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Analysis and Feasibility Study of Plant Disease using E-Nose
Conference Paper · November 2014
DOI: 10.1109/ICCSCE.2014.7072689
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2014 IEEE International Conference on Control System,Computing and Engineering,28 - 30 November 2014,Penang,Malaysia
Analysis and Feasibility Study of Plant Disease using
E-Nose
K.P. Prak Chang, A Zakaria, AS. Abdul Nasir, N. Yusuf, R. Thriumani, AY.M. Shakaff, AH. Adorn.
Centre of Excellence for Advanced Sensor Technology (CEASTech)
Universiti Malaysia Perlis, 02600 Arau, Perlis, Malaysia
E-Mail: khaarm.arm@gmail.com
Abstract-Agriculture is one of the main sources that contribute
to the economic development in the country. However, diseases
that attack the crops have given a little impact to the agricultural
production. Generally, plant pathologist has the difficulties to
detect the symptoms that relate to the plant disease. The plant
usually gets infected that are caused by different plant pathogens
such as bacteria, fungus and virus that attack the plant.
Therefore, some method of solution needs to be uncounted in
order to provide faster solution in detecting this disease. In order
to overcome this problem, the current study proposes detection
of plant disease using an electronic nose (e-nose) called PEN3 e
nose. This type of e-nose will detect the volatile organic
compounds that are produced by two types of plants which are
chilies and papaya. The data that have been collected from PEN3
e-nose are processed using principal component analysis (PCA)
and linear discriminant analysis (LDA) methods. In order to
validate the detection result, the bacteria sample will be stained
based on Gram staining procedure in order to observe the
structure and colony of the bacteria cells using a digital
microscope. Thus, this study shows that the PEN3 e-nose is able
to detect the presence of plant pathogenic bacteria on both chilli
and papaya plants.
Index Terms-Plant disease, pathogenic bacteria, e-nose,
principal component analysis, linear discriminant analysis.
I. INTRODU CTION
At present, technology of innovative device such as
electronic nose (e-nose) has been carried out and developed by
many researchers and scientist. E-nose is a smart device that
able to mimic the function of the human nose and translated by
intelligent algorithm. Currently, the e-nose technology was
applied for monitoring, recognition and classification in many
applications. The applications of e-nose are wide. These
include the used in agriculture, industry, commercial, military,
quality control, pharmaceutical, environmental and food
quality assessment [1][2]. Chemical sensing application in
agriculture has been utilizing e-nose for some time, especially
in providing quality assessment and rapid evaluation of agro
based product. Recently, the e-nose is gaining popularity due to
ability in providing early diagnose of infection and plant
disease [3].
Plant pathogenic bacteria cause many serious diseases of
plants throughout the world, which later affect the production
and caused economic losses in the agriculture industry
978-1-4799-5686-9/14/$31.00 ©2014 IEEE 58
worldwide. Therefore, health monitoring and detection of
disease in plants and trees is critical to the agricultural industry.
In general, there are small numbers of commercial equipment
available for detection of health conditions in plant. Currently,
many commercial farmers only use manual or camera-based
surveillance mechanisms to monitor the plants, which is time
consuming, labour-intensive, inaccurate and costly [4].
Therefore, this paper presents the analysis and feasibility study
of an e-nose device in order to detect the presence of plant
pathogenic bacteria in both chilli and papaya plants. In this
study, PEN3 e-nose is used to sniff and measure the volatile
organic compound produced by the bacteria from both cultured
samples of chilli and papaya plants.
IT. TYPES OF PLANT DISEASES
There are various types of disease that can be found from
both chilli and papaya plants. These include fungal, bacteria or
viral infection. Early realization of plant disease can facilitate
the disease control and as well as improve the productivity of
the plants. The most common types and symptoms of both
chilli and papaya diseases caused by bacteria (pathogen) are
summarized in the following sections.
A. Chilli Disease
1) Bacterial Soft Rot (Erwinia Carotovora pv Carotovora)
Bacterial Soft Rot is one of the disease found on chilli pods
which causes by Erwinia Carotoravopv carotovora bacterium.
This rod-shaped, negative gram soil-bacteria belongs to
Enterobacteriacaec family and identified as major cause of
soft rods of vegetables and fruits [5-7]. This disease
commonly known as post harvested disease as the soft rods
usually started in the carylx and peduncle of harvested chilli.
However, the infection from this bacteria also can occur via
wound (started by insect) on anyplace of the chilli pod.The
symptoms can be detected when the internal tissue at the
infected site become softens and turned into a watery mass,
releasing a foul odour eventually [8][9].
2) Bacterial Wilt (Ralstonia solanacearum)
Bacterium Ralstonia solanacearum is a rod-shaped gram
negative, soil-born bacterium from the genus of
betaproteobacteria. This bacterium at first known as Bacillus
solanacearum and after few revisions, it was classified as
Pseudomonas solanacearum. However, the latest studies has

established on the name of Ralstonia solanacearum [8][9].This
type of bacteria is responsible for the bacterial wilt disease in
many plants especially chilli [10]. This symptom can be seen at
the upper leaves during the warmest part of the day, which may
recover in the evening. The whole plant becomes completely
wilted or stunted and may die under conditions favourable to
the disease. The green colour for wilted leaves still maintain
until desiccated and fall as disease progresses [11]. Vascular
tissues in the lower stem of diseased plants show a dark brown
discoloration. Besides, ooze from lower stem in water is an
important indication of the present bacterial pathogen [12].
3) Bacterial Spot (Xanthomonas campestris pv vesicatoria)
Bacterial Spot is one of the highly damaging diseases which
affect the leaves, stems and fruits of chillies plants especially
on warm and moist environment. It is caused by the rod
shaped, negative gram bacteria known as Xanthomonas
campestris pv vesicatoria from Xanthomonadaceae family
[13]. The symptom of this disease firstly occurs on
undersurface of leaf in irregular, small, water-soaked spots
with dark brown centres and thin chlorotic borders. Then, the
spots enlarged and turned into black lesions with straw
coloured centre. Usually, lesions on the upper surface of leaves
are slight sunken while the under surface lesion are slightly
raised. Severely infected leaves will turn yellow and drop.
Light-browned, narrowed and elongated lesions may occurs on
stems of chilli plant. Fruit symptoms can be seen as water
soaked brown spots which crack as grow but looks like rough
in appearance [9][14].
B. Papaya Diseases
1) Bunchy Top (Rickettesiasp')
This disease is commonly found in papaya plants which cause
major limits of papaya production in the America tropics [IS].
This disease causes by Rickettesia sp. bacteria which is a non
motile genus, gram-negative and highly pleomorphic bacteria
that might present as coccior short-rods, in pair, isolated or
thread-like with the size usually in the range of 0.3-1.0Ilm
[16]. The symptoms of this disease mainly started by
occurrence of chlorosis on young leaves, water soaked spots
appears on stems and petioles of papaya plant and petioles
usually get rigid and shortened. Besides, the leaf of papaya
also turned thick and blades the cup of leaf downward. It is
also causes bunchy appearance to the plant result from
internodes shorten and discontinued growth.
2) Bacterial Canker (Erwinia sp)
Bacteria canker on papaya plant begins to occur during 1930s
in Java and identified 'Bacilluspapaya' bacteria as the cause
of this disease [17]. However, further study by few researchers
reported that the bacteria causing this disease typed to the
genus of Erwinia sp. [18-21]. The usual symptoms of this
disease include blackish, mushy, water-soaked lesions on
leaves, while firm, water-soaked cankers develop on the stem,
which leads to the destruction of papaya trees most of the
bacteria infected fruits shows symptoms with dark spots on
the fruit's skins and flesh with water-soaked area [21][22].
59
Ill. METHODOLOGY
A. Preparation ofthe Cultured Samples
In this study, the infected leaves, branch and fruits have
been taken from each of the chilli and papaya trees that are
planted in Pauh Plantation, Perlis. These samples have been
selected randomly from different parts of the trees in order to
find out whether these parts of the trees consist of the same
bacteria or not. Fig. l(a) and (b) represent the samples of the
leaves and fruits, respectively that have been taken from chilli
plant. Then, the samples are placed in the petri dish consisting
of two types of culture media which are Luria Broth (LB) agar
and sheep blood agar. Fig. 1 (c) and (d) show the samples of
LB agar and sheep blood agar, respectively. The bacteria will
be cultered on these two types of media cultures.
Luria broth (LB) agar is a nutrient-rich media commonly
used to culture the bacteria in the lab [23]. The addition of agar
to LB resulted in gel formation, which is suitable for bacteria
growing, as these bacteria are able to gather the nutrient from
the LB agar without undergo digestion of agar. Meanwhile,
sheep blood agar is one of the most commonly used growth
medium in bacteriology. This medium appears as an opaque,
red agar and is made up of a nutrient agar base with sheep
blood. It is a non-specific medium that will support the growth
of all but the most fastidious organisms.
After about four to seven days, the bacteria in the agar plate
starts to grow. Fig. I(e) represents the growth of the bacteria on
chillie (fruit) sample. Then, the bacteria will be isolated and
cultured again. For this time, a single colony of bacteria will be
taken from the bacteria that growth in the first plate and added
to the new plate on both agars as shown in Fig. l(t). Finally,
after about three days being transferred on the new plate, it can
be seen that a colony of bacteria has grown all over the agar
medium as shown Fig. I (g).
(d)

Fig. I. The process of culturing the bacteria.
B. Headspace measurement using E-Nose
Tn this study, the experimental work will be conducted
using PEN3 e-nose. The PEN3 e-nose has an array of 10
different metal oxide (MOX) sensors which has positioned into
a small chamber with a volume of I.S mL. The detection limit
of these sensors is in the range of 1ppm. The selectivity of the
sensors is determined by the sensing material, dopant material,
working temperature and the geometry of the sensors [24].
Table I show the list and characteristics of sensors used In
PEN3 e-nose [25].
TABLE I. LIST AND CHARACTERISTIC OF PEN3 SENSORS
[251
Number of Name Characteristic Level of
sensor sensitivity
I WIC Sensitive to aromatic Toluene ,
compounds IOmgkg-1
2 W5S Very sensitive and has broad NO"
range ofsensitivity. React to Imgkg-l
nitrogen oxides and sensitive to
negative signals
3 W3C Sensitive to Ammonia and used Benzene,
to sense aromatic compounds IOmgkg-1
4 W6S Sensitive to hydrogen H"
0.lmgkg-1
5 W5C Sensitive to alkanes, aromatic Propane,
compounds and less polar I mgkg-1
compounds
6 WIS Sensitive to methane and has CH3,
broad range 100 mgkg-1
7 WIW Sensitive to terpenes and H,S,
sulphur containing organic I mg kg-1
compound. Reacts to sulphur
compounds, H,S.
8 W2S Detects alcohol partially CO,
aromatic compounds. 100 mg kg-1
9 W2W Sensitive to aromatic H2S,
compounds and sulphur organic I mg kg-1
compounds
10 W3S Reacts to high concentrations Not
(>I00 mglkg) of methane Available
aliphatic compounds
60
The e-nose analytical system has a special sampling system
integrated, which by an automatic control (auto ranging)
prevents an overloading of the sensors and also leads to a better
and faster qualitative and quantitative analysis. The
combination of these 10 metal oxide gas sensors in an array
allows the detection the complex mixtures of gaseous
compounds from medium culture headspace [24][25]. Right
after the media cultured samples have been prepared; the
samples are subjected to e-nose for headspace measurement.
The measurement was repeat and replication several times. The
distinct odour or smell product by the bacteria is known as
'smellprint'. The smellprint are then stored in a digital database
so that the system can identifiedand recognized the "smell
print" for the latter experiment. Fig. 2 represents the testing
process using PEN3 e-nose.
Fig. 2. Bacteria headspace measurement using PEN3 e-nose.
C. Preparation ofthe Bacteria Slides and Data Acquisition of
Bacteria Images
Tn order to validate the availability of the bacteria inside the
cultured samples, the slides that consists the remaining parts of
cultured samples are then prepared in order to visualize the
appearance of the bacteria using a digital microscope. Tn order
to ease the visualization process, each slide sample has been
stained using a chemical dye so that the morphological
appearance of the bacteria can be seen. The procedure for
staining these agar samples is called Gram staining. Gram
staining is a quick procedure used to look for the presence of
bacteria in slide samples and to characterize the bacteria as
either Gram-positive or Gram-negative, based on the chemical
and physical properties of bacteria cells. The full descriptions
of the Gram staining procedure can be referred in [26].
Tn brief, during preparing the slide based on the Gram
staining procedure, the heat-fixed smear is flooded with a basic
purple dye called crystal violet. As the purple stain imparts its
colour to all cells, it is often referred to as a primary stain. Fig.
3 shows sample of slides that have been prepared through
Gram staining process. After the slides have been prepared,
each slide is examined fewer than 100X oil immersion
objective of Leica DLMA microscope. The images are then
captured at a resolution setting of SOOx600 pixels by using an
Tnfinity-2 digital camera. Fig. 4 shows a set of Leica DLMA
microscope, Tnfinity-2 digital camera and personal computer
interfaced together to acquire the bacteria images.

tl": L& Ar.,.1 (!I
(./C/wllt
Wit, 1)r
Fig. 3. Sample ofslides that have been prepared through Gram staining
process.
Fig. 4. A set ofLeica DLMA microscope, Intinity-2 digital camera and
personal computer interfaced together to acquire the bacteria images.
D. Data Analysis
Tn this study, JMP software has been used to analyze the
results obtained from the PEN3 e-nose. JMP software is a
computer program for statistics. This software is used in
various applications such as in quality control and engineering,
design of experiments and scientific research for exploratory
data analysis and visualization.
Data collected from the e-nose will be subjected to
principal component analysis to evaluate the integrity of
collected data and drift analysis. Later, the raw data is
processed using linear discriminant analysis (LDA). LDA is a
parametric classification method which reduced dimensional
space of the data, maximizes the inter-class separation and
minimized intra-class scatter. Tn this experiment, LDA is
adequate and preferable for data analysis since the normality
assumption is fulfilled. Besides, it also works especially when
the independent variables of each case of sensor data are
continuous quantities and highly correlated to each other's.
TV. RESULTS AND DISCUSSIONS
Tn this study, the data analyses that have been conducted
are divided into three different parts. The first part of data
analysis is by using the principal component analysis (PCA)
that follows the Gaussian conditional densities in order to
assess discrimination within the data set. The second part of
this section represents the prediction classifier using linear
discriminant analysis (LDA) method. The last part of this
section represents the validation part of the current study, by
61
examining the bacteria inside the slide samples using a digital
microscope.
Fig. 5 represents the PCA result for chilli sample, while
Fig. 6 represents the PCA result for papaya sample. The PCA
plot for combination of both chilli and papaya samples is
represented in Fig. 7. These PCA results have been produced
by using the JMP software. In this assessment, the collected
dataset are evaluated whether the data is acceptable or not.
After that, the results are plot and the sensor placements are
displayed in the pie chart. It has been observed in Fig. 5 that at
least four sensor has significant effect in discriminate complex
sample from infected chillies. Meanwhile, in Fig. 6, at least 6
sensors contribute toward discrimination of the odour produced
by bacteria. The volatiles from both infected samples are
clearly separated as seen in Fig. 7. Meanwhile in Fig. 8, the
separation of volatile organic compounds of different bacteria
was shown clearly. It is shown that sensor 1, sensor 2, sensor 7
and sensor 9 have higher sensitivity to bacteria sniffed compare
to other sensors.
1.0
.)
·4 .(IS
·6
·8-r----,----,-,----;-
---,----,-,--
--i -1.0
-4
·8 ·6 ·4 .)
Com onent! �59%)
·1.0 ·0.5 0.0 05
Com onent! �59 %)
Fig. 5. PCA plot ofbacteria cultured from infected chilli plant.
1.0
·4 -)
Component! (94.6 %)
·1.0 .(IS 0.0 OS
Component! (94.6 %)
Fig. 6. PCA plot ofbacteria cultured from infected papaya plant.
1.0
1.0

E�envalue 20406080
4.8151
2.3761
1.4373
0.8781
0.2857
01238
0.0556
0.0168
0.0114
0.0002
" 2
·
@
N
C
·
C
o
·
E
8 -2
-4
Chilli disease
.'
4 -2
.....
�
....
Papaya disease
"
..
Componenl1 (482%)
0.5
"
·
&
N
C 0 0
·
c
0
·
�
0
�.5
-10 -0.5 0.0 0.5
Componenl1 (482%)
Fig. 7. PCA plot of combined bacteria from infected papaya and chili plants.
340
335
330
:5 325
c
'"
()
320
315
52
Papaya disease
Chilli disease
310+---�--'---�---r--�---r--�---'��---.--�
-10 10 20
Canonical1
30 40
Fig. 8. LDA plot of combined bacteria from infected papaya and chili plants.
(d) Morphology ofbacteria:
Coccus/short rod-shaped bacteria Coccus/short rod-shaped bacteria
Fig. 9. Morphology ofbacteria from chilli (a, b) and papaya (c, d) bacteria
samples.
62
V. CONCLU SIONS
In this paper, the infected plant area that have been taken
from three different parts of the trees which are leaves, branch
and fruits have been analyzed using PEN3 e-nose and JMP
software. Based on the experiment that has been conducted, it
has been proven that the PEN3 e-nose is able to detect the
presence of plant pathogenic bacteria with high success rate for
both chilli and papaya plants. In addition, examination of
morphology of bacteria also provide an idea that bacteria
retained from chilli plant disease might present as bacilli (rod
shaped) while the bacteria retained from papaya plant disease
can be present as coccus (round-shaped/short rod-shaped
regions).
ACKNOWLEDGMENT
The authors gratefully acknowledge and thank the team
members in CEASTech and School of Mechatronic, UniMAP,
Perlis for their contribution in this research study. We also
would like to acknowledge the Malaysian Government for
providing the financial support of Research Grant Scheme
(9018-00042) under the Ministry of Higher Education.
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