https://orcid.org/0000-0001-6715-8986
Abstract
Dioecious plants show sexual dimorphism in their phosphorus (P) availability responses. However, the understanding of sex-specific strategies for P utilization and acquisition under varying soil moisture levels remains unclear. Here, we assessed a range of root functional traits, soil P properties, total foliar P concentration ([P]) and leaf chemical P fractions—inorganic P ([Pi]), metabolite P ([PM]), lipid P ([PL]), nucleic acid P ([PN]) and residual P ([PR])—as well as other leaf functional traits in female and male trees under different soil moisture levels (25% for high and 7% for low). Our results showed that females had larger specific root length under well-watered conditions, resulting in greater root foraging capacity. This led to a 36.3% decrease in soil active [Pi] in the rhizosphere and a 66.9 % increase in total foliar [P], along with all five foliar chemical P fractions ([Pi], [PM], [PL], [PN] and [PR]) compared with males. However, males exhibited significantly higher photosynthetic P utilization efficiency than females. Especially under low soil moisture levels, males exhibited a significant reduction in soil active organic P, coupled with a large increase in the exudation of soil phosphatases and carboxylates. Furthermore, the proportion of [PM] in total foliar [P] was 42.0% higher in males than in females. Mantel and Spearman correlation analyses revealed distinct coordination and trade-offs between foliar P fraction allocation and below-ground P acquisition strategies between the two sexes. Leveraging these sex-specific strategies could enhance the resilience of dioecious populations in forest plantations facing climate-induced variability.
摘要
雌雄异株植物对土壤磷有效性的响应表现出性别二态性。然而，关于不同土壤水分条件下雌雄异株植物性别特异的磷利用和吸收策略尚不清楚。因此，本研究测定了不同土壤水分条件下(高25%和低7%)雌雄胡杨(Populus euphratica)的根系功能性状、土壤磷组分、叶片总磷、叶片磷组分(无机磷、代谢磷、脂质磷、核酸磷和残余磷)以及其它叶片功能性状。结果表明，水分充足时，雌性的比根长更大，具有更强的根系觅食能力，导致雌性根际的土壤活性无机磷下降了36.3%，叶片总磷升高了66.9%，其他叶片磷组分也相应升高。然而，雄性的光合磷利用效率(PPUE)显著高于雌性，尤其在土壤水分不足时。相对于雌性，雄性土壤活性有机磷含量显著减少，土壤磷酸酶和羧酸盐含量显著增加，叶片代谢磷比例提高42.0%。Mantel和Spearman相关分析显示，胡杨叶片磷分配和地下磷获取策略之间存在性别特异的协调和权衡。在气候变化背景下，利用这些性别特异策略可以有效帮助雌雄异株植物种群的恢复。
INTRODUCTION
Phosphorus (P) is an indispensable element in plant metabolism, pivotal to many processes that underpin vegetative growth and enable adaptation to environmental stresses (Du et al. 2020; Hou et al. 2020). In terrestrial ecosystems, the bioavailability of P often acts as a bottleneck for plant development, necessitating strategic optimization of both P absorption efficiency and P utilization efficiency to ensure survival and continuation of growth trajectories (Jelincic et al. 2022; Lambers 2022; Poirier et al. 2022; Reichert et al. 2022). The spectrum of P forms accessible to plants ranges from readily assimilable orthophosphates to recalcitrant organic P (Po). The latter category, frequently embedded within complex soil matrices, requires mobilization via root exudates that facilitate mineralization—a process of paramount importance in P-impoverished ecosystems (Raven et al. 2018; Richardson et al. 2011). This mineralization not only epitomizes a pivotal adaptation mechanism but also unlocks a significant reservoir of P, thus ensuring nutrient cycling and ecosystem resilience (Gao et al. 2022; Wen et al. 2019; Xia et al. 2022).
Elastic allocation in foliar P fractions ensures the efficient use of absorbed P, reflecting another adaptive strategy to P scarcity (Han et al. 2021, 2022b; Hayes et al. 2022; Mo et al. 2019; Suriyagoda et al. 2022). Within plants, P is categorized into five chemical fractions: inorganic phosphate (Pi), metabolite P [PM], lipid P [PL], nucleic acid P [PN] and residual P [PR], the latter four constituting Po (Han et al. 2022a; Suriyagoda et al. 2022). The active Pi in the cytoplasm is fundamental to cellular metabolism, photosynthesis and respiration (Lambers 2022; Stigter and Plaxton 2015). During P scarcity, plants reallocate Pi to support growth, utilizing small metabolites and nucleic acids (Pratt et al. 2009; Veneklaas et al. 2012), while [PL] and [PR] maintain cellular functions and signaling (Matzek and Vitousek 2009; Nakamura 2017; Suriyagoda et al. 2022).
Soil P cycling and its bioavailability are profoundly modulated by variations in soil moisture levels, a critical factor in arid desert ecosystems where low precipitation and high evaporation prevail (Zhang et al. 2020). These regions are typified by a decelerated biogeochemical turnover of soil nitrogen (N) and P, resulting in inherently low-fertility substrates (Gong et al. 2015; Noy-Meir 1973; Yu and Wang 2018). As aridity intensifies, a concomitant decrease in plant nutrient acquisition ensues, largely due to restricted bioavailability (Cramer et al. 2009; Sardans and Penuelas 2012; Waraich et al. 2011). Drought also exacerbates this scenario by limiting the mobility of P within the soil matrix—impeding diffusion and mass flow—and by hampering the mineralization mechanisms essential for plant P acquisition (He and Dijkstra 2014; Lambers et al. 2008; Schimel et al. 2007). The resultant soil desiccation reduces the metabolic vigor of the soil microbial community and diminishes enzymatic activity, factors that are instrumental in P mineralization (Gao et al. 2020; He et al. 2023). The challenge is exacerbated by the alkaline nature of desert soils, often laden with calcium and magnesium ions that sequester P, thereby inhibiting its uptake and utilization by plants (Adnan et al. 2022). Consequently, in the context of the typical P shortage and fluctuating moisture regimes characteristic of desert soils, plants should fine-tune their P acquisition and utilization mechanisms, fostering resilience to these stringent environmental conditions.
Dioecious plant species, marked by pronounced sexual dimorphism, manifest distinct functional traits and reproductive investments between the sexes (Hultine et al. 2016; Randriamanana et al. 2014; Xia et al. 2023). Female plants bear higher resource demands, primarily attributable to their reproductive roles involving seed and fruit development (Juvany and Munne-Bosch 2015; Liu et al. 2021). Such resource allocation is especially critical for P, a key nutrient for seed and fruit production. Correspondingly, females often develop more acquisitive root architectures with extensive specific root lengths (SRLs), thereby broadening their foraging scope and enhancing P acquisition (Xia et al. 2020, 2024). On the contrary, male plants release more organic compounds into the soil. These exudates have notable impacts on P availability in the rhizosphere, either by directly mobilizing P (e.g. extracellular enzymes and organic acids) from soil Po and recalcitrant Pi, or serving as regulators (specific signals) for facilitating microbial growth and mycorrhizal pathways (Guo et al. 2023; Xia et al. 2024). Males also exhibit superior photosynthetic efficiency, a higher photosynthetic P use efficiency (PPUE), and maintain more robust photosystem II functionality under P stress compared with their female counterparts (Zhang et al. 2014). While previous studies have explored the relationship between total leaf P content and soil P availability in relation to plant sex (Xia et al. 2020), the complexity of how water scarcity influences leaf P fractions in dioecious species remains less understood. Crucially, associations between leaf P fraction allocation and below-ground P acquisition strategies under water scarcity warrant further investigation.
In the arid and saline–alkaline soils of Xinjiang, China, water scarcity and nutrient shortage pose significant challenges to plant survival. Populus euphratica, a dioecious species, thrives under these harsh desert conditions. We conducted an 18-year field experiment with varying water supply levels to examine root functional traits, soil P properties and the allocation patterns of foliar P fractions. A central aim was to elucidate the associations between root functionality and soil P properties as well as foliar P fractions. We hypothesized that: (i) Under high soil moisture conditions, females would exhibit enhanced morphological root foraging capabilities, enabling them to absorb more Pi and significantly increase each P fraction in their leaves. Therefore, females accumulate more total leaf P compared with males; (ii) Under low soil moisture levels, males can dissolve Pi and mineralize Po through root physiological traits and optimize the allocation of leaf P fractions to conserve P. Consequently, it is expected that males will adopt a more proactive strategy to mitigate P deficiency compared with females; (iii) Allocation of leaf P fraction and root functional traits exhibit sex-dependent correlations. This may be due to sex-specific differences in the acquisition and utilization of P forms.
MATERIALS AND METHODS
Study area
This study was conducted on the northwestern margin of the Tarim Basin in Xinjiang, China, characterized by calcic xerosol soil and a typical temperate desert climate. Meteorological data from the Arak meteorological station over the past 40 years (81°05ʹ E, 40°05ʹ N) reveals an average annual temperature of 10.8 °C, with an average annual precipitation of 50 mm. Seasonal temperature averages are 14.7 °C in spring, 24.1 °C in summer, 10.5 °C in autumn and −5.1 °C in winter, with corresponding annual rainfall of 10.5, 20.5, 10.4 and 8.6 mm, respectively.
Experimental design
We focused on P. euphratica trees established in 2003 within a vegetation restoration initiative along the upper reaches of the Tarim River (81°17ʹ E, 40°32ʹ–40°81ʹ N). A two-factorial design was employed, incorporating two tree sexes and two soil moisture levels across 12 replicated plots of 100 m² each, with a tree spacing of 1.2 m × 4.2 m, each containing 33 trees. Interplot buffer zones range from 10 to 15 m to mitigate edge effects (Supplementary Fig. S1). Initial irrigation was uniformly performed across all plots during the formative 5-year period. From 2009 to 2021, a dichotomous moisture regime was established, where half of the plots received consistent irrigation in March and April, constituting the well-watered treatment, while the remainder were subjected solely to ambient precipitation, characterizing the water deficiency treatment. The irrigation regimen ensured a gravimetric soil moisture content of 90% within the upper 60 cm soil profile, delivered over an 8-h daily cycle at a flow rate of 50 m³ h−1. At the beginning of April 2020 when P. euphratica began to flower, we identified the sex of each tree based on the flower morphology. After rigorous investigations, we found a disparity in the sex ratio (female to male), with a ratio of 0.86 in well-watered conditions and decreasing to 0.64 under conditions of water scarcity (Xia et al. 2022). For analytical robustness, three female and male trees from each treatment per plot were designated for replicate sampling (Supplementary Fig. S1).
Photosynthesis measurement
In April 2021, healthy P. euphratica female and male trees from each treatment were chosen for photosynthesis measurements. Three newly grown branches from the four cardinal directions at 10 m height were selected, using the third and fourth leaves from each selected branch. Measurements were repeated thrice per leaf for accuracy and averaged across trees per plot. The photosynthesis was measured using a portable open-system infrared gas analyzer (LI-6800, LI-COR, USA) during 8:30–10:30 a.m. on sunny days. All measurements were made using a 6 cm2 chamber (3 cm × 2 cm) under a saturating light of 1700 µmol m−2 s−1 photosynthetic photon flux density (red-blue light source; 6400-02B) at 400 µmol mol−1 CO2. ImageJ software was used to measure the leaf area for each leaf, facilitating mass-based (Amass:mass-based maximum photosynthetic carbon assimilation rate) photosynthetic rates (Hidaka and Kitayama 2013).
Leaf, root and soil collection
Leaves from three healthy trees per sex within each treatment were pooled as one replicate. The most recent fully expanded mature leaves with no visible damage or discoloration were collected. The samples were immediately submerged in liquid nitrogen for subsequent measurement of foliar P fractions. Additionally, subsamples were oven-dried and used for measuring leaf N and manganese (Mn2+).
Fine roots from the 0 to 80 cm depth were sampled for rhizosphere soil. Soil samples were mixed by sex within each plot and stored at −80 °C for analyses of alkaline and acid phosphatase activities, as well as soil P chemical properties. Following harvest, fine root samples were carefully selected and gently rinsed with deionized water to remove any adherent soil particles, ensuring sample integrity. Subsequently, the fresh weight of the root segments was accurately measured. These prepared root samples were then scanned using an Epson V700 scanner. The Win-RHIZO system by Régent Instrument Inc. was utilized to meticulously quantify total root length and volume, providing precise insights into root architecture. SRL was determined by subjecting samples to a rigorous drying process at 70 °C for 48 h. Root tissue density (RTD) was calculated as the ratio of the fresh weight of root segments to their volume.
Leaf analysis
Leaf mass per unit area (LMA) was calculated as oven-dried mass divided by area. Amass was calculated from Aarea and LMA. PPUE was calculated as the Amass divided by the total leaf P concentration. Plant N concentration was determined using a Kjeldahl Nitrogen Analyzer (K1160, Jinan Hanon Instruments Co. Ltd, China). The analyzer was set to a temperature of 950 °C for combustion, followed by a reduction to 640 °C per the method described by Bremner (1996). Leaf Mn2+ concentration analyses were conducted following drying, weighing and homogenization to standardize the samples. Mn2+ concentrations were quantified using an inductively coupled plasma optical emission spectrometer, specifically the Model 5300DV by PerkinElmer.
Determining P fractions in leaves was performed using the methods outlined by Yan et al. (2019). Briefly, leaf material was harvested and freeze-dried at −80 °C for 7 days. Freeze-dried materials were ground to a fine powder and divided into three subsamples. The first subsample was used to determine total P concentration. After digestion in a concentrated HNO3–HClO4 (3:1, v/v) mixture, the P concentration were determined using the molybdenum blue-based method (Ames 1966). The second subsample was used to determine leaf Pi. The subsample was treated with 1 mL of 1% (v/v) acetic acid by mechanical shaking (Precellys 24 Tissue Homogenizer; Bertin Instruments, Montigny-le-Bretonneux, France). Then, the homogenate was centrifuged at 12 000 rpm for 10 min at 4 °C to remove debris. The supernatant was acid digested with 3 mL of HNO3 and 1 mL of HClO4 to determine P concentration using the molybdenum blue-based method.
The third subsample was used to measure leaf Po. In brief, the subsample was extracted three times with 1 mL 12:6:1 chloroform:methanol:formic acid (CMF, v/v/v), followed three times with 1.26 mL 1:2:0.8 chloroform:methanol:water (CMW; v/v/v). The supernatant was transferred to a 10-ml tube, added 1.9 mL 1:9 chloroform:pure water (CWW, v/v), mixed thoroughly and centrifuged at 4000 g at 4 °C for 10 min. The upper and lower liquid was transferred to 10 mL glass flasks separately, labeled [PM] and [PL]. The remaining interfacial layer was rinsed with 1.44 mL of CMF:CMW:CWW (1/1.26/0.62 v/v/v) and the supernatant was transferred again, before transferring the layer to the metabolic P flask. A 1 mL of 85% methanol (v/v) was added to the remaining residue of the subsample, mixed thoroughly and then the supernatant was transferred into the metabolic P flask. The residue was extracted two times with 1 mL 5% trichloroacetic acid (TCA, v/v) in a 4 °C refrigerator for 1 h, mixed thoroughly every 10 min. The supernatant was then transferred into the metabolic P flask too. Finally, the residue of the subsample was extracted three times with 1 mL of 2.5% TCA (v/v) in a 95 °C hot water bath. The supernatant was measured for [PN] and the residue was analyzed for [PR]. All P fractions were evaporated at 45 °C under N2 gas and digested by HNO3–HClO4, then determined P concentration using molybdenum blue-based method. Low molecular mass P metabolites were obtained by subtracting Pi from metabolic P. The total amount of P recovered from the fractions was consistently over 90% of the P concentration directly measured in leaves.
Soil chemical analysis
Soil acid phosphatase (ACP) activity was determined following Xia et al. (2020). Briefly, 0.5 g of rhizosphere soil was mixed with 2 mL deionized water to form a suspension, and 0.5 mL of this suspension was placed into 2 mL Eppendorf tubes. To each tube, 0.1 mL of p-nitrophenylphosphate (pNPP) substrate (Sigma-Aldrich) and 0.4 mL acetate buffer (pH 5.2) were added. The mixture was incubated at 30 °C for 30 min with gentle shaking. The reaction was stopped by adding 0.5 mL of 0.5 mol L−1 NaOH and centrifuged at 12 000 g for 10 min. Controls had NaOH added before incubation. Absorbance was measured at 405 nm using a spectrophotometer. Soil alkaline phosphatase (ALP) activity was determined using the method described by Tabatabai (1994). Briefly, 1 g of fresh soil was placed into a 50-mL Erlenmeyer flask, before adding 4 mL of buffer (pH 11.0), 0.25 mL of toluene and 1 mL of 0.115 mol L−1 pNPP solution. The flask was gently shaken and then incubated at 37 °C for 1 h. Following this, 1 mL of 0.5 mol L−1 calcium chloride and 4 mL of 0.5 mol L−1 sodium hydroxide were sequentially introduced into the flask. After a brief swirling of the flask for a few seconds, the soil suspension was filtered through a folded filter paper (Whatman, No. 12). The absorbance of the resulting solution was measured at 400 nm using a spectrophotometer. Soil P properties were analyzed using the continuous extraction method pioneered by Hedley et al. (1982). The methodology involved adding 0.5 g of air-dried soil to a centrifuge tube and supplemented with 30 mL of distilled water and two 9 mm × 62 mm resin strips. The P in the resin strips was extracted using 0.5 mol L−1 HCl for 16 h, obtaining the resin-Pi fraction. Following removing the aqueous solution, 30 mL of 0.5 mol L−1 NaHCO3 at pH 8.5 was added and shaken for 16 h to extract the NaHCO3-P fraction. Finally, 30 mL of 0.1 mol L−1 NaOH was added to extract the NaOH-P fraction and shaken for 16 h. The resulting P fractions obtained were resin-Pi, NaHCO3-P and NaOH-P. Active [Pi] (Pi: resin-P, NaHCO3-Pi and NaOH-Pi) and active Po (Po: NaHCO3-Po and NaOH-Po) were used for analysis. Soil total P was determined by HClO4 digestion.
Statistical analyses
Before analysis of variance (ANOVA) analysis, we used the Shapiro–Wilk test to determine that the data conformed to a normal distribution. Two-way ANOVAs were utilized to assess the effects of sex identity, water management and their interaction on a range of parameters, including root functional traits, soil P properties, the concentrations and proportions of foliar P fractions and other leaf functional traits. We used a full-factor model and regarded sex and water management as fixed effects in two-way ANOVAs. Subsequent comparative analyses among various treatments for these parameters were conducted utilizing Tukey’s Honest Significant Difference tests following one-way ANOVAs at a significance level of P < 0.05. Before performing the principal coordinate analysis (PCoA) for a comprehensive evaluation, all datasets were normalized to mitigate the impact of scale discrepancies among assorted variables, thereby ensuring analytical precision. The application of PCoA, predicated on the Bray–Curtis dissimilarity index and facilitated by the ‘vegan’ package version 2.6-4 in R, was aimed at elucidating the divergent compositional patterns of foliar P fractions ([Pi], [PM], [PL], [PN] and [PR]) in response to variances in sex identity and water management regimes. In this context, the composite index derived from the principal coordinates along the initial two axes provided a robust framework for subsequent Tukey’s post hoc tests under the one-way ANOVA paradigm to delineate differences between sexes under water management. Additionally, the associations between concentration and allocation proportions of five foliar P fractions were examined through Spearman’s rank correlation coefficients in the ‘Hmisc’ package version 5.1-3. The method was suitable for evaluating a monotonic relationship between two variables. The ‘ggcor’ function in the ‘vegan’ package was performed to analyze and visualize Mantel tests, which could deal with complex data structures in matrices, illustrating correlations among root functional traits, foliar P fractions, soil P properties and other leaf functional traits. All statistical computations were performed by R software Version 4.0.2 (R Core Team 2024).
RESULTS
Soil P properties and root functional traits
Continuous irrigation significantly affected the soil’s physicochemical properties (Table 1). Under water deficiency conditions, we observed large decreases in soil moisture and available P, compared with soil pH, soil organic carbon (SOC), total N concentration, available N and total P concentration (Table 1). The activities of soil ACP and ALP, as well as soil active Pi and active Po in the rhizosphere, were significantly affected by water management practices (P < 0.001; Table 2; Fig. 1). Additionally, an interaction between water management practices and plant sex significantly impacted the concentration of soil active Pi (P < 0.01, Fig. 1a). These parameters mentioned above were significantly reduced under water deficiency conditions (P < 0.05, Table 2; Fig. 1). Under well-watered conditions, females exhibited a 36.5% lower concentration of soil active Pi than males (Fig. 1a) in the rhizosphere, while no sexual differences were observed under water deficiency conditions. In contrast, male plants exhibited significantly higher decreases in soil active Po (50.3%), and significantly increased activities of ACP (38.6%) and ALP (102.9%) than females under water deficiency (Fig. 1b; Table 2).
Soil properties in P. euphratica plantations under different water management practices
| Soil properties | Well watered | Water deficiency |
|---|---|---|
| Soil moisture (%) | 25.3 | 6.9 |
| pH | 8.4 | 8.7 |
| Total N (g kg−1) | 1.2 | 0.8 |
| SOC (g kg−1) | 40.1 | 32.1 |
| NO3-N (mg kg−1) | 6.4 | 4.6 |
| NH4-N (mg kg−1) | 2.2 | 1.3 |
| Total P (g kg−1) | 0.7 | 0.6 |
| Available P (mg kg−1) | 22.2 | 6.6 |
| Soil properties | Well watered | Water deficiency |
|---|---|---|
| Soil moisture (%) | 25.3 | 6.9 |
| pH | 8.4 | 8.7 |
| Total N (g kg−1) | 1.2 | 0.8 |
| SOC (g kg−1) | 40.1 | 32.1 |
| NO3-N (mg kg−1) | 6.4 | 4.6 |
| NH4-N (mg kg−1) | 2.2 | 1.3 |
| Total P (g kg−1) | 0.7 | 0.6 |
| Available P (mg kg−1) | 22.2 | 6.6 |
Soil properties in P. euphratica plantations under different water management practices
| Soil properties | Well watered | Water deficiency |
|---|---|---|
| Soil moisture (%) | 25.3 | 6.9 |
| pH | 8.4 | 8.7 |
| Total N (g kg−1) | 1.2 | 0.8 |
| SOC (g kg−1) | 40.1 | 32.1 |
| NO3-N (mg kg−1) | 6.4 | 4.6 |
| NH4-N (mg kg−1) | 2.2 | 1.3 |
| Total P (g kg−1) | 0.7 | 0.6 |
| Available P (mg kg−1) | 22.2 | 6.6 |
| Soil properties | Well watered | Water deficiency |
|---|---|---|
| Soil moisture (%) | 25.3 | 6.9 |
| pH | 8.4 | 8.7 |
| Total N (g kg−1) | 1.2 | 0.8 |
| SOC (g kg−1) | 40.1 | 32.1 |
| NO3-N (mg kg−1) | 6.4 | 4.6 |
| NH4-N (mg kg−1) | 2.2 | 1.3 |
| Total P (g kg−1) | 0.7 | 0.6 |
| Available P (mg kg−1) | 22.2 | 6.6 |
The activity of ACP and ALP, SRL, RTD and foliar Mn2+ concentration of Populus euphratica females (F) and males (M) under different water management practices
| Water management | Sex | ACP (mg pNP kg−1 h−1) | ALP (mg phenol kg−1 h−1) | SRL (m g−1) | RTD (g cm−3) | Foliar Mn2+ (mg kg−1) |
|---|---|---|---|---|---|---|
| Well watered | F | 44.5 ± 2.0a | 68.8 ± 2.1a | 13.5 ± 1.4b | 0.6 ± 0.1a | 56.7 ± 8.2b |
| Water deficiency | F | 24.6 ± 2.1c | 13.8 ± 1.7c | 19.5 ± 0.9a | 0.3 ± 0.1b | 53.3 ± 6.3b |
| Well watered | M | 41.8 ± 2.8a | 80.4 ± 5.9a | 8.5 ± 0.9c | 0.6 ± 0.1a | 44.9 ± 3.3b |
| Water deficiency | M | 34.1 ± 3.1b | 28.0 ± 1.5b | 12.3 ± 0.8b | 0.5 ± 0.1a | 78.4 ± 8.8a |
| Pwater | <0.001 | <0.001 | <0.001 | <0.05 | <0.05 | |
| Psex | 0.2 | <0.001 | <0.001 | <0.05 | 0.4 | |
| Pwater × sex | <0.05 | 0.7 | 0.3 | 0.3 | <0.05 | |
| Water management | Sex | ACP (mg pNP kg−1 h−1) | ALP (mg phenol kg−1 h−1) | SRL (m g−1) | RTD (g cm−3) | Foliar Mn2+ (mg kg−1) |
|---|---|---|---|---|---|---|
| Well watered | F | 44.5 ± 2.0a | 68.8 ± 2.1a | 13.5 ± 1.4b | 0.6 ± 0.1a | 56.7 ± 8.2b |
| Water deficiency | F | 24.6 ± 2.1c | 13.8 ± 1.7c | 19.5 ± 0.9a | 0.3 ± 0.1b | 53.3 ± 6.3b |
| Well watered | M | 41.8 ± 2.8a | 80.4 ± 5.9a | 8.5 ± 0.9c | 0.6 ± 0.1a | 44.9 ± 3.3b |
| Water deficiency | M | 34.1 ± 3.1b | 28.0 ± 1.5b | 12.3 ± 0.8b | 0.5 ± 0.1a | 78.4 ± 8.8a |
| Pwater | <0.001 | <0.001 | <0.001 | <0.05 | <0.05 | |
| Psex | 0.2 | <0.001 | <0.001 | <0.05 | 0.4 | |
| Pwater × sex | <0.05 | 0.7 | 0.3 | 0.3 | <0.05 | |
Different letters in the column indicate significant differences between treatments (P < 0.05, Tukey’s test). Values are expressed as means ± SE (n = 6). Statistically significant P-values are shown in bold. Abbreviations: Psex = sex effect, Pwater = water management practices, Pwater × sex = the interaction effect of water management practices and sex.
The activity of ACP and ALP, SRL, RTD and foliar Mn2+ concentration of Populus euphratica females (F) and males (M) under different water management practices
| Water management | Sex | ACP (mg pNP kg−1 h−1) | ALP (mg phenol kg−1 h−1) | SRL (m g−1) | RTD (g cm−3) | Foliar Mn2+ (mg kg−1) |
|---|---|---|---|---|---|---|
| Well watered | F | 44.5 ± 2.0a | 68.8 ± 2.1a | 13.5 ± 1.4b | 0.6 ± 0.1a | 56.7 ± 8.2b |
| Water deficiency | F | 24.6 ± 2.1c | 13.8 ± 1.7c | 19.5 ± 0.9a | 0.3 ± 0.1b | 53.3 ± 6.3b |
| Well watered | M | 41.8 ± 2.8a | 80.4 ± 5.9a | 8.5 ± 0.9c | 0.6 ± 0.1a | 44.9 ± 3.3b |
| Water deficiency | M | 34.1 ± 3.1b | 28.0 ± 1.5b | 12.3 ± 0.8b | 0.5 ± 0.1a | 78.4 ± 8.8a |
| Pwater | <0.001 | <0.001 | <0.001 | <0.05 | <0.05 | |
| Psex | 0.2 | <0.001 | <0.001 | <0.05 | 0.4 | |
| Pwater × sex | <0.05 | 0.7 | 0.3 | 0.3 | <0.05 | |
| Water management | Sex | ACP (mg pNP kg−1 h−1) | ALP (mg phenol kg−1 h−1) | SRL (m g−1) | RTD (g cm−3) | Foliar Mn2+ (mg kg−1) |
|---|---|---|---|---|---|---|
| Well watered | F | 44.5 ± 2.0a | 68.8 ± 2.1a | 13.5 ± 1.4b | 0.6 ± 0.1a | 56.7 ± 8.2b |
| Water deficiency | F | 24.6 ± 2.1c | 13.8 ± 1.7c | 19.5 ± 0.9a | 0.3 ± 0.1b | 53.3 ± 6.3b |
| Well watered | M | 41.8 ± 2.8a | 80.4 ± 5.9a | 8.5 ± 0.9c | 0.6 ± 0.1a | 44.9 ± 3.3b |
| Water deficiency | M | 34.1 ± 3.1b | 28.0 ± 1.5b | 12.3 ± 0.8b | 0.5 ± 0.1a | 78.4 ± 8.8a |
| Pwater | <0.001 | <0.001 | <0.001 | <0.05 | <0.05 | |
| Psex | 0.2 | <0.001 | <0.001 | <0.05 | 0.4 | |
| Pwater × sex | <0.05 | 0.7 | 0.3 | 0.3 | <0.05 | |
Different letters in the column indicate significant differences between treatments (P < 0.05, Tukey’s test). Values are expressed as means ± SE (n = 6). Statistically significant P-values are shown in bold. Abbreviations: Psex = sex effect, Pwater = water management practices, Pwater × sex = the interaction effect of water management practices and sex.
Soil active Pi (a) and Po (b) in the rhizosphere of P. euphratica females and males grown under varying water management practices. Values are means ± SE (n = 6). Two-way ANOVA was applied to evaluate the effects of different factors and their interactions. Abbreviations: S = sex effect, W = water management practices, S × W = sex × water management practices. The degrees of freedom are 1 and 20. Different lowercase letters above the bars indicate significant differences among treatments based on Tukey’s post hoc analysis (P < 0.05).
We found that irrigation markedly influences root functional traits (Table 2). The variation in SRL between sexes appeared unaffected by water management practices, with females consistently exhibiting higher SRL values than males (Table 2). Furthermore, both females and males significantly increased SRL, but only males significantly increased foliar Mn2+ concentration when subjected to water deficiency compared with well-watered scenarios (Table 2). RTD and foliar Mn2+ concentration were significantly greater in males than females under water deficiency conditions (Table 2).
Traits related to photosynthesis and total concentrations of foliar nutrient
Water management practices had a significant effect on the PPUE of both sexes (P < 0.001), with females and males showing 48.7% and 24.8% higher PPUE under water deficiency conditions, respectively (P < 0.05, Fig. 2a). Male trees exhibited 43.2% and 20.1% higher PPUE than female trees under well-watered and water deficiency conditions, respectively (P < 0.05, Fig. 2a). The interaction between sex and water management practices significantly affected Amass and Aarea (P < 0.05, P < 0.01, Fig. 2b and c). Under well-watered conditions, there were no significant differences in Amass and Aarea between sexes; however, under water deficiency conditions, male trees exhibited significantly higher Amass and Aarea compared with female trees by 35.7% and 34.6%, respectively (P < 0.05, Fig. 2b and c). The foliar total P concentration in P. euphratica positively correlated with soil moisture levels (R2 = 0.7, P < 0.001, Supplementary Fig. S2). The interaction between sex and water management practices significantly impacted the concentration of foliar total P (P < 0.001, Fig. 2e). Female trees displayed a 66.9% higher concentration of foliar total P under well-watered conditions than males (P < 0.05, Fig. 2e), while under water deficiency conditions, this difference was not significant. LMA, foliar N concentration and N:P ratio were unaffected by sex (Fig. 2d and f), while the foliar N:P ratio varied significantly under different water management practices (P < 0.05, Fig. 2g).
PPUE (a), Aarea (b), Amass (c), LMA (d), foliar phosphorus (P) concentration (e), foliar nitrogen (N) concentration (f) and foliar N:P ratio (g) for P. euphratica females and males grown under different water management practices. Abbreviations: S = sex effect, W = water management practices, S × W = sex × water management practices. The degrees of freedom are 1 and 20. Different lowercase letters above the bars indicate significant differences among treatments based on Tukey’s post hoc analysis (P < 0.05).
Concentration of foliar P fractions and their relative allocation
Water management practices significantly affected the concentration of all five foliar P fractions (Fig. 3a–e). We observed that reduced soil moisture significantly decreased the concentrations of these fractions in both sexes, with females experiencing a larger reduction (66.1% for [Pi], 86.8% for [PM], 58.33% for [PL], 56.0% for [PN] and 56.8% for [PR]) compared with males (38.3% for [Pi], 55.5% for [PM], 24.40% for [PL], 31.0% for [PN] and 20.5% for [PR]) (P < 0.05, Fig. 3a–e). Moreover, the interaction between sex and water management practices significantly affected the concentration of foliar Pi and Po (P < 0.001, Fig. 3a–e). Under well-watered conditions, female trees exhibited significantly higher concentrations across all foliar P fractions compared with their male counterparts, showing increases of 36.3% ([Pi]), 44.0% ([PM]), 39.9% ([PL]), 34.5% ([PN]) and 42.5% ([PR]). In contrast, no sex differences were observed under water deficiency conditions.
Concentrations of foliar phosphorus (P) fractions (a–e) and percentage of each P fraction of the total foliar P concentration (f–j) for P. euphratica females and males grown under different water management. Abbreviations: Pi = orthophosphate P, S = sex effect, W = water management practices, S × W = sex × water management practices. The degrees of freedom are 1 and 20. Different lowercase letters above the bars indicate significant differences among treatments based on Tukey’s post hoc analysis (P < 0.05). ‘r’ represents relative allocation.
While the relative allocation of inorganic P in leaf (rPi) and relative allocation of residual P in leaf (rPR) remained unaffected by water management practices (Fig. 3f and j), we noted distinct trends in other fractions under water deficiency conditions: a significant reduction in relative allocation of metabolic P in leaf (rPM) for females (64.8%) and males (32.7%) and an increase in relative allocation of lipid P in leaf (rPL) for both sexes (12.5% and 11.3%, respectively) (P < 0.05, Fig. 3g and h). A noteworthy sex-specific response was observed in relative allocation of nucleic P in leaf (rPN), with females showing a 19.4% increase under deficiency conditions (P < 0.05, Fig. 3i)—a response not mirrored in males. Remarkably, males exhibited a considerably 72.3% higher rPM under water deficiency conditions than females (P < 0.05, Fig. 3g). Principal coordinates analysis (PCoA) illustrated the impact of water management practices on the variations in concentrations of leaf P fractions, underscoring significant differences between female and male leaves under well-watered conditions (Fig. 4). However, under water deficiency conditions, no sex differences were found (Fig. 4).
PCoA score plots depicting variations in foliar P fractions among P. euphratica females and males grown under different water management practices. Values on PCoA axes indicate the percentages of total variation each axis explains. Scores of the first and second PCoA axes (PCoA 1 and PCoA 2) were used as a combined index to represent the foliar P fraction in one-way ANOVAs. Different lowercase letters indicate significant differences among treatments based on Tukey’s post hoc analysis (P < 0.05).
Relationships between root functional traits, soil P properties, foliar P fractions and other foliar functional traits
A consistent pattern emerged across both females and males, illustrating positive correlations between the concentration of foliar [P] and [Pi] as well as rPM, contrasted by negative correlations with rPR. Furthermore, foliar [PM] concentration was positively associated with [PN] concentration and rPM, while exhibiting negative correlations with rPR (P < 0.05, Fig. 5a and b). Similarly, the concentration of [PL] and [PN] displayed positive correlations with rPM and negative associations with rPR, whereas [PN] concentration also positively correlated with [PR] concentration but negatively with rPL (P < 0.05, Fig. 5a and b). The relative P allocation showed inverse relationships between rPi, rPM, rPN and rPR (P < 0.05, Fig. 5a and b). In females, foliar [P] concentration was positively correlated with concentrations of multiple foliar P fractions and rPi, while showing negative associations with rPL and rPR (P < 0.05, Fig. 5a). Contrarily, in males, foliar [P]’s concentration positive correlations were predominantly with [PN] concentration (P < 0.05, Fig. 5b).
![The relationships among the concentration of foliar P fractions and relative allocation of foliar P fractions in P. euphratica females (a) and males (b). Mantel tests between root functional traits and soil P properties and other foliar functional traits in P. euphratica females (c) and males (d). Gradient color indicates the size of the Spearman correlation coefficient, ranging from −1 to 1. Each circle represents the correlation between the two transverse and longitudinal parameters. Asterisks represent the level of significance. *P < 0.05, **P < 0.01, ***P < 0.001. Abbreviations: foliar [P] = total P concentration in leaf, Mn2+ = foliar Mn2+ concentration, Pi = orthophosphate Pi. ‘r’ represents relative allocation.](https://oup.silverchair-cdn.com/oup/backfile/Content_public/Journal/jpe/17/6/10.1093_jpe_rtae064/1/m_rtae064_fig5.jpeg?Expires=1747935221&Signature=iKc5owk2artjE64i-5RsY58Wv8c70lbFfOkgdrYcgy0r7Pu0sDnnyR1UWZfmX3iU~ZspnXV98LUSAfNVxsraQ7ZH-VTKiBY0RxXn9GLuvQypkcV-VbU43lBXQbM5UQkmf95usErPncswFChR-A1zNG1o7Zx4Ps2Wj9Cnha2EBNDpHDmxSnZSUSVE4JIP1eLLjCF0xPJTcAba8B-wzytzP-cqlCN4PXLW3cGDJBfDXG0czZz53hlhTJON9HibezSfaLAjGWvgUmHyeFG-wy2mSXcaTa2hgitwJnd9A0RYMqoz0KMERCNq9EbKiSq3OMbpJ3GkWaS4P0-qFFLveHIDEw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
The relationships among the concentration of foliar P fractions and relative allocation of foliar P fractions in P. euphratica females (a) and males (b). Mantel tests between root functional traits and soil P properties and other foliar functional traits in P. euphratica females (c) and males (d). Gradient color indicates the size of the Spearman correlation coefficient, ranging from −1 to 1. Each circle represents the correlation between the two transverse and longitudinal parameters. Asterisks represent the level of significance. *P < 0.05, **P < 0.01, ***P < 0.001. Abbreviations: foliar [P] = total P concentration in leaf, Mn2+ = foliar Mn2+ concentration, Pi = orthophosphate Pi. ‘r’ represents relative allocation.
For soil parameters, soil Pi concentration correlated positively with soil active Po concentration, ALP activity, foliar [P] concentration and Aarea, but negatively with SRL, the foliar N:P ratio and PPUE (Fig. 5c and d). Notably, sex-specific patterns were evident: soil active Po concentration negatively correlated with PPUE and LMA in females only, while soil active Pi concentration negative associations with foliar Mn2+ concentration were exclusive to males.
Mantel tests elaborated on the associations between relative foliar P fractions, other foliar functional traits and soil P properties, revealing sex-differentiated responses. For females, significant correlations were found between rPM and a series of parameters, including concentrations of soil active Pi and Po, ACP and ALP activity, foliar N:P ratio, PPUE, Aarea, Amass and LMA (Fig. 5c). In contrast, male-specific correlations for rPM predominantly involved soil active Pi concentration, ALP activity, foliar [P] concentration, Aarea and Amass (Fig. 5d).
DISCUSSION
This study systematically explores the sex-dependent strategies for P acquisition and utilization in P. euphratica under varying soil moisture levels. We identified a trade-off, with females optimizing Pi foraging under well-watered conditions, while males exhibiting enhanced Pi solubilization and Po mineralization under water deficiency conditions. Under high soil moisture levels, female trees exhibited not only increased total leaf P and enhanced concentrations of five specific foliar P fractions but also an associated improvement in net photosynthetic rate. Conversely, in low-moisture scenarios, male trees demonstrated a smaller reduction in concentrations of each foliar P fractions. They effectively mitigated photosynthetic reductions, improving PPUE through an augmented relative allocation to [PM]. Furthermore, the study reveals that leaf P allocation and root functional traits display sex-dependent correlations, likely indicative of intrinsic sexual dimorphisms in soil P acquisition and utilization strategies.
Sexual dimorphism in root P acquisition
Nutrient acquisition and utilization play crucial roles in plant growth, as highlighted by complex physiological and molecular interactions (López-Arredondo et al. 2014; Yuan and Liu 2008). In our study, under high soil moisture levels, the concentration of soil Pi in the rhizosphere of female plants was observed to be lower than in males (Fig. 1a), despite similar activities of ACP and ALP between the sexes (Table 2). These enzymes, ALP and ACP, are known to facilitate the mineralization of soil Po to meet plant P requirements (Dai et al. 2020), suggesting that the observed differences might be attributed to the intrinsic P foraging capabilities of the root systems, rather than P form transformation in the soil. The heightened dependence of female plants on P, possibly due to their higher reproductive costs, is substantiated by their consistently greater SRL across varying soil moisture levels and P availability (Table 2). These morphological traits, beneficial for nutrient uptake in nutrient-rich contexts (Eissenstat and Yanai 1997; Hodge 2006), enhance the interface between roots and soil, enabling more effective water and P acquisition in females and subsequently increased P accumulation in their leaves. However, this morphological specialization might make female plants more vulnerable in P-scarce environments.
Under low soil moisture levels, male plants demonstrate improved activities of ACP and ALP, coupled with higher foliar concentrations of Mn2+ (Table 2). This pattern suggests a robust capability in male plants to attenuate the reduction in soil P availability. Such physiological adaptability indicates compensatory growth, effectively reducing loss linked to diminished photosynthetic rates and concurrently lowering maintenance costs by reducing SRL (Zhao et al. 2024; Zhang et al. 2024; Xia et al. 2020). Such a strategy could also stem from molecular responses of male roots to environmental changes, such as adapting to varying moisture levels and P availability conditions by regulating critical genes involved in root growth and phosphatase expression (Kang et al. 2022). These physiological and molecular adjustments afford male plants longer root lifespans (Weemstra et al. 2016), slower growth rates (Yang et al. 2023) and stronger pathogen resistance (Freschet et al. 2018), providing males with growth and survival advantages in resource-limited environments.
Sexual dimorphism in leaf P utilization
Female plants demonstrate a high sensitivity to variations in soil moisture level compared with male plants. Our study reveals that, under high soil moisture levels, female plants exhibited higher concentrations of foliar P fractions than water deficiency (Fig. 2a–e), which correlates with enhanced area- and mass-based photosynthetic rates (Aarea and Amass), suggesting a potential improvement in energy transformation and the synthesis of photosynthetic products. Such enhancements likely underpin the observed robust growth and developmental vigor in female plants. Diving deeper into the sex-based distinctions in the relative allocation of foliar P fraction, we observed that females preferentially allocate P to nucleic acids under water deficiency (Fig. 2i), a strategy that aligns with their higher reproductive investment and is particularly pronounced under low soil moisture levels. This targeted allocation supports critical reproductive functions, such as rapid protein synthesis and turnover, necessary during stages of fruit and seed formation. Interestingly, analogous studies on Proteaceae species have highlighted a trade-off, showing species with high ribosomal RNA levels in mature leaves often exhibit lower PPUE (Sulpice et al. 2014). Conversely, male plants exhibit a more conservative decrease in the allocation of P to metabolites (Fig. 2g), which is vital for maintaining consistent Aarea, Amass and PPUE. This strategic allocation of P may contribute to relatively stable photosynthesis and superior PPUE (Hidaka and Kitayama 2011, 2013; Netzer et al. 2017).
Implications for sex-specific strategies of P-use and -acquisition
Plants must balance growth and reproduction to adapt to specific environments (Doust 1989; Liu et al. 2021; Obeso 2002). Previous studies have shown that habitats with female-biased sex ratios generally experience less stress (Hultine et al. 2016; Liu et al. 2021). In contrast, males tend to be more abundant than females under adverse environmental conditions (Hultine et al. 2016; Juvany and Munne-Bosch 2015; Liu et al. 2021). The trade-offs or covariation between root functional traits related to nutrient uptake strategies and leaf P fractions may differ based on sex. The correlation between leaf P fraction allocation and root functional traits exhibits a sex-dependent pattern, likely originating from the differential acquisition and utilization of soil P forms by male and female plants. Female plants demonstrate heightened sensitivity in the relative allocation of leaf P fractions, which may be attributed to their higher reproductive costs and dependency on resources. In contrast, male plants exhibit a more diversified strategy for P acquisition, potentially facilitated by enhanced rhizospheric physiological activities, thereby sustaining their performance in resource-constrained environments. Consequently, to harness ecological insights for habitat management, recognizing plant sex-specific nutrient strategies is vital. In restoration, prioritize nutrient-rich soils for females and robust males for challenging sites to balance sex ratios, enhancing ecosystem resilience. During management, monitor sex ratios as stress indicators, employing targeted water and P supplies for females, particularly in reproductive stages. Under climate change, with the intensification of extreme drought events, increasing the proportion of P. euphratica males may enhance below-ground ecological interactions, thereby supporting population sustainability and restoration efforts. These practices can inform sustainable management, ensuring ecosystem stability and biodiversity through nuanced, sex-aware approaches.
CONCLUSIONS
Our findings revealed distinct adaptive strategies between female and male trees, underscoring the complex interplay between plant sex and nutrient management strategies. Male trees responded to low soil water moisture levels by enhancing ACP and ALP activity in their roots, facilitating the conversion of soil active Po into Pi. This adaptation was complemented by a strategic allocation of leaf P toward the [PM] fraction, effectively reducing photosynthetic losses and enhancing PPUE. Conversely, females adopted a different approach under high soil moisture levels, optimizing root absorption of soil Pi. This resulted in a notable decrease in soil Pi within the rhizosphere of female plants, coupled with an increase in total leaf P and each specific foliar P fraction, which likely serves to increase photosynthetic capacity. The associations between below-ground acquisition strategies and allocation patterns of leaf P fractions may differ based on sex. Our study not only highlights the importance of considering sex in ecological and physiological studies of plants but also suggests that dioecious species may exhibit significant differences in nutrient management strategies, potentially leading to sex-specific adaptations to environmental stress.
Supplementary Material
Supplementary material is available at Journal of Plant Ecology online.
Figure S1: Schematic diagram of each sample plot distribution.
Figure S2: Relationships between foliar total P and soil moisture levels in Populus euphratica plantations across two soil water management practices (n = 12).
Table S1: Relationships between five foliar P fractions and two soil moisture levels in Populus euphratica plantations across two soil water management practices (n = 12).
Funding
This work was supported by the Key Laboratory of Humid Subtropical Eco-geographical Process (Fujian Normal University), Ministry of Education and Start-up Foundation for Advanced Talents of Anhui Agricultural University (rc372210).
Authors’ Contributions
Shengwei Si had the main responsibility for data collection, analysis and writing. Yue He contributed to data collection and analysis. Zongpei Li contributed to the interpretation of data and manuscript preparation. Zhichao Xia (the corresponding author) contributed to data analysis, writing and overall responsibility for experimental design and project management.
Conflict of interest statement. The authors declare that they have no conflict of interest.
REFERENCES
Author notes
Shengwei Si and Yue He contribute equally to this work.
