Journal of Arid Environments 199 (2022) 104703

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Journal of Arid Environments

journal homepage: www.elsevier.com/locate/jaridenv

Arbuscular mycorrhizal fungi and/or organic amendment enhance the

tolerance of prickly pear (Opuntia ficus-indica) under drought stress
Soufiane Lahbouki a, b, d, Raja Ben-Laouane a, b, Mohamed Anli a, b, Abderrahim Boutasknit a, b,
Youssef Ait-Rahou b, c, Mohamed Ait-El-Mokhtar b, Soumaya El Gabardi c, Allal Douira c,
Said Wahbi a, b, Abdelkader Outzourhit d, Abdelilah Meddich a, b, *
a
Center of Agrobiotechnology and Bioengineering, Research Unit Labelled CNRST (Centre AgroBiotech-URL-CNRST-05), "Physiology of Abiotic Stresses Team", Cadi
Ayyad University, Marrakech, 40000, Morocco
b
Laboratory of Agro-Food, Biotechnologies and Valorization of Plant Bioresources (Agrobioval), Faculty of Sciences Semlalia, Cadi Ayyad University, PO Box 2390,
Marrakech, 40000, Morocco
c
Laboratoire des Productions Végétales, Animales et Agro-industrie, Equipe de Botanique, Biotechnologie et Protection des Plantes, Faculté des Sciences, BP. 133,
Université Ibn Tofail, Kenitra, 14000, Morocco
d
Laboratory of Nanomaterials for Energy and Environment Physics Department, Faculty of Sciences Semlalia, Cadi Ayyad University, P.O. Box 2390, Marrakech,
40000, Morocco

A R T I C L E I N F O A B S T R A C T

Keywords: Drought stress is one of the most serious constraints that prevents plants from attaining their full potential,
Prickly pear particularly in arid and semi-arid regions. Among notable drought-tolerant plants, prickly pear stands as a plant
Arbuscular mycorrhizal fungi capable of tolerating daring water stress conditions, thanks to its water storage and CO2 uptake capabilities.
Vermicompost
However, its full potential remains constrained under drought stress. The use of biostimulants such as arbuscular
Drought stress
Cladodes growth
mycorrhizal fungi (AMF) and vermicompost (Vc) can help the plant to mitigate drought. The present study
Biochemical responses provides insights into the effects of an AMF, a vermicompost at a dose of 5%, and their combination on growth,
and biochemical responses of prickly pear cladodes under severe water stress (SWD), moderately watered (MD),
and well-watered (WW) treatments (15, 30 and 50% of field capacity (FC), respectively). AMF + Vc treatment
showed the highest values of AMF colonization and growth parameters under drought stress. The application of
these biostimulants increased physiological and biochemical parameters, under water deficit, and led to a decline
in malondialdehyde (MDA) and hydrogen peroxide (H2O2), notably on plants treated with AMF + Vc. Overall,
the results pointed out the role of AMF and vermicompost as strategic applicable tools in better mitigation of
drought stress in the prickly pear.

1. Introduction
The majority of global land area is situated under arid, semi-arid and
dry sub-humid climates, where agricultural productivity is affected by
several constraints especially drought stress (Zarei, 2018). In these re­
gions, plants are often subjected to periods of water deficit during their
growth and development (Talbi et al., 2020). Water scarcity negatively
affects physiological processes. mainly photosynthesis, transpiration,
cell turgidity and cell elongation, functioning, and productivity in the
plant (Chandra et al., 2021). Furthermore, water deficit can significantly
impact the plants antioxidant defense mechanism, as it can exacerbate
the production of reactive oxygen species (ROS) (Talbi et al., 2020). This
places the water deficit restriction and its management at the center of
agricultural concerns, and makes the efficient use of water a crucial need
(Vanham and Leip, 2020). Thence, encouraging the cultivation of
drought tolerant species requiring low water needs, presenting high
nutritional value, and social, economic, and environmental benefits can
help mitigate drought and water shortage (Nefzaoui, 2018).
Opuntia ficus-indica (L.) Mill is a cactus species that originates from
the American continent. It has been widely introduced into many parts
of the world including Africa, Mediterranean Europe, and Australia.
Opuntia ficus indica plays an important economic and ecological role in

* Corresponding author. Department of Biology, Laboratory of Agro-Food, Biotechnologies and Valorization of Plant Bioresources, Cadi Ayyad University, Faculty
of Sciences Semlalia, Bd My Abdallah BP 2390, 40000, Marrakesh, Morocco.
E-mail address: a.meddich@uca.ma (A. Meddich).

https://doi.org/10.1016/j.jaridenv.2021.104703
Received 5 August 2021; Received in revised form 22 November 2021; Accepted 22 December 2021
Available online 31 December 2021
0140-1963/© 2022 Elsevier Ltd. All rights reserved.
S. Lahbouki et al.
the agricultural systems of arid and semi-arid areas as it produces edible
fruits and cladodes, it can be used to stabilize soil and counter soil
erosion, it can be used as a forage substitute, and it is considered as a
dietary supplements and nutraceutical (Ventura-Aguilar et al., 2017;
Nefzaoui, 2018). Furthermore, O. ficus-indica has an extraordinary
ability to store water by deeply modulating photosynthesis (Ventur­
a-Aguilar et al., 2017), which reflects its ability to adapt to drought
conditions. This adaptation results in the acquisition of crassulacean
acid metabolism (CAM) associated with appropriate stomatal behavior
(Guralnick et al., 2020). These plants function by opening their stomata
during the dark period to absorb carbon dioxide, which is then stored in
the vacuoles in the form of malic acid (Scalisi et al., 2016; Ventur­
a-Aguilar et al., 2017), to be subsequently decarboxylated by the
Calvin-Benson cycle, and assimilated in the form of carbohydrates once
light is available. However, several studies have shown that the cactus
species can be damaged by severe drought stress, affecting yield,
growth, physiological, and biochemical parameters (Scalisi et al., 2016;
Campos et al., 2021). Hence, the application of biostimulants such as
bio-organic fertilizers and symbiotic microorganisms can increase the
ability of plants to withstand drought stress conditions.
Arbuscular mycorrhizal fungi (AMF) are one of the most important
microorganisms used to enhance plants drought tolerance. They are
obligate biotrophs that form symbiotic associations with the vast ma­
jority of land plants (Smith and Read, 2008). AMF widely occur in arid
and semi-arid regions (Mahmoudi et al., 2020), where drought nega­
tively affect the formation and function of mycorrhizal symbiosis
(Begum et al., 2019; Quiroga et al., 2020). In recent years, studies
indicated that AMF can increase plant growth and nutrients uptake,
decrease plants yield losses, reduce oxidative damage under stress
conditions (Toubali et al., 2020; Fattahi et al., 2021), improve regulation
of stomatal opening to get higher water-use efficiency and water status
by regulating plant aquaporin activities (Symanczik et al., 2020; Cheng
et al., 2021b).
Amendment of soil with bio-organic compounds such as vermicom­
post is among promising alternatives to improve physical and chemical
as well as biological soil properties, leading to the increase of growth
and yield improvement of plants under drought conditions (Demir,
2019; Rashtbari et al., 2020). Vermicompost is one of the most stabi­
lizing organic amendments produced by a non-thermophilic process.
Earthworms and microorganisms play the main role in this process (Cao
et al., 2021). Their interaction accelerates the decomposition of organic
matter, enhances microbial activity rates, and suppresses pathogens
(Huang et al., 2020). Furthermore, vermicompost plays a key role in
inducing plant drought tolerance through increasing nutrients uptake
and activation of antioxidant machinery (Rashtbari et al., 2020; Zeigh­
ami Nejad et al., 2020).
Several studies have shown that AMF inoculation and/or vermi­
compost supplementation had a positive effect on plants growth, phys­
iological and biochemical parameters (Toubali et al., 2020; Zeighami
Nejad et al., 2020; Fattahi et al., 2021). Until now, the effect of AMF
and/or vermicompost on cactus growth has been rarely studied (Ruiz
Rubio et al., 2012; Lahbouki et al., 2021). To our knowledge, no data are
available on the effect of combined application of AMF and vermicom­
post on drought tolerance of cactus and underlying mechanisms. In this
context, the aim of this experiment was to study the effect of AMF and
vermicompost applied separately and combined on the physiological
and biochemical mechanisms of O. ficus-indica cladodes subjected to
drought stress under greenhouse conditions.
2. Materials and methods
2.1. Biological, biostimulants materials and experimental design
The present study was carried out in a greenhouse (photon flux
density ranged from 500 to 750 μmol m− 2 s− 1) at the Faculty of Science
Semlalia, Cadi Ayyad University, Marrakech, Morocco. Plant material
2
Journal of Arid Environments 199 (2022) 104703
used in this experiment are mature cladodes of O. ficus-indica (34.11 ±
0.8 cm long, 13.02 ± 0.4 cm wide, and 7.5 ± 0.7 mm thick) obtained
from the Rhamna region, Morocco. The cladodes were dried for two
weeks in the shade in order to allow healing of the wounded areas. Next,
they were planted in 3 L polyethylene pots (one cladode per pot) con­
taining 2.5 kg of sterilized soil for 3 h at 180 ◦ C during three successive
days. Characteristics of the soil used are: pH: 8.60, EC: 0.19 dS m− 1,
NTK: 0.11%, TOC: 0.59%, OM: 1.02%, Polsen: 11.00 ppm, Na+: 990.00
ppm, K+: 890 ppm. Plants were watered regularly with the same amount
of water for a cultivation period of seven months.
The mycorrhizal consortium used was isolated from the rhizosphere
of O. ficus-indica plants in the Souihla Commune located 4 km North of
Marrakesh, Morocco. The AMF consortium contains 1034 spores/100 g
of soil distributed into 22 species. The species represent seven genera:
Glomus, Acaulosporam, Rhizophagus, Scutellospora, Diversispora, Clar­
oideoglomus and Gigaspora (Lahbouki et al., 2021). The mycorrhizal
consortium was multiplied by using Zea mays L. as a host plant under
controlled greenhouse conditions at the Faculty of Science Semlalia,
Cadi Ayyad University, Marrakesh as described by Meddich et al.
(2015). The infection process was performed by adding 10 g of colonized
roots to the soil around the cactus plant roots.
The organic amendment used in our experiment is a vermicompost
consisting of horse manure and wheat straw mixture vermicomposted by
earthworms (Eisenia fetida). The plants were supplied with autoclaved
vermicompost during three consecutive days at a rate of 5%. Physico-
chemical properties of the selected vermicompost are: pH: 7.70, EC:
4.89 dS m− 1, NTK: 0.62%, TOC: 28.00%, OM: 43.80%, Polsen: 290.00
ppm, Na+: 1940.00 ppm, K+:4960 ppm.
The experiment was conducted in a factorial arrangement based on a
completely randomized design with ten replicates per treatment of two
factors (biostimulants and water regimes). After four months of culti­
vation, drought stress was applied in the following manner: i) severe
water deficit (SWD): 15% of field capacity (FC), ii) moderately watered
(MD): 30% of FC, and iii) well-watered (WW): 50% of FC. The meth­
odology for water stress application is described by Meddich et al.
(2015). Each group composed of four biostimulants treatments:
1) T: Control treatment without any biofertilizer application
2) AMF: Plants inoculated with AMF consortium
3) Vc: Plants amended with vermicompost
4) AMF + Vc: Plants inoculated with AMF consortium and amended
with vermicompost
The data of mother and daughter cladodes for each treatment were
collected once a month during four months of the experiment for
growth, biochemical, and physiological analyses, performing cladodes
cores using a cork borer (10-mm diameter).
2.2. Root colonization, plant growth and water status
Plants were carefully harvested to avoid root damage or loss. The
root system was carefully washed with water to remove the adhered soil.
A fraction of the roots from the lateral root system was cleared with KOH
(10%) for 30 min, and subsequently stained with Trypan blue (Phillips
and Hayman, 1970). Stained root segments of 1 cm were examined
under a Zeiss Axioskop 40 microscope at 40–100 × magnification. The
frequency and intensity of mycorrhizal colonization of roots was
determined by the technique described by Trouvelot et al. (1986), and
was calculated as follows:
infected ​ root ​ segments ​
AMF ​ infection ​ frequency ​ (Fa)(%) = × 100
total ​ root ​ segments
(95n5 + 70n4 + 30n3 + 5n2 + ​ n1)
AMF ​ infection ​ intensity ​ (Ma)(%) =
total ​ root ​ segments
S. Lahbouki et al.
Where “n5” indicates to the number of roots with infection level of 5
(infection rate 90–100%). “n4” with infection level of 4 (infection rate
50–90%). “n3” with level 3 (infection rate at 10–50%). “n2” with level 2
(infection rate 1–10%). “n1” with level 1 (infection rate 0–1%).
Growth rate of cladodes was determined manually, using a
measuring tape to evaluate the length and width of each daughter
cladode and the number of new cladodes was recorded. The surface area
of the cladodes was determined by measuring the length (L) and width
(W), and the surface area was calculated using the following formula:
(width/2) (length/2) × π (Lahbouki et al., 2021).
To evaluate plant water status, several parameters were measured on
the cladodes for each treatment. Cladodes relative water content (RWC)
was estimated according to the method described by Barrs and Weath­
erley (1962). Cladodes cores were collected with the cork borer to
determine fresh weight (FW), dry weight (DW), and turgid weight (TW).
RWC was calculated according to the following formula:
(FW–DW)/(TW–DW) × 100. The thickness of the cladodes was
measured by a digital caliper for an additional information on the state
of the plant water.
2.3. Biochemical parameters
The concentration of total chlorophyll was determined according to
the method described by Arnon (1949). Photosynthetic pigments were
extracted from cladodes samples using acetone (80%). The extracted
material was centrifuged at 10,000×g for 10 min and the supernatant
absorbance was recorded at 645 and 663 nm using an UV visible spec­
trophotometer (UV–3100P).
To assess the total soluble sugar (TSS), frozen cladodes samples were
ground to a fine powder in liquid nitrogen. The total soluble sugar
content was determined by phenol-sulfuric acid method according to the
technique described by Dubois et al. (1956). Fine powder (0.15 g) of
each sample was homogenized in 4 mL of ethanol (80%). The super­
natant was collected and mixed with 0.25 mL of phenol (5%) and 1.25
mL of concentrated sulfuric acid. After 10 min, the TSS content was
determined by measuring the absorbance at 485 nm using
spectrophotometer.
Proline content was assessed by the method of Carillo et al. (2008).
Samples of each fresh cladode (100 mg) were homogenized in 4 mL of
ethanol (40%). Afterwards the homogenate was stored overnight at 4 ◦ C.
Solution of 0.5 mL was put into reaction with 1 mL of a solution con­
taining acetic acid (60%), ninhydrin (1%), and ethanol (20%) in a test
tube for 20 min at 90 ◦ C, followed by stopping the reaction by immersing
the tubes in an ice bath. The absorbance was read at 520 nm.
Hydrogen peroxide (H2O2) was determined as described by Velikova
et al. (2000). Briefly, fresh cladodes samples (0.25 g) were homogenized
in 5 mL of trichloroacetic acid (TCA) 10% (w/v), and centrifuged at 15,
000×g for 10 min. The supernatant (0.5 mL) was then mixed with 0.5 mL
of potassium phosphate buffer (10 mM, pH 7) and 1 mL of iodic potas­
sium (1 M). The absorbance values were taken at 390 nm after 1 h of
incubation in the dark.
The degree of lipid peroxidation was estimated by measuring the
content of malondialdehyde (MDA) as described by Rao and Sresty
(2000). Fresh frozen cladodes (0.25 g) were homogenized in 10 mL of
TCA 0.1% (w/v) and centrifuged at 18,000×g for 10 min. Two mL of
aliquot was mixed with 2 mL of TCA solution (20%) containing of thi­
obarbituric acid 0.5% (w/v). The mixture was heated at 100 ◦ C for 30
min, cooled in ice, and centrifuged at 10,000×g for 10 min. The
absorbance was measured at 532 nm. The content of MDA was expressed
as nmol MDA g− 1 of dry weight (DW).
To determinate the protein and antioxidant enzymes activities, the
cladodes powder samples (0.1 g) were homogenized in 5 mL solution
containing potassium phosphate buffer (0.1 M, pH 7.0), 0.1 g poly­
vinylpolypyrrolidone (PVPP), and ethylenediaminetetraacetic acid
(EDTA) 0.1 mM. The homogenate was centrifuged at 18,000×g, for 10
min at 4 ◦ C, and the extract was stored at − 20 ◦ C. The protein
3
Journal of Arid Environments 199 (2022) 104703
concentration was determined according to the method of Bradford
(1976) with bovine serum albumin as a standard. The absorbance was
taken at 595 nm.
Superoxide dismutase (SOD) activity was determined using the
method described by Beyer and Fridovich (1987). The reaction mixture
included 3 mL of sodium phosphate buffer (0.1 mM, pH 6), 75 μL of
methionine (55 mM), 300 μL of nitro-blue tetrazolium (0.75 mM) and
60 μL of riboflavin (0.1 mM). Each sample extract (100 μL) was mixed
with the prepared mixture. The absorbance was taken at 560 nm. SOD
activity was expressed in unit mg− 1 protein min− 1.
Peroxidase (POX) activity was determined using the method
described by Hori et al. (1997) based in following the change of ab­
sorption at 470 nm due to the oxidation of guaiacol. The activity was
measured for 1 min in an assay mixture (1 mL) containing phosphate
buffer (100 mM, pH 7.8), guaïacol 20 mM, H2O2 40 mM and 100 μL of
enzyme extract. POX activity was expressed in μmol guaïacol mg− 1
protein min− 1.
Polyphenol oxidase (PPO) activity was determined according to
Moore and Flurkey (1990). The reaction mixture contained catechol 20
mM in phosphate buffer (0.1 M, pH 7.8) and 100 μL of vegetal extract.
Activity was recorded as the decrease in absorbance at 420 nm and
expressed in μmol mg− 1 protein min− 1.
The extraction of total phenolic content (TPC) was performed on
finely powdered cladodes samples dried in oven at 75 ◦ C for 72 h. Five
grams of plants extract was immersed in 50 mL of methane (80%), and
shaken with an electric stirrer for 48 h. Thereafter, the obtained mixture
was filtered using a Buckner funnel and a Whatman No. 1 filter paper.
Total phenol content was determined spectrophotometrically by
recording the absorbance at 760 nm using Folin-Ciocalteu test
(Singleton and Rossi, 1965). Methanol extract 250 μL was reacted with
2.5 mL of distilled water, mixed with 250 μL of Folin-Ciocalteau reagent
in a test tube and incubated at room temperature for 3 min. Sodium
carbonate (Na2CO3) 250 μL was added and kept reacting for 90 min. TPC
was expressed as gallic acid equivalent per g of dry weight (mg GAE g− 1
DW).
2.4. Statistical analysis
The effects of drought treatments, AMF, and vermicompost was
examined in terms of variability based on average values of five repli­
cates ± standard error (S.E.) per treatment. Analysis of variance
(ANOVA) was carried out on drought treatments, with applied AMF,
vermicompost, and their combination to assess their effects using SPSS
23.0 software for Windows. The comparison between mean values in
each factor was estimated using Tukey’s test separately on a monthly
basis (4 months). Treatments’ effects were considered significant at P ≤
0.05.
3. Results
3.1. Effect of drought on plants AMF colonization, growth parameters and
water status
At the harvest, drought was revealed to negatively affect AMF
colonization (Table 1). In the absence of vermicompost, the values of the
AMF frequency and intensity of inoculated plants were 36 and 24%,
respectively under WW compared to SWD. The application of vermi­
compost increased these values under SWD (7 and 14%, respectively)
compared to AMF treatment. Concerning the production of new clad­
odes, a slight increase in the number of newly formed cladodes under
SWD in plants treated with vermicompost was recorded compared to
control treatment. (Table 1).
After the second month a significant decrease (P < 0.05) in the
surface areas of newly developed cladodes was observed (Fig. 1). Under
SWD, cladodes area was reduced in control treatments by 28%
compared to WW. However, the application of AMF, Vc or both
S. Lahbouki et al. Journal of Arid Environments 199 (2022) 104703

Table 1 3.2. Effect of drought on biochemical parameters

Effects of AMF and/or vermicompost on AMF infection frequency and intensity
and the number of newly formed cactus cladodes. The variation of total chlorophyll concentration in response to
Treatments Water AMF infection AMF infection Number of drought stress and biostimulants application is shown in Table 2. Under
regime frequency % intensity % cladodes SWD, this photosynthetic pigment significantly decreased after the third
T 15% 0.0 (0.0)d 0.0 (0.0)f 1.2 (0.7)f month of drought stress application in mother cladodes. However, the
30% 0.0 (0.0)d 0.0 (0.0)f 2.2 (0.4)b-f application of AMF and/or vermicompost increased the total chloro­
50% 0.0 (0.0)d 0.0 (0.0)f 2.0 (0.9)c-f phyll concentrations in mother and daughter cladodes compared to the
AMF 15% 62.2 (9.1)c 51.3 (3.5)e 1.3 (0.5)e-f
control plants under SWD.
30% 76.7 (3.6)ab 58.3 (4.4)cd 2.5 (0.55)a-e
50% 84.5 (6.5)a 63.4 (3.2)bc 2.2 (0.75)b-f Total soluble sugar and protein contents in cactus cladodes in
Vc 15% 0.00 (0.0)d 0.0 (0.0)f 1.7 (0.52)d-f response to drought and biostimulants applications are shown in Figs. 3
30% 0.00 (0.0)d 0.0 (0.0)f 2.8 (0.75)a-d and 4. During the drought stress application, TSS slightly increased in
50% 0.00 (0.0)d 0.0 (0.0)f 3.3 (0.52)ab daughter and mother cladodes under all water regimes. In addition,
AMF þ Vc 15% 66.7 (10.3)bc 58.4 (6.2)cd 1.5 (0.55)e-f
30% 81.2 (9.7)a 64.6 (4.4)ab 3.2 (0.75)a-c
biostimulants application further improved total soluble sugar content
50% 87.8 (8.9)a 69.8 (4.1)a 3.5 (0.84)a under SWD regime. However, the most important enhancement was
observed in mother cladodes treated by the dual application of AMF and
T: Control treatment; AMF: Plants inoculated with AMF consortium; Vc: Plants
Vc compared to the control under SWD regime (Fig. 3A). It was, 26, 32,
amended with vermicompost; AMF + Vc: Plants inoculated with AMF con­
sortium and amended with vermicompost. Data represent the means ± standard
25 and 28% in the first, second, third and fourth months, respectively.
error (SE) (n = 5). Means in the same column with different letters indicate In contrast, the protein content in mother cladodes decreased over­
significant differences at P ≤ 0.05. time under SWD (Fig. 4A). However, it showed an increase in daughter
cladodes (Fig. 4B). Moreover, the application of AMF and/or vermi­
significantly showed positive effects via increasing cladodes area by 46, compost had shown a significant positive increase (P < 0.05) on protein
57 and 64%, respectively compared to the control plants. The same content in mother and daughter cladodes compared to the control
trend was observed after the third and fourth months. plants.
One-month post drought application, no change was observed in The exposure of cactus plants to SWD regime resulted in an increase
RWC in mother and daughter cladodes in all treatments (Fig. 2A–B). in MDA and H2O2 contents in mother and daughter cladodes overtime
However, after the second month, a decrease in RWC was observed in particularly in non-inoculated and non-amended plants (Table 3).
mother cladodes under SWD compared to MD and WW regimes However, the application of AMF and/or vermicompost caused a sig­
(Fig. 2A), while daughter cladodes showed no significant change over nificant reduction in MDA and H2O2 contents under SWD, especially
the same period under all water treatments. However, a slight decrease after the third month. Moreover, the lowest MDA contents 38 and 34%
was recorded in RWC for daughter cladodes under SWD compared to were found in the mother and daughter cladodes respectively treated
WW after the third month of drought application (Fig. 2B). The appli­ with AMF + Vc compared to the control plants under SWD regime. As for
cation of AMF, vermicompost and their combination led to an increase H2O2 content, the lowest value was recorded in daughter cladodes
in RWC of mother and daughter cladodes under SWD compared to the treated by AMF + Vc treatment with a decrease of 34% compared to the
control during the third and fourth months. non-inoculated and non-amended plants (Table 3).
Additionally, a significant difference between mother and daughter Plants mother and daughter cladodes showed a significant increase
cladodes under SWD regarding thickness was recorded (Fig. S1). In fact, (P < 0.05) in antioxidant enzymes activities (SOD, POX and PPO) with
a significant decrease was observed during the four months of stress time, particularly in SWD regime (Table 4 and Fig. S2). Under SWD, the
application in mother cladodes under SWD, regardless of the bio­ applied biostimulants generally increased SOD, POX and PPO activities
stimulants application. In contrast, a thickness enhancement in daughter in mother and daughter cladodes compared to the control. After four
cladodes was recorded under SWD regime overtime. Furthermore, AMF months of drought stress, the highest values were recorded in AMF + Vc
inoculation and/or vermicompost supplementation improved this treatment in mother and daughter cladodes for SOD (37 and 41%), POX
parameter under SWD. (74 and 70%) and PPO (53 and 28%), respectively. Interestingly, the

Fig. 1. Kinetic of daughter cladodes area under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) in control plants, plants inoculated with
AMF consortium and/or amended with vermicompost, during four months of drought stress application. Values with different letters indicate significant differences
between treatments at p ≤ 0.05 by the Tukey’s test. Data represent the means ± standard error (SE) (n = 5).

4
S. Lahbouki et al. Journal of Arid Environments 199 (2022) 104703

Fig. 2. Kinetic of relative water content of mother (A) and daughter (B) cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) in
control plants, plants inoculated with AMF consortium and/or amended with vermicompost, during four months of drought stress application. Values with different
letters indicate significant differences between treatments at p ≤ 0.05 by the Tukey’s test. Data represent the means ± standard error (SE) (n = 5).

Table 2
Kinetic of total chlorophyll content of mother and daughter cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) during four
months of drought stress application.
Months Treatemts Mother Daughter

15% 30% 50% 15% 30% 50%

First month T 12.0 (0.6)de 12.7 (0.2)cd 12.8 (0.2)cd 10.2 (0.8)f 10.5 (0.2)ef 11.1 (0.3)d-f
AMF 12.3 (0.2)de 13.3 (0.1)c 13.3 (0.2)c 11.1 (0.1)ef 11.9 (0.2)c-e 12.3 (0.6)c-e
Vc 14.2 (0.2)b 14.5 (0.1)ab 14.5 (0.2)ab 12.4 (0.5)b-d 13.4 (0.2)a-c 13.4 (0.7)ab
AMF + Vc 14.4 (0.1)ab 15.0 (0.2)a 14.8 (0.1)ab 12.9 (0.8)a-c 13.9 (0.8)ab 14.1 (0.7)a
Second month T 11.2 (0.2)f 12.5 (0.2)e 12.6 (0.2)e 10.8 (0.3)f 11.3 (0.6)f 11.8 (0.3)ef
AMF 12.2 (0.2)e 13.1 (0.1)d 13.1 (0.1)d 11.9 (0.1)ef 12.6 (0.4)de 12.7 (0.3)c-e
Vc 13.7 (0.1)c 14.2 (0.1)bc 13.8 (0.1)bc 13.4 (0.7)b-d 13.5 (0.1)b-d 14.1 (0.4)b
AMF + Vc 14.0 (0.2)bc 15.0 (0.3)a 14.3 (0.1)b 13.9 (0.4)bc 14.1 (0.3)b 14.2 (0.3)a
Third month T 8.0 (0.5)g 11.9 (0.2)cd 12.3 (0.3)cd 11.0 (0.3)e 13.3 (0.7)d 13.4 (0.2)cd
AMF 9.7 (0.2)f 12.7 (0.1)b 12.9 (0.1)b 12.3 (0.4)d 15.4 (0.9)b 15.0 (0.7)b
Vc 10.7 (0.1)e 13.9 (0.2)a 13.9 (0.1)a 13.9 (0.6)bc 17.4 (0.2)a 17.3 (0.5)a
AMF + Vc 11.1 (0.2)de 14.1 (0.7)a 14.0 (0.1)a 14.2 (0.3)bc 18.0 (0.1)a 18.0 (0.7)a
Fourth month T 7.4 (0.3)h 11.5 (0.1)e 11.8 (0.1)de 11.5 (0.2)e 14.3 (0.1)c 14.3 (0.5)c
AMF 9.3 (0.3)g 12.4 (0.1)cd 12.7 (0.2)b-d 12.7 (0.2)d 15.9 (0.9)b 15.4 (0.3)b
Vc 10.2 (0.3)fg 13.4 (0.1)ab 13.3 (0.4)a-c 14.4 (0.6)bc 18.2 (0.7)a 18.3 (0.7)a
AMF + Vc 10.3 (0.6)f 13.8 (0.5)a 13.7 (0.2)a 14.9 (0.6)bc 18.9 (0.6)a 18.9 (0.5)a

T: Control treatment; AMF: Plants inoculated with AMF consortium; Vc: Plants amended with vermicompost; AMF + Vc: Plants inoculated with AMF consortium and
amended with vermicompost. Data represent the means ± standard error (SE) (n = 5). Means in the same column with different letters indicate significant differences
at P ≤ 0.05.

tested antioxidant activities were higher in mother cladodes than in plants.

daughter cladodes. In addition, no significant difference was recorded
under MD and WW in all treatments.
The results presented in Table S1 indicate that cactus plant exposure
to drought showed a significant increase in proline and TPC content.
Under SWD, an increase in proline and TPC contents was observed in
both mother and daughter cladodes, especially after the third month. In
addition, the application of AMF, Vc and AMF + Vc led to an increase in
both parameters compared to the control plants. However, the highest
values of proline and TPC content were recorded after the fourth month
in mother cladodes under SWD regime and treated with AMF + Vc, with
increases of 75 and 63% respectively, in comparison to the control
4. Discussion
Drought is one of the major stress factors limiting the plant growth
performance and production. Therefore, it is important to find
environmental-friendly techniques that can help plants to alleviate
drought stress effect. To our knowledge, this is the first study that
showed that AMF and/or vermicompost are crucial in boosting cactus
fitness and health under drought stress. Here, the focus was on growth
and biochemical parameters.
Our results showed that no AMF structures were observed in roots of

5
S. Lahbouki et al. Journal of Arid Environments 199 (2022) 104703

Fig. 3. Kinetic of total soluble sugar of mother (A) and daughter (B) cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) in
control plants, plants inoculated with AMF consortium and/or amended with vermicompost, during four months of drought stress application. Values with different
letters indicate significant differences between treatments at p ≤ 0.05 by the Tukey’s test. Data represent the means ± standard error (SE) (n = 5).

Fig. 4. Kinetic of protein content of mother (A) and daughter (B) cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) in control
plants, plants inoculated with AMF consortium and/or amended with vermicompost, during four months of drought stress application. Values with different letters
indicate significant differences between treatments at p ≤ 0.05 by the Tukey’s test. Data represent the means ± standard error (SE) (n = 5).

the non-inoculated cactus plants. Under drought, mycorrhizal root significantly increased root mycorrhizal colonization compared to single
colonization was negatively affected. This is in line with several studies AMF inoculation. Reduction of root colonization could be due to
conducted on maize Begum et al. (2019) and trifoliate orange (Cheng reduced development of AMF hyphae in the water-stressed soil (Cheng
et al., 2021a) subjected to drought stress. However, AMF + Vc treatment et al., 2021b). It is possible that a prolonged duration of drought stress or

6
S. Lahbouki et al. Journal of Arid Environments 199 (2022) 104703

Table 3
Kinetic of MDA and H2O2 content of mother and daughter cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) during four
months of drought stress application.
1 1
Months Treatemts MDA (nmol g− DW) H2O2 (nmol g− DW)

Mother Daughter Mother Daughter

15% 30% 50% 15% 30% 50% 15% 30% 50% 15% 30% 50%

First T 16.3 16.1 15.7 9.4 9.7 8.6 15.0 13.8 13.3 9.5 9.2 8.6
month (1.4)a (1.6)ab (2.2)a-c (1.7)a (1.6)a (1.4)a (1.1)a (0.9)ab (0.4)a-c (0.7)a (1.0)ab (0.8)a-c
AMF 13.3 13.5 13.5 8.1 8.6 8.2 13.4 11.3 11.3 8.1 8.4 8.1
(2.2)b-d (1.2)a-d (1.2)a-d (0.7)a (2.1)a (0.7)a (0.7)a-c (0.6)c (0.8)c (0.6)a-c (0.3)a-c (0.5)a-c
Vc 12.7 11.5 11.3 7.4 7.7 7.6 14.3 13.5 13.3 7.3 7.4 8.0
(2.6)b-d (0.6)b-d (1.3)cd (1.2)a (1.2)a (1.3)a (0.6)ab (1.0)a-c (1.2)a-c (0.8)bc (0.9)bc (0.3)a-c
AMF + Vc 9.3 10.5 9.0 7.2 7.1 7.4 13.5 12.8 12.3 6.8 7.1 7.4
(1.2)d (1.7)d (1.4)d (1.2)a (1.4)a (1.2)a (0.5)a-c (0.7)a-c (0.6)bc (0.6)c (0.6)c (0.7)bc
Second T 18.4 16.1 16.6 11.5 10.3 9.2 16.1 15.3 14.6 11.2 10.2 9.2
month (1.2)a (0.5)ab (0.6)ab (0.7)a (0.9)ab (0.6)ab (1.3)a (1.0)ab (0.5)a-c (0.6)a (0.8)ab (0.7)bc
AMF 14.5 12.3 13.5 9.4 9.8 8.6 15.3 13.1 12.6 9.2 9.3 8.5
(1.4)a-c (0.9)bc (1.2)bc (1.2)ab (1.7)ab (1.2)ab (0.9)ab (0.6)bc (1.1)c (0.5)bc (0.6)bc (0.9)bc
Vc 13.5 12.3 10.8 8.9 8.2 8.2 15.9 14.5 14.0 8.7 8.4 8.8
(1.4)bc (2.5)bc (1.7)c (1.1)ab (1.4)b (1.7)b (0.2)a (0.8)a-c (0.9)a-c (0.6)bc (0.6)bc (0.4)bc
AMF + Vc 11.5 11.7 10.0 8.0 7.7 8.0 15.5 14.2 13.2 7.8 7.8 8.0
(1.9)c (0.8)c (1.0)c (0.6)b (0.5)b (0.5)b (0.44)ab (0.7)a-c (0.8)bc (0.5)c (0.7)c (0.8)c
Third T 22.5 16.7 16.6 16.6 11.0 9.8 22.0 16.4 16.3 16.1 11.0 9.5
month (1.9)a (0.8)bd (1.2)b-d (0.7)a (1.7)bc (1.2)b-d (1.3)a (1.0)cd (0.5)cd (0.8)a (0.4)c-e (0.7)e-g
AMF 19.4 12.6 12.4 13.6 10.3 9.2 18.5 14.5 13.3 13.8 10.3 9.4
(1.0)b (1.2)d-f (1.3)ef (1.7)b (1.2)b-d (2.0)b-d (0.8)bc (0.8)d-f (0.5)f (0.7)b (0.4)c-f (0.5)e-g
Vc 17.9 11.8 11.5 12.0 9.3 8.6 19.0 16.1 14.8 11.8 9.9 9.0
(1.1)bc (1.5)f (1.9)f (3.2)bc (0.9)b-d (0.7)bc (0.8)b (1.0)c-e (1.2)d-f (0.8)c (0.4)d-g (0.6)fg
AMF + Vc 13.9 11.0 10.4 11.0 8.7 8.4 19.3 15.3 13.8 11.3 8.4 8.8
(1.4)c-f (1.5)f (2.3)f (2.9)bc (0.5)b-d (1.5)d (0.6)b (0.7)d-f (0.8)ef (0.7)cd (0.7)g (0.4)fg
Fourth T 25.6 15.2 15.2 21.7 11.5 10.3 25.3 17.3 16.7 21.7 11.8 10.4
month (1.1)a (0.7)c-e (2.1)c-e (0.4)a (0.9)d (1.2)d (0.8)a (0.5)c (0.6)c (1.2)a (0.4)d (0.6)de
AMF 21.5 12.2 12.2 19.6 11.0 9.4 21.9 15.0 13.8 19.2 10.8 9.6
(1.1)b (2.4)de (2.7)de (1.6)b (1.2)d (1.3)d (1.2)b (0.5)cd (0.8)d (0.6)b (1.5)de (0.6)de
Vc 19.3 11.6 11.0 16.8 10.2 8.9 23.1 15.9 15.4 16.7 10.2 9.3
(1.9)bc (1.3)de (1.3)e (1.9)bc (1.3)d (0.6)d (0.9)b (0.9)cd (1.2)d (0.4)c (0.7)de (1.1)e
AMF + Vc 16.9 10.7 10.3 15.5 9.4 8.6 22.3 15.5 14.2 15.5 9.6 8.9
(1.6)b-d (0.7)e (1.7)e (2.6)bc (0.7)d (0.5)d (0.7)b (0.7)cd (0.7)d (0.5)c (0.5)de (0.3)e

T: Control treatment; AMF: Plants inoculated with AMF consortium; Vc: Plants amended with vermicompost; AMF + Vc: Plants inoculated with AMF consortium and
amended with vermicompost. Data represent the means ± standard error (SE) (n = 5). Means in the same column with different letters indicate significant differences
at P ≤ 0.05.

pre-adaptation properties of AMF are involved in this reduction (Quir­
oga et al., 2020). Furthermore, drought stress reduced supply of car­
bohydrate by host plants leading to limitation of spore germination
(Cheng et al., 2021a).
The increase in mycorrhizal colonization by vermicompost applica­
tion may be due to several factors such as organic amendments
decreasing soil pH, which may further promote AMF colonization
(Demir, 2019). The formation of AMF structures is largely influenced by
soil management techniques. In fact, the increase in AMF colonization
may be related to the enhancement of soil characteristics by the ver­
micompost application (Demir, 2019). However, other studies reported
that the dual application of AMF and organic amendment could have an
inhibition effect on AMF colonization (Fernández-Gómez et al., 2012;
Toubali et al., 2020).
Conversely, the application of AMF and/or vermicompost can
compensate the effects of drought, mainly by improving the surface area
of new cladodes. Most researchers have indicated that cactus plants can
withstand two to three months of severe water stress due to water
retention in the cladodes, but after this period, many morphological and
physiological manifestations can begin to appear (Scalisi et al., 2016).
The beneficial effect of AMF on growth of cactus under water deficit
might be linked with greater uptake of water and essential nutrients
through fungi filaments (Begum et al., 2019; Fattahi et al., 2021). In
addition, the physical and biological characteristics of organic amend­
ments such as high porosity, aeration, drainage, and water retention
capacity increase nutrient uptake, which can help enhance plants
growth under drought stress (Demir, 2019; Rashtbari et al., 2020).
In this study, the production of a high number of daughter cladodes
and their development in dry conditions causes numerous physiological
negative effects like a decrease of RWC and photosynthetic pigments.
Our findings corroborate the results of Pimienta-Barrios et al. (2005)
who reported that the presence of high number of daughter cladodes
increases water loss, which affects the RWC in mother cladodes. The
combined effect of drought stress and new cladodes development may
reduce photosynthetic ability in the mother cladodes. Indeed, Pimien­
ta-Barrios et al. (2007) reported that under drought stress conditions,
cactus plant tends to store water in newly formed cladodes, probably as
a survival strategy and to provide more water to the most photosyn­
thetically active organs (daughter cladodes). In addition, Scalisi et al.
(2016) indicated that CO2 uptake and malic acid accumulation are
influenced by drought stress. This can affect photophosphorylation,
presumably leading to insufficient supply of ATP which consequently
affects growth (Lawlor and Cornic, 2002). Interestingly, we found that
the biostimulants application significantly increased RWC and chloro­
phyll content under drought stress as compared to plants grown in the
absence of the organic amendment and AMF inoculation. Previous
studies reported that mycorrhizae regulates plant water status by
increasing hydraulic conductivity through the expression and/or activ­
ity of plant root aquaporin (Symanczik et al., 2020; Cheng et al., 2021b).
Furthermore, enhancing water holding capacity is among the positive
effects of organic amendment application, which help improve plant
water uptake (Rashtbari et al., 2020). In addition, AMF and/or organic
amendment application enhance nutrients uptake especially magnesium
(Mg), an essential element in chlorophyll synthesis (Olayiwola and
Abiodun, 2019).
Our data showed that under drought stress, AMF and/or

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S. Lahbouki et al. Journal of Arid Environments 199 (2022) 104703

Table 4
Kinetic of SOD and POX activities of mother and daughter cladodes under different water regimes (15% field capacity (FC), 30% (FC) and 50% (FC)) during four
months of drought stress application.
1
Months Treatemts SOD (unit mg− protein min− 1) POX (μmol of guaïacol mg− 1
protein min− 1)

Mother Daughter Mother Daughter

15% 30% 50% 15% 30% 50% 15% 30% 50% 15% 30% 50%

First T 1.0 1.1 1.1 1.0 0.9 0.8 0.2 0.3 0.3 0.1 0.2 0.1
month (0.1)c (0.1)c (0.1)c (0.1)a-c (0.1)bc (0.1)c (0.0)b (0.1)ab (0.1)ab (0.0)c (0.1)c (0.0)c
AMF 1.4 1.5 1.5 1.3 1.2 1.0 0.3 0.3 0.3 0.1 0.2 0.1
(0.1)b (0.1)b (0.1)ab (0.2)ab (0.1)a-c (0.1)a-c (0.0)ab (0.1)ab (0.0)ab (0.0)c (0.1)c (0.1)ab
Vc 1.7 1.79 1.7 1.4 1.4 1.2 0.3 0.1 0.3 0.1 0.2 0.2
(0.2)ab (0.1)a (0.1)ab (0.1)a (0.1)a (0.2)a-c (0.0)ab (0.0)ab (0.0)ab (0.0)c (0.0)c (0.0)a
AMF + Vc 1.6 1.7 1.6 1.4 1.2 1.1 0.3 0.3 0.3 0.1 0.2 0.2
(0.1)ab (0.2)ab (0.1)ab (0.2)ab (0.1)ab (0.1)a-c (0.0)ab (0.1)ab (0.0)a (0.0)c (0.0)bc (0.1)a
Second T 1.3 1.2 1.2 1.2 0.9 0.9 0.3 0.3 0.3 0.2 0.2 0.2
month (0.1)c (0.3)c (0.2)c (0.1)a-c (0.1)c (0.1)c (0.1)c (0.0)c (0.0)c (0.1)d (0.1)d (0.0)a-d
AMF 1.8 2.3 2.0 1.4 1.2 1.0 0.3 0.4 0.4 0.2 0.2 0.2
(0.1)b (0.3)ab (0.2)ab (0.2)a-c (0.2)a-c (0.1)bc (0.0)bc (0.1)a-c (0.1)a-c (0.1)cd (0.0)cd (0.1)a-c
Vc 2.2 2.4 2.2 1.6 1.4 1.3 0.4 0.4 0.4 0.2 0.3 0.2
(0.2)ab (0.2)a (0.1)ab (0.1)a (0.1)a-c (0.1)a-c (0.0)a-c (0.1)a-c (0.1)a-c (0.0)d (0.1)b-d (0.0)ab
AMF + Vc 2.1 2.1 2.2 1.5 1.3 1.2 0.5 0.6 0.5 0.2 0.3 0.2
(0.2)ab (0.1)ab (0.2)ab (0.1)ab (0.2)a-c (0.1)a-c (0.1)a-c (0.1)a (0.1)ab (0.0)d (0.1)b-d (0.1)a
Third T 2.4 1.6 1.5 1.6 1.1 1.0 0.5 0.3 0.4 0.2 0.2 0.2
month (0.3)c (0.2)d (0.1)d (0.1)a-d (0.1)d-f (0.1)f (0.8)c-e (0.0)h (0.1)h (0.0)g (0.0)f (0.1)c-e
AMF 2.8 2.6 2.4 1.8 1.3 1.2 0.6 0.4 0.5 0.3 0.3 0.2
(0.1)b (0.2)bc (0.3)c (0.2)a-c (0.1)d-f (0.1)d-f (0.1)cd (0.1)de (0.0)de (0.0)e-g (0.0)e-g (0.1)b-d
Vc 3.3 2.8 2.6 1.9 1.4 1.4 0.7 0.5 0.5 0.4 0.3 0.2
(0.3)a (0.2)bc (0.2)bc (0.3)ab (0.1)b-f (0.1)d-f (0.0)b (0.1)d (0.0)de (0.0)c-f (0.0)d-g (0.0)b-c
AMF + Vc 3.0 2.5 2.4 2.0 1.4 1.2 0.8 0.6 0.6 0.5 0.3 0.3
(0.3)ab (0.2)bc (0.3)c (0.2)a (0.1)c-f (0.2)d-f (0.1)a (0.0)c (0.0)b (0.0)b-e (0.0)a (0.0)b
Fourth T 2.9 1.8 1.5 2.1 1.2 1.1 0.6 0.4 0.4 0.3 0.2 0.2
month (0.3)c (0.3)d (0.2)d (0.2)d (0.1)g (0.1)g (0.1)c (0.0)e (0.1)e (0.1)c (0.0)cd (0.1)c-e
AMF 3.5 2.7 2.5 2.45 1.4 1.3 0.8 0.5 0.5 0.4 0.3 0.3
(0.2)b (0.2)c (0.2)c (0.1)c (0.1)ef (0.01)f (0.1)b (0.0)d (0.0)d (0.0)ab (0.1)cd (0.0)cd
Vc 3.9 2.8 2.7 2.7 1.6 1.4 0.9 0.6 0.6 0.4 0.3 0.3
(0.2)a (0.2)c (0.2)c (0.1)ab (0.1)e (0.1)fg (0.1)b (0.1)cd (0.1)cd (0.1)ab (0.1)a-c (0.1)bc
AMF + Vc 3.9 2.7 2.4 2.9 1.5 1.3 1.1 0.6 0.6 0.5 0.4 0.3
(0.2)a (0.2)c (0.2)c (0.1)a (0.1)ef (0.1)fg (0.1)a (0.1)c (0.1)c (0.0)a (0.0)ab (0.0)bc

T: Control treatment; AMF: Plants inoculated with AMF consortium; Vc: Plants amended with vermicompost; AMF + Vc: Plants inoculated with AMF consortium and
amended with vermicompost. Data represent the means ± standard error (SE) (n = 5). Means in the same column with different letters indicate significant differences
at P ≤ 0.05.

vermicompost application increased the TSS and proline content, espe­
cially in mother cladodes, while protein content was enhanced in
daughter cladodes. This may be in order to maintain higher tissue hy­
dration and regulate osmolyte catabolism (sugar, proline and poly­
amins) playing a key role in maintaining physiological activities to
reduce the damage caused by ROS under drought stress (Zeighami Nejad
et al., 2020). Similar results were obtained in maize, and strawberry
plants treated with AMF and/or organic amendment (Begum et al.,
2019; Zuo et al., 2018). This is in accordance with studies, which re­
ported that sugar metabolism-related genes tend to be more expressed
within inoculated plants with AMF under drought stress (Ahanger et al.,
2014).
Under drought stress, cactus plants accumulated higher levels of
MDA and H2O2 at the end of the third month. These results prove the
ability of cactus plants to survive under severe drought conditions. This
is in agreement with previous studies (Scalisi et al., 2016; Campos et al.,
2021). Furthermore, mother cladodes accumulated higher levels of MDA
and H2O2 contents in comparison to daughter cladodes. This can be
explained by water transfer from mother to daughter cladodes. How­
ever, drought tolerance in plants is often associated with the improve­
ment of the antioxidant enzyme system (Chandra et al., 2021).
Biostimulants, such as AMF and/or organic amendment, play an
important role in improving antioxidant enzymes (i.e., SOD, POX and
PPO) as a defense system to scavenge the ROS (Zuo et al., 2018; Begum
et al., 2019). In this study, AMF and/or vermicompost induced SOD,
POX and PPO activities in mother and daughter cladodes under drought
stress. Nevertheless, more induction was observed in mother cladodes
especially, as it is the water source for daughter cladodes and also the
first to encounter drought stress. SOD is the first stage of plant defense
against ROS as it refines superoxide and converts it to H2O2 and O2
(Armand et al., 2016). The H2O2 produced by SOD is then refined
(Armand et al., 2016). H2O2 removing and sweeping is carried out by
POX and PPO enzymes (Rukmini et al., 2004). Thus, the damaging ef­
fects of ROS are reduced (Toubali et al., 2020). Similar results were
observed by Begum et al. (2019) and Zeighami Nejad et al. (2020), who
reported the effect of AMF and/or vermicompost on antioxidant activity
under drought stress.
In this study, results show that TPC has increased in plants under
SWD condition. These TPC are required to scavenge ROS and sustain the
membrane integrity against oxidative stress, either indirectly by
contributing to photo-protection or directly by participating as free
radical scavengers, quenching singlet oxygen, chelating metal ions or
providing a substrate for peroxidase enzymes (Luján-Hidalgo et al.,
2017; Chiappero et al., 2019). Furthermore, AMF and/or vermicompost
enhanced the TPC under drought. This increase might be attributed to
the improved uptake efficiency of macro- and micronutrients by AMF
through their hyphae at the root level (Ahanger et al., 2014), and the
contribution of vermicompost humic and fulvic acids to the generation
of phenolic and flavonoid compounds (Luján-Hidalgo et al., 2017).
5. Conclusion
Opuntia ficus-indica is a drought-tolerant plant species that is gener­
ally considered to be well adapted to arid and semi-arid regions.
Nevertheless, its growth, development, and biochemical responses can
drastically be affected by drought stress. In agriculture, the use of eco-

8
S. Lahbouki et al.
friendly biostimulants like AMF and vermicompost can significantly
improve the plants performance under drought conditions. Our results
suggested that biostimulants can positively affect prickly pear plants
growth and development. Application of AMF and/or vermicompost
alleviated the negative effect of drought stress by improving surface
area, and physiological parameters such as RWC and total chlorophyll
and proline content. The applied biostimulants significantly reduced
H2O2 and MDA contents and improved the antioxidant enzymes activity
and phenol content. Also, application of vermicompost increased
mycorrhizal colonization. In the light of these findings, further research
needs to be carried out to deepen knowledge on cactus plants morpho-
physiological and biochemical mechanisms under drought stress.
CRediT authorship contribution statement
Soufiane Lahbouki: Investigation, Data curation, Methodology,
Software, Formal analysis, Writing – original draft, Writing – review &
editing. Raja Ben-Laouane: Methodology, Software, Writing – original
draft, Writing – review & editing. Mohamed Anli: Investigation, Data
curation, Methodology, Software, Formal analysis. Abderrahim Bou­
tasknit: Investigation, Data curation, Formal analysis. Youssef Ait-
Rahou: Methodology, Software. Mohamed Ait-El-Mokhtar: Method­
ology, Software. Soumaya El Gabardi: Methodology, Software. Allal
Douira: Methodology, Software. Said Wahbi: Conceptualization, Proj­
ect administration, Validation, Supervision, Project administration.
Abdelkader Outzourhit: Conceptualization, Project administration,
Validation, Supervision, Project administration. Abdelilah Meddich:
Conceptualization, Project administration, Validation, Supervision,
Project administration, Writing – review & editing.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgments
This work was supported by “Introducing cactus plantations (Opuntia
spp.) and smart water management systems in marginal lands of Egypt
and Morocco to drive rural renaissance in the Mediterranean Region”
project (ID: ERANETMED3-204).
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.jaridenv.2021.104703.
References
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