Journal of Phytopathology and Pest Management 7(1): 109-120, 2020
pISSN: 2356-8577 eISSN: 2356-6507
Journal homepage: http://ppmj.net/
∗Corresponding author: Abdallah A. M. Ali,
E-mail: abdallahali@agr.aswu.edu.eg
109
Copyright © 2020
Effect of certain essential oils and biocides on
controlling marjoram root rot and wilt diseases
Abdallah A. M. Ali1*, Yosra Z. Abo-Shosha2, Mahmoud M. H. Hassanin3
1Plant Pathology Department, Faculty of Agriculture and Natural Resources, Aswan University, Aswan, Egypt
2Central laboratory of Organic Agriculture, Agricultural Research Centre, Giza, Egypt
3Ornamental, Medicinal and Aromatic Plant Diseases Research Department, Plant Pathology Research Institute, Agricultural Research Centre, Giza, Egypt
Abstract
Keywords: Majorana hortensis, Fusarium oxysporum, Fusarium semitectum,
Rhizoctonia solani, Plant Guard, Actamyl 70% wp, thyme oil.
The aim of this study was to determine the impact of certain essential oils (eucalyptus and
thyme essential oils) and biocides (Plant Guard and Rhizo-N) as fungicidal alternatives for
the control of root rot and wilt diseases of marjoram (Majorana hortensis L.) caused by
several fungi. Marjoram plants with root rot and wilt symptoms were obtained from Giza,
Beni-Suief, and Minia governorates, Egypt. Pathogenicity tests showed that all isolated
fungi (Fusarium semitectum, F. solani, F. oxysporum, F. roseum, Rhizoctonia solani, and
Macrophomina phaseolina) had the ability to infect plants and seedlings. Fusarium
oxysporum, followed by R. solani, was the most pathogenic fungus on the marjoram
seedlings, resulting in pre- and post-emergence damping-off. Additionally, the most
significant disease incidence percentages on marjoram plants after transplanting were
caused by F. oxysporum and F. semitectum. In vitro investigations were performed utilizing
eucalyptus and thyme essential oils indicated that the growth of the investigated fungi (F.
oxysporum, F. semitectum, and R. solani) was significantly inhibited. However, thyme was
the most efficient treatment, especially at a concentration of 6000 ppm, which completely
inhibited mycelial growth of R. solani and F. oxysporum. The effectiveness of eucalyptus
and thyme essential oils, Plant Guard, Rhizo-N, and Actamyl 70% wp was determined for
the control of target diseases under greenhouse conditions. The results showed that all
tested treatments significantly reduced the disease incidence caused by the investigated
fungi. Actamyl was the most efficient treatment. Thyme oil was an effective treatment
against R. solani and F. oxysporum in the second order following Actamyl, whereas Plant
Guard was effective against F. semitectum and F. oxysporum. Generally, Plant Guard was
the most successful treatment for enhancing plant growth of marjoram plants. These findings
demonstrate the potential of applying Plant Guard as an alternate fungicide against wilt and
root rot diseases of marjoram plants.
Ali et al., 2020
110
1. Introduction
Marjoram, or sweet marjoram (Majorana
hortensis L.), serves as one of the most
valuable medicinal as well as aromatic crops in
Egypt. Plantations of marjoram have expanded
in recent years in Egypt. Marjoram is a
perennial plant, usually planted in a nursery,
and seedlings (60 days old) are transplanted in
the field. Ordinarily, plants are cut twice per
year for a period of three years. The dried
leaves are extensively used in the food
industry. Marjoram essential oil has various
applications in pharmaceutical preparations. It
is used as a stimulating and antibacterial
component in toothpaste, as well as in drugs
against whooping cough. Additionally, it is
used as a carminative in the case of
gastrointestinal disorders (Bellakkadar et al.,
1988). Wilt and root rot diseases were recorded
in Egyptian marjoram plantations, which had a
negative effect on vegetative growth and plant
standing (El-Gebaly, 1998; Hilal & Helmy,
1998; Hilal et al., 1990). The incidence of these
diseases often rises year after year. A lack of
suitable agricultural practices, as well as poor
sanitation, may be contributing factors. The
present research aimed to evaluate the efficacy
of different fungicidal alternatives in
controlling root rot and wilt diseases of
marjoram caused by various fungi.
Specifically, the impact of eucalyptus and
thyme essential oils, as well as biocides such as
Plant Guard and Rhizo-N, compared to the
fungicide Actamyl 70%, was assessed.
2. Materials and methods
2.1 Isolation and identification of the causal
pathogens
During the 2019 season, naturally infected
marjoram plants with wilt and root rot diseases
were recorded in Giza, Beni-Suief, and Minia
governorates. The infected plants were at two
months after transplanting (free cut), six
months after transplanting (1st cut), and nine
months after transplanting (2nd cut). Disease
symptoms such as plant death, stunting,
dryness, yellowing, and wilting appeared in
irregular parts of the cultivation area. Samples
of infected plants were collected and
transferred to the laboratory. The infected
stems and roots were subjected to a procedure
to isolate the fungi. The procedure involved
washing the infected stems and roots using tap
water, cutting them into small fragments, and
surface sterilizing them with 2% NaOCl for 3
minutes. Following this, the fragments were
thoroughly rinsed with sterilized distilled
water multiple times and then dried using
sterilized filter papers. These fragments were
then placed onto Petri dishes containing potato
dextrose agar (PDA) medium and kept at a
temperature of 27°C. Daily checks were
carried out for one week to monitor any fungal
development. In order to determine the
prevalence of developed colonies for each
fungal isolate, a calculation was performed in
reference to the total isolates available.
Subsequently, the isolation of these fungi was
purified through the utilization of both single-
spore and hyphal tip methodologies. The
definitive identification of these purified fungal
specimens was conducted in accordance with
established protocols outlined by Gilman (1957),
Barnett and Hunter (1972), and Nelson et al.
(1983), with validation provided by the
Department of Mycology and Plant Disease
Survey at the Plant Pathology Research
Institute, ARC, Giza, Egypt. Furthermore, for
future investigations, pure cultures of the
isolated fungi were transferred onto PDA
slants and stored under controlled conditions
at a temperature of 5°C.
Ali et al., 2020
111
2.2 Pathogenicity tests
This experiment was performed under
greenhouse conditions. To prepare the
inoculum, each fungus was grown separately
on autoclaved sorghum medium (50 g of
cleaned sand, 100 ml of water and 100 g of
corn) in 500 ml glass bottles for 15 days at 28˚
C. A 5% formalin solution was used to sterilize
clay sand soil (1:1 w/w), which was then
allowed to dry for two weeks before being
used. Fungal inoculums were added to the soil
at a rate of 1% (w/w) and mixed thoroughly,
then placed in 25-cm-diameter pots and
watered one week prior to planting for fungal
colonization enhancement. Seeds of marjoram
were surface sterilized with 0.1% (v/v) of
sodium hypochlorite for 3 minutes, and then
they were rinsed with sterilized distilled water
and allowed to air dry. Fifty seeds were planted
per pot, and three replicates were used for each
treatment. Percentages of pre- and post-
emergence damping-off 15 and 60 days after
sowing were recorded. Additionally, the
pathogenicity of isolated fungi was tested on
marjoram transplants that were 60 days old
using infested pots. Five transplants were
planted in each pot, and treatments were
replicated three times. Infected marjoram
plants were recorded as percentages at 45 and
90 days from transplanting. In order to
compare with the original isolates, a new
isolation of fungi from diseased plants was
conducted.
2.3 Evaluation of antifungal activity of
essential oils in vitro
To study the efficacy of volatile essential oils
of eucalyptus and thyme on the mycelial
growth of the tested fungi, F. oxysporum, F.
semitectum and R. solani, three Petri dishes
containing PDA medium were inoculated
using 5 mm fungal growth discs of each
fungus. The inoculated Petri dishes were
inverted, and volatile oil at three
concentrations (1000, 3000, and 6000 ppm)
saturated 6 mm paper discs were placed at the
center of plate lids as described by Maruzzella
and Sicurella (1960). The dishes were
incubated at 27 ˚C and checked daily for
fungal development. The tested essential oils
were supported by Medicinal Plants Research
Department, Horticulture Research Institute,
Agricultural Research Center, Giza, Egypt.
Inhibition percentages in mycelial growth
were calculated when control plates (without
essential oils) were completely filled with the
fungal growth, according to the formula of
Pandey et al. (1982), as follows:
𝐺𝑟𝑜𝑤𝑡ℎ 𝑖𝑛ℎ𝑖𝑏𝑖𝑡𝑖𝑜𝑛 (%)=𝐺1 − 𝐺2
𝐺1 × 100
Where, G1 = growth in control, G2 = growth
in treatment.
2.4 In vivo studies
Under greenhouse conditions, essential oils of
eucalyptus and thyme (6 ml/l water), biocides
Rhizo-N and Plant Guard, as well as fungicide
Actamyl 70% wp at concentrations as
mentioned in Table (1) were tested for
controlling marjoram root rot and wilt
diseases. In this experiment, plastic pots (25
cm diam.) were used for planting, filled with
sterilized soil [clay sand soil (1:1)]. Soil
infestation with the tested fungi; F.
oxysporum, F. semitectum, and R. solani, was
carried out as described in pathogenicity tests.
The seedling roots were dipped for 20 minutes
in the treatments or in water (as an untreated
control), after which they were planted in the
infested soil. Fifteen marjoram seedlings (60
days old) were used for each treatment as well
as for the control (5 seedlings/ pot), and three
replicates were used for each. The
percentages of infection or survival plants,
Ali et al., 2020
112
as well as some plant parameters such as
root length, plant height, plant dry weight, and plant fresh weight, were recorded 90
days after transplanting.
Table 1: Fungicide and biocides used in the current study.
Commercial name
and formulation
Common name
(Active ingredients)
Application
rate/L.W
Composition
Actamyl 70% wp
(Powder)
Thiophanate methyl
1 g/l
Dimethyl [1,2 phenylenebis
(iminocarbonothioyl)] bis [carbamate]
Plant Guard (Liquid)
Trichoderma
harzianum
4 ml/l
Trichoderma harzianum; 30 × 106 spores/ml
Rhizo – N (Powder)
Bacillus subtilis
4 g/l
Bacillus subtilis; 30 × 106 cell/g
2.5 Statistical analysis
The present investigation was designed as a
factorial experiment with three replicates in a
completely randomized design (Snedecor and
Cochran 1989). Statistical analysis was
performed using MSTAT-C version (4) with
the Least Significant Difference (L.S.D.) test
set to 0.05.
3.
Results
3.1 Isolation, identification and percentages
of the isolated fungi frequency
Table (2) presents the results of isolated fungi
from infected marjoram plants showing
typical symptoms of root rot and wilt disease
and obtained from different locations in Giza,
Beni-Suief, and Minia governorates, Egypt.
The study identified three genera of fungi,
including Macrophomina, Rhizoctonia, and
Fusarium, with F. oxysporum and F. solani
being the most frequently isolated fungi
(43.0% and 29.06%, respectively), followed
by R. solani (11.28 %). In contrast, F. roseum
recorded the lowest mean occurrence
percentage (2.67%). Fusarium roseum and F.
semitectum were isolated only from plant
samples from Giza, as well as M. phaseolina
from Minia governorate.
Table 2: Frequency (%) of fungi isolated from infected marjoram plants collected from Giza,
Beni-Suief and Minia governorates, Egypt.
The isolated fungi
Location
Mean
Giza
Beni-Suief
Minia
Fusarium oxysporum
32.05
49.00
47.95
43.00
F. solani
37.92
41.20
8.06
29.06
Rhizoctonia solani
0.00
9.80
24.04
11.28
F. semitectum
22.03
0.00
0.00
7.34
Macrophomina phaseolina
0.00
0.00
19.95
6.65
F. roseum
8.00
0.00
0.00
2.67
Total
100
100
100
-
3.2 Pathogenicity tests
3.2.1 Percentages of pre- and post-emergence
damping-off
Table (3) presents the results of the
pathogenicity test of fungi isolated from
collected marjoram plant samples. The test was
conducted by measuring pre- and post-
emergence damping-off percentages 15 and 60
days from planting, as well as healthy survival
seedlings. All the tested fungi showed
significant differences in percentages of pre-
and post-emergence damping-off compared
with the uninoculated control, subsequently
Ali et al., 2020
113
decreasing healthy survival seedlings.
Fusarium oxysporum and R. solani were the
most pathogenic fungi, as they recorded the
highest percentages of pre- (36% and 32%,
respectively) and post-emergence damping-off
(26% and 24%, respectively), with non-
significant differences between each other in
this respect. Furthermore, they recorded the
highest percentage of survival seedling
reductions (62% and 52%, respectively). On
the contrary, F. solani recorded the lowest
percentages of reduction in survival (26%) and
pre-emergence damping-off (10%). Fusarium
oxysporum and F. semitectum were the most
pathogenic species and more virulent than F.
solani.
Table 3: Percentages of pre- and post-emergence damping-off caused by isolated fungi in
marjoram plants at 15 and 60 days after planting.
Reduction* (%)
Survival (%)
Damping-off (%)
The tested fungi
Post- emergence
Pre-emergence
62.0
38.0
26.0
36.0
Fusarium oxysporum
52.0
48.0
24.0
32.0
Rhizoctonia solani
48.0
52.0
22.0
30.0
F. semitectum
40.0
60.0
22.0
18.0
F. roseum
38.0
62.0
16.0
22.0
Macrophomina phaseolina
26.0
74.0
16.0
10.0
F. solani
-----
100.0
0.0
0.0
Uninoculated control
-----
-----
6.5
6.9
L.S.D. at 5%
*Reduction percentages in survivals relative to the control treatment.
3.2.2 Percentages of root rot or wilt incidence
Table (4) displays the results of the infection of
marjoram plants by all tested fungi. The plants
were transplanted in infested soil, and all tested
fungi were able to infect the plants, causing
wilt or root rot diseases, as opposed to the
uninoculated control. Infection percentages
increased as plant ages increased from 45 to 90
days from transplant. Fusarium oxysporum,
followed by F. semitectum, was the most
virulent fungus, as they recorded the highest
percentages of disease incidence 45 days after
transplanting (46.7% and 20.0%, respectively)
and 90 days after transplanting (73.3% and
26.7%, respectively). Moreover, they recorded
the highest percentages of survival plant
reductions (73.3% and 26.7%, respectively).
On the contrary, both M. phaseolina and F.
roseum recorded the lowest reduction
percentage of survival plants (13.3%),
resulting in 6.7% disease incidence after 45
days from transplanting and 13.3% disease
incidence after 90 days from transplanting.
3.3 Antifungal activity of essential oils on
mycelial growth of the tested fungi in vitro
Table (5) presents the results of the significant
inhibition of three concentrations (1000, 3000,
and 6000 ppm) of thyme and eucalyptus oils
on mycelial growths of all tested fungi. Thyme
oil was the best treatment, especially when it
was applied at a concentration of 6000 ppm,
which completely suppressed the growth of R.
solani and F. oxysporum. Rhizoctonia solani
was the most affected fungus by the tested oils,
followed by F. oxysporum. However, the
percentage reduction in linear growth was
increased by increasing oil concentrations.
They reached 74.44% and 100% at 6000 ppm
in the case of eucalyptus and thyme, respectively,
with R. solani. In contrast, F. semitectum was
the least affected fungus, as it recorded 60%
and 92.22% reduction in growth at 6000 ppm
in the case of eucalyptus and thyme, respectively.
Ali et al., 2020
114
Table 4: Percentages of infected marjoram plants, 45 and 90 days from transplanting in infested
soil under greenhouse conditions.
Reduction* (%)
Survival (%)
Disease incidence (%)
The tested fungi
90 days
45 days
73.3
26.7
73.3
46.7
Fusarium oxysporum
20.0
80.0
20.0
6.7
Rhizoctonia solani
26.7
73.3
26.7
20.0
F. semitectum
13.3
86.7
13.3
6.7
F.roseum
13.3
86.7
13.3
6.7
Macrophomina phaseolina
20.0
80.0
20.0
6.7
F. solani
-----
100.0
0.0
0.0
Uninoculated control
-----
-----
11.4
5.8
L.S.D. at 5%
*Reduction percentages in survivals relative to the control treatment.
Table 5: Effect of three concentrations of two essential oils on linear growth of the tested fungi.
Essential
oils
Fungi
Linear growth (cm)
Control
(cm)
1000
(ppm)
Reduction*
(%)
3000
(ppm)
Reduction*
(%)
6000
(ppm)
Reduction*
(%)
Eucalyptus
Rhizoctonia solani
9.0
5.9
34.44
4.4
51.11
2.3
74.44
Fusarium
oxysporum
9.0
6.2
31.11
5.6
37.78
3.2
64.44
F. semitectum
9.0
7.1
21.11
6.3
30
3.6
60
Thyme
R. solani
9.0
5.7
36.67
3
66.67
0
100
F. oxysporum
9.0
6.4
28.89
3.1
65.56
0
100
F. semitectum
9.0
6.9
23.33
3.8
57.78
0.7
92.22
*Reduction relative to the control. L.S.D.at 5 % for: Fungi (F) =1.13, Essential oils (O) = N.S., Concentrations (C) = 0.10, F
× O = 0.20, F × C = 0.25, O × C = 0.13, F × O × C = 0.36.
3.4 Effect of essential oils, biocides and
fungicide on disease incidence and some plant
parameters under greenhouse conditions
In this experiment, the efficacy of essential oils
(eucalyptus and thyme), biocides (Rhizo N and
Plant Guard), and fungicide (Actamyl 70%
wp) on disease incidence and some plant
parameters 90 days from planting in infested
soil separately with F. oxysporum, F.
semitectum, and R. solani were investigated.
These treatments were used as dipping of
seedling roots for 20 minutes.
3.4.1 The impact on disease incidence 90 days
from planting
Data in Table (6) indicates that percentages of
disease incidence decreased, and survival
plants increased with all tested treatments.
Compared to the untreated control, all
treatments significantly decreased disease
incidence. However, Actamyl 70% wp gave
the lowest percentage of disease incidence of
all the tested fungi. Thyme oil came in the
second order, after Actamyl with R. solani and
F. oxysporum, since they caused 6.7% and
13.3% disease incidence, respectively.
Subsequently, the percentages of survival
plants resulting from thyme treatment with R.
solani and F. oxysporum were 93.3% and
86.7%, respectively. While Plant Guard came
in the second order, after Actamyl, with F.
semitectum and F. oxysporum, both of which
caused 13.3% disease incidence, the
percentage of survival plants was 86.7%.
Eucalyptus oil was the least effective
treatment with F. oxysporum, since it gave
40% disease incidence.
Ali et al., 2020
115
Table 6: Effect of essential oils, biocides and fungicide on disease incidence (%), 90 days from planting under
greenhouse conditions.
Treatments
Disease incidence (%)
F. oxysporum
F. semitectum
R. solani
Disease incidence
(%)
Survival
(%)
Disease incidence
(%)
Survival
(%)
Disease incidence
(%)
Survival
(%)
Essential oils
Eucalyptus
40.0
60.0
26.7
73.3
13.3
86.7
Thyme
13.3
86.7
26.7
73.3
6.7
93.3
Biocides
Plant Guard
13.3
86.7
13.3
86.7
20.0
80.0
Rhizo-N
33.3
66.7
26.7
73.3
20.0
80.0
Fungicide
Actamyl 70% wp
6.7
93.3
0.0
100
0.0
100
Untreated control
66.7
33.3
53.3
46.7
73.3
26.7
L.S.D. at 5%
20.2
---
21.5
---
10.2
---
3.4.2 The impact on plant height 90 days
from planting
Data in Table (7) show that all treatments
increased plant height more than the control
treatment. The increases were significant only
by using Actamyl 70% treatment with all
tested fungi, Plant Guard with (F. oxysporum
and F. semitectum), Rhizo-N with (F. semitectum)
and eucalyptus with (R. solani). The best
treatment after Actamyl which gave the
superiority of increasing plant height was
Plant Guard with F. oxysporum and F.
semitectum (30.67% and 44.16%, respectively).
However, eucalyptus was the best treatment with
R.
solani
which gave a 32.87% increase in plant height
.
Table 7: Effect of essential oils, biocides and fungicide on plant height, 90 days from planting under greenhouse
conditions.
Treatments
Plant height (cm)
F. oxysporum
Increase (%) *
F. semitectum
Increase (%) *
R. solani
Increase (%) *
Essential oils
Eucalyptus
18.7
24.67
16.0
3.90
19.0
32.87
Thyme
17.8
18.67
16.0
3.90
14.8
3.50
Biocides
Plant Guard
19.6
30.67
22.2
44.16
14.5
1.40
Rhizo-N
18.1
20.67
20.2
31.17
14.8
3.50
Fungicide
Actamyl 70% wp
20.8
38.67
24.9
61.69
18.3
27.97
Untreated control
15.0
---
15.4
---
14.3
---
L.S.D. at 5%
4.5
---
3.6
---
3.9
---
* Increases relative to control
3.4.3 The impact on root length 90 days
from planting
Data in Table (8) demonstrates that all
treatments resulted in root length increases
compared to the untreated control (water only).
However, increases were significant only in
the treatment of Actamyl and thyme with all
tested fungi, Plant Guard with (F. oxysporum
and F. semitectum) and the percentage of
increases ranged between 89.5% and 17.5%.
The highest percentage of increase (89.5%)
resulted from the treatment of Plant Guard
with F. semitectum, while the lowest
percentage of increase (17.5%) resulted from
eucalyptus with F. semitectum.
3.4.4 The impact on plant fresh weight 90
days from planting
Data in Table (9) exhibit that most treatments
caused increases in plant fresh weight, and the
percentage ranged from 54.54% to 6.82%.
Increases were significant only in the treatment of
Ali et al., 2020
116
Plant Guard with F. oxysporum, which recorded the
highest percentage of increase (54.54%). In
contrast, thyme oil, Plant Guard, and Rhizo-N
with (R. solani) and Rhizo-N with (F.
semitectum) had no effect on plant fresh weight
since the percentage increase was 0.0%.
Table 8: Effect of essential oils, biocides and fungicide on root length, 90 days from planting under greenhouse
conditions.
Treatments
Root length (cm)
F. oxysporum
Increase (%) *
F. semitectum
Increase (%) *
R. solani
Increase (%) *
Essential oils
Eucalyptus
7.7
28.3
6.7
17.5
7.7
28.3
Thyme
9.3
55.0
9.3
63.16
10.3
71.7
Biocides
Plant Guard
9.3
55.0
10.8
89.5
7.2
20.0
Rhizo-N
7.3
21.7
7.3
28.1
7.3
21.7
Fungicide
Actamyl 70% wp
9.3
55.0
9.3
63.2
9.3
55.0
Untreated control
6.0
---
5.7
---
6.0
---
L.S.D. at 5%
2.0
---
2.9
---
2.0
---
* Increases relative to control
Table 9: Effect of essential oils, biocides and fungicide on plant fresh weight, 90 days from planting under
greenhouse conditions.
Treatments
Plant fresh weight (gm)
F. oxysporum
Increase (%) *
F. semitectum
Increase (%) *
R. solani
Increase (%) *
Essential oils
Eucalyptus
5.0
13.64
4.4
7.32
3.5
9.38
Thyme
4.7
6.82
4.4
7.32
3.2
0.0
Biocides
Plant Guard
6.8
54.54
4.7
14.63
3.2
0.0
Rhizo-N
4.7
6.82
4.1
0.0
3.2
0.0
Fungicide
Actamyl 70% wp
5.3
20.45
6.2
51.21
3.5
9.38
Untreated control
4.4
---
4.1
---
3.2
---
L.S.D. at 5%
1.4
---
4.5
---
1.2
---
* Increases relative to control
3.4.5 The impact on plant dry weight 90
days from planting
Data in Table (10) demonstrate that some
treatments caused increases in plant dry weight.
However, the increases were significant only in the
treatments of Actamyl 70% wp with (F. oxysporum
and F. semitectum) and Plant Guard with (F.
oxysporum). In contrast, Rhizo-N with all
tested fungi, Plant Guard with (F. semitectum
and R. solani), thyme with (F. oxysporum and
F. semitectum) and eucalyptus with (F.
semitectum) had no effect on plant dry weight
since the percentage increase was 0.0%.
Table 10: Effect of essential oils, biocides and fungicide on plant dry weight, 90 days from planting under
greenhouse conditions.
Treatments
Plant dry weight (gm)
F. oxysporum
Increase (%) *
F. semitectum
Increase (%) *
R. solani
Increase (%) *
Essential oils
Eucalyptus
2.5
13.64
2.2
0.0
1.9
18.75
Thyme
2.2
0.0
2.2
0.0
1.9
18.75
Biocides
Plant Guard
3.4
54.54
2.2
0.0
1.6
0.0
Rhizo-N
2.2
0.0
2.2
0.0
1.6
0.0
Fungicide
Actamyl 70% wp
2.8
27.27
3.1
40.91
1.9
18.75
Untreated control
2.2
---
2.2
---
1.6
---
L.S.D. at 5%
0.3
---
0.4
---
0.6
---
* Increases relative to control
Ali et al., 2020
117
4. Discussion
Marjoram (Majorana hortensis L.) is
considered one of the most important
medicinal and aromatic plant crops in Egypt.
Plantations of marjoram have been affected by
root-rot and wilt diseases, which have resulted
in reduced plant vegetative growth, plant
stand, and essential oil yield (El-Gebaly, 1998;
Garbagnoli & Gaetan, 1994; Hilal et al., 1990).
Isolation trials from the diseased plant samples
confirmed the presence of soil-borne fungi
associating with the infected plant tissues.
However, Fusarium spp. was recorded at a
higher frequency compared with the other
fungi. Such findings go in accordance with
Hilal et al. (1990) and El-Gebaly (1998), who
found soil-borne diseases of marjoram in
Egypt and identified their causal fungal
pathogens. The most prevalent isolate was
identified as F. oxysporum. The same result
was obtained by Garbagnoli and Gaetan
(1994), who identified the causal agent of
marjoram wilt as F. oxysporum according to
symptomology, cultural, and morphological
characteristics. Fusarium semitectum, F.
solani, F. roseum, M. phaseolina, and R. solani
were also identified. Some of the isolated fungi
in the present investigation were formerly
identified on marjoram (El-Gebaly, 1998;
Hilal et al., 1990), mint and rue (El-Shazly,
1996), and rosemary (Conway et al., 1997).
The pathogenicity tests of the isolated fungi
showed that F. oxysporum, and R. solani were
the most pathogenic fungi on marjoram
seedlings, causing pre- and post-emergence
damping-off, whereas in transplanted
marjoram plants, F. oxysporum and F.
semitectum showed the highest disease
incidence. These results are somewhat similar
to those obtained by El-Gebaly (1998). The
inhibitory effects of eucalyptus and thyme
essential oils at three concentrations (1000,
3000, and 6000 ppm) against the mycelial
growth of the tested fungi were confirmed,
since decreases were significant in most cases.
However, thyme oil was superior in its
inhibitory effect than eucalyptus oil,
additionally; the high concentration (6000
ppm) was the most effective than the others.
On the other hand, R. solani was the most
sensitive fungus affected by the oils, followed
by F. oxysporum, while F. semitectum was the
least affected. According to Farag et al.
(1989), Linskens and Jackson (1991),
Chauhan and Singh (1991), Zedan et al.
(1994), Zygadlo et al. (1994), and Halawa
(2004), a number of essential oils, including
these ones, have antifungal effects. Some
compounds of thyme oil are responsible for its
antifungal effects, such as Thymol and
carvacrol (Agarwal et al., 1979). However, the
antifungal effect of thyme may be explained
by the idea that it penetrates the cell wall,
causing damage to the lipoprotein cytoplasmic
membrane, which allows the cytoplasm to
escape (Zambonelli et al., 1996). Faghih-
Imani et al. (2020) reported the highest level
of antifungal effect of thyme essential oil
against F. graminearum and F. culmorum, the
causal pathogens of crown and root rot on
wheat. The authors also mentioned that
scanning the vegetative growth of pathogenic
fungi under a light microscope showed
destructive changes in the hyphae as a result of
thyme use. The obtained results are also in
agreement with Sarhan (2020) who found that
the use of thyme essential oil, followed by
eucalyptus, significantly inhibited the linear
growth of the tested fungi; F. moniliforme, F.
solani, and R. solani, the causal agents of
soybean root rot diseases compared with the
control. Among the twelve tested essential
oils, thyme showed the best antifungal effect
against the phytopathogenic fungi F.
oxysporum and Bortytis cinerea (Palfi et al.,
2019). Despite the smallest dosage applied 50
μl/10 ml of PDA agar medium, it was
Ali et al., 2020
118
completely capable of inhibiting the mycelial
growth of both phytopathogenic fungi. Under
greenhouse conditions, using essential oils,
biocides and fungicide as dipping treatments
for 20 minutes on marjoram transplants
significantly reduced disease incidence with
all the tested fungi compared to the untreated
control. However, the fungicide Actamyl 70%
wp was the most effective treatment, which
gave the lowest percentage of disease
incidence with all fungi under study compared
to the untreated control. Plant Guard had
superiority after Actamyl against F.
semitectum and F. oxysporum. These findings
are in harmony with Sarhan (2020), who found
that Plant Guard and Vitavax-200 treatments
gave the highest reduction percentage of
soybean root rot and damping-off under
greenhouse conditions. Thyme oil came in the
second order after Actamyl with R. solani and
F. oxysporum. Faghih-Imani et al. (2020)
revealed that thyme oil had superiority in
reducing the disease incidence and severity of
Fusarium species in inoculated wheat plants.
The positive effect of these treatments may be
explained by the suppression of wilt and root
rot pathogens after transplantation as a result
of the antifungal effects of their compounds,
which reduce or inhibit the spread of
infections. Abo-Elyousr et al. (2014) noticed
strong antagonistic effects of four isolates of
Trichoderma harzianum and five isolates of T.
longibrachiatum against Alternaria porri, the
causal pathogen of onion purple blotch. The
authors also reported that microscopic scans
revealed the growth of Trichoderma spp. over
the hyphae of A. porri with coiling,
surrounding, and lysis of the hyphae. On the
other hand, the results of Hilal et al. (1990;
1994; 2003), Helmy et al. (2001), and Abo El-
Ela (2003) correspond to our findings about the
effectiveness of the tested treatments against
the causal agents of root rot and wilt diseases
of marjoram. For marjoram plant growth,
Actamyl and Plant Guard were superior to
other treatments in most cases for improving
plant growth parameters, i.e., root length, plant
height, fresh and dry weight. It is possible that
these treatments have beneficial effects due to
their reduction in disease infection and
severity, and improve nutrient absorption.
Somewhat similar results were recorded on
enhancing plant growth with various
treatments by Nada (1997), Abo El-Ela
(2003), Hilal et al. (2003), Shafie (2004),
Halawa (2004), Mahmoud Amany (2004), and
Hassanin (2007).
5. Conclusion
The current study evaluated the efficiency of
some biocides and fungicidal alternatives in
the management of marjoram wilt and root rot
diseases. Actamyl 70% wp, Plant Guard and
thyme oil significantly decreased disease
incidence than the other treatments compared
to the untreated control with all the tested
fungi. Furthermore, Actamyl and Plant Guard
improved plant growth parameters, i.e., plant
height, root length, fresh and dry weight than
the other treatments in most cases. These
results indicate the possibility of using Plant
Guard (biocide) as a fungicidal alternative
against marjoram wilt and root rot diseases.
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