Available via license: CC BY 4.0
Content may be subject to copyright.
Page 1/21
Exploring root architecture and rhizosphere biology in
fourteen winter wheat varieties released in Chile from
1965 to 2020
Paula Paz-Vidal
Dalma Castillo-Rosales
María Dolores López
Iván Matus Tejos
Felipe Noriega
Maurico Schoebitz ( mschoebitz@udec.cl )
Universidad de Concepción: Universidad de Concepcion https://orcid.org/0000-0003-3953-8425
Research Article
Keywords: Andisol, Carboxylate, Arbuscular mycorrhizal, Triticum aestivum
Posted Date: February 21st, 2023
DOI: https://doi.org/10.21203/rs.3.rs-2595474/v1
License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full
License
Page 2/21
Abstract
Background and Aims. The study aims to explore the impact of advancements in wheat genetics on root structure
and rhizosphere biology, which are still not fully understood. Specically, we investigated various factors including
the exudation of carboxylates, colonization by arbuscular mycorrhizal fungi, microbial activity, and root architecture
in winter wheat varieties that have been released between 1965 and 2020.
Methods. To conduct our study, we sowed fourteen different winter wheat varieties with four replicates on acidic
Andisol at eld conditions. Complete root systems and soil samples were extracted using a tractor-mounted
hydraulic sampler tube of 3.5 cm diameter, which reached a depth of 60 cm.
Results. In this sense, succinate showed a signicant increase by 21%. Mycorrhizal colonization was inversely
proportional to P concentrations and all varieties showed higher microbial activity at anthesis. The longest roots
were found in varieties released after the year 2000, but no signicant differences were found in other root
architecture parameters. There was no clear pattern observed in root architecture or biological activity as a function
of the year of release. Plant genetics moderated root architecture, carboxylate exudation, microbial activity, and
mycorrhizal colonization, all of which are affected by high P concentrations.
Conclusions. This study investigated plant-microorganism interactions, often overlooked due to root system
analysis challenges. Older wheat varieties showed higher carboxylate exudation. We identied wheat varieties with
potential for improved root systems and crop eciency.
Introduction
Winter wheat (
Triticum aestivum
L.) is a vital cereal crop with widespread global signicance, playing a crucial role
in food security. In Chile, the genetic improvement of winter wheat started in 1959 at the Instituto de Investigaciones
Agropecuarias (INIA) Carillanca, with the creation of F1 hybrids that incorporated the dwarng genes of the Norin 10
cultivar (Del Pozo et al. 2021). This cereal is the most extensively planted type of wheat in the world (Qin et al.
2019).
In the pursuit of improving wheat production, various genetic modications have been made to the crop, including
selection of aerial organ traits and application of agrochemicals. This has resulted in the creation of new wheat
varieties with improved yield and productivity (McGrail and McNear 2021). However, little consideration has been
given to the impact of these advancements on root exudation and interactions with arbuscular mycorrhizal fungi
(AMF).
The release of modern wheat varieties has been accompanied by a decrease in root biomass over time (Aziz et al.
2017). This trend is likely due to the incorporation of dwarng genes in the 1960s (Matus et al. 2012; McGrail and
McNear 2021) and the application of inorganic fertilizers in excess of the plant's needs. These factors result in a
high availability of nutrients, so the plant does not need to invest resources in developing its root system to intercept
these nutrients (Jacquiod et al. 2022; McGrail and McNear 2021).
The relationship between plants and microorganisms is shaped by the variations in root morphology and traits
within the plant genotype, such as root length and axial development, as well as the characteristics of the
microorganisms themselves (Campos et al. 2018; de Souza Campos et al. 2021). Plant cultivars have functional
traits in their root systems that enable them to eciently obtain phosphorus (P) from the soil. These include root
exudation of carboxylates, colonization by AMF (Ober et al. 2021), and modications in root morphology, such as
Page 3/21
increased depth, density, and diameter. Deeper, denser, and larger diameter roots lead to higher yields due to their
greater eciency in nutrient acquisition (Severini et al. 2020; Wen et al. 2022). P is a nutrient that tends to bind to
soil clays, hindering its mobility and having to be absorbed by the roots through the diffusion mechanism (de Souza
Campos et al. 2021), where AMF colonization favors this process, occurring mainly under P-limited conditions since
high P application limits AMF community structure (Qin et al. 2020) and the action of some enzymes essential for P
mineralization (Liu et al. 2023). On the other hand, when P is scarce, plants increase the release of organic
compounds such as carboxylates to dissolve inorganic P in the soil (Pi). This can also be achieved indirectly by
signaling to the AMF through the distribution of photosynthates (Wen et al. 2022), This is further reinforced by the
work of enzymes such as acid phosphatase (P-ase), which play a role in the mineralization of organic phosphorus
(Po) (Liu et al. 2023; Nazareno et al. 2020). P can also be obtained from organic matter. However, in soil with low
organic P, inorganic sources are a signicant source of P. In Andisols, P is bound to aluminum and iron. Furthermore,
Chilean Andisols have high levels of total P, organic matter, and acidity. Andisol are the least extensive soil order
(Soil Survey Staff 2014) and occupy only 0.7% of the earth’s land surface or just below 963,000 km2. In Chile,
volcanic soils support the bulk of agricultural and forestry production, covering more than 5.3x106 hectares and
representing nearly 50–60% of the country’s arable land (Borie and Rubio 2003).
The study of carboxylates is becoming increasingly important due to their effectiveness in dissolving P. It has been
demonstrated that certain genetically modied plants are capable of increasing the release of organic anions in
environments with a shortage of P (Wang et al. 2013a). The eciency of P solubilization by carboxylates is given
by: citrate > oxalate > malate, being citrate and malate most studied in wheat (Wang et al. 2013c; Wang et al. 2017).
As a result, these exudates improve the uptake of nutrients that are otherwise dicult to dissolve and also stimulate
the growth and activity of the soil microorganisms. This, in turn, enhances root exploration and leads to better
nutrient absorption (Bünemann et al. 2018; Kuzyakov et al. 2000). However, this rhizosphere biological activity can
increase or decrease depending on the nutritional needs of crop phenological stages (Wang et al. 2017) which are
closely related to root development and growth (Dharmateja et al. 2021).
Research has shown that during the anthesis stage, there is a rise in the number of AMF in wheat roots. This is
because the leaves are growing rapidly and producing a high rate of photosynthesis, generating essential
carbohydrates for the nutrition of the mycorrhizae. On the other hand, during the physiological maturity stage, the
leaves start to age and photosynthesis decreases, leading to a decrease in AMF colonization (Naseer et al. 2022;
Zhu et al. 2017). The goal of this study was to compare and evaluate fourteen winter wheat varieties that were
released in Chile from 1965 to 2020, with a focus on their rhizospheric biological activity and root and aerial
development at the tillering, anthesis, and physiological maturity stage, under conventional eld conditions.
Materials And Methods
Site and soil description
This study was conducted during the 2021–2022 season at INIA Experimental Station Santa Rosa (36°31'S,
71°54’W), located in the Mediterranean climate region of central Chile. The soil belongs to the Diguillín series,
coming from modern volcanic ashes of the Andisol order, its texture is silt loam and the soil is classied as Typic
Haploxerands (Soil Survey Staff 2014; Stolpe 2006). The chemical soil properties (0–20 cm) are as follows: organic
matter: 5.8%; pH: 5.6; available N: 32 mg kg− 1; P-Olsen: 30 mg kg− 1; available K: 278 mg kg− 1.
Page 4/21
Wheat varieties
Fourteen winter wheats (
Triticum aestivum
L.) varieties released in Chile between 1965 and 2020 were selected. The
wheat varieties and their respective release years: Druchamp (1965), Melifen (1974), Manquefen (1977), Talafen
(1982), Laurel (1987), Lautaro (1990), Tukan (1993), Kumpa (2002), Bicentenario (2010), Maxwell (2012), Pionero
(2013), Rocky (2015), Kiron (2017), Chevignon (2020). Are considered modern cultivars because they were released
after 1960 (Matus et al. 2012) and the semi-dwarf genes (Rht1 and Rht2) are incorporated into their germplasm (Del
Pozo et al. 2021).
Experimental procedure
Experimental design was a complete block with four replicates. Plots consisted of nine rows of 2 m in length and
0.20 m distance between rows. Sowing date was may-june and the seed dose was 180 kg ha− 1, previously
disinfected with 250 mL of Real®Top (BASF; 166.6 g L− 1 thiophanate-methyl, 8.3 g L− 1 pyraclostrobin and 83.3 g
L− 1 triticonazole) and 120 mL of Punto 600 FS (ANASAC Chile; 600 g L− 1 imidacloprid) were applied per 100 kg of
seed. Fertilizer application consisted of 250 kg ha− 1 of triple superphosphate before planting and 230 kg ha− 1 of
urea (46% N), which was applied at the four-leaf stage, tillering, and rst node. At the tillering stage, 100 kg ha− 1 of
sulpomag (22% K, 18% Mg, and 22% S) and 100 kg ha− 1 of muriate of potash (60% K) were also applied. Weed
control was carried out in preemergence with the application of 1 L ha− 1 of Bacara Forte, Bayer (120 g L− 1
ufenacet, 120 g L− 1 urtamone, and 120 g L− 1 diufenican) and in postemergence with the application of 4 g ha− 1
of Ally, Dupont (600 g kg− 1 metsulfuron-methyl) and 4 kg ha− 1 of MCPA (750 g L− 1 MCPA-dimethylammonium). For
fungal disease control, 6.25 mL L− 1 of Juwell Top (BASF; 150 g L− 1 phenpropimorph, 125 g L− 1 kresoxim-methyl,
and 125 g L− 1 epoxiconazole) and 0.8 L ha− 1 of Priori (Syngenta; 250 g L− 1 azoxystrobin) were applied. Plots were
furrow irrigated at the tillering, ag leaf just visible, early heading, and medium milk stages.
Measurements
Samples from each variety were collected at three phenological stages of wheat: tillering, anthesis, and
physiological maturity (Zadoks et al. 1974). The measurements taken during the study included: basal soil
respiration (from soil adhered to roots), activity of FDAse and acid phosphatase enzyme (P-ase), root and aerial
biomass at tillering, anthesis, and physiological maturity stages. At anthesis, carboxylate exudation was measured
in complete root systems at two different soil depths (from 20 to 60 cm), root architecture (area, diameter, volume,
length, and weight), and the percentage of AMF colonization.
Soil samples were collected from the central rows of each plot using a tractor-mounted hydraulic sampler tube of
3.5 cm diameter, which reached a depth of 60 cm (Fig. S1). The samples were divided into three fractions based on
their depths: 0–20 cm, 20–40 cm, and 40–60 cm.
FDAse Activity
The activity of uorescein diacetate hydrolase (FDAse) was analyzed using the modied methodology by Alef and
Nannipieri (1995). This method is based on a colorimetric approach and involves the hydrolysis of FDAse. We used
0.5 g of soil, taking duplicates and leaving one control with 60% conditioning. The soil sample was added to falcon
Page 5/21
tubes along with 4.45 mL of sodium phosphate buffer (pH 7.8) and 50 µL of FDAse. A control was made by adding
5 mL of sodium phosphate buffer (pH 7.8) to the falcon tube. Afterwards, the tubes were placed in a
thermostatically controlled bath at 25°C for 1 h and then removed and placed in an ice bath to slow down the
reaction. Then, 5 mL of acetone was added to the samples and controls, they were mixed in a vortex mixer, and
ltered through a funnel with Whatman N° 40 paper. The ltrate was measured in a spectrophotometer at an
absorbance of 490 nm, with a reference to the reagent blank (consisting of 1 mL of distilled water and 1 mL of
acetone).
Basal Soil Respiration
The basal soil respiration was analyzed using the closed system soil incubation methodology (Alef and Nannipieri
1995). 25 g of 60% WPFS-conditioned soil was placed in an incubation ask with a test tube containing 7.5 mL of
NaOH. The control procedure containing only 7.5 mL of NaOH. The incubation asks were tightly sealed and placed
in an incubation chamber at 22°C for 2 days. After that, 1 mL (1000 µL) of NaOH was extracted and 2 mL of BaCl2
was added. Finally, the solution was titrated with HCl until it reached the endpoint at the equivalence point (when the
pH reached 8.3).
Acid Phosphatase Enzyme Activity
For the analysis of phosphatase activity, we used Tabatabai and Bremmer assay (1969): 2 mL of MUB buffer (12.2
g hydroxymethyl aminomethane + 11.6 g maleic acid + 14 g citric acid + 6.28 g boric acid + 488 mL NaOH 1
M
and
brought to 1 L distilled water) pH 6.5 and 0.5 mL p-nitrophenyl phosphate (PNP) were mixed and incubated at 37°C
for 1 h, then cooled in a bath with ice for 10 min to stop the reaction. Next, 0.5 mL of 0.5
M
calcium chloride and 2
mL of 0.5
M
sodium hydroxide were added to the samples. Parallel blanks were also prepared by adding PNP after
the addition of calcium chloride and sodium hydroxide. Both samples and controls were then centrifuged at 3400
rpm/8 min. The relevant dilutions were made and readings in a spectrophotometer at 398 nm. The standard curve
was performed with PNP. Once the absorbance was obtained, it was interpolated on the standard curve, obtaining
the µmol of PNP g− 1 soil h− 1.
Root And Aboveground Biomass
The aboveground biomass was determined by measuring the dry matter. The aboveground part of the plant was
harvested and dried in kraft paper bags in an oven set at 70°C/48 h until reaching a constant weight. The total
biomass was weighed (Li et al. 2017). For root analysis at different depths, roots were washed to remove adhering
soil (Qin et al. 2019). Root parameters such as length, area, volume, and diameter will be determined using the
WinRhizo computerized system (Regent instruments Inc, Quebec, Canada) considering the volume of the sample
extractor tube and centimeters of depth. Appendices Fig. S2 to Fig. S15 show exemplary images of roots of each
wheat variety taken with the WinRhizo program.
Carboxylate Exudation
Complete root systems were collected from the rst 20 cm of soil. The roots were washed and incubated in 50 mL
CaSO4 (0.2
mM
) at pH 5.5 and shaken on an orbital shaker for 2 h. The solution was ltered with a sterile syringe
Page 6/21
containing a 0.22 µm lter (PureTech™, FineTech, Taichung, Taiwan) and frozen at -20°C. The frozen solution was
lyophilized in a lyophilization apparatus (OPERON, Korea) and resuspended in 200 µL of analytical grade water.
Carboxylates were quantied with an HPLC apparatus (Hitachi Primaide, MERCK, Darmstadt, Germany) and the
separation was performed on a reversed-phase column (Kromasil 100-5-C18, Nouryon, Göteborg, Suecia). The
mobile phase was carried out according to the method of Cawthray (2003). Citrate, malate, oxalate, and succinate
were used as standards and detected at 210 nm. Values were expressed as rate of carboxylates exuded per gram of
fresh weight per hour (µmol g− 1 FW h− 1) (Delgado et al. 2013).
Percentage Of AMF Colonization
The extent of AMF colonization in roots was determined using a method described by Nicolson (1995) and Read et
al. (1976). Root pieces of 1 cm were cut and washed with plenty of water to remove adhering soil, then claried with
KOH (2.5% w/v) at 120°C/15 min. After that, the KOH was removed and the roots were washed with water, then the
roots were covered with HCl (1% w/w) and left for 1 day to remove excess KOH. Subsequently, the HCl was
discarded, and the roots were washed with plenty of water and covered with trypan blue (0.05% w/v) for 1 day.
Finally, the dye was carefully discarded and the roots were washed again with water. With this method, AMF
structures (hyphae, arbuscules, and vesicles) were visualized in root tissues. For the quantication of root
colonization, we used the line intercept method, the roots were randomly distributed on a grid plate and observed
with a microscope at 40X, subsequently, all root intersections with horizontal lines were counted (Giovannetti and
Mosse 1980).
Statistical analysis
The statistical analysis was carried out using the INFOSTAT program. Normality and homogeneity assumptions
were checked using the Shapiro-Wilk and Levene tests. The results were analyzed using an analysis of variance
(ANOVA) and were considered statistically signicant at a p-value of less than 0.05. Mean comparison was
conducted using Fisher's LSD test. To analyze the percentage of AMF colonization at different depths (0–20 cm,
20–40 cm, and 40–60 cm), a Euclidean distance cluster analysis was applied.
Results
Soil biological properties
The analysis of FDAse activity provides information on the activity of the microbial community present in wheat
varieties at tillering, anthesis, and physiological maturity stages. Figure1a displays the experimental results of
FDAse compound measurement, which showed a statistically signicant increase (
p
<
0.05
) at tillering and anthesis
stages compared to physiological maturity. There was a decrease of 59% in FDAse activity during the physiological
maturity stage compared to the other two stages. However, no signicant differences were observed among the
fourteen wheat varieties at any of the three phenological stages (Fig.1b) or during the entire season (Fig.1c).
The analysis and comparison of basal soil microbial respiration in the different phenological stages, tillering,
anthesis and physiological maturity, were statistically signicant (
p
> 0.05) in the tillering and anthesis stages, being
40% higher than in the physiological maturity stage (Fig.2a). As for the varieties, Fig.2b shows signicant
differences (
p
> 0.05) among varieties in the tillering stage, with the variety Laurel (1987) the one that obtained the
highest basal soil respiration, being 35% higher than the rest of the varieties. However, the statistical test showed no
Page 7/21
signicant differences among varieties at the stages of anthesis and physiological maturity, nor during the whole
season (Fig.2c).
The measurement of the P-ase enzyme activity, represented by PNP in the three phenological stages, showed
signicantly different values (
p
> 0.05), where the anthesis stage was 31% higher than the tiller stage and 18%
higher than maturity stage physiological (Fig.3a). Figure3b shows the activity of the P-ase enzyme in each wheat
variety concerning their phenological stages, the varieties presented signicant differences (
p
> 0.05) only in the
tillering stage, with Pionero (2013) being approximately 46% higher than the other varieties. However, no signicant
differences were found between varieties in the stages of anthesis and physiological maturity, nor during the whole
season (Fig.3c).
Plant Analysis
Aboveground biomass
Regarding aerial biomass, as shown in Fig.4a, the weight of the plants at different phenological stages - tillering,
anthesis, and physiological maturity - is displayed. The stages of anthesis and physiological maturity were
signicantly different (
p
> 0.05) compared to the tillering stage, with a decrease of 93%. Figure4b presents the
weight of the wheat varieties at different phenological stages. Statistical analysis revealed signicant differences at
the anthesis and physiological maturity stages. Specically, at anthesis, the Druchamp (1965) variety was found to
be statistically different (
p
> 0.05), displaying a 35% increase in weight compared to the other varieties. Regarding
the state of physiological maturity, the Druchamp (1965) variety differed signicantly (
p
> 0.05), being around 33%
higher than the varieties Laurel (1987), Lautaro (1990), Tukan (1993), Bicentenario (2010), Pionero (2013), Rocky
(2015), Kiron (2017) and Chevignon (2020). Overall, the varieties showed a decrease in weight compared to the
Druchamp (1965) variety at both the anthesis and physiological maturity stages. However, no correlation was found
between the release years and the decrease in plant weight.
The measurement of plant height at various phenological stages revealed signicant differences (
p
> 0.05) at the
anthesis and physiological maturity stages, which were 87% taller than the tillering stage (Fig.5a). For wheat
varieties, signicant differences among varieties were observed at the anthesis and physiological maturity stages.
At both stages, Druchamp (1965) was the variety that obtained the greatest height, being signicantly different (
p
>
0.05) by approximately 30% from the other varieties (Fig.5b). Generally, at the anthesis and physiological maturity
stages, a decrease in height was observed in relation to the Druchamp (1965) variety. At the physiological maturity
stage, the varieties released up until the 1990s showed the greatest height, with a 15% increase compared to the
varieties from the 2000s. However, among this group, Chevignon (2020) was one of the tallest varieties, surpassing
even Tukan (1993).
Carboxylate Exudation
The root systems were found to primarily exude succinate as the main carboxylate, exuding an average of 76%
more than the other carboxylates, which was found to be statistically signicant (
p
> 0.05) (Fig.6a). Regarding the
exudation of each variety (Fig.6b), no signicant differences were found for succinate, malate and succinate
exudation, except oxalate, where the varieties Druchamp (1965) and Talafen (1982) showed signicant differences,
exuding approximately 89% less oxalate than the other varieties.
Page 8/21
Percentage Of AMF Colonization
Figure7a provides an overview of the signicant variations in AMF colonization percentages at different soil depths
(0–20 cm, 20–40 cm, and 40–60 cm) at anthesis stage. The results indicate that, in general, the highest AMF
colonization rate of 63% was observed at a soil depth of 20–40 cm, and it was found to be statistically signicant
(
p
> 0.05). At 40–60 cm depth, AMF colonization reaches 54%. However, only 6.5% AMF colonization was observed
at 0–20 cm depth. About this, Fig.7b showcases the concentration of P at various soil depths. The highest
concentration of P (30 ppm P-Olsen) was observed at a depth of 0–20 cm, while the lowest concentration (10 ppm
P-Olsen) was found at a depth of 40–60 cm. The P concentration was found to have signicant effects (
p
> 0.05)
on the percentage of AMF colonization.
Among the varieties, Laurel (1987) and Maxwell (2012) showed the highest colonization percentages at a soil depth
of 0–20 cm, and these results were found to be statistically signicant (
p
> 0.05). Druchamp (1965), Laurel (1987),
Lautaro (1990), Tukan (1993), Bicentenario (2010), Pionero (2013), and Chevignon (2020) showed signicant
differences (
p
> 0.05) and had 29% higher AMF colonization than the other varieties at 20–40 cm depth. At a soil
depth of 40–60 cm, the highest colonization percentages were achieved by the varieties Kiron (1987) and Laurel
(2017), and these results were found to be statistically signicant (
p
> 0.05) compared to the other varieties. Overall,
the varieties that showed the highest colonization percentages at a depth of 60 cm were Laurel (1987), Bicentennial
(2010), and Druchamp (1965), and these results were signicantly different (
p
> 0.05) from those obtained by
Talafen (1982), Kumpa (2002), and Rocky (2015) (Fig.7d).
Roots Analysis
The variety Chevignon (2020) had the longest root length up to a depth of 60 cm, with a length that was 39% greater
than that of the other varieties, and these results were found to be statistically signicant (
p
> 0.05) with respect to
Bicentenario (2010), Laurel (1987), Kiron (2017), Talafen (1982), Maxwell (2012), Druchamp (1965), Lautaro (1990),
Pionero (2013), Manquefen (1977) and Melifen (1974) (Fig.8a). The root length results for each variety represent
the combined length of all its individual roots. The varieties with the largest root area at a depth of 60 cm were
Chevignon (2020), Laurel (1987), and Bicentenario (2010). However, the ANOVA statistical test showed that there
were no signicant differences in root area compared to the other varieties (Fig.8b). The varieties Laurel (1987),
Bicentenario (2010), and Kumpa (2002) obtained the greatest root volume at 60 cm depth, but no statistically
signicant differences were found with respect to the other varieties (Fig.8c). Regarding root diameter at a depth of
60 cm, the varieties Manquefen (1977), Laurel (1987), and Bicentenario (2010) had the highest values. However,
statistical tests did not show signicant differences with respect to the other varieties (Fig.8d). At 60 cm depth, the
varieties Rocky (2015), Druchamp (1965), and Melifen (1974) showed the greatest root weight. However, statistical
analysis did not reveal any signicant differences among these varieties and the others (Fig.8e).
Discussion
Soil microbial activity
In regards to soil microbial activity, the activity of FDAse and microbial respiration were found to be signicantly
higher for all wheat varieties at the tillering and anthesis stages compared to the physiological maturity stage. The
results of a eld study by Chen et al. (2021) showed a decrease in soil respiration in wheat during anthesis and
physiological maturity. The soil microbial activity was found to be highest at the tillering and anthesis stages of the
Page 9/21
wheat varieties. This is because the highest nutrient uptake rate (N and P) occurs during these stages. The wheat
plant accumulates approximately 75% of its total N and 85% of its total P during anthesis, after which the nutrient
uptake rate decreases and the plant begins to remobilize nutrients from its roots to its aerial organs, such as
spikelets and grains. This observation aligns with a study conducted by Berardo and Reussi (2009) which found
that the rate of nutrient absorption decreases after anthesis. Plants have a strategy to obtain nutrients from the soil
through increased microbial activity. This activity is further enhanced by the rhizodeposition of previous and
established crops, where the quantity and quality of C inputs from rhizodeposition play a role in favoring the activity
of soil microorganisms (Chaparro et al. 2014; Powlson et al. 2008). One strategy that plants use to obtain nutrients
from the soil is the exudation of carbon by roots. This is because exudates serve as an important source of energy
for soil microorganisms. However, the amount and composition of these exudates will vary depending on the plant
species and its phenological stage (Gargallo-Garriga et al. 2018). In our study, we observed that the variety Laurel
(1987) had higher CO2 emission in the tillering stage, and this result was statistically signicant compared to the
other varieties. The variety Laurel (1987) is characterized by good tillering ability (Aguayo 1986), suggesting a
higher photosynthetic rate and greater C exudation by roots at the tillering stage. On the other hand, Meier et al.
(2023) showed that the addition of P in winter wheat increased the utilization of C sources by microorganisms and,
therefore, CO2 production.
During the early stages of a wheat plant's life cycle, there is a high demand for P. The uptake of P and the
development of roots occur before shoot growth (Romer and Schilling 1986). When the plant is in a P-decient
environment, it generates strategies to acquire this element, such as the modication of the root system or the
secretion of phosphatase enzymes (also released by P-solubilizing microorganisms), which are essential to
mineralize soil Po (de Souza Campos et al. 2021; Liu et al. 2023; Wu et al. 2021). However, our results indicated that
phosphatase activity was signicantly lower during the tillering stage. In general, the results of phosphatase activity
were high compared to those obtained by Schoebitz et al. (2020). Unlike that study, our wheat crops received a high
dose of triple superphosphate fertilizer prior to sowing and initially had a high concentration of P in the soil. In
relation to this, a study by Liu et al. (2023) determined that high P inputs (200 kg ha− 1 yr− 1) reduce the relative
abundance of genes involved in the P cycle because high doses of P block the signal transduction pathway among
the main genes encoding the regulation of the P starvation response (
phoR
and
phoP
) and regulating the high-
anity phosphate-specic transport system (
pst
). However, their results showed that P-ase genes (
phoN
,
aphA
, and
olpA
) increased with long-term P inux and alkaline phosphatase (P-al) genes (
phoA
and
phoD
) decreased. On the
other hand, Meier et al. (2023) cultivated wheat varieties in an Andisol with low and high P concentrations (4 and 30
mg P kg− 1), with water stress and abundant irrigation, and the results obtained differentiated among genotypes.
The results of P-ase activity differed among the wheat varieties, as seen in the study conducted by Meier et al.
(2023). The Druchamp (1965) variety displayed stable P-ase activity regardless of changes in P doses and water
stress, unlike the Rancofén variety, which showed the highest P-ase activity in low P conditions. These ndings
indicate that the impact of high P doses on P-ase activity varies among different cultivar varieties (de Souza
Campos et al. 2021).
Analysis Of Aboveground Plant Biomass And Root Architecture
Our ndings reveal that the release of wheat varieties between the 1960s and 1990s resulted in a 33% reduction in
weight and a 25% decrease in height, while the varieties released in the 2000s experienced a 8% reduction in weight
and a 17% decrease in height. However, concerning the plant weight of wheat varieties, no gradual decrease was
found in the years of release, as shown by Lo Valvo et al. (2018) in wheat varieties released between the years 1918
Page 10/21
and 2011 in Argentina. Our ndings on plant height align with the results reported by Del Pozo et al. (2021), where
they observed that the height of the plants decreased with the year of release of the wheat variety. Similarly, a study
by Matus et al. (2012) showed that the height of modern wheat cultivars (released after the 1960s) was reduced by
26% when compared to older cultivars. This is a consequence of the introduction of semi-dwarf genes (Rht1 and
Rht2) (Matus et al. 2012), which were incorporated starting in the 1960s and produced greater genetic gain in grain
yield, including height reduction (Del Pozo et al. 2021).
Our ndings show that the root length at 60 cm depth was greatest in the varieties released in the 2000s. This is
because, during the tillering stage, plants allocate more of their photoassimilates towards developing their root
system, before focusing on the growth of the above-ground parts. After the anthesis stage, root growth slows down
while the above-ground parts continue to grow rapidly until the stage of physiological maturity. (Cabeza and
Claassen 2017). This is consistent with the ndings of Siddique et al. (1990), who observed that modern wheat
varieties had a root penetration rate that was twice that of an older wheat cultivar. However, it's important to note
that these results can vary greatly based on the specic plant genotype and soil nutrient conditions. The root
system characteristics of 182 wheat varieties were studied by Dharmateja et al. (2021) in hydroponics under P-
limited and non-P-limited conditions. They found that the parameters of root length, area, volume, and diameter
showed signicant genetic variation. Under P-limited conditions, the total length, total volume, and total root area
increased, while the primary root length and average root diameter decreased. Our crops were under high doses of
inorganic fertilizers, generating a zone of high nutrient availability. Therefore, the plants would not need to invest
resources in the development of their root system to acquire nutrients (McGrail and McNear 2021). These results
show the response of the root architecture of different wheat varities to a high dose of inorganic fertilization in the
eld.
Carboxylates And AMF Colonization
Carboxylates can solubilize soil P by three mechanisms, chelating metal ions that immobilize P, displacing P from
adsorption sites, and changing soil pH (Wang et al. 2013a). Citrate and malate have been the most documented
carboxylates in wheat due to their eciency to solubilize inorganic soil P (de Souza Campos et al. 2021; Wang et al.
2013a; Wang et al. 2017). However, our results showed that, in general, all wheat plants exuded signicantly more
succinate, followed by oxalate, and to a lesser extent, malate and citrate. A study by Wang et al. (2015) evidenced
that in P-decient environments, plants exude higher amounts of carboxylates, on the contrary, in a high P
environment, plants limit the release of carboxylates due to the signicant photosynthate cost (5–25% of total
carbon) involved in exudation (Campos et al. 2018; Wen et al. 2022).
Wang et al. (2013c) demonstrated that 200 kg P ha− 1 fertilization reduced citrate release, but citrate increased the
eciency of phosphate fertilizer at low fertilization rates (40 kg P ha− 1) by increasing crop growth and P uptake.
Also, Wang et al. (2017) found that the exudation of citrate, malate, and succinate in the soil by wheat plants was
correlated with the levels of P applied and days after emergence, nding higher carboxylates at 29 days to 42 days
after emergence, and higher carboxylate accumulation at fertilization of 48 kg P ha− 1 compared to no fertilization.
Our results showed that wheat plants exuded higher amounts of succinate, followed by oxalate and then citrate and
malate. This is in contrast to the ndings of Wen et al. (2019), who reported lower carboxylate exudation in wheat.
However, our results are consistent with those of de Souza Campos et al. (2022), who observed higher exudation of
oxalates and lower amounts of citrate and malate in wheat genotypes grown in pots with an acid Andisol at low P-
Olsen concentration.
Page 11/21
Tukan (1993), Pioneer (2013), and Rocky (2015) exuded the highest amounts of carboxylates. However, in general,
varieties from 60 to 90 showed higher carboxylate exudation (15%) compared to those from 2000. Our study
revealed that there was no correlation between morphological traits and carboxylate exudation in the Tukan (1994),
Pioneer (2013), and Rocky (2015) varieties. This nding is consistent with the results of Iannucci et al. (2021), who
observed that despite having similar root characteristics, two durum wheat varieties (ancient and modern) showed
differences in carboxylate exudation. This is because varieties of the same species may differ in their ability to
exude carboxylate depending on their nutritional strategies (Iannucci et al., 2021). In addition, carboxylate exudation
may depend on the phenological stage of the plant (Chaparro et al. 2014). However, Tukan (1993) is considered an
inecient genotype in P acquisition due to its low tolerance to Al in acidic Andisol and decient in P (Seguel et al.
2017). Low Al tolerance, is associated with low citrate exudation (Dong et al. 2004). Our ndings indicate that
Tukan (1993) exuded a signicantly higher amount of citrate compared to other varieties. The varying results in
previous studies suggest that the exudation of carboxylates is inuenced by the plant genotype and its interaction
with the environment. Additionally, multiple processes that take place over time and space affect the plant's
requirements (Seguel et al. 2017).
The relationship between high levels of P and low colonization by AMF has been widely documented. This
relationship is due to the high energy costs for the plant (Kobae et al. 2016; Qin et al. 2020; Wen et al. 2019). A
deeper explanation may be demonstrated by Wang et al. (2013b) who showed that the
Ta-PHR1-A1
gene encoding
PHR1
protein in wheat was involved in Pi signaling in response to Pi starvation, furthermore, upon expression, it
stimulated root growth and thus enhanced P uptake. Das et al. (2022) demonstrated that rice
PHR2
(Phosphate
Starvation Response) protein is an important regulator of phosphate starvation response in AMF. In their research
they discuss that, at high phosphate levels,
PHR2
is repressed by
SPX
proteins (Lv et al. 2014) in the cytoplasm,
preventing
PHR2
translocation into the nucleus and consequently, preventing
PHR2
binding to the promoters of
phosphate starvation-induced genes and genes involved in AMF signaling, exuding a lower amount of
strigolactones by the plant and expressing a lower amount of genes sensing AMF Myc-factors. In our eld
experiment the depth of the soil samples has an impact on the level of AMF colonization, which is inuenced by the
soil´s P level.
Conclusion
Our ndings reveal that there is substantial genotypic variation among winter wheat varieties with regards to their
rhizosphere biology and root architecture. This suggests that there are varieties with more ecient nutritional
strategies, such as carboxylate exudation, microbial activity, and root growth, which are not necessarily dependent
on the year of their release. Our ndings indicate that there is no clear pattern of variation in root architecture
parameters (such as volume, area, diameter, and length) and rhizosphere biological activity with respect to the year
of release of the wheat variety. This eld experiment shows that there is no direct correlation between root
architecture characteristics and rhizosphere biological activity. Instead, the level of biological activity in the
rhizosphere and root architecture is inuenced by various factors including P fertilization, environmental conditions,
and the crop's growth stage. Further research is required to examine the root architecture, microbial activity and
nutrient utilization eciency under low levels of fertilization.
Declarations
Acknowledgements. This work was supported by ANID FONDECYT Regular Project No.1220425 and 1201950.
Page 12/21
Funding. This work was supported by ANID FONDECYT Regular Project No.1220425 and 1201950.
Author Contributions. All authors contributed to the study conception and design. Material preparation, data
collection and analysis were performed by Paula Paz-Vidal, Dalma Castillo-Rosales, María Dolores López, Iván
Matus, Felipe Noriega and Mauricio Schoebitz. The rst draft of the manuscript was written by Paula Paz-Vidal and
all authors commented on previous versions of the manuscript. All authors read and approved the nal manuscript.
Data Availability. The datasets generated during the current study are available from the corresponding author on
reasonable request
References
1. Aguayo L (1986) Laurel INIA, nuevo cultivar de trigo invernal, para la zona centro sur [on line]. Investigación y
Progreso Agropecuario Quilamapu 27: 6-9. doi: https://hdl.handle.net/20.500.14001/40897. (Accessed 11 dec
2022).
2. Alef K, Nannipieri P (1995) Enzyme activities, in Methods in applied soil microbiology and biochemistry.
Academic Press Limited Chapter 7: 311-373.
3. Aziz M, Palta J, Siddique K, Sadras O (2017) Five decades of selection for yield reduced root length density and
increased nitrogen uptake per unit root length in Australian wheat varieties. Plant Soil 413:181–192.
https://doi.org/10.1007/s11104-016-3059-y
4. Berardo A, Reussi N (2009) PAUTAS PARA EL MANEJO DE LA FERTILIZACIÓN EN TRIGO. 2° NUTRICIÓN DE
CULTIVOS Y PASTURAS. doi:
https://www.laboratoriofertilab.com.ar/Trabajos/2009.Manejo%20de%20la%20fertilizacion%20en%20Trigo.pdf
. (Accessed 04 Jan 2023).
5. Borie F, Rubio R (2003) FÓSFORO TOTAL Y ORGÁNICO EN SUELOS VOLCÁNICOS CHILENOS. Gayana Bot 60:
69-73. http://dx.doi.org/10.4067/S0717-66432003000100011
. Bünemann E, Bongiorno G, Zhanguo B, Creamer R, De Deyn G, de Goede R, Fleskens L, Geissen V, Kuyper T,
Mäder P, Pulleman M, Sukkel W, Willem J, L B (2018) Soil quality – A critical review. Soil Biol Biochem 120:
105–125. https://doi.org/10.1016/j.soilbio.2018.01.030.v
7. Cabeza R, Claassen N (2017) Sistemas radicales de cultivos: extensión, distribución y crecimiento. Agro Sur
45(2): 31-45, 201. https://doi.org/10.4206/agrosur.2017.v45n2-04
. Campos P, Borie F, Cornejo P, Lopez-Raez JA, Lopez-Garcia A, Seguel A (2018) Phosphorus Acquisition
Eciency Related to Root Traits: Is Mycorrhizal Symbiosis a Key Factor to Wheat and Barley Cropping? Front
Plant Sci 9: 752. https://doi.org/10.3389/fpls.2018.00752
9. Cawthray GR (2003) An improved reversed-phase liquid chromatographic method for the analysis of low-
molecular mass organic acids in plant root exudates. J Chromatogr A 1011: 233-240.
https://doi.org/10.1016/s0021-9673(03)01129-4
10. Chaparro JM, Badri DV, Vivanco JM (2014) Rhizosphere microbiome assemblage is affected by plant
development. ISME J 8: 790-803. https://doi.org/10.1038/ismej.2013.196
11. Chen S, Wang J, Zhang T, Hu Z, Zhou G (2021) Warming and straw application increased soil respiration during
the different growing seasons by changing crop biomass and leaf area index in a winter wheat-soybean
rotation cropland. Geoderma 391: 114985. https://doi.org/10.1016/j.geoderma.2021.114985
Page 13/21
12. Das D, Paries M, Hobecker K, et al (2022) PHOSPHATE STARVATION RESPONSE transcription factors enable
arbuscular mycorrhiza symbiosis. Nat Commun 13: 477. https://doi.org/10.1038/s41467-022-27976-8
13. de Souza Campos P, Meier S, Morales A, Borie F, Cornejo P, Ruiz A, Seguel A (2022) Root traits distinguish
phosphorus acquisition of two wheat cultivars growing in phosphorus-decient acid soil. Rhizosphere 22:
100549. https://doi.org/10.1016/j.rhisph.2022.100549
14. de Souza Campos PM, Borie F, Cornejo P, Meier S, Lopez-Raez JA, Lopez-Garcia A, Seguel A (2021) Wheat root
trait plasticity, nutrient acquisition and growth responses are dependent on specic arbuscular mycorrhizal
fungus and plant genotype interactions. J Plant Physiol 256: 153297.
https://doi.org/10.1016/j.jplph.2020.153297
15. Del Pozo A, Jobet C, Matus I, Mendez-Espinoza AM, Garriga M, Castillo D, Elazab A (2021) Genetic Yield Gains
and Changes in Morphophysiological-Related Traits of Winter Wheat in Southern Chilean High-Yielding
Environments. Front Plant Sci 12: 732988. https://doi.org/10.3389/fpls.2021.732988
1. Delgado M, Zúñiga-Feest A, Alvear M, Borie F (2013) The effect of phosphorus on cluster-root formation and
functioning of Embothrium coccineum. R. et J Forst. Plant Soil: 373:765–773. https://doi.org/10.1007/s11104-
013-1829-3
17. Dharmateja P, Kumar M, Pandey R, Mandal PK, Babu P, Bainsla NK, et al (2021) Deciphering the change in root
system architectural traits under limiting and non-limiting phosphorus in Indian bread wheat germplasm. PLoS
ONE 16 (10): e0255840. https://doi.org/10.1371/journal.pone.0255840
1. Dong P, Peng X, Yan X, (2004) Organic acid exudation induced by phosphorus deciency and/or aluminum
toxicity in two contrasting soybean genotypes. Physiol Plant: 190–199. https://doi.org/10.1111/j.1399-
3054.2004.00373.x
19. Gargallo-Garriga A, Preece C, Sardans J, Oravec M, Urban O, Penuelas J (2018) Root exudate metabolomes
change under drought and show limited capacity for recovery. Sci Rep 8: 12696.
https://doi.org/10.1038/s41598-018-30150-0
20. Giovannetti M, Mosse B (1980) An Evaluation of Techniques for Measuring Vesicular Arbuscular Mycorrhizal
Infection in Roots. New Phytol 84: 489-500. https://doi.org/10.1111/j.1469-8137.1980.tb04556.x
21. Iannucci A, Canfora L, Nigro F, De Vita P, Beleggia R (2021) Relationships between root morphology, root
exudate compounds and rhizosphere microbial community in durum wheat. Appl Soil Ecol 158: 103781.
https://doi.org/10.1016/j.apsoil.2020.103781
22. Jacquiod S, Raynaud T, Pimet E, Ducourtieux C, Casieri L, Wipf D, Blouin M (2022) Wheat Rhizosphere
Microbiota Respond to Changes in Plant Genotype, Chemical Inputs, and Plant Phenotypic Plasticity. Front Ecol
Evol 10: 903008. https://doi.org/10.3389/fevo.2022.903008
23. Kobae Y, Ohmori Y, Saito C, Yano K, Ohtomo R, Fujiwara T (2016) Phosphate Treatment Strongly Inhibits New
Arbuscule Development But Not the Maintenance of Arbuscule in Mycorrhizal Rice Roots. Plant Physiol 171:
566-579. https://doi.org/10.1104/pp.16.00127
24. Kuzyakov Y, Friedel JK, Stahr k (2000) Review of mechanisms and quantication of priming effects. Soil Biol
Biochem 32: 1485–1498. https://doi.org/10.1016/S0038-0717(00)00084-5
25. Li W, Wen X, Han J, Liu Y, Wu W, Liao Y (2017) Optimum ridge-to-furrow ratio in ridge-furrow mulching systems
for improving water conservation in maize (Zea may L.) production. Environ Sci Pollut Res Int 24: 23168–
23179. https://doi.org/10.1007/s11356-017-9955-8
2. Liu L, Gao Z, Yang Y, Gao Y, Mahmood M, Jiao H, Wang Z, Liu J (2023) Long-term high-P fertilizer input shifts
soil P cycle genes and microorganism communities in dryland wheat production systems. Agric Ecosyst
Page 14/21
Environ 342: 108226. https://doi.org/10.1016/j.agee.2022.108226
27. Lo Valvo P, Miralles D, Serrago R (2018) Genetic progress in Argentine bread wheat varieties released between
1918 and 2011: Changes in physiological and numerical yield components. Field Crop Res 221: 314-321.
https://doi.org/10.1016/j.fcr.2017.08.014
2. Lv Q, Zhong Y, Wang Y, Wang Z, Zhang L, Shi J, Wu Z, Liu Y, Mao C, Yi K, Wu P (2014) SPX4 Negatively
Regulates Phosphate Signaling and Homeostasis through Its Interaction with PHR2 in Rice. Plant Cell 26:
1586–1597. https://doi.org/10.1105/tpc.114.123208
29. Matus I, Mellado M, Pinares M, Madariaga R, del Pozo A (2012) Genetic progress in winter wheat cultivars
released in Chile from 1920 to 2000. Chil J Agric Res 72: 303–308. https://doi.org/10.4067/s0718-
58392012000300001
30. McGrail R, McNear D (2021) Two centuries of breeding has altered root system architecture of winter wheat.
Rhizosphere 19: 100411. https://doi.org/10.1016/j.rhisph.2021.100411
31. Meier S, de Souza P, Palma R, Morales A, Hirzel J, Aponte H, Cartes P, Ondrasek G, Seguel A (2023) Assembly
between wheat cultivars and soil microorganisms modulates phosphorus and water use eciency. Rhizosphere
25: 100631. https://doi.org/10.1016/j.rhisph.2022.100631
32. Naseer M, Zhu Y, Li FM, Yang YM, Wang S, Xiong YC (2022) Nano-enabled improvements of growth and
colonization rate in wheat inoculated with arbuscular mycorrhizal fungi. Environ Pollut 295: 118724.
https://doi.org/10.1016/j.envpol.2021.118724
33. Nazareno M, Ruscitti M, Arango M (2020) Micorrizas arbusculares. Biología y aplicaciones en el sector agro-
forestal. Libro de cátedra 1: 11-89. https://doi.org/10.35537/10915/99599
34. Nicolson T (1995) The Mycotrophic Habit in Grasses: Vesicular-arbuscular Endophytes and Other Fungi
Associated with the Roots of Members of the Gramineae (Doctoral dissertation, University of Nottingham).
35. Ober ES, Alahmad S, Cockram J, Forestan C, Hickey LT, Kant J, Maccaferri M, Marr E, Milner M, Pinto F, Rambla
C, Reynolds M, Salvi S, Sciara G, Snowdon RJ, Thomelin P, Tuberosa R, Uauy C, Voss-Fels KP, Wallington E, Watt
M (2021) Wheat root systems as a breeding target for climate resilience. Theor Appl Genet 134: 1645-1662.
https://doi.org/10.1007/s00122-021-03819-w
3. Powlson DS, Riche AB, Coleman K, Glendining MJ, Whitmore AP (2008) Carbon sequestration in European soils
through straw incorporation: limitations and alternatives. Waste Manag 28: 741-746.
https://doi.org/10.1016/j.wasman.2007.09.024
37. Qin X, Feng F, Wen X, Siddique K, Liao Y (2019) Historical genetic responses of yield and root traits in winter
wheat in the yellow-Huai-Hai River valley region of China due to modern breeding (1948–2012). Plant Soil 439:
7–18. https://doi.org/10.4067/s0718-58392012000300001.
3. Qin Z, Zhang H, Feng G CP, Zhang J, Li X, Gai J (2020) Soil phosphorus availability modies the relationship
between AM fungal diversity and mycorrhizal benets to maize in an agricultural soil. Soil Biol Biochem 144:
107790. https://doi.org/10.1016/j.soilbio.2020.107790
39. Read DJ, Koucheki HK, Hodgson J (1976) Vesicular-arbuscular mycorrhiza in natural vegetation systems. I. The
occurrence of infection. New Phytol 77: 641–653. https://doi.org/10.1111/j.1469-8137.1976.tb04657.x
40. Romer W, Schilling G (1986) Phosphorus requirements of the wheat plant in various stages of its life cycle.
Plant Soil 91: 221–229. https://doi.org/10.1007/BF02181789
41. Schoebitz M, Castillo D, Jorquera M, Roldan A (2020) Responses of Microbiological Soil Properties to
Intercropping at Different Planting Densities in an Acidic Andisol. Agronomy 10(6): 781.
Page 15/21
https://doi.org/10.3390/agronomy10060781
42. Seguel A, Cornejo P, Ramos A, Von Baer E, Cumming J, Borie F (2017) Phosphorus acquisition by three wheat
cultivars contrasting in aluminium tolerance growing in an aluminium-rich volcanic soil. Crop Pasture Sci 68:
305–316. https://doi.org/10.1071/CP16224
43. Severini A, Wasson A, Evans J, Richards R, Watt M (2020) Root phenotypes at maturity in diverse wheat and
triticale genotypes grown in three eld experiments: relationships to shoot selection, biomass, grain yield,
owering time, and environment. Field Crop Res 255: 107870. https://doi.org/10.1016/j.fcr.2020.107870
44. Siddique K, Belford R, Tennant D (1990) Root:shoot ratios of old and modern, tall and semi-dwarf wheats in a
mediterranean environment. Plant Soil 121: 89–98. https://doi.org/doi.org/10.1007/BF00013101
45. Soil Survey Staff (2014) Keys to Soil Taxonomy, USDA-Natural, Resources Conservation Service, Washington,
DC. 12th edn.
4. Stolpe N (2006) Descripciones de los principales suelos de la VIII región de Chile. 15-17.
47. Tabatabai MA, Bremmer J (1969) Use of p-nitrophely phosphate for assay of soil phosphatase activity. Soil
Biol: 301-307. https://doi.org/10.1016/0038-0717(69)90012-1
4. Wang E, Ridoutt B, Luo Z, M P (2013a) Using systems modelling to explore the potential for root exudates to
increase phosphorus use eciency in cereal crops. Environ Model Softw 46: 50-60.
https://doi.org/10.1016/j.envsoft.2013.02.009
49. Wang J, Sun J, Miao J, Guo J, Shi Z, He M, Chen Y, Zhao X, Li B, Han F, Tong Y, Li Z (2013b) A phosphate
starvation response regulator Ta-PHR1 is involved in phosphate signalling and increases grain yield in wheat.
Ann Bot 111(6): 1139-1153. https://doi.org/10.1093/aob/mct080
50. Wang Y, Almvik M, Clarke N, Eich-Greatorex S, Øgaard A, Krogstad T, Lambers H, Clarke J (2015) Contrasting
responses of root morphology and root-exuded organic acids to low phosphorus availability in three important
food crops with divergent root traits. AoB PLANTS 7: plv097. https://doi.org/10.1093/aobpla/plv097
51. Wang Y, Xu H, Kou J, Shi L, Zhang C, Xu F (2013c) Dual effects of transgenic Brassica napus overexpressing CS
gene on tolerances to aluminum toxicity and phosphorus deciency. Plant Soil 362: 231–246.
https://doi.org/10.1007/s11104-012-1289-1
52. Wang Y, Krogstad T, Clarke N, Øgaard AF, Clarke JL (2017) Impact of phosphorus on rhizosphere organic anions
of wheat at different growth stages under eld conditions. AoB PLANTS 9.
https://doi.org/10.1093/aobpla/plx008
53. Wen Z, Li H, Shen Q, Tang X, Xiong C, Li H, Pang J, Ryan MH, Lambers H, Shen J (2019) Tradeoffs among root
morphology, exudation and mycorrhizal symbioses for phosphorus-acquisition strategies of 16 crop species.
New Phytol 223: 882-895. https://doi.org/10.1111/nph.15833
54. Wen Z, White PJ, Shen J, Lambers H (2022) Linking root exudation to belowground economic traits for resource
acquisition. New Phytol 233: 1620-1635. https://doi.org/10.1111/nph.17854
55. Wu A, Fang Y, Liu S, Wang H, Xu B, Zhang S, Palta J, Siddique K, Chen Y (2021) Root morphology and
rhizosheath acid phosphatase activity in legume and graminoid species respond differently to low phosphorus
supply. Rhizosphere 19: 100391. https://doi.org/10.1016/j.rhisph.2021.100391
5. Zadoks J, Chang T, Konzak C (1974) A Decimal Code for the Growth Stages of Cereals. Weed Res 14: 415-421.
https://doi.org/10.1111/j.1365-3180.1974.tb01084.x
57. Zhu Y, Lv G, Ying-Long C, Xiao-Fang G, Yi-Nan P, Zhi-Ye W, Ai-Tian R, X Y-C (2017) Inoculation of arbuscular
mycorrhizal fungi with plastic mulching in rainfed wheat: A promising farming strategy. Field Crops Res 204:
Page 16/21
223-230. https://doi.org/10.1016/j.hpj.2020.06.003
Figures
Figure 1
(a) Mean values of FDAse activity of the soil (µg of FDAse g dry soil-1) present at tillering, anthesis, and
physiological maturity phenological stages, (b) mean values of FDAse activity (µg of FDAse g dry soil-1) of each
wheat variety at tillering, anthesis and physiological maturity phenological stages, and (c) mean values of FDAse
activity (µg of FDAse g dry soil-1) of each wheat variety during the whole season. Means ± SEM is indicated (n=56).
Different letters indicate signicant differences among phenological stages, as revealed by Fisher's LSD test
(
p
>0.05).
Figure 2
(a) Mean values of soil microbial respiration of the soil (µg CO2 g-1 h-1) at tillering, anthesis, and physiological
maturity phenological stages, (b) mean values of soil microbial respiration (µg CO2 g-1 h-1) of each wheat variety at
tillering, anthesis and physiological maturity phenological stages and (c) Mean values of soil microbial respiration
(µg CO2 g-1 h-1) during the whole season. Means ± SEM is indicated (n=56). Different letters indicate signicant
differences among varieties and phenological stages, as revealed by Fisher's LSD test (
p
>0.05).
Page 17/21
Figure 3
(a) Mean values of acid phosphatase enzyme activity of the soil (µmol PNP g-1 soil h-1) at the phenological stages
of tillering, anthesis, and physiological maturity, (b) mean values of acid phosphatase enzyme activity of the soil
acid phosphatase (µmol PNP g-1 soil h-1) of each wheat variety at tillering, anthesis, and physiological maturity
phenological stages, and (c) mean values enzyme activity of the soil acid phosphatase (µmol PNP g-1 soil h-1) of
each wheat variety during the whole season. Means ± SEM is indicated (n=56). Different letters indicate signicant
differences among varieties and phenological stages, as revealed by Fisher's LSD test (
p
>0.05).
Figure 4
(a) Mean plant weight values (g) at tillering, anthesis, and physiological maturity phenological stages and (b) mean
plant weight values of each wheat variety at tillering, anthesis, and physiological maturity phenological stages.
Means ± SEM is indicated (n=56). Different letters indicate signicant differences among varieties and phenological
stages, as revealed by Fisher's LSD test (
p
>0.05).
Page 18/21
Figure 5
(a) Mean values of plant height at tillering, anthesis, and physiological maturity phenological stages and (b) mean
values of plant height of each wheat variety at tillering, anthesis, and physiological maturity phenological stages.
Means ± SEM is indicated (n=56). Different letters indicate signicant differences among varieties and phenological
stages, as revealed by Fisher's LSD test (
p
>0.05).
Figure 6
(a) Total carboxylate content exuded per complete root systems of each wheat variety (µmol g-1 FW h-1) (b) mean
values of oxalate, succinate, malate, and citrate exudation per complete root systems of each wheat variety (µmol g-
Page 19/21
1 FW h-1). Mean ± SEM is indicated (n=56). Different letters indicate signicant differences between varieties as
revealed by Fisher's LSD test (
p
>0.05).
Figure 7
(a) Mean values of AMF colonization (%) in wheat varieties at depths of 0-20 cm, 20-40 cm, and 40-60 cm, (b) mean
values of AMF colonization percentage (%) in wheat varieties in relation to phosphorus concentrations analyzed at
different depths 0-20 cm, 20-40 cm, and 40-60 cm, (c) Euclidean distance analysis of percent AMF colonization (%)
in each wheat variety at depths of 0-20 cm, 20-40 cm and 40-60 cm and (d) mean values of percent AMF
colonization (%) in wheat varieties at 60 cm depth. Means ± SEM is indicated (n=42). Different letters indicate
signicant differences among varieties and depths, as revealed by Fisher's LSD test (
p
>0.05).
Page 20/21
Figure 8
(a) Mean values of root length of wheat varieties at a depth of 60 cm (cm), (b) mean values of root area of wheat
varieties at a depth of 60 cm (cm2), (c) mean values of root volume of wheat varieties at a depth of 60 cm (cm3), (d)
mean values of root diameter at a depth of 60 cm (mm) and (e) mean values of root weight of wheat varieties at a
depth of 60 cm (g). Means ± SEM is indicated (n=42). Different letters indicate signicant differences between
varieties as revealed by Fisher's LSD test (
p
>0.05).
Supplementary Files
This is a list of supplementary les associated with this preprint. Click to download.
