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Daftar isi
170
J. HPT J.
Tropika
HPT Tropika Vol. 19, No. 2,ISSN:
2019:1411-7525
170-178
Vol. 19, No. 2, September 2019 E-ISSN: 2461-0399
Pages: 170–178 DOI : 10.23960/j.hptt.119170-178

ANTIVIRAL ACTIVITY OF CASHEW NUTSHELL EXTRACT

AGAINST Cowpea mild mottle virus ON SOYBEAN

Wuye Ria Andayanie1, Wahidin Nuriana1, & Netty Ermawati2

1
Department of Agrotechnology, Faculty of Agriculture, Universitas Merdeka Madiun, Indonesia
Jl. Serayu 79 Madiun 6311
2
Politeknik Negeri Jember, Department of Agriculture and Production, Indonesia
Jl. Mastrip Jember
E-mail: wuye.andayanie@gmail.com

ABSTRACT

Antiviral activity of cashew nutshell extract against Cowpea mild mottle virus on soybean. Cowpea mild mottle virus
(CPMMV) is one of the important viruses infecting soybeans. The CPMMV disease management usually done by controlling
the vector with insecticides that cause negative impact to the environment and non-target insects. Therefore, it is important
to find safer alternative to control the disease such as the use of plant extracts. Cashew nutshells (CNS) are waste during
cashew processing and contain several antimicrobial substances. This study aimed to evaluate the potential of CNS extracts
to inhibit CPMMV infection. The experiment was designed with a randomized block design with 4 replications and each
treatment unit consisted of 10 plants. The application method consisted of spraying the leaves before and after virus inoculation
and mixing the sap with CNS extract. CNS extract concentrations tested were 0.75; 1.5; 3.0; 6.0%. The result showed that CNS
extract applications can decrease the disease incidence and increase the incubation period. However, plants treated with CNS
extracts at concentration of 3.0% and 6.0% before virus inoculation and mixing CNS extracts at concentration of 1.5; 3.0 and
6.0% with CPMMV sap showed that the virus titer was detected negative by ELISA using CPMMV antiserum. The results
indicated that the CNS extract has potency as antivirus for CPMMV. All treatment combinations can suppress viral infections
without phytotoxicity, except for CNS 6.0%.
Key words: antiviral, cashew nutshell extract, Cowpea mild mottle virus (CPMMV), soybean

INTRODUCTION is insufficient for the needs of farmers (Saleh, 2007;

Andayanie & Adinurani, 2014; Andayanie et al., 2017).
The soybean (Glycine max L.) productivity of Today, the disease control strategy is using the chemical
Indonesia is quite low and unable to meet domestic insecticide against the virus vectors, but it is considered
needs, so the government continues to import soybean to be ineffective and has a negative impact on the
about 2.5 million tons every year. Some of the factors environment, humans, and biological resources
causing the low productivity of soybeans are climate (Andayanie et al., 2019).
factors, planting area and attack of pests and diseases. Plant systemic resistance to viruses can be
The mild mottle disease caused by Cowpea mild mottle activated by applying inducing agents from plant
virus (CPMMV). The mosaic disease caused by extracts. Gunaeni et al. (2015) reported that the active
Soybean mosaic virus (SMV) and stunt disease by compound in the extracts of the japanese glorybower
Cucumber mosaic virus soybean strain (CMVs) are (Clerodendrum japonicum Thunb.) and periwinkle
important viruses of soybeans in Indonesia. Symptoms (Catharanthus roseus L.) can increase resistance to
of CPMMV infection can not be distinguished from Pepper leaf curl virus infection in red chili. In addition,
mosaic symptoms caused by SMV. This disease can chrysanthemum (Tanacetum vulgare L.) flower extract
reduce soybean yields around 14–18% and difficult to has antiviral activity against Cucumber mosaic virus
control (Andayanie et al., 2011). This is because the and Potato Y virus in tomato plants (Petrov et al., 2016).
disease is spread by vectors, such as whitefly (Bemisia Cashew nutshell (Anacardium occidentale L.) is a
tabaci Genn.) and transmitted through soybean seeds waste in the process of cashew seeding, has many
(Tavassoli et al., 2008). Young soybean plants are more antimicrobial activities such as antibacterial and
susceptible to CPMMV than adult plants. The disease antifungal (Garcia et al., 2018; Sudjaroen et al., 2018).
control strategies were carried out by using resistant This extract also has activity as an insecticidal,
varieties, even though the number of resistant varieties molluscicidal, and herbicidal (Towaha & Achmadi, 2011;
Andayanie et al. Antiviral Activity of Cashew Nutshell Extract 171

Andayanie & Ermawaty, 2019; Andayanie et al., 2018). inoculation method and inoculated mechanically to the
Cashew nutshell contains anacardic acid compound Willis soybean variety at 10 days after planting (DAP),
(76.93%), cardols (12.75%), cardanol (4.66%), the infected plants then nurse in the screen house. On
2- methyl cardanol (3.81%) (Andayanie et al., 2019). the 21 days after inoculation (DAI), the infected plants
Anacardic acid (6-pentadecylsalicylic acid) is a showed specific symptom of infection such as mosaic
derivative of salicylic acid substituted by a long alkyl with vein clearing, and chlorosis. The CPMMV isolate
chain (Aderiye et al., 2015). Salicylic acid triggers an with specific antiserum CPMMV (DSMZ) was detected
induce the activity of Pathogenesis-Related (PR) by positive reaction on DAS ELISA. The young soybean
proteins, such as peroxidase, chitinase, β-1.3 glucanase, leaves that showed specific symptom of infection
and β-1.4 glucosidases, prevent the viral multiplication, cultivated at 28–42 days after planting and stored at
spread, and localize the viral infection, although it needs 4 oC.
a retention period for systemic resistance after induced
by the extract (Murphy et al., 2001; Elbeshehy, 2017). Extraction and Treatment of Cashew Nutshell.
Salicylic acid derivative compounds in cashew nutshell Cashew nutshells were dried to a moisture content of
have not been reported for their antiviral effect on plant 15–20% and ground to produce a 400 Mesh powder.
viruses. Therefore, this study aimed to evaluate the Cashew nutshell powder (500 g) was moistened with
potential of CNS extracts to inhibit CPMMV infection n-hexane and ethanol (3:1) for 4 hours to produce the
in soybean plants. mass. Percolation was done by moving the mass
gradually into the percolator and adding hexane and
MATERIALS AND METHODS ethanol (3:1) solvents, then incubated at room
temperature for 24 hours. The extract evaporated at
Research Site. The study was conducted from May 45 oC with a low-pressure vacuum (500–550 mmHg)
to October 2018 in the Laboratory of Plant Disease, to produce concentrated extracts.
Faculty of Agriculture, Universitas Merdeka Madiun.
Cashew nutshell was obtained from cashew producer Incubation Period and Symptoms. The incubation
in Pondok Village, Ngadirojo District, Wonogiri Regency. period was calculated from CPMMV inoculation until
the initial symptoms appear on the plant.
Experimental Design. The experiment was arranged
in a randomized block design (RBD) consisting of 14 Disease Incidence and Disease Severity. The
levels of treatment: 12 experimental treatments, disease incidence and disease severity were observed
CPMMV infected plants as positive control and healthy weekly for each plants starting from 21 days after
plants as a negative control. Each treatment was CPMMV inoculation until 49 DAI.
repeated 4 times and each treatment unit consisted of
10 plants. The 12 types of experimental treatment Disease incidence. The disease incidence was
consisted of cashew nutshell extract treatment before calculated using following formula:
CPMMV inoculation, after CPMMV inoculation and n
mixing the extract with CPMMV sap, with the DI   100%
N
concentration of 0.75; 1.5; 3.0 and 6.0%. DI : disease incidence
n : number of plants showing CPMMV infection
Planting the Test Plants. The Willis soybean variety symptoms
and Chenopodium amaranticolor, respectively 2 and N : number of observed plant
4 weeks after planting were used as test plants. The
plants were planted in polybags containing a mixture of Disease severity. The disease severity was calculated
soil and manure at 2:1 ratio and maintained in a screen using following formula:
house.
 (n  v)
DS   100%
Inoculum Multiplication. The source of inoculums NV
were obtained from the Laboratory of Plant Disease, DS : disease severity
Faculty of Agriculture, Universitas Merdeka Madiun n : number of infected leaves with specific score
since June 2018. The inoculums were isolated from the v : disease severity score
CPMVV infected soybean in Ngawi. The inoculums N : number of observed leaves/plant
then purified in C. amaranticolor using the single spot V : highest severity score
172 J. HPT Tropika Vol. 19, No. 2, 2019: 170-178

The scores of disease severity were performed Serological Detection of CPMMV. The CPMMV
based on Arif & Hassan (2002) with minor modifications detection was carried out at 28 DAI by DAS ELISA
based on symptom patterns as follows (Figure 1): method. This method used CPMMV antiserum with
1 = healthy plants, mild yellow blotches (mottles) procedures according to the recommendations of the
2 = mild yellow blotches, no wrinkle (10–30%) antiserum maker (DSMZ, Braunschweigh, Germany)
3 = clear yellow blotches, slightly mosaic (30–50%) CPMMV infected plants without extract treatment
4 = Severe mosaic symptom (51–70%), leaf distortion, (positive control) and healthy plants (negative control)
plant dwarf were used as a comparison. Each treatment used 10
5 = Severe mosaic symptom (>70%), plant dwarf or plants and from each plant 4 leaf samples were taken.
dead The virus titer or virus accumulation was quantified by
ELISA Reader (Bio-Rad Model 550) at a wavelength
Disease Development and Inhibition. The total area of 405 nm.
under the disease development curve (AUDPC = Area
Under Disease Progress Curve) were calculated at 21, Data Analysis. The obtained data were analyzed by
28, 35, 42, and 49 DAI based on the formula (Strange, analysis of variance (ANOVA) and the effect of
2003):
AUDPC   1

n Y  YI1 
t i 1  t i 
significantly different treatments was followed by
Duncan’s multiple range test at a significance level
i 1 2 of 5%. In order to obtain homogenous data, prior
analizing, the data of disease incidence and disease
Y1 = data on i-th observation severity was transformed using arcsin x  0 .5 .
Y i+1 = data on i-th + 1 observation
t i+1 = time at i-th observation RESULTS AND DISCUSSION
ti = time at i-th +1 observation
n = total number of observation Selection of Cashew Nutshell Extract
Concentration. Application of cashew nutshell extract
The percentage of systemic disease inhibition (P) with various concentrations showed different abilities
CPMMV on the soybean due to the cashew nutshell in suppressing CPMMV infection. Observation of the
extract were calculated by: disease incidence was started since the early symptoms
appear on the plants treated by cashew nutshell extract
AUDPC of possitive control - AUDPC experiment
P  100% at 21 DAI. On the positive control, the yellow blotch
AUDPC of possitive control was first appeared at 17 DAI, then developed into a
mild mosaic and very severe mosaic. Plants with the
The percentage of CPMMV local inhibition in the treatment of cashew nutshell extract at a concentration
C. amaranticolor induced by the cashew nutshell of 0.75; 1.5% before CPMMV inoculation and cashew
extract treatment was calculated by: nutshell extract treatment at a concentration of 0.75;
ab 1.5; 3.0% after CPMMV inoculation showed clear
P  100%
a yellow spots at 21 DAI, then developed into a light
a = number of local spot on the tested plants without mosaic. This symptom appeared lighter compared to
nutshell extract treatment the positive controls. The results revealed that cashew
b = number of local spot on the tested plants with nutshell nutshell extract was able to suppress the development
extract treatment of symptoms due to CPMMV. The positive controls had
a shorter incubation period than the experimental plants

Figure 1. The score of disease severity. (A) score 1, (B) score 2, (C) score 3, (D) score 4, (E) score 5.
Andayanie et al. Antiviral Activity of Cashew Nutshell Extract 173

with cashew nutshell extract. Plants treated with cashew concentration and CPMMV sap were not showed any
nutshell extract at a concentration of 3.0; 6.0% before escalation in disease severity percentage from 28 DAI
inoculation with CPMMV and extract mixing treatment, till 49 DAI (Table 2). Cashew nutshell extract as a
each at a concentration of 1.5; 3.0; 6.0% with sap resistance inducer was not made the plant completely
CPMMV and negative controls, revealed healthy leaves resistant, but it increases the plant resistance to
until the end of the observation time so that they did not CPMMV by inhibiting disease development. This is
have an incubation period. However, plants treated with presumably due to the operation of the systemic defense
cashew nutshell extract at a concentration of 6.0% genes, which inhibit the disease incidence in soybean
before inoculation with CPMMV revealed mild plants. Plant defense genes required inducers and
phytotoxicity in the sprayed part of the extract. Cashew sufficient time to work effectively.
nutshell extract treatment was able to suppress the
incidence of disease lower than positive control with a The Inhibitory Activity of Cashew Nutshell
range of 0.11 to 5.92% at the end of the observation Extract. Based on the preliminary research, the
time (49 DAI) (Table 1). bioactive component in cashew nutshell extract
The inoculation of 6.0% cashew nutshell extract consisted mainly of anacardic acid
in CPMMV sap to soybean plants did not show any (6-pentadecylsalicylic acid) of 76.93% which is a
development of disease incidence and severity at 21 derivative of salicylic acid (Andayanie et al., 2019).
DAI. Percentage of disease incidence since 28 DAI in This bioactive compound was assumed to play a role in
cashew nutshell extract treatment at concentrations 3.00 increasing resistance to CPMMV infection. According
and 6.00%, compared to before CPMMV inoculation to Andayanie et al. (2011) and Gunaeni et al. (2015),
was not showing any significant symptoms improvement. disease incidence and intensity of soybean mosaic
The same result was also found in the treatments of mix disease were related to the infection time, infected plant
cashew nutshell extract at concentrations of 3.00 and age, environment, and induction of plant resistance by
6.00%, respectively with CPMMV sap. The symptoms various external treatments. The japanese glorybower
were consistent until 49 DAI. The same trends were leaf extract also has salicylic acid derivative compounds
also observed on the disease severity. Plant treated with and plays a role for systemic acquired resistance (SAR)
mix of cashew nutshell extract with 3.00 and 6.00% through salicylic-phenyl propanoid acid. The salicylic

Table 1. The effect of cashew nutshell extract application on disease incidence in soybean
Extract Observation (DAI) (%)
Extract treatment
concentration (%) 21 28 35 42 49
Before CPMMV inoculation 0.75 1.74 cd 2.05 c 2.13 cd 2.38 c 2.38 c
1.50 1.38 cd 1.45 c 1.85 cd 2.10 c 2.10 c
3.0 1.13 cd 1.13 cd 1.13 cd 1.13 cd 1.13 cd
6.0 0.41 d 0.41 d 0.41 d 0.41 d 0.41 d
After CPMMV inoculation 0.75 2.75 c 3.12 b 5.92 b 5.92 ab 5.92 ab
1.50 2.40 c 2.86 b 4.86 bc 5.03 b 5.03 b
3.0 2.15 c 2.40 bc 2.68 bc 2.72 c 2.72 c
6.0 1.29 cd 1.32 c 1.75cd 1.87 c 1.87 c
Mixed with 0.75 1.21 cd 1.82 c 2.01 cd 2.30 c 2.30 c
CPMMV sap 1.50 1.10 cd 1.28 c 1.28 cd 1.28 c 1.28 c
3.0 0.38 d 0.38 d 0.38 d 0.38 d 0.38 d
6.0 0.15 d 0.15 d 0.15 d 0.15 d 0.15 d
Negative control - 0.11 d 0.11 d 0.11 d 0.11 d 0.11 d
Positive ccontrol - 8.86 a 12.35 a 18.14 a 32.7 a 32.7 a
CV (%) 10.91 11.85 13,02 13.62 13.63
DAI: days after inoculation; CV: coefficient of variation; the same letter at each column showed the no significance
different based on Duncan multiple range test at significance level of 5%.
174 J. HPT Tropika Vol. 19, No. 2, 2019: 170-178

acid content in red chili pepper plants treated with Local inhibition on indicator plants
japanese glorybower and periwinkle was higher than (C. amaranticolor) and systemic on host plants from
control on the plants infected by the curly virus. Salicylic cashew nutshell extract have an average value of 74.83
acid triggers the formation of Pathogenesis-Related and 56.35%, respectively. Extract treatment before
(PR) protein and prevents the multiplication, spread, and inoculation of CPMMV at concentrations 3.0 and 6.0%,
localization of viruses (Kristyaningrum et al., 2015; after inoculation of CPMMV at concentrations of 6.0%,
Gunaeni et al., 2015). and mix of cashew nutshell extract and CPMMV sap
at concentrations (1.5; 3.0; 6.0%) had the percentage
Inhibition Properties of Cashew Seed Extract of local and systemic inhibition above the average. This
Towards CPMMV. Currently, there were no reports showed that the treatment was able to induce resistance
about cashew nutshell extract in activating defense against CPMMV in soybean. Besides that, the mix of
control genes in plants as a reaction to pathogenic cashew nutshell extract at 6.0% concentration and/
infections. However, some applications of cashew CPMMV sap had the lowest AUDPC value (5.25) and
nutshell extract as an inducer showed it capability to the highest percentage of local and systemic inhibition
induce plant resistance to CPMMV in soybean. The was 97.85 and 85.21%, respectively. The AUDPC value
application of cashew nutshell extract as an induction was followed by the treatment of cashew nutshell extract
against CPMMV infection has a lower value of area at a concentration of 1.5% in CPMMV suction and
under disease progress curve (AUDPC) than positive cashew nutshell extract at concentrations (6.0 and 3.0%)
control or plants that were inoculated with CPMMV with prior inoculation of CPMMV respectively 32.29;
without any extract application. The higher the AUDPC 39.82; 50.44.
value, the lower the inhibition percentage locally and
systemically. The largest AUDPC value for cashew nut Virus Detection by ELISA. The positive control
shell extract treatment after CPMMV inoculation was treatment confirmed the absorbance value of ELISA
in concentration of 0.75%, with AUDPC value of 115.78 (NAE) > 2 times the NAE of the negative control. CNS
indicating a local inhibition of 30.32 and a systemic of extract treatment (3.0 and 6.0%) before CPMMV
18.18 (Table 3). inoculation and CNS extract mixture (1.5; 3.0; 6.0%)

Tabel 2. The effect of cashew nutshell extract application on disease severity in soybean

Extract Observation time (DAI) (%)

Extract treatment
concentration (%) 21 28 35 42 49
Before CPMMV inoculation 0.75 2.68 bc 3.28 bcd 4.45 cde 4.45 bcd 4.45 bcd
1.5 2.11 c 2.63 cd 2.83 de 3.07cde 3.07 cde
3.0 1.14 d 1.22 de 2.22 de 2.22 def 2.22 def
6.0 1.08 d 1.77 de 1.98 de 1.98 ef 1.98 ef
After CPMMV inoculation 0.75 3.25 b 4.84 bc 7.52 bc 7.52 ab 7.52 ab
1.5 2.77 b 4.31 bc 5.01 bcd 5.06 abc 5.08 abc
3.0 2.75 bc 3.16 bcd 4.45 cde 4.93 abc 4.95 abc
6.0 1.42 cd 2.27 cd 2.51 de 2.70 de 2.70 de
Mixed with 0.75 1.96 c 2.75 cd 2.80 de 2.95 cde 2.95 cde
CPMMV sap 1.5 1.50 cd 2.30 cd 2.57 de 2.57 de 2.57 de
3.0 0.83 d 1.92 de 1.92 de 1.92 ef 1.92 ef
6.0 0.00 e 0.61 e 0.61 e 0.61 g 0.61 g
Negative control - 0.12 d 0.23 e 0.23 e 0.34 g 0.34 g
Positive control - 10.90 a 22.35 a 25.11 a 31.24 a 31.27 a
CV (%) 6.98 8.01 9.46 7.93 8.09
DAI: days after inoculation; CV: coefficient of variation; the same letter at each column showed the no significance
different based on Duncan multiple range test at significance level of 5%.
Andayanie et al. Antiviral Activity of Cashew Nutshell Extract 175

Tabel 3. The potency of cashew nutshell extract as plant resistance inducer towards CPMMV infection in soybean

Extract Inhibition (%)

Extract treatment AUDPC (Days)
concentration (%) Local Systemic
Before CPMMV inoculation 0.75 87.42 65.23 48.70
1.5 82.28 68.95 46.25
3.0 50.44 83.60 58.49
6.0 39.82 85.18 66.05
After CPMMV inoculation 0.75 115.78 30.32 18.18
1.5 108.56 48.59 25.84
3.0 95.49 63.71 40.20
6.0 56.10 76.25 52.68
Mixed with 0.75 58.65 74.12 54.17
CPMMV sap 1.5 32.29 87.43 66.92
3.0 10.07 92.36 70.34
6.0 5.25 97.85 86.21
Negative control - 2.43 99.26 99.53
Positive control - 198.70 0.00 0.00
Average (%) 55.11

with CPMMV sap showed healthy plants without mild from cashew nutshell extract has a close relationship
yellow blotch on 8 test plants until the end of observation with salicylic acid which is an inhibitor and antiviral
for each treatment and had absorbance values < 2 times against Rotaviruses that cause diarrhea. Mirabilis
NAE of negative controls and were detected negative jalapa leaf extract was also able to increase salicylic
with CPMMV antiserum, except CNS extract treatment acid content to activate the formation of PR protein and
(6.0%) after CPMMV inoculation and mixing CNS has a role for induced systemic resistance against Bean
extract (0.75%) with CPMMV sap showing healthy common mosaic virus (Kurniingsih & Damayanti, 2012).
plants and mild yellow blotch. This healthy-looking plant Bioactive compounds from natural antivirals will have
with mild yellow blotch had an absorbance value > 2 effectiveness if the inducer was applied shortly after
times NAE from negative control and positive detection. challenge inoculation with the virus. Plants with low viral
Whereas twelve out of five treatments above showed concentrations have high peroxidase activity, as reported
clear yellow and slightly mottled symptoms and were in tobacco resistance response from TMV infection
detected positive with CPMMV antiserum. However, (Matthews, 1992; Madhusudhan et al., 2011).
the treatment of cashew nutshell extract at a
concentration of 6.0% before and after the inoculation CONCLUSION
of CPMMV caused symptoms of phytotoxicity (Table
4). Plants appear healthy without visible symptoms of This research succeeded in proving the
mild yellow blotch and do not react with CPMMV effectiveness of cashew nutshell extract and its potential
antiserum allegedly due to the influence of bioactive content as an antiviral that was able to suppress
compounds from the application of these extracts that CPMMV infection in soybeans. From the tested
could reach the active area of the ribosome, thus treatments and concentrations, the mix of cashew
preventing the removal of the protein envelope from nutshell extract at a concentration of 6.0% with sap
the virus. The bioactive compound in the CNS extract CPMMV has the lowest AUDPC value with the highest
was anacardic acid with a concentration of 76.93%, percentage of local and systemic inhibition and the lowest
while the remaining 21.22% consisted of cardols CPMMV titer. Although the virus titer in the treatment
(12.75%), cardanol (4.66%) and 2-methyl-cardols did not have a significant difference with the virus titer
(3.81%) (Andayanie et al., 2019 ). According to Cecílio of mix cashew nutshell extract at a concentration of
et al. (2012) and Aderiye et al. (2015) anacardic acid 3.0% with CPMMV sap and the cashew nutshell extract
176 J. HPT Tropika Vol. 19, No. 2, 2019: 170-178

Table 4. The ELISA absorbance value (EAV) of selected treatment from CNS extract as resistance inducer
Extract
ELISA result*
Extract treatment concentration Symptom
(%) EAV Reaction
Before CPMMV inoculation 0.75 Clear yellow blotch on the 0.621 b** +
leaves, no wrinkle observed
1.5 Clear yellow blotch on the 0.606 b +
leaves, no wrinkle observed
3.0 Healthy leaves 0.490 c -
6.0 Healthy leaves, mild 0.458 cd -
phytokinesis on the sprayed
area
After CPMMV inculation 0.75 Clear yellow blotch on the 0.634 b +
leaves, no wrinkle observed
1.5 Clear yellow blotch on the 0.628 b +
leaves, no wrinkle observed
3.0 Clear yellow blotch on the 0.622 b +
leaves, no wrinkle observed
6.0 Leaves appear healthy, mild 0.619 b +
yellow blotch observed, mild
phytokinesis on the sprayed
area
Mixed with 0.75 Leaves appear healthy, mild 0.596 bc +
CPMMV sap yellow blotch observed
1.5 Healthy leaves 0.513 c -
3.0 Healthy leaves 0.451 cd -
6.0 Healthy leaves 0.440 cd -
Negative control - Healthy leaves 0.296 d -
Positive control - Severe mosaic symptom, leaf 1.045 a ++
distortion, plant dwarf
Coefficient of variation (%) 21.758
*
The EAV of negative control = 0.296, EAV of positive control= 1.045; The reaction was positive if the EAV of
the sample in twice than EAV of negative control (positive e” 0,592); + (visually light yellow); ++ (visually dark
yellow); “ (visually clear);** The same letter at each column showed the no significance different based on Duncan
multiple range test at significance level of 5%.

at a concentration of 6.0% before inoculation of Education Basic Research scheme. The author would
CPMMV. However, the treatment of cashew nutshell also like to acknowledge: 1) Head of the Virology
extract concentration of 6.0% prior to the inoculation Laboratory, Universitas Merdeka Madiun; 2) The Head
of CPMMV caused phytotoxicity. Therefore for of Bioscience Laboratory, Politeknik Negeri Jember; 3)
application in the field, it is necessary to use cashew The Head of Ngale Beans and Tuber Crops Research
nutshell extract at concentrations lower than 6.0% Installation in Ngawi who facilitated this research.
before CPMMV infection.
REFERENCES
ACKNOWLEDGMENT
Aderiye BI, David OM, & Atere VA. 2015.
Acknowledgments are conveyed to the DRPM Administration of cashew extracts in the
Ministry of Research, Technology and Higher Education treatment of some infections and diseases. Adv.
of the Republic of Indonesia who have funded this Med. Plant Res. 3(3): 75–86.
research and publication through the Leading Higher
Andayanie et al. Antiviral Activity of Cashew Nutshell Extract 177

Andayanie WR, Sumardiyono YB, Hartono S, & Yudono Garcia NZT, Barbosa GF, Matias R, Pedrinho DR, Bono
P. 2011. Incidence of soybean mosaic disease in JAM, & Martini D. 2018. Antifungal potential of
East Java Province. Agrivita J. Agricultural cashew nut shell liquid in the control of plant
Science. 33(1): 15–21. pathogens. Biosci. J. 34(1): 95–103.
Andayanie WR & Adinurani PG. 2014. Seleksi galur Gunaeni N, Wulandari AW, & Hudayya A. 2015.
dari populasi F4 kedelai yang tahan terhadap Pengaruh bahan ekstrak tanaman terhadap
penyakit mosaik (Soybean mosaic virus) dan pathogenesis related protein dan asam salisilat
berdaya hasil tinggi. J. HPT Tropika. 14(2): 152– dalam menginduksi resistensi tanaman cabai
159. merah terhadap virus kuning keriting. J. Hort.
25(2): 160–170.
Andayanie WR, Santosa V, & Rahayu M. 2017.
Resistance to Soybean mosaic virus with high Kristyaningrum VT, Martosudiro M, & Hadiastono T.
yield on F7 soybean lines. Int. J. Agric. Biol. 2015. Ekstrak bayam duri (Amaranthus spinosus
19(2): 226–232. L.) sebagai penginduksi ketahanan tanaman cabai
besar (Capsicum annuum L.) terhadap infeksi
Andayanie WR, Ermawati N, & Iswati R. 2018. Use
Cucumber mosaic virus (CMV). JHPT. 3(1):
tillage system and botanical herbicide of cashew
61–66.
nut shell extract on losses nutrient and organic
matter in the sloping land. AIP Conf. Proc. 2024. Kurnianingsih L & Damayanti TA. 2012. Lima ekstrak
020018 (2018). tumbuhan untuk menekan infeksi Bean common
mosaic virus pada tanaman kacang panjang. J.
Andayanie WR & Ermawati N. 2019. Developmental
Fitopatol. Indones. 8(6): 155–160.
effect of cashew nut shell extract against nymphal
instar of silver leaf whitefly (Bemisia tabaci Madhusudhan KN, Vinayarani G, Deepak SA, Niranjana
Genn.). IOP C. Ser. Earth Env. Sci. 293 012039. SR, Prakash HS, Singh GP, Sinha AK, & Prasad
BC. 2011. Antiviral activity of plant extracts and
Andayanie WR, Nuriana W, & Ermawati N. 2019.
other inducers against Tobamoviruses infection
Bioactive compounds and their their antifeedant
in bell pepper and tomato plants. Int. J. Plant
activity of cashew nut (Anacardium occidentale
Pathol. 2(1): 35–42.
L.) shell extract against Bemisia tabaci,
(Gennadius, 1889) (Hemiptera: Aleyrodidae). Matthews REF. 1992. Fundamentals of Plant Virology.
Acta Agric. Slov. 113(2): 281–288. Academic Press, California.
Arif M & Hassan S. 2002. Evaluation of resistance in Murphy AM, Gilliland A, Wong CE, West J, Singh DP,
soybean germplasm to Soybean mosaic & Carr JP. 2001. Signal transduction in resistance
potyvirus under field condition. Online J. Biol. to plant viruses. Eur. J. Plant Pathol. 107(1):
Sci. 2(9): 601–604. 121–128.
Cecílio AB, de Faria DB, Oliveira PdeC, Caldas S, de Petrov NM, Stoyanova M, & Valkova M. 2016. Antiviral
Oliveira DA, Sobral MEG, Duarte MGR, Moreira activity of plant extract from Tanacetum vulgare
CPdeS, Silva CG, & de Almeida VL. 2012. against Cucumber mosaic virus and Potato virus
Screening of Brazilian medicinal plants for Y. J. BioSci. Biotechnol. 5(2): 189–194.
antiviral activity against rotavirus. J.
Saleh N. 2007. Sistem produksi kacang-kacangan untuk
Ethnopharmacol. 141(3): 975–981.
menghasilkan benih bebas virus. Iptek Tanaman
Elbeshehy EKF. 2017. Inhibitory activity of different Pangan. 2(1): 66–76.
medicinal plants extracts from Thuja orientalis,
Strange RN. 2003. Introduction to Plant Pathology.
Nigella sativa L., Azadirachta indica and
John Willey and Sons Ltd, New York.
Bougainvillea spectabilis against Zucchini
yellow mosaic virus (ZYMV) infecting Citrullus Sudjaroen Y, Thongkao K, & Suwannahong K. 2018.
lanatus. Biotechnol. Biotechnol. Equip. 31(2): Antioxidant, antibacterial and cytotoxicity
270–279. activities of cashew (Anacardium occidentale)
nut shell waste. Int. J. Green Pharm. 12(1):
S229–S234.
178 J. HPT Tropika Vol. 19, No. 2, 2019: 170-178

Tavassoli M, Shahraeen N, & Ghorbani S. 2008. Towaha J & Ahmadi NR. 2011. Pemanfaatan cashew
Detection and some properties of Cowpea mild nut shell liquid sebagai sumber fenol alami pada
mottle virus isolated from soybean in Iran. PJBS. industri. Buletin RISTRI. 2(2): 187–198.
11(23): 2624–2628.