VOL.6, NO.3.indd
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145
Seed Health, Quality Test, and Control of Seed-borne
Fungi of Some Improved and Local Cultivars of Rice
(Oryza sativa L.) in Kano, Northwestern Nigeria
Abstract
This research was carried out to evaluate the health
and quality of rice seed. The germination of seed and
presence of rice seed related fungi were recorded and
used to evaluate the eff ect of seed dressing chemicals
on germination and vigor index over untreated rice
seeds. Seven cultivars commonly grown in Kano,
Kano State, Northwestern Nigera “FARO 52” (WITA),
“FARO 44” (SIPI), “FARO 60” , (improved varieties),
“Kwandala” , “Jamila” , Ex-china, and “JIF” (local
varieties) were used in this study. The seed dressing
chemicals used were Apron Star 42 WS, Dress
Force 42WS and ZEB-Care 80%WP. This study was
performed under three main tests, i.e dry inspection,
blotter tests, agar plate and microscopic examination.
The highest number of healthy seeds (94.16%) was
recorded from “JIF” variety and lowest (64.77%) from
“Jamila” . The highest number of deformed seeds
was observed from variety “FARO 44” whereas
the lowest noted on “JIF” . The identifi ed fungi were
Fusarium spp., Bipolaris oryzae, Aspergillus fl avus,
Curvularia lunata, Aspergillus niger, and Nigrospora
oryzae., Rhizoctonia spp. and Rhizopus spp. Highest
seed infection was recorded for A. fl avus, A. niger,
and Fusarium spp., and the least with C. lunata and
N. oryzae. Treated seeds with Zeb-care (Mancozeb
80% WP), increased their vigor index over untreated
by 62.78% and can be recommended as seed
dressing chemical for optimun control of rice seed-
borne pathogens.
Keywords: quality seed, seed germination, vigor
index, seed-borne fungi, seed dressing
chemical
Bolanle Tolani EdunA, Yahuza LurwanuA*, Mustapha SunusiB, Ali A. SulaimanA
A Crop Protection Department, Bayero University, PMB 3011, Kano, Nigeria
B Crop Science Department, Federal University, PMB 7156, Dutse, Jigawa State-Nigeria
*Corresponding author; email: ylurwanu.cpp@buk.edu.ng
RESEARCH ARTICLE
Introduction
Rice is considered as one of the major staple cereal
foods and forms a signifi cant component of the energy
resource of the human race. Rice is an economically
and scientifi cally important crop and mostly produced
and consumed in Asia and African countries (Islam
et al. 2000; Dede et al. 2019). Currently, 35–40 %
more yield productivity is required to meet global
demands (FAO 2017). The available rice stock in
these countries is stored to adjust the demands and
supply for both seed and food (Islam et al., 2000).
To date, the seed of other cereals crops like wheat,
maize, and barely have received adequate attention
for storage and post-harvest, however, the research
for rice is still not completely yet (Misra et al., 1995).
Storage facilities in Asia and Africa are still far from
satisfactory. The farmers keep their stock in various
ways, some of the means of storage are out-of-
date; therefore, weak and practically unserviceable.
Increasing rice production and subsequent reduction
of its international importation and endemic plant
pathogens continue to be a challenge in safeguarding
plant health in Nigeria. Therefore, early and accurate
diagnosis and pathogen surveillance is the most
important approach to tackle this matter. The
assessment of seed health standard in rice is very
important for farmers and food security as it is a fi rst
line approach in managing seed-borne diseases of
plants.
Furthermore, the quality of planted seeds has a
critical infl uence on the ability of crops to become
established and to realize their full yield potential and
value (McGee 1995). The importance of seed quality
in understanding the full potential of a variety is well
Andika
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146 Bolanle Tolani Edun, Yahuza Lurwanu, Mustapha Sunusi, Ali A. Sulaiman
known. The three signifi cant aspects of seed quality
are genetic and physical purity, high germination
percentage and vigor, and free from seed-borne
diseases and insects (Seshu and Dadlani 1989).
Seed vigor is recognized as an essential seed quality
parameter distinct from germination (Seshu et al.,
1988). Seed-borne diseases may not only introduce
new pathogens to aff ect the quantity or quality of the
crop yield but also contaminate the soil permanently
(Anselme, 1981).
Fungi are commonly identifi ed as causal agents
of many seed-borne diseases (Nsemwa and
Wolff hechel 1999). Studies from Biruma et al. (2003)
and Kanobe et al. (2004) showed a wide range of
seed-borne fungi pathogens on the farmer’s stored
rice seeds, of which many farmers mainly rely on
to propagate the next planting session. Most seed-
borne diseases caused by the fungi pathogens are
disastrous as they may decrease seed germination,
causing seed discoloration and produce toxins that
may be detrimental to man and domestic animals.
Distribution of some seed- borne fungi associated
with rice seeds has been reported in many countries,
including Nigeria (Suleiman and Omafe 2013);
Pakistan (Butt et al. 2011); Egypt (Madbouly et al.
2012); Bangladesh (Ora et al. 2011); Cameroon
(Nguefack et al. 2007) and Chad Republic (Serferbe
et al. 2016).
The main objectives of this study were i) to identify
seed-borne fungi associated with rice and evaluate
its role for seedling abnormalities ii) to investigate the
possibility of combining germination and seed health
tests to assess the infl uence of seed treatments on
germination and seedling vigor.
Materials and Method
Experimental Site
The experiment was carried out at the Research
and Teaching laboratory and screen house of the
Department of Crop Protection, Faculty of Agriculture,
Bayero University, Kano, (11° 58” N, 8° 25” E and 47m
above sea level) Northwestern , Nigeria.
Source of Experimental Materials
The study was conducted using seven rice varieties
commonly grown in the local area. These are “FARO
52” (WITA), “FARO 44” (SIPI), “FARO 60” (improved
varieties), “Kwandala” , “Jamila” , Ex-china, and “JIF”
(local varieties). Out of the seven cultivars, four rice
seed samples collected from Green spore Agric.
Limited (“FARO 44” , “FARO 52” , “FARO 60” and
“Jamila” ) while the remaining three varieties (“JIF”
, “Kwandala” , and Ex-china) collected from rice
farmers in Kura and Bichi local government areas,
respectively. The three seed dressing chemicals
used were Apron Star 42 WS, Dress Force 42WS,
and ZEB-Care 80%WP.
Detection of Seed-borne Fungi using Blotter Method
To perceive the seed-borne fungal pathogens
associated with the seeds using blotter method was
used following the international rules for seed testing
as used by Ahmed et al. (2013). modifi ed from ISTA to
perceive the seed-borne fungal pathogens associated
with the seeds. In this method, three layers of blotting
papers (WhatmanTM fi lter paper No.1) were soaked in
sterile distilled water and placed at the bottom of the
90 mm diameter plastic Petri dishes. Four hundred
seeds from each sample were taken randomly and
then put on the moist fi lter paper at the rate of 25
seeds per plate. Incubation was conducted at 28 ± 2°C
under 12/12 alternating cycles of light and darkness
in the incubation room for seven days. During the
incubation period, Petri dishes were checked and
wetting of fi lter paper done as the need arise. After
incubation, seeds were examined under the stereo-
microscope for the presence of seed-borne fungi.
Number of infected seeds expressed in percentage.
Two selected rice varieties within the highest disease
incidence were tested on agar plate method and
seed treatment.
Identifi cation of Fungi
Each seed was observed under stereo-microscope
at ×16 and × 25 magnifi cations to identify the seed-
borne fungi. The most associated fungi were detected
by observing their growth characters on the incubated
seeds and identifi ed following the identifi cation keys
from (Ahmed et al., 2013). Temporary slides were
also prepared and observed under a compound
microscope for proper identifi cation. The fungi
were identifi ed up to species level, following the
identifi cation keys of (Fakir et al., 2002). The results
presented as percent incidence for the individual
fungal pathogen indentifi ed.
Agar Plate Method
In this technique, one hundred seeds tested for each
variety and replicated ten times. Surface sterilized
seeds as above were plated (10 seeds/Petri dishe)
on the potato dextrose agar (PDA) medium. The
plated seeds were incubated for seven days at 28 ±
2 °C in darkness. At the end of the incubation period,
fungi developed from the seeds on the agar medium,
and then were sub-cultured into fresh medium for
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147
sporulation. The fungi were examined and identifi ed
based on colony characteristics, and morphology of
fruiting bodies under a compound microscope after
seven days incubation.
Examination of Incubated Materials
The slide of various fruiting structures of the fungi
was prepared and observed under a compound
microscope (× 100) for identifi cation following (Barnett
and Hunter, 1998).
Dry Inspection of Seed Samples
Four hundred seeds from each sample were visually
inspected and graded into nine categories. The
categories were: (1) good seeds, (2) spotted seeds,
(3) discolored seeds, (4) deformed seeds, (5) varietal
mixture (6) broken seeds, (7) insect damaged
seeds, (8) chaff y seeds and (9) inert matter. The
result of diff erent seed categories was expressed in
percentage.
Fungicide Seed Treatment
Seeds were rinsed with sterile distilled water in a clean
plastic bowl to remove impurities before chemical
seed treatment. Fungicides were applied to the
seeds following the manufacturer‘s recommendation.
The three fungicides used are Apron Star 42 WS
(20% w/w thiamethoxam + 20% w/w metalaxyl-M +
2% w/w difenoconazole), Dress Force 42WS (20%
Imidacloprid + 20% metalaxyl-M + 2% tebuconazole)
and ZEB-Care 80%WP (mancozeb 80%WP). 0.025g
of each chemical used in treating 10g of rice seeds
of “Jamila” and “FARO 44” varieties with a high
percentage of disease incidence that had been
selected for this experiment. The control consisted of
untreated rice seeds, that were allowed to dry after
treatment.
Germination of Treated-Seeds in Plastic Pots
Planting media used in the screen house was
sterilized sandy soil so as to meet ISTA regulations for
grain size, pH, and conductivity (only recommended
for seeds >5mm). Plastic pot (30 cm diameter) 10 L
in volume and about 16.5 cm depth was used. The
treated seeds have been supplied with suffi cient
water to prevent from drought condition.
Vigor Test
A vigor test was done using the sand pot method
(ISTA 2001). Shoot and root length were measured
using a ruler (30 cm length) after 14 days of sowing.
Fifteen seedlings (30 seedlings per treatment) were
randomly selected for the measurement of shoot and
root length. The seedling vigor was determined using
the formula of (Abdul-Baki and Anderson, 1972):
Vigor index = (mean of root length + mean of shoot
length) × % of seed germination.
Data Analysis
All the collected data was analyzed using descriptive
statistics and presented as percentage (%).
Results and Discussion
Dry Inspection of Rice Seed Samples
The highest number of visually healthy seed (94.16
%) recorded from “JIF” variety and lowest (64.77 %)
from “Jamila” , respectively (Table 1). The highest
number of deformed seed (1.8 %) recorded from
variety “FARO 44” , and lowest (0.47 %) recorded
for “JIF” . The highest number of apparently healthy
seed and lowest deformed seeds were observed in
the seeds of “JIF” variety (Table 1), suggesting it as
the best among all the varieties.
Survey and determination of collected seed health
status from diff erent regions in Bangladesh also have
been conducted by previous studies. Fakir et al.
(2002) recorded that collected seeds in Bangladesh
consist of 91.20 to 98.89 % pure seed, 3.72 to
37.71 % spotted seed, and 8.46- 15.50 % deformed
seed. Following study from Fakir et al. (2003) also
reported a wide variety of components among the
collected seed samples of rice from diff erent regions
of Bangladesh. Another study from Uddin (2005)
showed that farmer’s seeds (variety Upazilla) in
Noakhali can be categorized into healthy seed
(44.33 to 59.42 %), spotted seed (27.84 to 44.77 %),
discolored seed (3.93 to 8.94 %), partly fi lled seed
(0.43 to 2.35 %), deformed seed (0.91 to 3.98%),
unfi lled seed (0.001 to 0.68%), varietal mixture (0.26
to 2.22%), other plant parts (0.001 to 0.36%), inert
matter (0.50 to 0.34%) and insect damaged seed
(0.05 to 0.75%). To date, seed health survey at Bogra
district, Bangladesh showed that seed from farmer’s
storages can be grouped into good seed, spotted
seeds, deformed seeds, discolored seeds, chaff y
grains and insect damaged seeds recording: 77.84
%, 16.32 %, 3.22 %, 1.03 %, and 0.56 % respectively
(Akter and Hossain, 2016).
Fungal Pathogens Identifi ed from Rice Seed Samples
The present study has also focused on the survey of
seed-borne fungi associated with rice varieties mostly
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148 Bolanle Tolani Edun, Yahuza Lurwanu, Mustapha Sunusi, Ali A. Sulaiman
grown in Kano state, Nigeria. Some of the fungal
species were isolated and identifi ed with the blotter
method while others were obtained by the agar plate
method. During our investigation, eight species of
fungi identifi ed on the seeds of seven rice varieties
(Table 2). The identifi ed fungi were Fusarium spp.,
Bipolaris oryzae, Aspergillus fl avus, Curvularia
lunata, Aspergillus niger, and Nigrospora oryzae.,
Rhizoctonia spp., and Rhizopus spp. Highest seed
infection recorded for A. fl avus, A. niger, Fusarium
spp., followed by Rhizoctonia spp and Rhizopus
spp., while B. oryzae, C. lunata, N. oryzae recorded
the lowest percentage of seed-borne infection. The
Aspergillus species, the one recorded with high
frequency, tends to produce toxic substances apart
from the deteriorating eff ect of root rotting. This result
was in consistance with fi ndings of (Javaid et al.,
2002; Ibiam et al., 2006), that reported the presence
of Aspergillus species but, not as main pathogen.
Many of the isolated fungi from our study have been
reported to be associated with seeds of other crops
(Tsopmbeng and Fomengia, 2015). Some of them
are also known to cause seed rot, decrease seed
germination and cause pre and post damping off
as well as seedling death (Al-Kassim and Monawar,
2000) when Fusarium spp and Aspergillus fl avus are
found in high frequency. This high frequency of the
detected seed-borne pathogen from our study has
been reported in previous studies on the seeds of rice
in India (Archana and Prakash 2013) and Pakistan
(Khan et al., 1988; Javaid et al., 2002). Soil borne
mycofl ora associated with rice and their infl uence on
growth was also recorded in Abakaliki, Ebonyi State in
Southeastern Nigeria (Utobo et al., 2011). Although,
Table 1. Dry inspection of rice seed samples
Diff erent grades of seeds (%)
HS DS SS BS IDS CS DES VM IM
Variety
Ex-CHINA 66.29 18.46 6.50 0.00 1.92 0.9 1.11 1.04 3.78
FARO52 86.35 1.52 9.13 0.55 0.00 0.078 1.33 0.00 0.26
FARO44 90.53 1.20 3.18 2.16 0.00 1.13 1.8 0.00 0.00
“JAMILA” 64.77 2.10 28.4 1.18 0.00 1.7 1.6 0.00 0.25
FARO60 82.76 1.10 12.99 0.13 0.00 1.31 1.69 0.00 0.02
“KWANDALA” 77.72 0.96 19.13 0.33 0.00 0.95 0.55 0.00 0.36
“JIF” 94.16 0.99 2.49 0.63 0.057 1.04 0.47 0.00 0.16
Note: HS = healthy seed = good looking seed free from spots and abnormalities, DS = discolored seed, SS = spotted
seeds, BS= broken seeds, IDS = insect damaged seeds, CS = chaff y seeds, DES = deformed seeds, VM = varietal
mixture and IM = inert matter
Table 2. Fungal pathogens identifi ed from rice seed samples
S/No. Variety Fungi identifi ed
1 “KWANDALA” Fusarium spp.,Rhizopus spp.,A. fl avus, A.
niger
2 “JIF” Bacterial growth, Fusarium spp., A. fl avus,
Aspergillus niger.
3 “FARO 60” Rhizopus spp., A.niger, A. fl avus, Rhizoc-
tonia spp.
4 “FARO 44” A. fl avus, A. niger, Fusarium spp, Bipolaris
oryzae, Curvularia lunata, Rhizoctonia
spp., Rhizopus spp.
5 “FARO 52” Rhizoctonia spp., Fusarium spp., A. fl avus,
A. niger, Rhizopus spp., Curvularia lunata.
6 “JAMILA” Bacterial growth, Rhizopus spp., N. ory-
zae, A. fl avus, A. niger, Fusarium spp., C.
lunata.
7 Ex-CHINA Bacterial growth
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149
no such kind of soil borne pathogens related to rice
crops were recorded in our study area.
Quality Test: Purity Analysis of Rice Seed Samples
A seed purity analysis was conducted and concluded:
100 % pure seed in “FARO 44” rice variety, and the
lowest pure seed (95.18%) was recorded in variety
Ex-China (Table 3). The lower purity of cultivar Ex-
China compared to other cultivars was caused by
lousy storage practices by the farmers while high
purity “FARO 44” was obtained from applied seed
technology in commercial company. Another study
from Chowdhury’s (2012) about seed quality status
reported that HYV has the highest percentage of
purity. Haque et al. (2007) also found 99.01 % pure
seed from the trained farmers and minimum (96.19
%) in untrained farmers. Percentage of seed purity
was determined by Uddin (2005) in Upazilla under
Noakhali district that ranged from 95.59 to 99.39
%. In convex with our fi ndings, Fakir et al. (2002)
categorized the percentage of pure seeds ranging
from 91.20 to 98.89 % collected from trained farmer’s
stored rice seed.
Germination Test Using Blotter Method of Rice Seed
Samples
The result in Figure 1 shows the mean percentage of
germination of diff erent rice seeds, ranged from 0 %
to 90.50%. The highest germination was recorded in
variety “JIF” (90.50%), while the lowest germination
found in variety Ex-China (0%). The variety “Jamila”
has the highest percentage incidence of fungal
mycelia growth examined using blotter method
(14.25%), and variety Ex-China recorded the lowest
rate (1.25%). This research clearly showed that “JIF”
has the highest germination percentage even though
it has been sourced from farmer-stored seed within
the study area. The zero germination in the variety Ex-
China could be a result of poor storage conditions that
lead to the development of seed-borne pathogens,
insects, mechanical damage at both handling and
storage, or aging which renders the seed to be not
viable or dormant. Some of these fungal pathogens
are known to cause seed rot, decreasing of seed
germination, pre and post damping off , and seedling
death (Al-Kassim and Monawar, 2000). According to
Lamrani et al. (2013), Alternaria padwickii colonizes
variety of seeds thus reducing the percentage
germination and causes seed rot. Species of the
genus Curvularia, in particular, C. lunata has been
reported to infect the embryo of the seeds, therefore,
reducing the percentage germination of rice seeds
(Imolehin, 1987; Bautista and Opina, 1991). C. lunata
was reported in diff erent rice varieties and identifi ed
as one of seed-borne pathogens (Butt et al., 2011;
Utobo et al. , 2011; Ashfaq et al., 2015).
Eff ect of Seed Treatment on Seed Vigor Index
The variety “FARO 44” and “Jamila” were selected as
test sample of seed dressing trial using three diff erent
chemicals due to the facts they are most commonly
grown variety in Kano. “FARO 44” is an improved
variety while “Jamila” is local. Another consideration
is these both varieties have higher mycelia growth
than others and this indicator is suitable to measure
the increasing of vigor index based on the chemical
treatment.
The vigor index of treated seeds using Zeb-care was
increased up to 62.78% over the untreated control
(Table 4). The control (untreated variety) from “Jamila”
has a vigor index score of 928.71, while “FARO 44”
has the vigor index of 904.11. Variety “Jamila” treated
with Zeb-care has 991.49 vigor index and it indicated
that there was an increase of vigor index with 62.78%
followed by Zeb-care treatment on “FARO 44” with
an increase of 31.78%. The lowest increase in vigor
index (5.28%) was found in variety “FARO 44” treated
with the Apron star (Table 4). The variety “FARO 44”
treated with Dress force shown a negative value
of -6.11 over the untreated which directly implies
that there is a decrease in vigor index over control.
Table 3. Seed quality and purity test among the rice seed samples
Diff erent components under purity analysis (%)
Variety Pure seed Other seed Inert matter
Ex-CHINA 95.18 1.04 3.78
“FARO 52” 99.74 0.00 0.26
“FARO 44” 100.00 0.00 0.00
“FARO 60” 99.98 0.00 0.02
“JAMILA” 99.75 0.00 0.25
“JIF” 99.84 0.00 0.16
“KWANDALA” 99.64 0.00 0.36
Journal of Tropical Crop Science Vol. 6 No. 3, October 2019
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150 Bolanle Tolani Edun, Yahuza Lurwanu, Mustapha Sunusi, Ali A. Sulaiman
Following the obtained values, the effi cacy of seed
dressing chemicals was hypothesized benefi t the
variety “Jamila” more. The Zeb-care dressing
chemical displayed a more positive eff ect (+31.78
and +62.78) on both cultivars (“Jamila” and “FARO
44”) followed by Apron star (+5.28 and +31.17) and
lastly Dressed force (-6.11 and +22.40) respectively.
Similar result also has been reported on previous
studied by (Bhuiyan et al., 2005; Hossain and Hossain
2012) whereas vigor index increased in vegetable
treated seed using BAU Bio-fungicide. Furthermore,
Shultana et al. (2009) also evaluated wheat seeds
treated with Bavistin and showed higher vigor index
on 2843 followed by BAU Bio-fungicide treated seed
on 2661.
Conclusion
Based on the result obtained in this experiment, it can
be concluded that farmers are strongly advised to use
any of the following varieties “JIF” , “Jamila” and
“FARO 44” in Kano State as it has been tested and
found to be suitable in both seed quality and purity, it
will also help to improve rice production and reduce
the threats due to seed-borne fungal diseases. The
use of Zeb-care treated seed, which is often neglected
by farmers, is encouraged as the seed dressing
fungicide (Zeb-care 80WP) signifi cantly increased
seed germination and seedling vigor.
Acknowledgement
The authors thank Mr. Muhammad Nura Abdulkadir
of Plant Pathology Laboratory, Crop Protection
Department, Bayero University, Kano for laboratory
assistance.
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