at
treatments


No. of Days
)ate of Sampling After
Treatment

First Treatment
4-18-75 15
5- 3-75 30
5-19-75 46
6- 2-75 60
7- 2-75 90
8- 1-75 120
9-30-75 180
Second Treatment
10-16-75 15
10-31-75 30
11-15-75 45
12- 1-75 61


fferent sampling dates after two Temik 15(



Aldicarb Residues (ppm)

1.5 g 3.0 g 6.75 g



0.03 0.03 0.01
< 0.02 < 0.02 0.2,
< 0.02 < 0.02 0.1'
0.05 0.03 0.7'
0.02 0.09 0.51
< 0.02 < 0.02 0.1)
< 0.02 < 0.02 < 0.0!


< 0.02 0.02 0.1(
< 0.02 0.10 0.3-
0.04 0.04 0.31
0.04 0.13 0.71


2-28-76 179 (Composite of all rates < 0.02)


2-methyl-2 (methylsufinyl) propionalde- 45 g around each mat of Giant Cavendish
hyde 0-(methylcarbamoyl) oxime and bananas appreciably reduced the popula-
2-methyl-2 (methylsolponyl) propional- tion of plant parasitic nematodes in the
dehyde 0(methylcarbamoyl) oxime in or soil but not as much in the roots.
on the raw bananas was 0.3 ppm. After a Increased fruit production was not yet
thorough evaluation of all pertinent data apparent during the 2-year period, thus
on acute oral and dermal LD50 (lethal necessitating a more prolonged period of
dose) studies, etc., the Environmental 3 to 5 years of data collection from
Protection Agency (EPA) concluded that which the efficacy of the nematicide can
the tolerance of 0.3 ppm will protect the be fully assessed. It was observed, how-
public health. ever, within the short period that treated
Temik 15G Aldicarb is absorbed plants had less incidences of tip-over.
through the roots up to the fruit. The Subsequent field experiments can be
highest dosage level used in the experi- made to determine optimal dosages and
ment was shown to be the maximum frequency and time of application not
quantity of Temik 15G to apply to be only to reduce nematode population
within the residue tolerance level set by levels but ultimately increase fruit pro-
EPA. duction.

SUMMARY AND RECOMMENDATIONS Plants treated with 45 g of Temik


lan. & June 1978









-u~y = ...7 ,.-y-..**-.-s


tne C.r.A. uosages lower man me on
above which also reduced the nematod
populations in the soil left residues i


LITER

DAVIDE, R. G. 1973. Evaluation of tl
cognita affecting banana. Philipp. I
DAVIDE, R. G. and A. L. ELOJA. 197
of application of Temik 10G for t
topathol. 9:67-75.
FEAKIN, S. D. ed. 1972. Pest control i
seas Plant Research, College House
GUERENT, R. 1972. Relation entire le
croissence du bananier. Fruits 27:3
KEETCH, D. P., B. E. REYNOLDS an
and post-plant nematicides for the
Citrus and Sub-tropical Fruit Rei
cultural Abstracts 46: November 1
PEREZ, ROSALINDA A. and M. B. C.
gical effects of Meloidogyne incogn
THORNE, G. 1961. Principles of Nemi
York, Toronto, London. 533 p.


Lme iJuLa u- iUL ma Ulill uti appiuvU
tolerance level.



TURE CITED

ree nematicides for control of Meloidogyne
gr. 57:187-197.
SEvaluation of the different methods and r
e control of nematodes on abaca. Philipp. I

bananas. Pans Manual No. 1. Center for O
Wrights Lane, London W8 5SJ, England. 12
populations de Radopholus similis Cobb e
1-337.
L. A. MITCHELL. 1976. An evaluation of
control of plant parasitic nematodes in bana
-arch Institute, Nelspruit, South Africa. H(
76.
ITILLO. 1973. Development and histopathi
ta on celery. Philipp. Phvtopathol. 9:46-52.
ology. McGraw-Hill Book Company, Inc., I


__








-77


STUDIES ON THE WHITE RUST DISEASE OF KANGKONG
IPOMOEAA AQUA TICA FORSK.)

MA. VICENTA C. GACUTAN and TRICITA H. QUIMIO

Graduate student, Department of Botany, Diliman, Quezon City and Assistant
Professor, Department of Plant Pathology, College of Agriculture, U. P. at Los Banos.
College, Laguna, respectively.
Sincere thanks are due to Dr. F.R. Uyenco, Dept. of Botany, U.P. Diliman, for the
use of the laboratory during the conduct of this study.


ABSTRACT

The histopathology of Ipomoea aquatica Forsk. infected with white
rust organism was studied and the morphology of the organism critically
examined for proper identification. The sporangia germinated both by
production of germ tube and by releasing flagellated zoospores. Symptoms
on the inoculated leaves appeared 2 weeks after inoculation although inter-


Saw., than A. ipomoeae-panduranae (i

Ipomoea aquatica Forsk. or "kang-
kong", is a common vegetable crop in the
Philippines as well as in Southeast Asia. It
is usually grown all throughout the year
in flooded areas although there are varie-
ties that grow on dry upland soils. Like
any other vegetable crops, this plant is
proned to several diseases which are
apparently much neglected. In the Phil-
ippines, the only disease reported
on this crop was the root-knot caused by
Meloidogyne incognita (Benigno and
Quebral, 1977).
The white rust disease may be general-
ly regarded as a disease of limited impor-
tance. However, where stems, petioles
and leaves are harvested, any disfigura-
tion or distortion due to the disease
seriously reduces the market value of
the crop.
Two species of Albugo had been repor-
ted to cause white rust disease of
Ipomoea. One is A. ipomoea-panduranae
(Schw.) Swing (Singh and Bedi, 1965)


hw.) Swing.

and the other one is A. ipomoea-aquaticae
Saw. (Ho and Eddie, 1969). They differ
very slightly in some morphological
aspects yet considered quite distinct
species.

This paper reports on some aspects of
the histopathology of the diseased plants
and the morphology of the causal
organism leading to the proper identifi-
:ation of the causal fungus attacking
"kangkong" in the Philippines.

MATERIALS AND METHODS

Source of infected plants
The infected plants were taken from a
cangkong field at the U.P. campus,
)iliman, Quezon City and a backyard
cangkong garden at the UPLB housing
Lrea. The healthy plants used for inocula-
:ion were taken from the same areas.
Germination and inoculation
Spores scraped from infected plants








Philippine Phytopathology


were used for inoculation. The sporangia
were first germinated in a Syracuse dish
containing distilled water. The process
of germination was observed with the
use of a compound microscope. The
resulting zoospore suspension, together
with some ungerminated sporangia, was
dropped on healthy leaves, wetting both
surfaces. The inoculated plants were
then covered with a plastic bag for the
first 24 hours after inoculation.

Histopathology
Different stages of the infected leaves,
stems and petioles were cleared in satu-
rated chloral hydrate solution. Infected
portions were then cut with a freezing
microtome at 50-100 u thick and
mounted in cotton blue lactophenol.

RESULTS AND DISCUSSION

Germination and inoculation
Germination started with the forma-


tion of a small papillae through which the
protoplasmic contents would eventually
be released (Fig. 1). As the protoplasm
was extruded, differentiation of the zoo-
spores would bring about the bursting
of the vesicle, releasing them simulta-
neously. Occasionally, zoospore differen-
tiation occurred within the sporangium
and motile zoospores would be released
individually. Kajomchaiyakul and Brown
(1976) reported the same type of germi-
nation on A. tragopogi. The zoospores
would swim around for some time before
encysting and germinating into a short
germ tube.
Sporangia of the Albuginaceae nor-
mally germinate indirectly producing
motile biflagellate zoospores (Webster,
1970). Direct germination has been ob-
served, however, in several species of
Albugo including A. ipomoea-aquatica
(Eddie & Ho, 1970) and A. ipomoea-
panduranae (Harter & Weimer, 1929).
As observed in this study. sooraneia








C


Fig. 1. Stages in the germination of the sporangium, and formation of zoospores. Germi-
nated in water and observed at 1 hour interval.


Vol. 14


:
-i :,; :
..?









-UJ. x UUJIC j. JL I


111~ ruor u~~r ul rr~6~u6


IaiU UlllllllalticU ulCUy UIi UiV bUllilC
of the host.


Infection
Inoculated leaves showed symptoms
Af infection 1 to 2 weeks after inocula-
tion with zoospore and sporangia sus-
pension. Pustules formed were scattered
throughout the leaf, indicating localized
infection (Fig. 2). Foliage symptoms
First erupted through the lower epidermis
suggesting stomatal penetration. Natu-
rally-infected plants showed indication
of systemic mode of infection by pro-
iucing symptoms and signs first on the
base of the blade then along the midrib






























A
Fig. 2. White rust symptoms on Ipomoea
white rust pustules on the lower ai
infected young leaves.


anu lmiiany uu laogv laturi, veins krrg. J).
Ho and Eddie (1969) reported cases of
both systemic and localized infection
caused by the white rust fungus.
Normally the general shape and size of
the leaves were not seriously affected
although the upper surface of the leaf
often appeared wrinkled and yellow. In
case of leaf bud infection, the organs
appeared totally wrinkled and malformed.
Stems were also infected producing galls
and swellings, eventually cracking open.
Histopathology
Penetration of the fungus was directly
through the sub-epidermal cells by the
germ tube produced by the germinating






























B
luatica produced by artificial inoculation. A
I upper surfaces of the leaves. B. puckering o








Philippine Phytopathology


Fig. 3. Naturally-infected plant showing
symptoms starting from the lower
base of the leaf to the midrib.


sporangia (Fig. 4). Penetration by zoo-
spores nor stomatal penetration was not
observed. Kajornchaiyakul and Brown
(1976) reported that either zoospores of
A. tragopogi penetrated the stomata and
became encysted within the substomatal
cavity or they encysted in the water and
their germ tube seeked the stomatal
opening. Eddie and Ho (1970) also ob-
served penetration of A. ipomoea-aqua-
tica through the stomates by the formation
of appresoria and infection pegs. Direct
penetration, as observed in the present
study, however, had been reported by
Webster (1970).
Different stages of intercellular myce-
lial development were noted in the tissues
of the host as early as 3 days after inocu-
lation. Fig. 5 shows sub-epidermal hyphal
growth indicating early stage of infection.
Hyphal growth was intercellular and


Fig. 4. A germinating sporangium directly penetrating the sub-epidermal
cells giving rise to intercellular hyphae.


Vol. 14








wnlTe LnusT ulsease c


Fig. 5. Early stage of infection shoe
epidermal portion of the leaf


several globose haustoria were produced
along the entire hyphal length (Fig. 4).
As the disease advanced, disintegration
of tissues occurred as well as chloroplast
in-balance resulting to the early yellow-
ing of infected tissues which was the early
symptom of infection. At a latter stage,
the hyphae formed a mat of closely
intertwining mycelia just beneath the
lower epidermis. On this layer were
formed the initials of the developing
club-shaped conidiophores (Fig. 6). The
increasing growth of conidiophores and
sporangia exerted pressure on the epi-
dermis causing it to bulge into white


ring aggregation of hyphae at sub-



epidermal layer brought about dispersion
of the spores by either wind or water.
Sexual reproductive structures were
observed only on the stem tissues but
never on the leaves. They were confined
mostly on the pith together with ramifying
mycelium. Hyperplasia of infected cells
caused swelling and enlargement of the
diseased stem.
The oogonium and oosphere had a
smooth wall but as fertilization took
place with an attached antheridium (Fig.
9a), the oospore wall became warty (Fig.
Qh Ev Pntinllu pithpr thp nnonnial wall


Y









4hilippine mhytopathology


Fig. 6. A mat of intertwining hyphae giving rise to conidiophore initials.
*w ^" ,

^) ^ ^ ^9


74








a. & June 1978 White Rust Disease of Kangkong to

































Fig. 8. Close-up of a group of sporangial chains showing disjunctor cells
(arrows) holding the sporangia in chain.


A closer examination on the morpho-
agy of the present organism showed
he following features: Sporangia 17.2
o 25.0 x 14.0 to 23.4 microns, wall of
uniform thickness and without distinct
quatorial thickness; conidiophore 31.2
o 5.15 x 14.2 to 21.9 microns; oospore
vall irregularly tuberculate or warty,


nows u e mIUUlMpa.ilvc IliealuricllIiili allnu
descriptions of the two species of Albugo
reported to cause white rust of Ipomoea
zquatica. The figures show that the
present organism under study more
closely resembled A. ipomoeae-aquatica
than A. ipomoeae-panduranae.









































Fig. 9. Stages in oospore develop
tube. B. Warty oospore insi


ent. A Clavate antheridium (arrows) attack
! a smooth-walled oogonium. C. Mature war








White Rust Disease of I


ble 1. Comparative characteristics of three


Size
Criterion A. ipomoeae-panduranae /


nidiophore 15 x 30


species of Albugo from Ipomoea aquatica


u ) and description
ipomoeae-aquaticae A. sp. organism
under study)

.1-48.1 x 16.5-22.0 31.2-51.5 x 14.2-
21.9


14-20 x 12-18 17.1-24.8 x 16.5- not distinct 17.2-
21.5 25.0 x 14.0-23.4
)ospore irregular curved ridges, irregularly tubercu- irregularly tuber-
papillate 25-55 diam late 52.3-74.3 diam culate 49.9-68.6
diam


LITERATURE CITED

IENIGNO, D. A. and F. C. QUEBRAL. 1977. Host index of plant diseases in the Philip-
pines. Handbook. 183 p.


. 'ri,.LrJr j&. 1.. uilu u. J. vyv m wUVic ..U-
and their control. Bulletin 99, Wisconsi
O, B. W. C. and H. H. EDDIE. 1969. W1
moea aquatic in Hongkong. Plant Dis.
.AJORNCHAIYAKUL, P. and J. F. BROV
affecting infection of sunflower by 1
66:91-95.
INGH, H. and K. BEDI. 1965. Some aspect
logy of Ipomoea aquatica Forsk. i
(Schw.) Swingle. Phytopath Zeitschi. 5
HIRUMALACHAR, M. J., M. D. WHITE]
and oospore formation in Cystopus (Al
TEBSTER, J. 1970. Introduction to fungi. C


kgr. Expt. Sta., 53-56.
- _- IAJ


eptr. 53:959-962.
i. 1976. The infection process and factors
bugo tragopogi. Trans. Brit. Mycol. Soc.

of the morphology, anatomy, and embryo-
'ected with Albugo ipomoea-panduranae
201-215.
kD and J. S. BOYLE. 1949. Gametogenesis
ugo) evolvuli Bot. Gaz. 83:487-491.
abridge Univ. Pres. 424 p.


77








R


RESISTANCE IN SOYBEANS [GL YCINE MAX (L.) Merr.]
TO ROOT-KNOT NEMATODES AND STATISTICAL ANALYSIS OF
CORRELATIONS OF ASSESSMENT PARAMETERS

M.B. CASTILLO, AVELINA P. RODIL and B. K. LIM

Associate Professor and former Graduate Students, Department of Plant Patholo
College of Agriculture University of the Philippines at Los Banos, College, Laguna.
A portion of this study was the M. S. thesis of the second author.
The authors thank Mr. R. Navarro of the Department of Agronomy, College
Agriculture, U. P. at Los Banos and Mr. B. Legaspi of the Bureau of Plant Industry (BI
Los Banos, Laguna for the seeds of the varieties used in this study.

ABSTRACT

Twenty-eight soybean varieties exhibited varying degrees of resistance
to mixed populations of Meloidogyne incognita, M. acrita, M. arenaria, and
M. javanica in pots.
Nine varieties singly infected by M. incognita, M. arenaria and M.
javanica also differed in degrees of resistance to each species. Based on
resistance ratings or averages of computed indices of the different assessment
parameters, namely numbers of galls produced, nematodes recovered/g of
roots, egg masses/g of roots, and eggs/egg mass, and percentage yield reduc-
tion, Tainung #3, Strain #12 and Strain #99 were consistently the most
resistant varieties to the three species. Except percentage yield reduction, all
parameters correlated positively with resistance rating to M. incognita.
Number of galls produced and number of nematodes recovered/g of roots
also correlated positively with resistance ratings to both M. arenaria and
M. javanica. Positive correlation also existed between percentage yield re-
duction and resistance rating to M. javanica. All the assessment parameters
that correlated positively with resistance rating to particular species were
equally efficient, except number of eggs/egg mass which was less efficient
There were positive correlations between number of galls produced by either
of the three species and number of nematodes recovered/g of roots. Corre-
lations of other combinations of parameters varied with species.

The potential importance of the root- crops, like soybeans, are constraints t
knot nematode, Meloidogyne incognita, the use of nematicides and crop rotation
on soybeans in the United States has been Thus stress in research has been laid o
reported (Kinloch, 1974). In the Philip- the identification of resistant varieties
pines, surveys (Castillo, 1971; Castillo, (Holston and Crittenden, 1951; Critter
1975) and experiments (Valdez, 1968; den, 1955; Kinloch and Hinson, 197
Castillo, 1976) also indicated the im- and 1974).
portance of Meloidogyne spp. on the Many soybean varieties had show
crop. promise of resistance to the locallU


Tirr-- *ni--i._-











and Ballon, 1974; Castillo, unpublished were pooled in 13-cm diam. clay pots in
dnttl+ Th. n- -n+n ;--r

t species.


same way as it was pot. Nematode-infested s(
nematode-resistant inoculum because it was
m and Castillo, in to egg masses still attached
tion to be derived isolated egg masses, isolh
ture mass screening larvae in an earlier experid
cnot nematodes. Castillo, in press2).
Five seeds of each vari
ID METHODS four times, were planted p
ing the mixed nematode
ig soybean varieties pots were randomly arrai
ed cultures of four elevated wire cage outside t
ecies The seedlings were thinne
'enty-eight soybean pot, 6 days after seeding
promise of resistance insects and fungal diseases
:reenings to one or by foliar spraying, when(
tode species (Meloi- with a combination of n
elected. Nematode- 16.8% EC and benoi
elected. Nematode-
:rved as check. at manufacturers' recom
Moisture was supplied to
used. Single egg needed, but no fertilization


d) was


erior
eces,
and


e on
lean i
s'). 1
[ld be
resistai

MAT

!ening
resist
-knot
'arietih
-ties tt
irelimi
e root
n/ sr
eptiblh
eemato


from rnntQ nf rnunpaon oiplr ,,it Wau lU aln rateu lor galling, using a 1 to


ULait, anlu elery, respectively, were
separately maintained for several genera-
ions on potted tomato (VC 11-1) in the
reenhouse. These nematode cultures
rere used as source of inocula.
Preparation of nematode inocula and
planting. Soil of each nematode cul-
ire was thoroughly mixed and 10 300-cc
imples obtained. These samples were
processed for nematodes using the routi-
ary sieving-Baermann funnel technique.
ased on the average numbers of nema-
)des recovered per sample, the amount
F soil from each culture that would
contain approximately 250 active nema-
)des of the particular species was com-
uted. Subsequently, the four cultures

'Lim, B. K. and M.B. Castillo. Scre


representative replicate of each variety
'as selected and from this a 1-g root
simple was randomly obtained. The root
imples were stained in acid fuchsin and
eared in lactophenol. To assess the
lative nematode infection and repro-














iing soybeans for resistance to reniform


UC ils iU Lu LUIIII
It also aimed to d


'J,
.OS


1111 ULU= Lluservaalulls. .......Y -v ictivau, vi r ,ttjlt Vi ULU lUUl


-4. 1- r _ ____ nrlrf~C11fl n thP M-fi-l r in -1- +r fill -1,h


Resistance in So


1n








Philippine Phytopathology


duction, the numbers of nematodes in- Individual galls of the resistant varieties,
side the roots, egg mass/g of roots and however, were usually discernible. In
eggs/egg mass were determined with the addition, randomly-obtained 1-g root
_'J _r J:-* *.' ~ _;---.-- "I.--A : nm 1 f^-m Ann~l^ ~ n* <. , n4-:nsr


--:A ,,


nnl and


teasing it apart with egg masses
:s. with a di:
soybean varieties to composite
not nematode species pooled soi
for nemat(
Nine varieties that funnel te
to the mixed cultures quantified
acrita, M. arenaria, scope. Yie
he preceding experi-
ark 63 was likewise Analysi
as the nematode- Except th
Five seeds of each from the ,
ed in each 13-cm all the d,
staining baked soil. analyzed a
hinned to three/pot, the variety
,. with DMF
based on
used and inoculation inoculated
)nth old cultures of inoculated
aria and M javanica probable
masses of each nema- of the v,
introduced close to for the dil
Id seedlings of the root gallin
Three replicate pots roots, nur


300-cc
>m the
icessed
rmann
atodes
micro-
d.
Fata.
overies
icated,
;tically
:ans of
o level
a were
ons in
e non-
>f the
entials
indices
lamely
m 1 g
roots,


inoculated with each nematode species
were randomly arranged in an elevated
wire cage and the plants cared for in the
same manner as in the preceding experi-
ment.

Data collected. Eleven weeks after
inoculation, fhe roots were washed and
the relative lumber of galls in roots of
all plants in each pot determined by
visual observation. Difficulties were en-
countered in counting the usually
large and overlapping galls in Clark 63.


each variety and considering the average
of these indices as the resistance rating
of the variety to the particular nematode
species. Based on the resistance ratings,
the degrees of resistance of the varieties
were compared.
Correlations of resistance ratings and
the assessment parameters and of
different combinations of parameters.
Lesser number of parameters than that
used in this study could be used in practi-
cal mass screening of varieties for resist-


Sdisse
iistanc
h of t
Variet,
libited
M. in,
1 M. j
nt we:
eluded
ceptib]
iety
n. cli
: seed
'eek af
Venmat
cedurn
incog?
-e used
e spe
ts of
'erent


VY U


Vol. 14








I DP..tO.n in iRnvhanne


ance to root-knot nematodes. Thus, it is
essential to determine which of these
parameters are the most efficient in
determining resistance ratings and, there-
fore, degree of varietal resistance. To
attain this objective, correlation of
resistance rating and each of the assess-
ment parameters used was determined by
statistical analysis of r values (correlation
coefficients). Analyses of correlation of
the different parameters were also made.


RESULTS AND DISCUSSION

Screening of promising soybean varieties
for resistance to mixed cultures of four
root-knot nematode species
Based on root gall rating and nematode
recovery and reproduction in roots, the
28 soybean varieties exhibited varying
degrees of resistance to the mixed cul-
tures of M. incognita, M. acrita, M.
arenaria and M. javanica (Table 1).
Gall ratings. Except in CES XVI-103,
Taichung E 32 and UPL SY-2, the gall
ratings of the promising varieties were
significantly lower than in the nematode-
susceptible Clark 63, which exhibited
severe galling. Only light to moderate
galling were exhibited by the promising
varieties other than CES XVI-103,
Taichung E 32 and UPL SY-2.

Nematode recoveries from roots.
Numerically lower numbers of nematodes
were recovered, from roots of the promi-
sing varieties than from roots of Clark
63. Among the promising varieties, Strain
# 12, CES XVI-16, CES XVI-24, PIN,
and CES XVI-112 had the least numbers
of nematodes in their roots (less than 10,
compared to 405 in Clark 63).
Nematode reproduction. The num-
bers of egg masses found in roots and the
numbers of eggs/egg mass numerically


differed among varieties, with Clark 63
having the most numbers. Except in CES
144 W, CES XVI-106, CES 434, Taichung
E 32, and UPL SY-2, 10 or less egg masses
were found in the promising varieties,
compared to 46 in Clark 63. Bethel,
SL-6, Strain # 12, and Strain # 15 had
the least numbers of eggs/egg mass (less
than 30, compared to 452.9 in Clark
63).

These findings show that reduced
root galling and nematode recovery and
reproduction were generally associated
with resistance. However, it is difficult to
rank the degrees of varietal resistance to
the mixed nematode cultures, based on
all the parameters used. This is because
deviations from the normal trend of
direct relationships among the parameters
occurred in certain varieties and it is not
known which parameter should be given
more importance. For instance, CES IV-
14 W and CES XVI-37 had comparatively
low gall ratings, but the numbers of ne-
matodes recovered from their roots,
numbers of egg masses or numbers of
eggs/egg mass were higher than in some
of the varieties with higher gall ratings.
These deviations could probably be attri-
buted not only to the differences in host
specificities of the nematodes, but also
to the differences in reactions of the
varieties to each of the nematode species.
Resistance in nine soybean varieties to
each of three root-knot nematode species
General observations. Based on the
nematode-susceptible Clark 63, reduced
galling and lower nematode recoveries
from roots and soil, number of eggs/egg
mass and percentage yield reduction
were likewise generally associated with
resistance in the nine varieties used,
although not consistently shown by
statistical analyses (Table 2). Whether
or not the differences in nematode re-


A V 4 M









Philippine Phytopathology


Table 1. Gall rating of promising soybean varieties grown in potted soil containing
mixed cultures of Meloidogyne incognita, M. acrita, M. arenaria and M. javanica
and nematode recovery and reproduction 2 months after planting


Number of
nematodes Number of Number of
Gall recovered/ egg masses/g of eggs/egg
Variety rating g of rootsY rootsY mass

Bethel 2.0 a 18 2 6.5
CES IV-14 W 2.3 abc 186 42 154.0
CES XV-4 2.0 a 35 2 53.0
CES XVI-16 2.3 abc 10 3 88.0
CES XVI-24 PIN 2.0 a 10 3 49.0
CES XVI-33 WIN 2.0 a 63 3 135.0
CES XVI-37 2.1 a 103 4 283.0
CES XVI-42 PIN 2.5 abc 95 10 211.0
CES XVI-103 4.0 d 60 6 250.0
CES XVI-105 PIN 3.0 50 5 80.0
CES XVI-106 2.9 c 274 22 118.0
CES XVI-112 2.8 bc 6 3 227.0
CES 434 2.1 a 84 17 116.0
Ford 2.0 a 57 1 65.0
K 475 2.4 abc 26 2 63.0
L 114 2.0 a 21 4 35.0
SL-6 3.0 c 146 3 27.0
Strain # 12 2.0 a 4 1 7.0
Strain # 15 2.0 a 71 4 28.0
Strain # 78 2.0 a 15 4 88.0
Strain # 99 2.0 a 12 3 34.0
Taichung E 32 4.2 d 186 26 146.0
Tainung # 3 2.0 a 73 9 63.0
Tainung # 4 2.0 a 25 7 68.0
Taiwan 2.4 abc 16 8 95.0
TK # 5 2.0 a 110 7 45.0
Wayne 2.0 a 11 4 90.0
UP SY-2 4.0 d 148 21 167.8
Clark 63
(Susceptible check) 4.2 d 405 46 452.9

WApproximately 250 larvae of each nematode species were initially present in the
mixed cultures.
XMeans of four replicates. Data were based on a 1 to 5 severity scale, determined
from relative percentages of galled portions of root systems as follows: 1 = 0% (no galls);
2 = 1-25% (light galling); 3 = 26-50% (moderate galling); 4 = 51-75% (severe galling);
and 5 = over 75% (very severe galling). Different letters indicate significant differences at
the 5% level with DMRT.

YBased on only one representative replicate.

ZBased on counts from 10 randomly-obtained egg masses, when there were 10 or
more egg masses, otherwise on counts from all egg masses found.


Vol. 14










Resistance in Soybeans


Table 2. Galling of resistant soybean varieties grown in potted soil singly infested with
Meloidogyne incognita, M. arenaria and M. javanica, nematode recovery and
reproduction and yield reduction 11 weeks after inoculation X


Number of nematodes
Number of recovered from: Number of Number of Percentage
galls egg masses/ eggs/egg yield
Variety produced 1 g roots 300 cc soil g of roots massy reductions


76.6 b
174.0 c
9.3 a
34.3 ab
6.0 a
11.6 a
34.0 ab
9.0 a
4.5 a


74.7 be
130.7 d
84.7 bc
97.7 cd
12.0 a
51.7 ab
51.0 ab
23.0 a
23.0 a


M. incognita
72
20
120
64
96
88
20
152
112


179.0 c 382.3 e 272
M. arenaria


145.0 c
200.0 d
17.0 a
67.3 b
11.7 a
10.3 a
17.3 a
10.5 a
57.3 b


223.0 d
273.0 e
46.0 a
177.6 c
83.3 ab
37.6 a
56.3 a
35.5 a
115.0 b


10.0 de
13.0 e
3.0 ab
9.0 cd
2.0 a
4.0 ab
5.3 ab
6.0 bc
6.0 bc


93.7 ab
122.0 ab
160.0 bc
114.3 ab
227.7 e
196.7 cd
110.3 ab
89.0 a
148.0 abc


19.3 f 345.0 e


11.3 cd
25.0 e
2.7 a
11.6 cd
3.0 a
3.0 a
10.3 bcd
10.0 bc
15.3 d


K 475
L 114
Strain # 12
Strain # 15
Strain # 78
Strain # 99
Tainung # 3
Tainung # 4
TK#5
Clark 63
(check)


K 475
L 114
Strain # 12
Strain # 15
Strain # 78
Strain # 99
Tainung # 3
Tainung # 4
TK# 5
Clark 63
(check)


K 475 112.0 ab
L 114 232.0 cd
Strain # 12 8.3 a
Strain # 15 84.3 ab
Strain # 78 145.0 bc
Strain # 99 27.0 a
Tainung # 3 102.3 ab
Tainung # 4 67.7 ab
TK # 5 145.7 bc
Clark 63
(check) 300.0 d


85.0 ab
253.3 c
12.0 a
56.3 ab
67.3 ab
42.7 ab
69.0 ab
86.6 b
103.0 b

408.6 d


56.6 a
106.6 ab
201.0 bc
295.3 cd
275.5 cd
101.0ab
171.0 abc
288.5 cd
218.6 be


6.0 ab 357.0 d


5.0 a
13.0 b
1.0 a
3.0 a
2.0 a
3.0 a
4.0 a
3.0 a
4.0 a


204.0 bcd
33.6 a
175.0 bcd
193.0 bcd
219.0 cd
96.7 ab
55.0 a
100.0 ab
107.0 abc


5.0 a 249.0 d


XInoculum level/pot was 50 nematode egg masses. Except data on number of
nematodes recovered from 300 cc soil which were not replicated, data are means of
three replicates. Different letters indicate significant differences at the 5% level with
DMRT.
YBased on counts from 10 randomly-obtained egg masses, when there were 10 or
more egg masses, otherwise on counts from all egg masses found.
ZBased on yield differences between plants grown in baked soil and plants grown in
nematode-infested soil (non-inoculated control).


227.5 d 278.0 e 420
M. javanica


19.2 a
24.7 ab
18.1 a
23.0 ab
29.2 ab
18.7 a
29.9 ab
35.9 ab
28.9 ab

48.5 b


9.6 a
23.2 ab
16.2 ab
23.3 ab
25.1 ab
21.2 ab
12.4 a
24.7 ab
30.5 ab

37.2 b


19.2 a
29.3 a
15.1 a
25.8 a
29.1 a
28.5 a
24.1 a
32.4 a
29.2 a

57.4 b


Jan. & June 1978








flL 1~. flt....t -- At. 1 -


coveries from the roots could be attri- rates of development among the species
buted to failure of some nematodes to could have attributed for these inconsis-
enter the roots and/or to death of some tencies. Further investigations, however,
nematodes after entering could not be are needed to confirm this observation.
determined from the experiment. In all the three species, data on galling,
Data on number of egg masses/g of nematode recovery from roots, number
roots were not consistent in the three of egg masses, and number of eggs/egg
nematode species. It is difficult to ascer- mass significantly varied among the
tain if this inconsistency could be attri- resistant varieties. Nematode recoveries
buted to the differences in the rates of from the soil were also apparently varia-
development and reproduction of the ble in those varieties, although signifi-
species, since the determination of the chance of differences could not be deter-
number of egg masses was not synchro- mined because the data were obtained
nized with the egg-laying stages of the from only one replicate. These observa-
three species in the different varieties. It tions suggest that, although percentage
is probable, however, that in susceptible yield reductions did not significantly
and less resistant varieties, egg masses differ among the resistant varieties, vary-
were produced earlier than in the more ing degrees of resistance to M. incognita.
resistant varieties and the eggs hatched M. arenaria and M. javanica existed in
before the numbers of egg masses were these varieties.
determined. Lim and Castillo (in press3) Comparison of the degrees of varietal
reported that M. incognita reached the resistance to each nematode species. -
egg-laying stage within 21 to 25 days and Based on resistance ratings or averages
32 to 37 days after larval inoculation of computed indices for galling, nema
of Clark 63 and L114, respectively; ee .gg. _. ..


was noted in the latter up to 60 days
from inoculation. In the present study,
M. incognita egg masses were signifi-
cantly more in Clark 63 than in any of
the resistant varieties, 11 weeks after
inoculation. It is probable that most of
the egg masses in Clark 63 were already
those of the second generation nema-
todes, while egg masses in the resistant
varieties could still be those of the first
generation nematodes. On the other
hand, lower numbers of M. arenaria egg
masses were observed in Clark 63 than in
K475, L114, Strain # 15, and TK # 5;
the numbers of M. javanica egg masses
significantly vary among the varieties,
including Clark 63. Differences in the

3Lim, B.K. and M. B. Castillo. Inte


uil vancusti vaIncu iuiilg IlneIClalue
species (Table 3).
Tainung # 3, Tainung # 4, Strain # 12,
and Strain # 99 were the four most
resistant varieties to bothM. incognita and
M. javanica. Except Tainung # 4, which
ranked fifth, these varieties were also
the most resistant to M. arenaria. The
rankings of K 475, Strain # 15, Strain
# 78, and TK # 5 varied with nematode









rtions of Meloidogyne incognita and Roty


.1 .








Iesis5anve in ooy u~uba


tle -. Resistance ranng of soybean vanenl
arenaria and M. javanica based on it
roots, production of nematode egg
reduction a


Nematode Egg ms
Galling recovery product
Variety index index index

M. incot

nung # 4 2.00 1.20 2.50
ain # 12 2.00 3.00 1.50
ain # 99 2.00 1.80 1.50
nung # 3 3.00 1.80 1.50
# 5' 2.00 1.20 2.50
ain # 78 2.00 1.20 1.00
175 4.00 3.00 4.50
ain # 15 3.00 4.20 3.50
.14 6.00 4.80 5.00
rk 63
'check 6.00 6.00 6.00
M. arer
ain # 99 1.50 1.20 1.20
ain # 12 1.50 1.20 1.20
inung # 3 1.50 1.20 3.60
ain # 78 1.50 1.80 1.20
inung # 4 1.50 1.20 3.00
175 4.50 4.80 4.20
# 5 3.00 2.40 4.80
ain # 15 3.00 3.60 4.20
L14 6.00 6.00 6.00
irk 63
(check) 6.00 6.00 1.80
M. java
rain # 99 1.50 2.25 3.00
inung # 3 2.25 2.25 3.0C
rain # 12 1.50 1.50 3.0C
inung # 4 2.25 3.00 3.00
475 2.25 2.25 3.0C
rain # 15 2.25 2.25 3.0C
K # 5 3.75 3.00 3.0(
rain # 78 3.75 2.25 3.0C
114 5.25 4.50 6.0(
ark 63
(check) 6.00 6.00 3.0C

alndices for galling, nematode recovery
eld reduction were determined from the st;
ble 2 for numbers of galls produced, nema
ots, and eggs/egg mass, and percentage yi.
bitrary value of 6 to the highest mean ar


ro cuTn U j ivlullu~ugynl nJ1lugi a, ti.
ices of galling, nematode recovery from
masses and eggs and percentage yield




Egg Yield Resistance
a production reduction rating
index index (avg)

ita
1.20 4.50 2.28
3.00 3.00 2.50
4.20 3.00 2.50
8.80 4.50 2.52
2.40 4.50 2.52
6.00 4.50 2.94
1.80 3.00 3.26
1.80 4.50 3.40
1.80 4.50 4.42

6.00 6.00 6.00
'ia
2.25 4.50 2.13
3.75 4.50 2.43
3.00 3.00 2.46
5.25 4.50 2.85
5.25 4.50 3.09
1.50 3.00 3.60
3.75 4.50 3.69
5.25 4.50 4.11
2.25 4.50 4.95

6.00 6.00 5.16
ica
2.25 3.00 2.40
1.50 3.00 2.40
4.50 3.00 2.70
2.50 3.00 2.70
4.50 3.00 3.00
4.50 3.00 3.00
3.00 3.00 3.15
5.25 3.00 3.45
1.50 3.00 4.05

6.00 6.00 5.40

egg mass production, egg production, and
istical means (differentiated by letters) in
ades recovered/g of roots, egg masses/g of
d reduction, respectively, by assigning an
equally-divided lower values (determined


a. & June .wi








- **-*rk----~- -' V--~~


species. L 11 t wa1 s WUlSsl tU uy uLUe iaL4
resistant to the nematodes, among the
resistant varieties.
Strain # 12, Strain # 99 and TK # 5
were also earlier found (Lim and Castillo,
in press') resistant to the reniform nema-
tode (Rotylenchulus reniformis) in a field
screening. In the same screening, K 475
was moderately resistant, while Strain
# 78, Tainung # 3 and Tainung #4 were
moderately susceptible. Since root-knot
and reniform nematodes are perhaps
the most important nematode pests of
field legumes and vegetables in the


resistance ratings to m. incognita, M.
arenaria and M. javanica (Table 4). Num-
ber of egg masses/g of roots and number
of eggs/egg mass also correlated positive-
ly with resistance rating to M. incognita
at 1% and 5% levels, respectively, but not
with resistance ratings to M. arenaria and
M. javanica. Percentage yield reduction
correlated positively (1% level) with
resistance rating to M. javanica, but not
to M. incognita and M. arenaria. These
observations show that under the condi-
tions of the experiment, the parameters
that can be used to identify resistance to


useful in nematode control ana breeding uL LOUL", mIUIIVi UI Vr a, ,.1
roots, and number of eggs/egg mass;
programs. to M. arenaria, number of galls produced
The use of resistance rating, based on and number of nematodes recovered/g of
computed indices of more than one roots; and to M. javanica, number of galls
assessment parameters, to compare the produced, number of nematodes re-
degrees of varietal resistance to nema- covered/g of roots and percentage yield
todes was first attempted in reniform reduction. Based on analyses of signifi-
nematode-resistant soybean varieties (Lim chance of differences between correlation
and Castillo, in press') and on other coefficients, all the parameters that
results of the present experiment, the correlated positively with resistance
principles involved in this method could rating to particular species were equally
be employed in objective rankings of efficient assessment parameters, except
varietal resistance in crops to other ne- number of eggs/egg mass which was less
matode pathogens. It appears, however, efficient (Table 4). Apparently, the
that better comparisons of varietal resis- varieties used in this study are not
+nn-- n -l ltiT1 -F.m hA mnta .___f_-rn_ _- --rr f. a. ^ h. A u *--,--i,-n Al


rogeneous populanon or vanletles arenari


the assessment parameters and of differ-
ent combinations of parameters. Statis-
tical analyses, based on r values (correla-
tion coefficients), showed highly signi-
ficant positive linear correlations of
number of galls produced and number of
nematodes recovered/g of roots with


nematode-resistant soybean varieties (Lim
and Castillo, in press') and on other
crops resistant to root-knot nematodes
(Christie, 1949; Madamba, et at, 1965).
Highly significant positive linear corre-
lations existed between number of galls
produced by either of the three nema-


I- -








Resistance in Soybeans


Table 4. Computed r values (correlation coefficients) of assessment parameters and resis-
tance rating of soybean varieties to Meloidogyne incognita, M. arenaria and
M. javanica of different combinations of parameters Y


Parameter

Number of Number of Number Percentage Resistance
Parameter nematodes egg masses/ of eggs/ yield rating
recovered/ g of roots egg mass reduction
g of roots


Number of galls produced

Number of nematodes
recovered/g of roots

Number of egg masses/g
of roots
Number of eggs/egg mass
Percentage yield reduction


Number of galls produced
Number of nematodes
recovered/g of roots
Number of egg masses/g
of roots

Number of eggs/egg mass
Percentage yield reduction


Number of galls produced 0.9257**
Number or nematodes
recovered/g of roots
Number of egg masses/
a of roots


Number of eggs/egg mass
Percentage yield reduction


M. incognita
0.8075** 0.9162** 0.3760NS 0.4197NS 0.9183**a

0.8802** 0.7040* 0.6005NS 0.9288**a


0.3940NS 0.5614NS
0.5575NS


0.9260**a
0.6366*b

0.5788NSb


M. arenaria

0.9653** 0.4933NS 0.0189NS 0.3253NS 0.8989**a

0.5377NS 0.0008NS 0.2873NS 0.9239**a


0.2967NS 0.0241NS 0.5969NSb
0.7092* 0.2748NSb

0.5490NSb


M. javanica
0.6180NS 0.1693NS 0.7412'


0.9216**a


0.5758NS 0.1663NS 0.8553** 0.9398**a


0.0002NS 0.1884NS 0.4456NSb

0.2769NS 0.4146NSb
0.8105**a


YCompared to the tabulated r values (0.6320 and 0.7650, for 5% and 1%, respec-
tively); NS = not significantly different; = significantly higher at the 5% level and ** =
significantly higher at the 1% level.

ZFor each nematode species, different letters indicate significance of difference in
r values, following Fisher's (1954) procedure.


Jan. & June 1978








Philippine Phytopathology


tode species and number of nematodes egg masses/g of roots (1% level) and with
recovered/g of roots (Table 4). Differen- number of eggs/egg mass (5% level),
ces in varietal interactions with the three number ofM. arenaria eggs/egg mass with
nematode species were indicated by percentage yield reduction (5% level),
correlation analyses of other combinations number of galls produced by M. javanica
of parameters. Positive correlations also with percentage yield reduction (5% level),
existed between number of galls produced and number ofM. javanica recovered/g of
by M. incognita with number of egg roots with percentage yield reduction
masses/g of roots, number ofM. incognita (1% level).
recovered/g of roots with number of



LITERATURE CITED

CALINGA, R. H. and F. B. BALLON. 1974. Studies on the pathogenic reactions of
different varieties of vegetables to Meloidogyne incognita. Philipp. J. Plant Ind.
39:107-114.
CASTILLO, M. B. 1971. Reniform nematode, Rotylenchulus sp., in mungo, soybean
and peanut soils at the UPCA Central Experiment Station. Philipp. Phytopathol.
7:61-63.
CASTILLO, M. B. 1975. Plant parasitic nematodes associated with mung bean, soybean
and peanut in the Philippines. Philipp. Agr. 59:91-99.
CASTILLO, M. B. 1976. Relative susceptibility, resistance and tolerance of selected
crops to inoculation with Meloidogyne spp. in pots. Philipp. Phytopathol. 12:17-23.
CHRISTIE, J. R. 1949. Host-parasite relationships of the root-knot nematodes, Meloi-
dogyne spp. III. The nature of resistance in plants to root-knot nematodes. Hel-
minthol. Soc. Wash. Proc. 16:104-108.
CRITrENDEN, H. W. 1955. Root-knot nematode resistance of soybeans. Phytopatho-
logy 45: 347 (Abstr.)
FISHER, R. A. 1954. Statistical methods for research workers. Oliver and Boyd,
Edinburgh. 354p.
HOLSTON, E. M. and H. W. CRITTENDEN. 1951. Resistance in soybeans to root-knot
nematodes. Phytopathology 41:562 (Abstr.).
KINLOCH, R. A. 1974. Response of soybean cultivars to nematicidal treatments of soil
infested with Meloidogyne incognita. J. Nematol. 6:7-11.
KINLOCH, R. A. and K. HINSON. 1972. The Florida program for evaluating soybean
[Glycine max (L.) Merr.] genotypes for susceptibility to root-knot nematode
disease. Soil and Crop Sci. Soc. Flor. Proc. 32:173-176.
KINLOCH, R. A. and K. HINSON. 1974. Comparative resistance of soybeans to Me-
loidogyne javanica. Nematropica 4:17-18 (Abstr.).
MADAMBA, C. P., J. N. SASSER and L. A. NELSON. 1965. Some characteristics of the
effects of Meloidogyne spp. on unsuitable host crops. N. C. Agr. Exp., Tech. Bull.
169. 34 p.
VALDEZ, R. B. 1968. Survey, identification and host-parasite relationships of root-
knot nematodes occurring in some parts of the Philippines. Philipp. Agr. 51:802-
824.


Vol. 14



























T.

ABSTRA(

oper, Nilaparvat


ig, 1977; Fernando, et at, 1977;
hap, 1977; Stapley, 1975).
n the other hand, the brown plant
'er's ability to transmit grassy stunt











brown planthopper differ in their abili
to transmit grassy stunt.

MATERIALS AND METHODS

Nymphs of three biotypes of t
brown planthopper provided by t
Entomology Department, IRRI, we
allowed to feed on grassy stunt-diseas
Plants of Taichung Native 1 (TN1) for
or 4 days. After the acquisition fee
ing, the insects were subjected to a da
serial transmission test. The insects we
transferred individually to 7-day (
healthy TN1 seedlings in test tubes
1 insect/seedling per tube and covert

Table 1. Transmission characteristics <
biotypes


Active Latent
transmitters period
Biotype (%) (days)

1 12.1 10.2
2 10.1 9.4
3 11.4 10.2

aNone of the differences for each

Each tube contained a small amount
water for the seedling. The insects we
confined there for 24 hours for seedli
inoculation. The insects were th
transferred to inoculate a new set
seedlings. The transfer was repeated
daily intervals until all insects died. Aft
each transfer, the inoculated seedlin
were removed from the test tubes, trar
planted in clay pots, and kept in ti
greenhouse for the symptoms to develop
The seedlings were periodically spray
with insecticides to keep them insect fre


S examined for grassy stunt symptoms. 1
seedling infection was used as an indi
tor of the infectivity of the insect dur
the time of inoculation. The test i
repeated with 4,690 insects, more tl
1,500 of each biotype, and 27,374 i
S culated seedlings.

RESULTS AND DISCUSSION

Various aspects of the ability of
three biotypes to transmit grassy sti
were compared (Table 1). The act
S transmitters ranged from 2 to 27%
the insects tested in the replicate te
The average percentage of active tra


rice grassy stunt disease by Nilaparvata lu



Retention Disease- Infect
period transmitting seedlii
(days) days (%) (no./ini

21.6 54.9 9.
20.2 56.2 9.
20.8 64.3 9.

laracter are statistically significant.

F mitters did not differ significantly am<
S the three biotypes. The latent period
: the causal agent in the insect varied fr
S 4 to 37 days. The differences in lat
f period among three biotypes were i
t statistically significant. All biotypes w
able to retain their infectivity al
molting; hence, all three biotypes I
transstadial passage. The retention per
of the infective insects varied from 4
48 days after acquisition feeding. But
average retention period was simt
among the three biotypes. The disea











100% of the period from the day the insect tests were observed and recorded. The life
became infective until the insect's death, spans of the insects varied from 3 to 57
But the average percentage of disease- days. The nymphal stadia varied from 5
transmitting days did not differ signifi- to 15 days. Some insects did not molt
:antly among the three biotypes. During because they died early; others molted
the insect's life span, an infective insect five times to become adults. The inter-
infected as many as 30 seedlings. But the vals between two molts varied from 1 to
average infected seedlings per infective 11 days. But the three brown planthopper
insect did not differ significantly among biotypes used in the transmission study
the three biotypes. Consequently, the did not differ significantly in those deve-
three biotypes had similar ability to lopmental features (Table 2).
transmit grassy stunt by the above trans- The field incidence of grassy stunt is
mission features.

able 2. Developmental features of the brown planthopper biotypes used in the study
of grassy stunt transmission

Molting
Life span Nymphal stadia Molts interval
3iotype (days) (days) (no.) (days)

1 26.1 10.1 2.9 2.9
2 24.9 9.3 2.8 2.9
3 24.3 10.6 3.1 2.9

aNone of the differences for each feature are statistically significant.

The developmental features of the determined by factors such as popula-
brown planthopper influence some of the tion of insect vector, availability of the
above transmission features. For instance, disease source, and varietal susceptibility
the life span of the insect affects the to the insect vector and to the disease.
retention period. If insects of a biotype The present results suggest that the abi-
die earlier, that biotype has a shorter lity of the brown planthopper to transmit
retention period. Similarly, the trans- grassy stunt in the field is not the major
stadial passage can be determined only factor causing variations in disease inci-
when the insect molts. Therefore, the dence at least, not among the three
developmental features of 1,072 brown current biotypes in the Philippines.


LITERATURE CITED

ABEYGUNAWARDENA, D. V. W., C. M. BANDARANAYAKA, and C. B. KARANDA-
WELA. 1970. Virus diseases of rice and their control. Trop. Agr. 126:1-13.
ANJANEYULU, A. 1974. Identification of grassy stunt, a new virus disease of rice in
India. Curr. Sci. 43:416-417.








.Itlppine K Eny upiuLuI'J'uy


in Taiwan. Paper presented at the brown planthopper symposium, IRRI, Los Banos
Philippines, April 1977. 45 p. (mimeo.)
FERNANDO, H. E., D. SENADHEERA, Y. ELIKEWELA, H. M. DE ALWIS, and C
KIDAGAMAGE. 1977. Varietal resistance to the brown planthopper, Nilaparvate
lugens (Stal), in Sri Lanka. Paper presented at the brown planthopper symposium
IRRI, Los Banos, Philippines, April 1977. 15 p. (mimeo.)
GOPALAKRISHNAN, R., N. GOPALAN, K. M. GEORGE, and S. N. SHAMUGHAN
1973. Grassy stunt epidemic in Kerala. Agri. Res. J. Kerala 11:84.
HARAHAP, Z. 1977. Breeding for resistance to the brown planthopper and grassy stun
Indonesia. Paper presented at the brown planthopper symposium, IRRI, Lo
Banos, Philippines, April 1977. 15 p. (mimeo.)
HSIEH, S. P. Y. and R. J. CHIU. 1970. The occurrence of grassy stunt in Taiwan (il
Chinese, English Summary). P1. Prot. Bull. 12:136-140
IRRI (International Rice Research Institute). 1970. Annual report for 1969. Los Banoi
Philippines. 266 p.
IRRI (International Rice Research Institute). 1976. Annual report for 1975. Los Banos
Philippines. 479 p.
LING, K. C. and V. M. AGUIERO. 1967. Breeding for efficient transmitting colony c
Nilaparvata lugens, vector of rice grassy stunt virus.
LING, K. C. 1977. Transmission of rice grassy stunt by the brown planthopper, p. 73-8&
In The rice brown planthopper, compiled by Food and Fertilizer Technolog
Center for the Asian and Pacific Region, Taipei, Taiwan, Republic of China.
PATHAK, M. D., C. H. CHENG, and M. E. FORTUNO. 1969. Resistance to Nephotetti
impicticeps and Nilaparvata lugens in varieties of rice. Nature 223:502-504.
RIVERA, C. T., S. H. OU, and T. T. IIDA. 1966. Grassy stunt disease of rice and it
transmission by the planthopper Nilaparvata lugens Stal. Plant Dis. Reptr. 54:45W
456.
SOEPRIAMAN, Y., K. SUNENDAR, L. T. PALMER, O. MOCHIDA, and T. SURYANA
1976. An endemic of frassv stunt virus of rice in Indonesia. PaDer Dresented at th


Iu









lilipp. Phytopathol. 14:93-98
received for publication: 10 August, 1978

POPULATION DYNAMICS OF PLANT PARASITIC NEMATODES. II.
ROTYLENCHUL US RENIFORMIS ON MONOCULTURED
BUSH SITAO IN NEMACUR-TREATED AND
NON-TREATED FARMER'S FIELD

R. H. CALINGA and M. B. CASTILLO

Respectively, Senior Plant Pathologist, BPI, Economic Garden, Los Batos, Laguna
nd Associate Professor, Department of Plant Pathology, College of Agriculture, U. P.
t Los Bafios, College, Laguna.

ABSTRACT

Changes in Rotylenchulus reniformis population on bush sitao in
Nemacur-treated and non-treated beds were determined during wet and dry
season plantings in a relatively well-drained farmer's field in Cale, Tanauan,
Batangas.
Nematode population density was relatively constant or changed only
slightly during the first few weeks of cropping. Density drastically peaked on
the 5th week from seedling emergence during the wet season and on the
4th week during the dry season. Reductions in densities resulted after the
peak periods and, during the dry season, the reduction on the 5th week was
followed by a steady rise until a second peak was reached on the 7th week,
then by a steady decline towards the end of the cropping period.
During the wet season, the suppressive effect of preplant soil applica-
tion of Nemacur at the rate of 4 kg a. i/ha lasted only up to 7 weeks from
seedling emergence and nematode control was not reflected by crop yield.
Increasing the application rate to 19.3 kg a.i./ha gave better and consistent
nematode control throughout the second cropping (dry season) and a corres-
ponding yield increase of 28.4%.

Nematode assays before and after the 1st month, then drastically increased


:s or poputanon oullu-up oi me renm-
m nematode, Rotylenchulus renifor-
, on susceptible moderately suscep-
le and resistant crops (Castillo, et at,
76). Recently, observations of monthly
:tuations in the population of the
natode on mung bean, cowpea and
bean related the trends in popula-
i build-up with root growth and nema-
.e development and reproduction (Cas-
o, et al, in press'). Generally, popula-
a densities changed only slightly during

'Castillo, M. B., M. B. Arceo and J.
rasitic nematodes. I. Rotylenchulus renifor
elds of field legumes. In press at Philipp. Ag


s oemg ooserveu at last narvesL. Imce
Snematode assays were made at shorter
tervals, the relative periods during
rich the densities actually peaked on
e three crops could not be ascertained.
The present study was conducted to
termine the changes in R. reniformis
population on bush sitao at various
riods during wet and dry season plant-
gs in a farmer's field. It was also con-
Icted to determine the duration of the
maticidal effect of Nemacur, and its

Litsinger. Population dynamics of plant
in a poorly drained soil and its effect on











consequent influence on crop yield

MATERIALS AND METHODS

Experimental area
An upland farmer's field in Cale
Tanauan, Batangas, measuring appro.
ximately 365 sq. m and previously
planted to eggplant was used in the
experiment. The clay loam soil was rela
tively well drained, with pH 6.
First cropping period
The field was prepared by means of
hand tractor and divided into three 4.5
27.0 m blocks, separated by 1.0-m walk
Eight 1.5 x 6.0 m beds, half a metei
apart, were constructed per block. Du
ring land preparation, Nemacur [ethyl
3-methyl-4 (methylthio) phenyl (1-methyl
ethyl) phosphoramidate ] a systemic ne
maticide, was incorporated into the soil ol
four adjacent beds of each block at the
rate of 72 g/9 sq m (4 kg a. i./ha). Three
days after nematicidal application (Jun(
2, 1977), bush sitao (Vigna sinensis x V
sesquipedalis) was planted in four row:
(30 cm apart) per bed (three seeds/hill
with 30-cm distance between hills).
Green pods were primed whenever
necessary and the total harvest fronr
each bed was recorded. Final harvesting
was done on July 31, 1977.
Second cropping period
The field was again prepared in the
same manner as during the 'first cropping
with care being practiced to maintain
the integrity of each bed. During land
preparation, Nemacur was again incor.
porated into the soil of the same beds
receiving this treatment during the first
cropping period. This time the applica.
tion rate of the nematicide was increased
to 347 g/9 sq. m (19.3 kg a. i./ha). A


week after nematicidal application (Nov
ember 17, 1977), bush sitao was planted
in the same manner as in the first crop
ping period.
Priming of green pods and recording
of harvest from each bed were also done
Final harvesting was done on January
23, 1978.
Care of experimental plants
Handweeding was done 3 weeks aftei
planting. No fertilization was made bui
control of insect pests was provided by
foliar spraying with Azodrin 168, al
manufacturer's recommended rate, when.
ever necessary. Plants were purely rain.
fed.
Determination of nematode population
densities
The initial densities of plant parasitic
nematodes were determined after land
preparation immediately before planting
for the first cropping. Sampling for
nematodes was done by randomly col
lecting 10 approximately 50-cc soil sub.
samples from each bed. The subsamples
collected from each bed were pooled anc
a composite 400 cc soil obtained. During
the first cropping period, subsequent
nematode assays were made at 2, 4, 5, 7
and 8 weeks after seedling emergence
Nematode densities were monitored al
weekly intervals starting from 1 week
after emergence up to 9 weeks during
the second cropping period. Sampling
for nematodes was done in the same
manner as during the determination
of initial densities, except that the
subsamples were collected close to
plant roots and the composite sample
obtained from each bed consisted of
400 cc soil and 1 g roots.
Active nematodes were extracted
from soil samples through the routine






































'erimental area. of R. reniformis population at w4
.*.- --1 -~( J^ - - - .' _


emacur-treatea ana non-arearea oeas
Id corresponding crops yields during
vo cropping periods
Rotylenchulus predominated over the
other genera of plant parasitic nema-
>des during the two cropping periods.
he species was identified asR. reniformis,
ised on measurements of diagnostic
lorphological characters. Changes in
populations of the other genera did not
)pear to be related with cropping time
id, thus, data on these nematodes
ere excluded.
First cropping period. The seedlings
nerged in about 3 days after seeding for
ie first cropping. Nematode determina-
ons at different periods showed that
, reniformis population in non-Nemacur-
eated beds was very low 2 weeks
'ter emergence, increased only slightly


lowed that the population of the nema-
ide in non-Nemacur-treated beds was
latively constant during the first 3
eeks, then drastically peaked during
ie 4th week (Fig. 1). The population
as reduced to less than 50% during the
h week then rose up again during the
h week until a second peak (lower
lan the first) was reached on the 7th
eek from emergence. Thereafter, it
eadily declined up to the 9th week or
hen the crop was finally harvested.
)il application with Nemacur at the rate
' 19.3 kg a. i./ha reduced the R. renifor-
is population throughout the cropping
:riod. Recovery from Nemacur-treated
ids averaged only 10.4% of the recovery
om non-treated beds. Relatively higher
coveries from Nemacur-treated beds
ere obtained during the last 3 weeks of


~_ _









I


Recovery of Rotylenchulus reniformis at various periods from N
itao during wet and dry seasons. During the wet season, Nemac
Iry season, 19.3 kg a. i./ha.




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