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Antifungal activities of essential oil from Moroccan endemic Anthemis

tenuisecta and seed emergence
A. Mziouid a, B. Chebli a,⇑, M. Berrabah b, N. Heimeur a, E.H. Mayad c
a
Mechanics, Process, Energy and Environment Laboratory, National School of Applied Sciences, IBN ZOHR University, PO Box: 1136/S, Agadir, Morocco
b
Laboratory of Solid Mineral and Analytical Chemistry, Department of Chemistry, Faculty of Sciences, Mohammed Premier University, Oujda, Morocco
c
Laboratory of Plant Biotechnology, Faculty of Sciences, Ibn-Zohr University, B.P. 28/S, Agadir, Morocco

a r t i c l e i n f o a b s t r a c t

Article history: Anthemis tenuisecta is an endemic Moroccan Asteraceae. The in vitro antifungal activity of Anthemis
Received 17 February 2020 tenuisecta essential oil (EO) using agar dilution and disk diffusion assays was evaluated against three
Received in revised form 8 March 2020 fungi isolated from decayed strawberries. Also, its efficiency to prolong the shelf life of raspberries and
Accepted 27 March 2020
strawberries was tested. Moreover, the seed emergence of this species was evaluated. The results con-
Available online xxxx
tributed to value this species regarding the antifungal potential of its EO as a natural alternative to chem-
ical fungicides, either as a model for synthesizing new effective bioactive products that suggest its
Keywords:
conservation for sustainable use.
Anthemis tenuisecta
Antifungal activity
Ó 2020 Elsevier Ltd. All rights reserved.
Essential oil Selection and peer-review under responsibility of the scientific committee of The Third International
Strawberries Conference on Materials and Environmental Science.
Raspberries
Seed emergence

1. Introduction uated against three fungi isolated from decayed strawberries. Fur-
thermore, its efficiency to extend the shelf life of raspberries and
The family Asteraceae comprises the largest number of strawberries was tested. Moreover, in vitro A. tenuisecta seed emer-
described species, belonging to 1600–1700 genera [1]. The Anthe- gence was evaluated for a useful approach of conservation and sus-
mis genus contains more than 210 species and is ranked as the tainable use.
second-largest genus in this family [2]. In North Africa, the Anthe-
mis genus (Asteraceae) is represented by 28 species, 16 of which
2. Materials and methods
exist in Morocco with an endemism ration close to 68% [3]. The
species of Anthemis genus prefer dry and are distributed naturally
2.1. Plant materials and EO extraction
on calcareous substrates [4]. Within this genus, many species have
been used in folk medicine for various healing properties [5].
Aerial parts of A. tenuisecta were collected when flowering
Anthemis tenuisecta is an annual plant, not yet described for its
between March and May 2015 from Tin Mansour area, cited near
medicinal properties. It is an endemic species of Morocco with a
Agadir, Morocco (latitude: 30°60 5800 , longitude: 9°330 600 ). A voucher
restricted distribution in the Middle Atlantic and the Haut Atlas
specimen has been deposited in the herbarium of the laboratory of
[6]. Chemical composition and antibacterial activity of A. tenuisecta
biotechnology, the National School of Applied Sciences, Ibn Zohr
EO were previously reported of this species from Safi Moroccan
University, Agadir, Morocco. The air-dried plant was hydrodisil-
region [7].
lated for approximately 3 h using a Clevenger type apparatus.
The aim of this study is the valorization of A. tenuisecta that is an
The A. tenuisecta EO yield was 0.4% (w/w). The EO were dried over
unexplored species. Indeed, this work provides more information
anhydrous sodium and kept in the dark at 4 °C until use.
on the biological activities of A. tenuisecta EO that has not been
reported earlier, to find new natural bioactive products. So, the
antifungal activity using agar dilution and disk diffusion was eval- 2.2. Antifungal tests

2.2.1. Fungal cultures

⇑ Corresponding author. Botrytis cinerea, Penicillium sp. and Rhizopus sp. were isolated
E-mail address: b.chebli@uiz.ac.ma (B. Chebli). directly from naturally decayed strawberries. Isolation of fungi

https://doi.org/10.1016/j.matpr.2020.03.686
2214-7853/Ó 2020 Elsevier Ltd. All rights reserved.
Selection and peer-review under responsibility of the scientific committee of The Third International Conference on Materials and Environmental Science.

Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
2 A. Mziouid et al. / Materials Today: Proceedings xxx (xxxx) xxx
was realized in 3 replicates using potato dextrose agar medium
(PDA) amended with gentamycin (10 mg/L) to stop the growth of
bacteria. The plates were incubated at 25 ± 2 °C for 5 days. Devel-
oping fungal cultures were repeatedly purified before their identi-
fication. Pure cultures of the three fungi were maintained on PDA
and stored at 4 °C.
2.2.2. In vitro antifungal assays
The in vitro antifungal activity of A. tenuisecta EO was carried
out against mycelial growth of the three fungal cultures using
the agar dilution method [8] and the disk diffusion assay [9]. Both
antifungal assays were assessed with 6 concentrations of A. tenui-
secta EO: 0.125, 0.25, 0.5, 1, 2 and 4 mL/mL.
For the agar dilution method, a stock solution was prepared by
dissolving pure EO in sterile solution 0.2% Tween 80 (v/v). To
obtain the desired concentrations, it consisted of mixing appropri-
ate amounts of stock solution with autoclaved potato dextrose agar
medium (PDA) maintained at 45 °C just before pouring into Petri
dishes. Then, a mycelial plug, 6 mm in diameter, was excised from
pure fungal culture and placed at the center of each Petri dish. Con-
trols consisted of un-amended PDA. The plates were sealed with
parafilm and incubated at 25 °C.
For volatile effect assay, the Petri dishes were filled with sterile
PDA medium and then inoculated with mycelial plugs, cut from
pure culture fungi. After solidification, the plates were inverted.
According to each concentration, different amounts of pure EO
were deposited on sterile filter paper discs, which were then
placed on the inner surface of the Petri dish lids. The dishes were
sealed with parafilm and incubated upside-down at 25 °C. Controls
consisted of sterile distilled water-soaked filter paper in the lid of
the Petri dish.
For both methods, the test was stopped when control Petri
dishes were covered by mycelial culture. For each concentration,
three plates were used. The percentage inhibition of mycelial
growth of the three tested fungi by the EO, compared to the con-
trols, was calculated according to the following equation:
Inhibition % ¼ 100ðC  TÞ=C
where C was the diameter of fungal growth on the control and T
was the diameter on the test plate.
2.2.3. Transfer experiments
For both antifungal assays, the lowest concentration that com-
pletely inhibited the growth of the fungus was considered the min-
imum inhibitory concentration (MIC). For assessing the fungistatic
nature of the A. tenuisecta EO on the target fungus, the fungal discs
from concentration plate with no growth were re-inoculated into
fresh medium and revival of their growth at 25 °C under monitor-
ing during five to ten days [10]. A fungicidal effect (MFC) was
where no mycelial growth occurred after additional incubation.
2.3. Evaluation of A. tenuisecta EO on the shelf life of raspberries and
strawberries
The experiment was realized according to [11]. Three concen-
trations: 200, 400 and 600 ppm of A. tenuisecta EO were prepared
using 0.4% Tween 80 and sterile distilled water under vigorous stir-
ring. Batches of ten raspberries and six strawberries were tested
separately per replicate. The EO emulsions were sprayed on fresh
fruits at room temperature. After spraying, berries were allowed
to dry for 15 min and then were stored at 4 °C. Visual decay of
raspberries and strawberries was evaluated for 12 days and the
contamination levels (percentage of berries showing decay signs)
were daily taken during storage. For treatments and controls (un-
treated), three replicates were made. At the end of storage, the
Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
inhibition percentages of postharvest decay were calculated fol-
lowing the formula:
Inhibition % ¼ ðS  BÞ
where, S and B were the decay percentages on the control sample
and treated berries, respectively. The correlations between the inhi-
bition percentages and treatment concentrations were evaluated.
2.4. A. tenuisecta seed emergence
2.4.1. Seed morphology and weight
Fresh seeds were separated from flowers using a dissecting nee-
dle. The seeds were visually checked to eliminate nonviable ones
(thin and empty seeds). The length and width of fresh seeds were
measured of 20 different flowers and the seed weight was deter-
mined with five replicates of 25 seeds each. The seeds were kept
in air-dried plants stored under ambient laboratory conditions.
2.4.2. Seed emergence tests
The percentage of seed emergence was tested at four different
age levels: S1 (Seeds collected one month after the plant collec-
tion), S2 (Seeds collected from dried plants 6 months after collec-
tion) S3 (Seeds collected from dried plants 12 months after
collection) and S4 (Seeds collected from dried plants 24 months
after collection). The seed emergence was evaluated on cotton,
peat and native soil (sieved to remove stones) which were auto-
claved at 121 °C for 45 min, placed into plastic boxes and moist-
ened with distilled water. The test was conducted under
laboratory light and temperature, and distilled water was added
as need. Three replicates of 25 to 35 seeds each were made. Seed
emergence was recorded for a one month observation period. To
determine the effect of photoperiod on seed emergence, the seeds
were also incubated in complete darkness at the same conditions
(Petri dishes wrapped in aluminum foil) and were checked only
when incubation was terminated. After sterilization with 1%
sodium hypochlorite solution for 1 min and rinsed three times
with distilled water, some germinated seeds were transferred
aseptically to an agar medium [12] with 1% (w/v) sucrose and
0.8% (w/v) agar, pH 5.7, autoclaving at 121 °C for 20 min. The test
on the agar medium was kept at 22 °C under a photoperiod of
16 h/8h light/dark with cool-white fluorescent (40 mmol m2 s1).
The final seed emergence percentage (SEP) was determined follow-
ing the formula:
SEP ¼ ðnumber of emerged seeds=number of seeds per sampleÞ
 100
2.5. Statistical analysis
All assays were carried out in triplicate and were repeated
twice. All data were evaluated with one-way ANOVA using SPSS
16.0 software. The means comparisons were performed using
Newman & Keuls tests. The results are presented as mean val-
ues ± standard deviation with different letters which are statisti-
cally significant at 5% level probability.
3. Results and discussions
3.1. Antifungal tests
The inhibition percentages obtained in antifungal activity of dif-
ferent concentrations of A. tenuisecta EO using agar dilution and
disk diffusion techniques are presented in Table 1.
 A. Mziouid et al. / Materials Today: Proceedings xxx (xxxx) xxx 3

Table 1
The inhibition percentages of A. tenuisecta EO on mycelial growth of B. cinerea, Rhizopus sp. and Penicillium sp.

Pathogens Antifungal assays Concentrations (mL/mL) IC50 (mL/mL) R2

0.125 0.25 0.5 1 2 4
B. cinerea Agar dilution 0 ± 0.00f 0 ± 0.00f 5 ± 0.00f 43 ± 6.56c 100 ± 0.00 a 100 ± 0.00 a 1.18 0.896
Disk diffusion 14 ± 1.73 e 19 ± 1.73 e 34 ± 7.94 d 76.33 ± 1.15b 100 ± 0.00 a 100 ± 0.00 a 0.68 0.846
Rhizopus sp. Agar dilution 0 ± 0.00 e 0 ± 0.00 e 0 ± 0.00 e 18 ± 1.73 d 95 ± 0.00b 100 ± 0.00 a 1.36 0.912
Disk diffusion 0 ± 0.00 e 0 ± 0.00 e 0 ± 0.00 e 29 ± 3.60c 100 ± 0.00 a 100 ± 0.00 a 1.27 0.9
Penicillium sp. Agar dilution 0 ± 0.00 g 0 ± 0.00 g 14 ± 1.73 e 22.33 ± 2.51 d 32.33 ± 2.51c 40.67 ± 4.04b >4 0.912
Disk diffusion 0 ± 0.00 g 7.33 ± 2.51f 19.67 ± 2.51 d 29 ± 1.73c 38 ± 3.60b 49 ± 5.29 a >4 0.909

Mean value of three replicates ± Standard deviation.

Letters indicate statistically difference to 5% level probability according to Newman and Keuls test.
R2: Correlation coefficient.

As can be seen in this table, there was no inhibitory effect
against fungi at many concentrations. However, there were strong
correlations between increasing the EO concentration and a
decrease of mycelial growth of the three tested fungi in both anti-
fungal assays. In the agar dilution method, the EO completely
inhibited B. cinerea and Rhizopus sp. at 2 and 4 mL/mL, respectively.
In disk diffusion assay, no mycelial growth was shown of B. cinerea
and Rhizopus sp. at 2 mL/mL. For Penicillium sp., the inhibitory effect
of EO at the highest concentration was lower than 50% in both anti-
fungal methods. The IC50 values of B. cinerea and Rhizopus sp. in
disk diffusion assay were lower than those of agar dilution. Also,
Penicillium sp. was better inhibited in the disk diffusion technique
at all concentrations. So, the A. tenuisecta EO exhibited better anti-
fungal activity using disk diffusion assay against the three studied
fungi.
Table 2 shows the MIC and MFC of A. tenuisecta EO obtained for
Rhizopus sp. and B. cinerea. By both antifungal assays, MFCs for B.
cinerea were lower/or equal to 4 mL/mL, whereas those for Rhizopus
sp. were strictly higher than 4 mL/mL. In general, it can be con-
cluded that B. cinerea was the most sensitive fungus to the A. tenui-
secta EO and Penicillium sp. was the most resistant.
The EO of A. tenuisecta exhibited significant antifungal activities
which might be attributed to their major constituents. The chem-
ical composition of A. tenuisecta EO used in this study was previ-
ously determined by our laboratory team using gas
chromatography coupled with mass spectrometry. The major com-
ponents detected in the EO were: carvacrol (19.52%), bicyclo[2.2.1]
heptan-2-one, 1,7,7-trimethyl-, (1R)- (16.76%), 5-hepten-2-one, 6-
methyl- (16.46%) and camphene (13.18%) which are reported as
antifungal agents or a responsible for increasing the antifungal
activity [13–16]. Additionally, the synergistic interaction of the
EO components contributed probably to the antifungal activity [9].
The resistance of Penicillium sp. may be ascribed to the active
enzymes which some species of Penicillium genus produce that
may deactivate the effective antifungal compounds of EO [17]. It
is may be also possible that the hyphae of Rhizopus sp. and B.
cinerea were more sensitive to A. tenuisecta EO compared to that
of Penicillium sp.
Table 2
Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC)
of A. tenuisecta EO.
Pathogens Antifungal assays Concentrations (mL/
mL)
MIC MFC
Botrytis cinerea Agar Dilution 2 4
Disk Diffusion 2 4
Rhizopus sp. Agar Dilution 4 >4
Disk Diffusion 2 >4
MIC: fungistatic concentration.
MFC: fungicidal concentration.
The best inhibitory effect of A. tenuisecta EO was given in disk
diffusion assay. These findings agreed with other studies on Melissa
officinalis and Pulicaria mauritanica EOs [9,18]. The results of this
comparison could be attributed to the low-molecular-weight and
low solubility of EO, and the lipophilic nature of both fungal struc-
ture and EO that easily diffuse into the cell membrane [19].
Previous studies have tested the antifungal activity of EOs
against B. cinerea and species from Rhizopus and Penicillium genus.
The vapors of Solidago Canadensis L. EO inhibited 78% at 3 mL/mL%
of mycelial growth of B. cinerea [20]. The MICs of three species
from the Satureja genus were 300 mL/mL, 300 mL/mL and 600 mL/
mL, against B. cinerea, R. Stolonifer, and P. digitatum, respectively
[21]. The inhibition rate of Pulicaria mauritanica EO reached 100%
and 87.36% at 2 mL/mL against P. expansum and R. stolonifer, respec-
tively [9]. The Warionia saharae EO inhibited at 2 mL/mL 85.51% and
82.89% of P. expansum and R. stolonifer, respectively [22]. Thus, it
can be suggested that the EO of A. tenuisecta have a great in vitro
antifungal effect against the three tested fungi.
3.2. Evaluation of A. tenuisecta on the shelf life of raspberries and
strawberries
The postharvest decay percentages daily measured of EO trea-
ted berries and their controls (untreated) during 12 days at cold
storage are shown in Fig. 1 (raspberries) and Fig. 2 (strawberries).
Until the seventh-storage day, 400 and 600 ppm EO treated
raspberries showed no postharvest decay, while those untreated
were half infected. At day 12, control sample was completely
infected, whereas the decay rates of 200, 400 and 600 ppm EO trea-
ted raspberries were 23.33%, 10% and 10%, respectively.
As the Fig. 2 showed, no decayed strawberries were obtained
when A. tenuisecta EO were applied at 400 and 600 ppm until the
fifth-storage day. At day 8, the decay rate of untreated strawberries
was 66.67% while both 400 and 600 ppm EO treated fruits pre-
sented only 5.67%. At the end-storage day, A. tenuisecta EO has pre-
served 45% of 400 and 600 ppm treated strawberries.
For both treated berries, no statistical difference was observed
between A. tenuisecta EO concentrations (p < 0.05) unless
200 ppm EO treated strawberries that were equal to the control.
It can be also marked the potential correlation between A. tenui-
secta EO concentration and the inhibition percentage of the decay
rate.
Substitution of synthetic fungicides treatments by the applica-
tion of EOs for controlling post-harvest pathogens is gaining
increasing interest [17].
In vitro studies of the A. tenuisecta EO indicated its potential as a
natural antifungal agent against the most prevalent phy-
topathogenic fungi found on strawberries [21,23]. However, many
limiting factors within in vivo conditions such as the high fungal
resistance [24], a problem solubility of EO [25], the interactions
between phenolic compounds and the food matrix [26], toxicity

Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
4 A. Mziouid et al. / Materials Today: Proceedings xxx (xxxx) xxx

Fig. 1. The daily evolution of the decay rate of A. tenuisecta EO treated raspberries during 12 days at 4 °C. The treated raspberries with 200 ppm (-r-), 400 ppm (-j-) and
600 ppm (–▲–) of EO. The sample of untreated raspberries used as control (  ). Mean of three replicates (number) ± SD (vertical bar). The correlation coefficient between
EO concentrations and the inhibition percentages of decay at the end storage day R2 = 0,945.

Fig. 2. The daily evolution of the decay rate of A. tenuisecta EO treated strawberries during 12 days at 4 °C. The treated strawberries with 200 ppm (-r-), 400 ppm (-j-) and
600 ppm (–▲–) of EO. The sample of untreated strawberries used as control (  ). Mean of three replicates (number) ± SD (vertical bar). The correlation coefficient between
concentrations and the inhibition percentages of decay at the end storage day R2 = 0,941.

effect of some EO compounds on cell fruit integrity [11,27] and
easy volatility of carvacrol [28]. Thus, further in vivo studies to
investigate the efficiency of A. tenuisecta EO as a bio-treating pesti-
cide of fruit were necessary. The results demonstrated that the A.
tenuisecta EO significantly reduce the postharvest decay in raspber-
ries and strawberries during the shelf life at cold storage. However,
the A. tenuisecta EO preserved raspberries better than strawberries
indicating that the efficiency of EO is also related to the fruit char-
acteristic [29]. In addition, due to the organoleptic properties of
fresh fruit, the efficiency of EO at lower concentrations is highly
required.
Other works have shown the efficiency of EOs or their compo-
nents to extend the shelf life of fresh fruits. Allyl isothiocyanate
at 5 mL/l and methyl jasmonate at 22.4 mL/l improved the posthar-
vest shelf life of fresh raspberries during 10 days of storage at 10 °C
[30,31]. The EOs of Thymus vulgaris and Mentha piperita at 200 ppm
were able to reduce the postharvest decay of treated strawberries
by 40% compared to the control during 14 days at 4 °C [11]. The

Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
 A. Mziouid et al. / Materials Today: Proceedings xxx (xxxx) xxx 5

percentages of decayed cherry tomatoes treated by 200 mL/mL of
Cicuta virosa EO was reduced by 10% compared to the control
[32]. The EOs of Satureja khuzistanica and Thymus daenensis at
2000 mL/L were efficient for controlling anthracnose in avocado
postharvest [33]. Thus, the A. tenuisecta EO seems to be a promising
eco-friendly alternative that could be used in the agro-food
industry.
3.3. A. tenuisecta seed emergence
3.3.1. Seed morphology and weight
A seed was conical with a length of 1.5–2 mm a wide of 0.4–
0.5 mm. The seed number per flower ranged from 5 to 22. The fresh
seed mass was 0.18 ± 0.05 mg.
3.3.2. Seed emergence tests
The seed emergence percentages of A. tenuisecta in three seed-
ling supports and at four different age levels are summarized in
Table 3.
At the first age level, there was no seed emergence in both seed-
ling supports. The best seed emergence percentages were at the
third age level (12 months of storage). As can be observed, no sta-
tistical difference was observed between peat and native soil of A.
tenuisecta (p < 0.05) and also between the second and third age
levels.
The pictures of A. tenuisecta seed emergence at different seed-
ling supports are presented in Fig. 3.
The stimulation of seed germination by alternating tempera-
tures is also common in species from arid zones [34,35], thereby;
the seed emergence tests were carried out under laboratory condi-
tions. The results indicate that A. tenuisecta exhibits seed dormancy
at the first age level. As it is an aromatic plant, the dormancy at the
start of storage may be due to the concentration of phenolic com-
pounds in the plant which had probably to be reduced to remove
the dormancy of seeds [36]. Although the endemism of A. tenui-
secta and its restricted distribution area, the results of seed emer-
gence in peat showed that the germination of A. tenuisecta seeds is
not strictly linked to its native soil. Moreover, the disinfection and/
or specific treatments were not primary for obtaining seeds able to
germinate. A prior study reported that without any treatment the
seed emergence percentage of Prealpine endemic Asteraceae spe-
cies Telekia speciosissima was less than 10% [37]. Also, the percent-
ages of seed germination by alternating temperatures of Baccharis
dracunculifolia and Zinnia peruviana two native Asteraceae species
of Argentina were 43% and 44%; respectively [38]. In the darkness,
no seed emergence was recorded. This indicates that alternating
light is necessary to ensure the germination of A. tenuisecta seeds
as other small-seeded species [39].
Propagation from seed is a crucial step for the ex-situ conserva-
tion [40], the present work demonstrates that ex-situ propagation
of A. tenuisecta is possible from seeds. Moreover, disinfection pro-
cedures and phytohormones can generate better percentages of
seed emergence, as another study in endemic Asteraceae species
Santolina semidentata reported [41].

Table 3 4. Conclusions
The percentages of A. tenuisecta seed emergence at four different age levels.
Due to consumer demands of agricultural products without
Seed ages Percentage of seed emergence (%)
chemical residues and climate change impacts, the concept of sus-
Cotton Peat Native soil
tainable production was gaining interest all around the world that
S1 0 ± 0.00c 0 ± 0.00c 0 ± 0.00c the discovery of natural controlling agents has been growing value.
S2 19.67 ± 6.51b 36.67 ± 5.77 a 38.45 ± 7.86 a
A. tenuisecta EO revealed to be a potential natural antifungal agent.
S3 22.45 ± 3.67b 37.67 ± 3.57 a 39.67 ± 5.62 a
S4 2.33 ± 1.53c 0.33 ± 0.58c 1.9 ± 1.4c Due to its inhibition effect against three phytopathogenic fungi and
efficiency to extend the shelf life of fresh berries, this EO is sug-
Mean value of three replicates ± Standard deviation.
gested as an alternative to chemical fungicides, either as a model
Letters indicate statistically difference to 5% level probability according to Newman
and Keuls test.
for synthesizing new effective bioactive products. Furthermore,
the investigation of other biological properties is also supported

Fig. 3. The A. tenuisecta seed emergence at different seedling supports, agar medium (A and B), peat (C and D) and native soil (E).

Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
6 A. Mziouid et al. / Materials Today: Proceedings xxx (xxxx) xxx
to promote the biotechnological use of this plant. Moreover, the
hopeful results of in vitro seed emergence encourage establishing
a strategy for its conservation which builds a real model for the
sustainable production of fruits and vegetables.
CRediT authorship contribution statement
A. Mziouid: Conceptualization, Methodology, Investigation,
Writing - original draft, Formal analysis. B. Chebli: Conceptualiza-
tion, Supervision, Writing - review & editing, Validation. M. Berra-
bah: Investigation, Resources, Formal analysis. N. Heimeur:
Resources, Investigation. E.H. Mayad: Formal analysis, Writing -
review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing finan-
cial interests or personal relationships that could have appeared
to influence the work reported in this paper.
References
[1] V.A. Funk, A. Susanna, T.F. Steussy, H.E. Robinson, Classification of compositae,
Syst. Evol. Biogeogr Compos. (2009).
[2] C. Oberprieler, Phylogenetic relationships in Anthemis L. (Compositae,
Anthemideae) based on nrDNA ITS sequence variation, Taxon 50 (2001)
745–762.
[3] J. El Oualidi, H. Khamar, M. Fennane, M. Ibn Tattou, S. Chauvet, M.S. Taleb,
Checklist des endémiques et spécimens types de la flore vasculaire de l’Afrique
du Nord, Doc Inst Sci Univ. Mohammed V-Agdal Rabat, 2012.
[4] A. Uzel, A. Guvensen, E. Cetin, Chemical composition and antimicrobial activity
of the essential oils of Anthemis xylopoda O. Schwarz from Turkey, J.
Ethnopharmacol. 95 (2004) 151–154.
[5] J. Bellakhdar, Medicinal Plants in North Africa and Basic Care, Handb. Mod.
Herb. Med, Fennec Casablanca, 2006.
[6] H. Rankou, A. Culham, S.L. Jury, M.J. Christenhusz, The endemic flora of
Morocco, Phytotaxa 78 (2013) 1–69.
[7] E.F. Hanbali, F. Mellouki, M. Akssira, B. Ma, Composition and antimicrobial
activity of essential oil of Anthemis tenuisecta ball, J. Essent. Oil Bear. Plants 10
(2007) 499–503.
[8] B. Chebli, M. Achouri, L.A. Idrissi Hassani, M. Hmamouchi, Chemical
composition and antifungal activity of essential oils of seven Moroccan
Labiatae against Botrytis cinerea Pers: Fr, J. Ethnopharmacol. 89 (2003) 165–
169.
[9] M. Znini, G. Cristofari, L. Majidi, J. Paolini, J.M. Desjobert, J. Costa, Essential oil
composition and antifungal activity of Pulicaria mauritanica Coss., against
postharvest phytopathogenic fungi in apples, LWT-Food, Sci. Technol. 54
(2013) 564–569.
[10] V. Hervieux, E.S. Yaganza, J. Arul, R.J. Tweddell, Effect of organic and inorganic
salts on the development of Helminthosporium solani, the causal agent of
potato silver scurf, Plant Dis. 86 (2002) 1014–1018.
[11] K.D. Vu, R.G. Hollingsworth, E. Leroux, S. Salmieri, M. Lacroix, Development of
edible bioactive coating based on modified chitosan for increasing the shelf life
of strawberries, Food Res. Int. 44 (2011) 198–203.
[12] T. Murashige, F. Skoog, A revised medium for rapid growth and bio assays with
tobacco tissue cultures, Physiol. Plant. 15 (1962) 473–497.
[13] M.B. Reddy, P. Angers, A. Gosselin, J. Arul, Characterization and use of essential
oil from Thymus vulgaris against Botrytis cinerea and Rhizopus stolonifer in
strawberry fruits, Phytochemistry 47 (1998) 1515–1520.
[14] G. Arras, M. Usai, Fungitoxic activity of 12 essential oils against four
postharvest citrus pathogens: chemical analysis of Thymus capitatus oil and
its effect in subatmospheric pressure conditions, J. Food Prot. 64 (2001) 1025–
1029.
[15] K.S. e Silva, B.R. da S Neto, P.F. Zambuzzi-Carvalho, C.M. de Oliveira, L.B. Pires,
L. Kato, A.M. Bailão, J.A. Parente-Rocha, O. Hernández, J.G. Ochoa, Response of
Paracoccidioides lutzii to the antifungal camphene thiosemicarbazide
determined by proteomic analysis, Future Microbiol. 13 (2018) 1473–1496.
[16] M.U. Yamaguchi, A.P. Barbosa da Silva, T. Ueda-Nakamura, B.P. Dias Filho, C.
Conceição da Silva, C.V. Nakamura, Effects of a thiosemicarbazide camphene
derivative on Trichophyton mentagrophytes, Molecules 14 (2009) 1796–1807.
[17] D. Sivakumar, S. Bautista-Baños, A review on the use of essential oils for
postharvest decay control and maintenance of fruit quality during storage,
Crop Prot. 64 (2014) 27–37.
Please cite this article as: A. Mziouid, B. Chebli, M. Berrabah et al., Antifungal activities of essential oil from Moroccan endemic Anthemis tenuisecta and
seed emergence, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2020.03.686
[18] Y. El Ouadi, M. Manssouri, A. Bouyanzer, L. Majidi, H. Bendaif, H. Elmsellem, M.
A. Shariati, A. Melhaoui, B. Hammouti, Essential oil composition and antifungal
activity of Melissa officinalis originating from north-Est Morocco, against
postharvest phytopathogenic fungi in apples, Microb. Pathog. 107 (2017) 321–
326.
[19] J. Tian, X. Ban, H. Zeng, B. Huang, J. He, Y. Wang, In vitro and in vivo activity of
essential oil from dill (Anethum graveolens L.) against fungal spoilage of cherry
tomatoes, Food Control 22 (2011) 1992–1999.
[20] S. Liu, X. Shao, Y. Wei, Y. Li, F. Xu, H. Wang, Solidago canadensis L. essential oil
vapor effectively inhibits Botrytis cinerea growth and preserves postharvest
quality of strawberry as a food model system, Front. Microbiol. 7 (2016) 1179.
[21] M. Farzaneh, H. Kiani, R. Sharifi, M. Reisi, J. Hadian, Chemical composition and
antifungal effects of three species of Satureja (S. hortensis, S. spicigera, and S.
khuzistanica) essential oils on the main pathogens of strawberry fruit,
Postharvest Biol. Technol. 109 (2015) 145–151, https://doi.org/10.1016/
j.postharvbio.2015.06.014.
[22] M. Znini, G. Cristofari, L. Majidi, A. El Harrak, J. Paolini, J. Costa, In vitro
antifungal activity and chemical composition of Warionia saharae essential oil
against 3 apple phytopathogenic fungi, Food Sci. Biotechnol. 22 (2013) 113–
119.
[23] V.H. Tournas, E. Katsoudas, Mould and yeast flora in fresh berries, grapes and
citrus fruits, Int. J. Food Microbiol. 105 (2005) 11–17.
[24] S. Rupp, R.W. Weber, D. Rieger, P. Detzel, M. Hahn, Spread of Botrytis cinerea
strains with multiple fungicide resistance in German horticulture, Front.
Microbiol. 7 (2017) 2075.
[25] F. Demirci, K. Guven, B. Demirci, M.Y. Dadandi, K.H.C. Baser, Antibacterial
activity of two Phlomis essential oils against food pathogens, Food Control 19
(2008) 1159–1164.
[26] M.M. Yooussef, Q. Pham, P.N. Achar, M.Y. Sreenivasa, Antifungal activity of
essential oils on Aspergillus parasiticus isolated from peanuts, J. Plant Prot. Res.
56 (2016) 139–142.
[27] J. Hadian, M. Ghasemnezhad, H. Ranjbar, M. Frazane, M. Ghorbanpour,
Antifungal potency of some essential oils in control of postharvest decay of
strawberry caused by Botrytis cinerea, Rhizopus stolonifer and Aspergillus niger,
J. Essent. Oil Bear. Plants. 11 (2008) 553–562.
[28] G.B. Martínez-Hernández, M.L. Amodio, G. Colelli, Carvacrol-loaded chitosan
nanoparticles maintain quality of fresh-cut carrots, Innov. Food Sci. Emerg.
Technol. 41 (2017) 56–63.
[29] J.G. Lopez-Reyes, D. Spadaro, M.L. Gullino, A. Garibaldi, Efficacy of plant
essential oils on postharvest control of rot caused by fungi on four cultivars of
apples in vivo, Flavour Fragr. J. 25 (2010) 171–177.
[30] C.Y. Wang, Maintaining postharvest quality of raspberries with natural volatile
compounds, Int. J. Food Sci. Technol. 38 (2003) 869–875.
[31] K. Chanjirakul, S.Y. Wang, C.Y. Wang, J. Siriphanich, Effect of natural volatile
compounds on antioxidant capacity and antioxidant enzymes in raspberries,
Postharvest Biol. Technol. 40 (2006) 106–115.
[32] J. Tian, X. Ban, H. Zeng, J. He, B. Huang, Y. Wang, Chemical composition and
antifungal activity of essential oil from Cicuta virosa L. var. latisecta Celak, Int. J.
Food Microbiol. 145 (2011) 464–470.
[33] A. Sarkhosh, A.I. Vargas, B. Schaffer, A.J. Palmateer, P. Lopez, A. Soleymani, M.
Farzaneh, Postharvest management of anthracnose in avocado (Persea
americana Mill.) fruit with plant-extracted oils, Food Packag. Shelf Life 12
(2017) 16–22.
[34] A. Mahmoud, A.M. El Sheikh, S.A. Baset, Germination of two halophytes:
Halopeplis perfoliata and Limonium axillare from Saudi Arabia, J. Arid Environ. 6
(1983) 87–98.
[35] R.J. Probert, The role of temperature in the regulation of seed dormancy and
germination, Seeds Ecol. Regen. Plant Communities. 2 (2000) 261–292.
[36] R.D. Williams, R.E. Hoagland, The effects of naturally occurring phenolic
compounds on seed germination, Weed Sci. 30 (1982) 206–212.
[37] G. Brusa, R. Ceriani, B. Cerabolini, Seed germination in a narrow endemic
species (Telekia speciosissima, Asteraceae): implications for ex situ
conservation, Plant Biosyst. 141 (2007) 56–61.
[38] G. Galíndez, P. Ortega-Baes, M.I. Daws, S. Sühring, A.L. ScoPel, H.W. Pritchard,
Seed mass and germination in Asteraceae species of Argentina, Seed Sci.
Technol. 37 (2009) 786–790.
[39] S.-K. Shen, F.-Q. Wu, G.-S. Yang, Y.-H. Wang, W.-B. Sun, Seed, germination and
seedling emergence in the extremely endangered species Rhododendron
protistum var. giganteum—the world’s largest Rhododendron, Flora-Morphol.
Distrib. Funct. Ecol. Plants 216 (2015) 65–70.
[40] E. Khurana, J.S. Singh, Ecology of seed and seedling growth for conservation
and restoration of tropical dry forest: a review, Environ. Conserv. 28 (2001)
39–52.
[41] A. Gomes, R.C. Pimpão, S. Fortalezas, I. Figueira, C. Miguel, C. Aguiar, L.
Salgueiro, C. Cavaleiro, M.J. Gonçalves, A. Clemente, Chemical characterization
and bioactivity of phytochemicals from Iberian endemic Santolina semidentata
and strategies for ex situ propagation, Ind. Crops Prod. 74 (2015) 505–513.