Effects of different karst fissures and rainfall distribution on the biomass, mineral nutrient elements, antioxidant substances, and photosynthesis of two coniferous seedlings

Abstract

Background

Studying the physiological growth status of Pinus yunnanensis Franch and Pinus elliottii Engelm. seedlings under different karst fissure thicknesses and rainfall distributions is of great significance for the management, vegetation restoration, and tree species selection in karst rocky desertification areas. In this study, we used a two-factor block experiment and set different rainfall durations, namely reduced rainfall duration (I_3d), natural rainfall duration (I_6d), and extended rainfall duration (I_9d); Different karst small habitats, i.e., stone-free soil (S₀), less stone and more soil (S_1/4), and half stone and half soil (S_1/2), are simulated at these three levels. Analyze the changes in physiological growth and photosynthetic characteristics in two coniferous seedlings under different treatments with different karst thicknesses.

Results

The results showed that with the increase of karst thickness, the growth volumes of height and diameter of P. yunnanensis seedlings, the biomass of various organs, and the accumulation of K⁺, Ca²⁺, Na⁺, and Mg²⁺ showed a significant change pattern of first increasing and then decreasing (P < 0.05); P. elliottii seedlings show a gradually decreasing trend (except for Ca²⁺). The biomass accumulation of each organ in two coniferous seedlings showed that leaves > stems > roots. The K⁺, Ca²⁺, and Mg²⁺ content in various organs of P. yunnanensis seedlings showed that leaves > roots > stems, while Na⁺ shows the order of roots > leaves > stems. The accumulation of mineral elements in various organs of P. elliottii seedlings is manifested as roots > stems > leaves and the accumulation of mineral elements in both coniferous seedlings is manifested as Ca²⁺ > Mg²⁺ > K⁺ > Na⁺. Root length, root volume, root surface area, root diameter, SOD, POD, SP, photosynthetic pigment content, fluorescence parameters, and gas exchange parameters of P. yunnanensis seedlings gradually increase with the increase of karst thickness (except for the 9-day rainfall duration), while those of P. elliottii seedlings gradually decrease. The light saturation point of P. yunnanensis seedlings is highest under the I_6dS_1/2 treatment, while that of P. elliottii is highest under the I_3dS₀ treatment.

Conclusions

In summary, prolonging rainfall duration has an inhibitory effect on the growth of two types of coniferous seedlings. Increasing karst thickness inhibits the growth of P. elliottii seedlings, and to some extent, promotes the growth and development of P. yunnanensis seedlings. I_6dS_1/4 and I_3dS₀ treatments have the best growth effects on P. yunnanensis and P. elliottii seedlings. Therefore, we give priority to P. yunnanensis as the tree species for vegetation restoration or rocky desertification management in karst areas. Our study reveals the role of limestone-filled different karst fissures in mitigating the effects of drought as “containers” for plant growth. These findings help us understand the response of plants to drought stress and provide valuable insights for vegetation restoration in karst environments affected by global climate change. Therefore, further experiments with various karst fissure sizes are necessary to test the universality of the reactions of various plants under different karst fissures.

Introduction

Karst rocky desertification (KRD) is a major area with a fragile ecology and environment, seriously threatening the sustainable development of society and economy [1,2,3]. According to reports, most of the land in the Mediterranean region is threatened by karst development, and Lebanon is one of the countries severely affected by karst, with karst areas accounting for 70% of its land area [4]. With the development of karst landforms, soil erosion in western Ireland has accelerated, and the loss of water resources has become increasingly barren in the local soil [5]. Research has shown that due to the presence and development of karst, soil erosion in Cuba has exceeded a certain range, with an average variation between 12.3-13.7t ha^− 1a^− 1, and the risk of soil erosion is particularly high, which greatly hinders social and economic development [6]. This is also the case in China, especially in the karst areas of southwestern China. The deterioration caused by land degradation and rocky desertification of the ecological environment is particularly prominent, affecting the survival and development space of residents [7,8,9].

In addition, soil resources in karst areas are extremely scarce, with scarce soil and exposed bedrock embedded and distributed together [10]. Due to the different thicknesses of karst fissure layers, there are also differences in moisture, nutrients, oxygen, and space in the fissure soil, resulting in heterogeneity in the vertical direction [11]. Against the backdrop of global climate change, the karst areas in southwestern China have shown a trend of unchanged total rainfall and a significant decrease in daily rainfall throughout the year [12]. As the intensity of a single rainfall increases, the soil is prone to erosion and soil erosion becomes more severe, exacerbating the vertical heterogeneity of soil niche and further affecting plant growth. Studies have shown that karst cracks store 27% of total precipitation, exceeding the contribution of surface soil (21%) to vegetation [13, 14]. However, soils filled or formed in surface karst cracks can partially reduce soil erosion rates [15], and the presence of cracks enhances surface roughness and soil compaction ability, promoting plant growth [16]. Studies have shown that drought can lead to a corresponding decrease in biomass allocation in the aboveground parts, while the biomass input in the roots will increase accordingly, thereby absorbing more water and nutrients [17]. Some graduate students have also found that under mild water stress, plants increase the proportion of biomass input in photosynthetic components, thereby maintaining their ability to assimilate CO₂ and produce organic matter. Only after a certain degree of drought, plants increase the input of root biomass and reduce the input of aboveground parts to maintain normal plant growth [18]. In addition, rainfall patterns also have an impact on biomass accumulation and distribution. Relevant studies have shown that when rainfall time is prolonged, the accumulation of aboveground biomass in Fraxinus malacophylla H seedlings is inhibited, while the accumulation of underground biomass is promoted to a certain extent, leading to an increase in root biomass accumulation and thus maintaining plant life activities.

Plant roots play an important role in global ecosystems. The root system is an important organ for maintaining vegetation, especially forest growth and development, and connects underground and above-ground ecological processes through the flow of matter and energy [19]. Biomass and productivity are key factors in global and regional carbon cycling [20], especially in fine roots [21]. Roots can protect soil from erosion, and root architecture (root tissue density, root length, root diameter, root surface area, and root volume) effectively reduces soil detachment rate and reduces the erodibility of surface soil in karst mountainous areas [22].

In addition, the uptake of mineral elements (K⁺, Ca²⁺, Na⁺, Mg²⁺) by plants, especially the uptake of root nutrients and water, is affected by environmental stress, leading to delayed plant growth and development [23]. K⁺ has important effects on the activation of enzymes, ion balance, cell turgor pressure, water balance, and carbohydrate transport in plants [24]. Excessive or insufficient Ca²⁺ can affect related physiological and biochemical processes, ultimately leading to plant damage. A study has found that plant leaves develop slowly or curl up at the leaf edges when lacking Ca²⁺ [25]. In karst rocky desertification areas, there are already many carbonates with relatively high Ca²⁺ content, and the Ca²⁺ content varies in different karst habitats. Therefore, exploring the Ca²⁺ content in different karst habitats and precipitation treatments is of great significance. Excessive accumulation of Na⁺ can lead to many secondary effects, such as plant growth restriction, inhibition of leaf photosynthesis, and transpiration [26, 27]. Moreover, excessive Na⁺ content can lead to osmotic stress, ion toxicity, and uneven ion distribution, disrupting plant cell structure, affecting the balanced absorption of mineral nutrients by plant roots, and disrupting the physiological metabolism of plant cells. At the same time, it can also induce the production of a large amount of reactive oxygen species and oxygen free radicals in the plant body, leading to oxidative stress and severely limiting plant growth and development [23]. Mg²⁺is considered to be the most abundant free divalent cation in plant cells, playing an important role in stabilizing cell macromolecular structure, maintaining enzyme activity, balancing intracellular reactive oxygen species and ions, and serving as a ligand and activator for over 300 enzymes [28]. It is also an important component of chlorophyll and plays a crucial role in plant chloroplast photosynthesis [29]. At the same time, plants have evolved various mechanisms and strategies to cope with the damage caused by different environmental stresses, including absorbing water through roots and transporting it to leaves to maintain water conditions, accumulating compatible solutes such as proline (Pro) and mineral nutrients, regulating the toxicity of Na⁺ to plants, and maintaining ion distribution balance [30]. Plants eliminate excess reactive oxygen species and alleviate oxidative stress damage by enhancing the activity of antioxidant enzymes in their bodies, such as superoxide dismutase (SOD) and peroxidase (POD) [31]. It is also possible to enhance plant stress resistance by activating and inducing intracellular defense genes, such as salt over sensitive (SOS) signaling pathways, by upregulating the expression levels of antioxidant enzymes and SOS-related genes [32]. However, the transport system of K⁺ and Na⁺ binding is a key determinant of plant salt tolerance [33]. Na⁺ competes with K⁺ for binding to root cell sites and transport proteins, leading to translocation, deposition, and partitioning within the plant body. Studies have shown that Chenopodium quinoa W. seedlings adopt self-protection mechanisms such as increasing soluble sugar and proline content, enhancing antioxidant enzyme activity, and reducing malondialdehyde (MDA) content to adapt to salt stress [36]. Primula forbesii F. seedlings can resist salt damage by increasing soluble protein, proline content, superoxide dismutase, and peroxidase activity [35]. It is unknown how the two types of coniferous seedlings can resist the stress of different karst fissures through mineral elements and antioxidant systems.

The photosynthetic physiological characteristics of plants can reflect their adaptability to habitat, and studying the light energy utilization efficiency of plants is the key to exploring their productivity [36]. Plants in karstic rocky desertification areas are subjected to a variety of natural and anthropogenic factors, which limit their photosynthetic efficiency. As a result, most plant photosynthetic productivity has not reached the ideal state, which is not conducive to the comprehensive management of rocky desertification. Therefore, exploring the photosynthetic physiological characteristics and rainfall response patterns of plants in different karst habitats is the key to improving plant productivity in rocky desertification areas. In addition, the photosynthetic light response curve of plants reveals the corresponding relationship between net photosynthetic rate and photosynthetically active radiation. The physiological parameters obtained from the photosynthetic light response curve (including apparent quantum efficiency, maximum net photosynthetic rate, light compensation point, light saturation point, and dark respiration rate) can directly or indirectly reflect the physiological and ecological processes of plants [37]. Moreover, the soil thickness in karst areas affects the photosynthetic physiology of plants in terms of soil moisture, nutrients, and underground space availability [38], which in turn affects plant growth. Stomata are physiologically important as they act as water pathways in the physiological processes of photosynthesis, respiration, and transpiration. The photosynthesis of plants is significantly affected by water, and although the physiological effects of water deficiency are well documented, it remains a highly important issue in the case of imbalanced precipitation distribution and different karst fissures.

Pinus yunnanensis Franch is a unique species in southwest China, a pioneer tree species for natural regeneration of barren mountains in the Yunnan-Guizhou Plateau, and a major commercial tree species in Yunnan Province of China, accounting for about 52% of the forest area in Yunnan Province. P. yunnanensis is crucial for the economic and environmental sustainability of Yunnan forestry. It is widely distributed in the rocky desertification areas of eastern Yunnan, China, and has the characteristics of fast growth and drought resistance. It is a pioneer tree species for afforestation in barren mountains in the southwestern karst region. Pinus elliottii Engelm originally from the United States, has the advantages of a fast growth rate, high-fat production, and strong adaptability. It is an excellent timber and landscaping tree species. It was introduced to China in the 1930s and has now become one of the main afforestation tree species for greening barren mountains and ecological protection in 15 provinces in southern China [39]. Therefore, based on the fact that karst rocky desertification areas have large karst fissures and are relatively arid. How do P. yunnanensis seedlings grow as local tree species in such harsh environments? How to adapt to the growth of the region through its physiological regulatory mechanisms is not yet clear. As an exotic tree species, it is unknown whether and how P. elliottii grows normally in rocky desertification areas. The purpose of this study is to (1) elucidate how the biomass of various organs, root characteristics, mineral element accumulation, antioxidants, and photosynthesis of two coniferous seedlings in different karst habitats. (2) Reveal the trade-off strategies for the growth of each organ of two coniferous seedlings under this stress condition. Therefore, this article studied the physiological growth status of two coniferous seedlings under different degrees of rocky desertification, providing a theoretical reference for the selection of tree species for vegetation restoration in rocky desertification areas. In addition, the impact of rainfall duration combined with different degrees of rocky desertification on two coniferous tree species was also introduced, further improving the efficiency of rocky desertification management.

Materials and methods

Research site and planting materials

The research was conducted in a greenhouse at Southwest Forestry University in China (Kunming, Yunnan, 25°03′N、102°46′E). The research area is located in the monsoon climate zone of the subtropical plateau, with an average altitude of 1954 m, few frost periods, and a warm climate; The annual average temperature is 16.5 ℃, the annual precipitation is 1035 mm, the relative humidity of the air is 23%~67%, the atmospheric CO₂ concentration is 400–412 ppm, and there is sufficient light. The plant materials used are 2-year-old seedlings of Pinus yunnanensis Franch. and Pinus elliottii Engelm. provided by the state-owned Huayuan Forest Farm in Yiliang County, Yunnan Province, China. The tested soil and rocks are red soil and karst limestone, respectively, both taken from Jianshui County, Yunnan Province, China, which has typical karst landforms. The physical and chemical properties of red soil are shown in Table 1.

Table 1 Physical and chemical properties of the tested soil

Experimental design

This study used a two-factor randomized block experiment, setting two factors: karst habitat and rainfall duration. The two-factor randomized block experimental design is an experimental design method that randomly assigns experimental subjects to different treatment groups to control experimental errors and reduce the impact of experimental errors. At the same time, this experimental design method can also determine the degree of influence of different factors on the experimental results by comparing different treatment groups, thereby providing a reference for subsequent experimental research. Through pre-experiments and referencing the design of Wang et al. [40], we divided the treatment of karst small habitats into three levels: stone-free soil (S₀), less stone and more soil (S_1/4), and half stone and half soil (S_1/2), to simulate these three levels of karst habitats; The container ensures that the total volume of each habitat is the same (upper diameter 30 cm, height 20 cm). S₀ habitat is the whole soil. On the top 3/4 of the S_1/4 habitat is the soil layer, and the bottom 1/4 is the karst fissure layer. The upper part half of the S_1/2 habitat is the soil layer, and the lower part half is the karst fissure layer. According to Liu [41], the annual average number of consecutive days without effective precipitation in Yunnan in the southwestern region is about 6 days. The processing of rainfall time intervals is divided into 3-day rainfall intervals (I_3d), 6-day rainfall intervals (I_6d), and 9-day rainfall intervals (I_9d) to simulate three levels of rainfall allocation. According to the average rainfall of 123.58 mm in Kunming City, China from June to December [42], the irrigation amount was calculated based on natural rainfall and rainfall distribution, and the total rainfall of the three rainfall intervals was kept consistent, thus achieving simulation of total rainfall and rainfall distribution (Table 2). There are 9 treatments for each tree species, totaling 18 treatments. Each treatment is repeated 3 times, with 10 seedlings per replicate, resulting in a total of 540 coniferous seedlings. The experiment was conducted in April 2023 by transplanting seedlings into containers, with one seedling per pot. The seedlings were subjected to seedling refining treatment from April to May, and the experimental research was conducted from June to the end of December 2023.

Table 2 Different rainfall allocation

Sample determination and methods

Plant sampling

On December 22nd, 6 seedlings were randomly selected from each treatment for destructive sampling. Clean the roots with clean water and absorb moisture with filter paper. Place the roots, stems, and leaves of the same plant in envelope bags and mark them accordingly. The roots, stems and leaves were put into an oven with a temperature of 105 ℃ for sterilization for 30 min, and then adjusted to 80 ℃ for drying to constant weight. The biomass (g) of the roots, stems, and leaves of two coniferous seedlings was obtained by weighing each organ.

Growth rate

At the beginning of the experiment, ground diameter (mm) was determined with a vernier caliper, and seedling height (cm) was determined with a straightedge for the 2 conifer seedlings in each treatment. At the end of the experiment, the height increment of each treated seedling was measured to calculate the increment of seedling height = measured height - initial height, and the increment of ground diameter = measured ground diameter - initial ground diameter, root shoot ratio = root biomass/aboveground biomass. The sum of the biomass of each organ is the total biomass, and the proportion of root, stem, and leaf biomass is calculated based on the biomass of each organ and the total biomass.

Root morphology characteristics

Immediately use the EPSON Scan root scanner to scan the cleaned roots, and then use WinRHIZO analysis software to obtain the morphological characteristics of the roots, including total root length (RL), root surface area (RS), root diameter (RD), and root volume (RV). Furthermore, the specific root length (cm/g) = root length/root biomass, specific root area (cm²/g) = root surface area/root biomass, and root tissue density (g/cm³) = root biomass/root volume were calculated [43].

Physiological indicators

After weighing, the plant samples were crushed using a grinder and sieved through a 1.0 mm sieve. An accurate 0.2 g of the sample was weighed and digested with H₂SO₄-H₂O₂. The K⁺, Ca²⁺, Na⁺, and Mg²⁺contents in each organ were determined using plasma chromatography [44]. The antioxidant system, including soluble protein (SP), peroxidase (POD), superoxide dismutase (SOD), malondialdehyde (MDA), and proline (Pro), was determined according to the steps of the Suzhou Greese Biotechnology Co., Ltd. reagent kit.

Photosynthetic pigments and fluorescence parameters

Measurement of photosynthetic pigment content (mg/g): Take 0.2 g of ground leaf powder, put it into a mortar, add a small amount of SiO₂, and then add 2–3 mL of 95% ethanol. Grind the homogenate and make up to 25 mL with 95% ethanol [45]. Pour the pigment extraction solution into a colorimetric dish and measure the absorbance values at wavelengths 665, 649, and 470 nm, using 95% ethanol as the control. Calculate the chlorophyll a (Chla), chlorophyll b (Chlb), carotenoids (Car), chlorophyll a + b, and chlorophyll a/b of the leaves [45].

Chlorophyll fluorescence parameter determination: Select the coniferous leaves under each growth point of Pinus yunnanensis Franch and Pinus elliottii Engelm, and after dark adaptation for 30 min, use the LI-6800 portable photosynthetic analyzer to obtain the initial fluorescence value (Fo), maximum fluorescence value (Fm), and variable fluorescence (Fv) under dark conditions; Chlorophyll fluorescence parameters include maximum fluorescence value (Fm’) under light conditions, maximum photochemical quantum efficiency (Fv/Fm) of PSII under dark adaptation, the potential activity of PSII (Fv/Fo), excitation energy capture efficiency of PSII reaction center (Fv ‘/Fm’), non-photochemical quenching coefficient (NPQ), photochemical quenching coefficient (qP), and electron transfer rate (ETR).

Photosynthetic gas exchange and response curve

The photosynthetic indexes of P. yunnanensis and P. elliottii needles were measured by Li-6800 portable photosynthesis tester (LI-COR, USA) in each treatment.

Measurement of photosynthetic gas exchange parameters: from 9:00 am to 11:30 am on a sunny day at a light intensity of 1800µmol·m^− 2·s^− 1, CO₂ concentration of 400µmol·mol^− 1. Measure the net photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), and intercellular CO₂ concentration (Ci) of P. yunnanensis and P. elliottii needles under different treatments at a temperature of 25 ℃.

Measurement of light response curve: At 9:00–11:30, set the light intensity gradient to be 1800, 1600, 1400, 1200, 1000, 800, 600, 400, 300, 200, 150, 100, 50, 30, and 0µmol·m^− 2·s^− 1 in sequence, CO₂ concentration set at 400µmol·mol^− 1.

Determination of CO₂ response curve: CO₂ concentration gradients are set at 400, 300, 200, 100, 50, 20, 10, 400, 400, 600, 800, 1000, 1200, 1400, 1600, and 1800µmol·mol^− 1, PAR value set to 1800µmol·m^− 2·s^− 1.

Referring to Ye [46]analysis, two types of needle parameters were obtained: apparent quantum efficiency (AQY), maximum net photosynthetic rate under light (L_Pnmax), light saturation point (LSP), and light compensation point (LCP); Maximum net photosynthetic rate (Pnmax) and carboxylation efficiency(α)、Dark respiration rate (Rd), photorespiration rate (Rp), CO₂ saturation point (CSP), CO₂ compensation point (CCP). In the setting of Li-6800 environmental parameters, such as the setting of light intensity, we set the corresponding light intensity based on the maximum light intensity of plants through pre-experiments. The setting of parameters such as temperature, humidity, and carbon dioxide concentration is based on the instructions for using Li-6800. Fluorescence parameters and photosynthetic curves can be simultaneously measured for relevant indicators using Li-6800.

Statistical analysis

Utilizing Amos 23.0 Construct a structural equation model to explore the degree of influence between physiological growth and photosynthetic characteristics of two types of coniferous seedlings. A structural equation model (SEM) is a method for establishing, estimating, and testing causal relationship models, which combines path analysis and confirmatory factor analysis. It can study both observable variables and unobservable variables (latent variables); It can also study the direct effects between variables and the indirect effects between variables; And allow for measurement errors in each variable. Organize and calculate data using Excel 2010, and perform multiple comparisons and two-factor analysis of variance using SPSS 25.0 (SPSS Inc., Chicago, IL, USA). Origin 2021.0 (OriginLab Co., Northampton, MA, USA) and conducted graphical statistical analysis. Fuzzy comprehensive evaluation analysis is conducted to determine which treatment yields the best results for two coniferous seedlings.

Results

Growth and biomass

Growth rate

According to Table 3, there were no significant differences (P > 0.05) in seedling height, diameter, and needle length between P. yunnanensis and P. elliottii under different karst habitats and rainfall distribution before the experiment began, indicating that there was not much difference in growth among all seedlings before the experiment began. However, in terms of seedling height and needle length, P. elliottii is significantly higher than P. yunnanensis, while the ground diameter is lower than P. yunnanensis. From Fig. 1, there was a significant difference in needle length between the two types of coniferous seedlings under the interaction between rainfall duration and karst habitat, except for the needle length of P. elliottii, while the other growth rates show significant differences (P < 0.05) under different rainfall duration, karst habitat, and their interaction. Specifically, under the same rainfall duration, the growth of P. yunnanensis shows a significant pattern of first increasing and then decreasing with the increase of karst cracks, reaching its maximum in the S_1/4 karst habitat. However, extending the rainfall duration (I_9d) has a certain inhibitory effect on the growth of P. yunnanensis. Under I_3d treatment, the height growth of seedlings under the S_1/4 karst fissure reached 9.105 cm, the ground diameter increased by 15.530 mm, and the needle length increased the fastest (21.958 cm). Under the I_6d treatment, the growth rate of seedling height under the S_1/4 karst fissure was 1.631 times that of the S₀ habitat, with a ground diameter of 1.307 times and a needle length of 1.100 times. Under the I_9d treatment, the height of seedlings under the S_1/4 karst fissure increased by 0.561 cm compared to S₀, with a ground diameter of 0.902 mm and a needle length of 1.633 cm. However, the performance of P. elliottii is not entirely the same. The growth of P. elliottii significantly decreases with the increase of karst cracks under the same rainfall duration (P < 0.05), and also decreases with the increase of rainfall duration. The growth rates of seedling height, ground diameter, and needle length were highest in the 3d rainfall duration and the whole soil habitat, with values of 36.117 cm, 10.853 mm, and 12.267 cm, respectively. The growth of P. elliottii is significantly inhibited by extending the duration of rainfall.

Table 3 Initial growth of P. Yunnanensis and P. Elliottii

Fig. 1

Effects of different karst habitats and rainfall distribution on the growth of two coniferous seedlings. Note Different uppercase letters in the figure indicate significant differences (P < 0.05) between different karst habitats under the same rainfall interval duration, while different lowercase letters indicate significant differences (P < 0.05) between different rainfall intervals under the same karst habitat

Biomass

As shown in Fig. 2, the biomass of various organs and aboveground biomass of P. yunnanensis seedlings showed significant differences (P < 0.01) under different karst habitats, rainfall distribution, and their interaction, while the root shoot ratio did not show significant differences under the interaction of karst habitats and rainfall distribution. In addition to the root biomass and root shoot ratio of P. elliottii, there are substantial differences in stem, leaf, and total biomass under different karst habitats, rainfall distribution, and their interactions. Under the duration of 3d rainfall, the biomass accumulation of roots, stems, and leaves of P. yunnanensis seedlings increased by 4.718 g, 3.517 g, and 5.823 g in the S_1/4 karst habitat treatment compared with the S₀ karst habitat, respectively. Under 6 days of rainfall, the biomass of roots, stems, and leaves was highest under the S_1/4 karst habitat treatment, with values of 14.167 g, 15.249 g, and 21.858 g, respectively. Under 9 days of rainfall, the biomass accumulation of roots, stems, and leaves increased by 6.634%, 9.065%, and 1.520% in the S_1/4 karst habitat treatment compared with the S₀ karst habitat, respectively. Under the same rainfall duration, the root-shoot ratio of P. yunnanensis shows a trend of first increasing and then decreasing with the deepening of karst fissures (Fig. 2-E). However, under the same rainfall duration, the biomass of various organs and total biomass of P. elliottii show a significant decrease with the deepening of karst cracks; under the same karst habitat, there is a substantial decrease in biomass with the extension of rainfall duration. The increase in rainfall duration significantly inhibits the accumulation of biomass in various organs, while the decrease in rainfall duration significantly promotes the accumulation of biomass in P. elliottii. Under the S_1/4 treatment, the root-shoot ratio of P. elliottii seedlings was the highest, indicating that P. elliottii allocates more biomass to roots for the normal growth of seedlings, reflecting the growth strategy of seedlings (Fig. 2-E). In the S₀ karst habitat, the biomass of roots, stems, and leaves during 9d rainfall are 3.112 g, 1.603 g, and 9.369 g lower than the 6-day rainfall duration, respectively. In the S_1/4 karst habitat, the biomass of the root, stem, and leaf decreased by 36.176%, 22.639%, and 141.954% in the 9-day rainfall duration compared to 6 days of rainfall, respectively. In the S_1/2 karst habitat, the biomass of roots, stems, and leaves was the lowest after 9 days of rainfall, with values of 7.297 g, 10.314 g, and 5.268 g, respectively. In conclusion, S_1/4 karst fissures (mild rocky desertification) promote the accumulation of biomass in all organs of P. yunnanensis, while the accumulation of biomass in all organs of P. elliottii is inhibited by karst fissures.

Fig. 2

Effects of different karst habitats and rainfall distribution on the biomass of two coniferous seedlings. Note Different uppercase letters in the figure indicate significant differences (P < 0.05) between different karst habitats under the same rainfall interval duration, while different lowercase letters indicate significant differences (P < 0.05) between different rainfall intervals under the same karst habitat. Same below

Root system characteristics

As shown in Fig. 3, the root morphology of the two coniferous seedlings showed significant changes (P < 0.05) under different karst fissures and rainfall durations. For P. yunnanensis seedlings, there were significant differences in root morphology between karst cracks and rainfall duration treatments (P < 0.01), while only root length had no significant difference under their interaction (P > 0.05). The root length, root surface area, root volume, and root diameter of P. yunnanensis all increase with the deepening of karst fissures under the same rainfall duration, and the best effect is achieved after 6 days of rainfall, with values of 3557.752 cm, 3964.692cm², 552.708cm³, and 4.590 mm, respectively. Specifically, the root length, root surface area, root volume, and root diameter of P. yunnanensis are 1.856 times, 2.880 times, 4.369 times, and 2.461 times higher in S_1/2 karst fractures than in S₀ karst habitats, respectively, under a rainfall duration of 3 days. Under a rainfall duration of 6 days, the root length, root surface area, root volume, and root diameter of P. yunnanensis seedlings increased with the increase of karst habitat, while the specific root length and specific root area showed a trend of first decreasing and then increasing. However, the root tissue density showed a decreasing trend. However, root length, root surface area, root volume, and root diameter were inhibited under a rainfall duration of 9 days, but they also increased with the increase of karst fractures. For P. elliottii seedlings, there were significant differences in root morphology in karst fissures and rainfall duration treatment (P < 0.01), and there were significant differences in other indicators except for root surface area and specific root area under the interaction of both treatments (P < 0.05). The root characteristics decrease with the deepening of karst cracks under the same rainfall duration, and the best rainfall effect of 3d is observed under each karst crack. Specifically, root length, root surface area, root volume, and root diameter of P. elliottii increased by 19.622%, 48.207%, 120.599%, and 92.373%, respectively, under all-soil habitats (S₀) for 3d rainfall duration compared to 6d rainfall duration; However, the root length, root surface area, root volume, and root diameter were 434.044 cm, 773.848cm², 159.699cm³, and 1.066 mm lower under a 9-day rainfall duration than under a 6-day rainfall duration, respectively. In summary, increasing the thickness of karst cracks promotes the growth of root characteristics in P. yunnanensis seedlings, which also indicates that P. yunnanensis can adapt to certain karst habitats. However, prolonged rainfall duration (9 days) or high-frequency rainfall (3 days) are not conducive to the growth of P. yunnanensis root systems. However, the root system characteristics of P. elliottii are radically different. Deepening karst cracks have a significant inhibitory effect on root characteristics, and high-frequency rainfall has the best effect on root growth.

Fig. 3

Effects of different karst habitats and rainfall distribution on the root morphology of two coniferous seedlings

Tissue element contents

As shown in Fig. 4, the accumulation of mineral elements in different organs of the two coniferous seedlings showed significant changes (P < 0.05) under different karst fissures and rainfall durations. For P. yunnanensis seedlings, the content of K⁺, Ca²⁺, Na⁺, and Mg²⁺ in various organs showed extremely significant differences (P < 0.01) under different karst fissures and rainfall durations. Under the interaction of the two, except for significant differences in leaf K⁺, Ca²⁺, and Mg²⁺ content, there was no significant difference in the accumulation of other contents (P > 0.05). Under the same rainfall duration, the accumulation of mineral element content in various organs shows a significant change pattern of first increasing and then decreasing with the deepening of karst fractures (P < 0.05). Under the same karst fissure, the accumulation of mineral element content in various organs also shows a significant change pattern of first increasing and then decreasing with the increase of rainfall duration (P < 0.05) (except for the K⁺, Ca²⁺, and Mg²⁺ content in leaves in the S_1/4 karst fissure). The values of K⁺/Na⁺ in roots, stems, and leaves varied between 1.522 ~ 2.252, 4.100 ~ 11.579, and 7.045 ~ 20.716 under different treatments, respectively. The K⁺/Na⁺ values in stems and leaves were relatively high, indicating that P. yunnanensis seedlings have a certain salt tolerance. The accumulation of K⁺, Ca²⁺, and Mg²⁺ content in various organs is as follows: leaves > roots > stems, while Na⁺ is as follows: roots > leaves > stems. For P. elliottii seedlings, under the same rainfall duration, the content of K⁺, Na⁺, and Mg²⁺ in each organ showed a significant decrease with the deepening of karst cracks (P < 0.05), while the content of Ca²⁺ showed a changing pattern of first increasing and then decreasing, and the effect was best under a 3-day rainfall duration. The K⁺/Na⁺ values in the roots were highest in the karst habitat of S_1/2 and rainfall duration of 6 days (6.330), while the K⁺/Na⁺ values in the stems and leaves were highest in the habitat of S₀ with a rainfall duration of 9 days, with values of 7.727 and 9.979, respectively. Moreover, the accumulation of K⁺, Ca²⁺, Na⁺, and Mg²⁺ content in various organs is manifested as root > stem > leaf, with mineral element content more inclined towards root accumulation. In summary, the accumulation of mineral elements in various organs of the two types of coniferous seedlings showed a trend of Ca²⁺ > Mg²⁺ > K⁺ > Na⁺. The overall accumulation of mineral elements in various organs of P. yunnanensis seedlings was higher than that of P. elliottii.

Fig. 4

Effects of different karst habitats and rainfall distribution on mineral elements in two coniferous seedlings. Note The bar graph in the figure represents the changes in mineral element content in P. yunnanensis seedlings, while the line chart represents the changes in P. elliottii seedlings. The significant difference in the upper left corner represents P. yunnanensis, and the one in the upper right corner represents P. elliottii

Antioxidant system

Figure 5 shows significant differences (P < 0.05) in SOD, POD, SP, MDA, and Pro of P. yunnanensis and P. elliottii under rainfall duration, karst habitat, and their interaction. However, there was no significant difference (P > 0.05) in the SOD of P. elliottii under their interaction. For P. yunnanensis seedlings, under the same rainfall duration, the activities of SOD and POD antioxidant enzymes and soluble protein content significantly increased with the deepening of karst cracks (P < 0.05). Under a rainfall duration of 3 days, SOD, POD, and SP all reached their maximum values in the S_1/2 karst habitat, with values of 821.938 U·g^− 1, 146.833 U·g^− 1, and 54.057 mg·g^− 1, respectively. Under 6 days of rainfall, the activities of SOD, POD, and SP in the S_1/2 karst habitat were significantly higher than those in the S0 habitat by 8.325%, 31.684%, and 82.289%. Under a 9-day rainfall duration, the activities of SOD, POD, and SP were the lowest in the S_1/2 karst habitat. Both MDA and Pro activities were highest under the 9-day rainfall duration S_1/2 karst habitat treatment, with values of 19.309 U·g^− 1 and 61.395 U·g^− 1, respectively, indicating that the structure and function of the biofilm of P. yunnanensis seedlings were damaged under this treatment. For P. elliottii seedlings, under different treatments, the activities of SOD, POD, and SP significantly decreased with the increase of karst cracks and rainfall duration (P < 0.01). All indicators were highest under the S₀ treatment with a rainfall duration of 3 days, with values of 434.394 U·g^− 1, 146.833 U·g^− 1, and 62.450 mg·g^− 1, respectively. Both MDA and Pro activities were highest under the S_1/2 karst habitat treatment with 9 days of rainfall, with values of 26.239 U·g^− 1 and 71.799 U·g^− 1, respectively. This value is higher than that of P. yunnanensis seedlings, indicating that the structure and function of the biofilm in P. elliottii seedlings have been severely damaged. However, the levels of SOD, POD, and SP in Pinus elliottii E. seedlings are lower than those in P. yunnanensis seedlings, indicating that P. elliottii seedlings have poorer ability to resist karst cracks and rainfall duration compared to P. yunnanensis In summary, P elliottii has greater biofilm destruction than P. yunnanensis, and its antioxidant enzyme activity is lower than P. yunnanensis.

Fig. 5

Effects of different karst habitats and rainfall distribution on the activity enzymes of two coniferous seedlings. Note A scale above 0 represents the enzyme activity of P. yunnanensis, while a scale below 0 represents the enzyme activity of P. elliottii

Photosynthetic pigments and fluorescence parameters

Photosynthetic pigment content

As shown in Fig. 6, the significant differences in photosynthetic pigment content between the two coniferous seedlings were different for different karst cracks and rainfall duration (P < 0.05). For P. yunnanensis seedlings, the content of Chla, Chlb, Car, and Chla + b showed extremely significant differences (P < 0.01) under different karst fissures and rainfall durations, while there was no significant difference (P < 0.05) under the interaction of the two. Under the same rainfall duration, the content of photosynthetic pigments increases with the deepening of karst cracks. Increasing karst cracks can to some extent promote the synthesis of photosynthetic pigments. The content of Chla, Chlb, Car, and Chla + b reaches their maximum under the S_1/2 karst habitat treatment with a rainfall duration of 6 days, with values of 1.043mg·g^− 1, 0.302mg·g^− 1, 0.208mg·g^− 1, and 1.345mg·g^− 1, respectively. For P. elliottii seedlings, there were significant differences in photosynthetic pigment content (P < 0.05) under different karst cracks, rainfall duration, and their interaction. Under the same rainfall duration, the content of photosynthetic pigments shows a significant trend of first increasing and then decreasing with the deepening of karst cracks. S_1/4 karst crack treatment has a certain promoting effect on the synthesis of photosynthetic pigment content in P. elliottii seedlings while prolonging rainfall duration inhibits the synthesis of photosynthetic pigments. Under different treatments, the Chlb synthesis of the two coniferous seedlings is the same, but the synthesis of Car in P. elliottii is significantly higher than that in P. yunnanensis (Fig. 6-C). In summary, mild (S_1/4) and moderate rocky desertification (S_1/2) are beneficial for the synthesis of photosynthetic pigments in P. yunnanensis, while the photosynthetic pigment content in P. elliottii increases under mild rocky desertification (S_1/4) treatment, while it decreases under moderate rocky desertification (S_1/2) treatment.

Fig. 6

Effects of different karst habitats and rainfall distribution on photosynthetic pigments of two coniferous seedlings

Chlorophyll fluorescence parameters

The effects of karst cracks and rainfall duration on the fluorescence parameters of two types of coniferous seedlings are shown in Fig. 7. For P. yunnanensis seedlings, different karst cracks, rainfall duration, and their interaction have a significant limit effect on all indicators (P < 0.01). Under the rainfall duration of 3d and 6d, Fm, Fv, Fo/Fv, Fv/Fm, Fv ‘/Fm’, qP, ETR, and NPQ all significantly increased with the deepening of karst fissures (P < 0.05), indicating that the photosynthetic system of P. yunnanensis seedlings was less or not damaged with the deepening of karst fissures during 3d and 6d rainfall. The increase in their values also indicates that the photosynthetic capacity of plants is enhanced, utilizing their photosynthesis. On the contrary, under the 9-day rainfall duration, these indicators showed a downward trend, indicating that the treatment of karst cracks during the 9-day rainfall duration is not conducive to the improvement of plant photosynthetic capacity. For P. elliottii seedlings, under the same rainfall duration, Fo/Fv, Fv/Fm, Fv ‘/Fm’, qP, ETR, and NPQ show a significant decreasing trend with the increase of karst fissures. Under the full soil (S₀) treatment, the 3d rainfall duration P. elliottii, Fo/Fv, Fv/Fm, Fv ‘/Fm’, qP, ETR, and NPQ increased by 208.498%, 43.169%, 55.692%, 53.315%, 9.081%, and 90.092%, respectively, compared to the 6-day rainfall duration; The duration of rainfall for 9 days is significantly lower than that for 6 days, with values of 0.552, 0.221, 0.274, 0.240, 30.092, and 0.261 respectively. In summary, under normal rainfall duration (6d) and reduced rainfall duration (3d), the increase of karst cracks to some extent promotes the photosynthesis of P. yunnanensis seedlings, thereby enabling the plant to produce more organic matter to provide energy for its growth and development. However, the photosynthetic system of P. elliottii seedlings is relatively complete in the whole soil habitat with a duration of 3d rainfall, and plant photosynthesis is the strongest, synthesizing the most organic matter. Therefore, reducing the duration of rainfall is beneficial for the growth of P. elliottii seedlings. Extending the duration of rainfall has an inhibitory effect on the growth and development of P. yunnanensis and P.elliottii.

Fig. 7

Effects of different karst fissures and rainfall distribution on fluorescence parameters of two coniferous seedlings

Photosynthetic characteristics

Gas exchange parameters

According to Fig. 8, the changes in photosynthetic gas parameters of the two types of coniferous seedlings are different under different treatments. As for P. yunnanensis seedlings, there were significant differences in Tr, Pn, Ci, and Gs under different karst cracks, rainfall duration, and their interaction (P < 0.01). Under the rainfall duration of 3 days and 6 days, Tr, Pn, and Gs all significantly increased with the deepening of karst fractures. Under the rainfall duration of 9 days, they showed a significant change pattern of first increasing and then decreasing (P < 0.05). At the rainfall duration of 6 days and S_1/2 karst fractures, Tr, Pn, and Gs reached their maximum values, which were 0.335 mol·m⁻²·s⁻¹, 14.358µmol·m⁻²·s⁻¹, and 0.273 mol·m⁻²·s⁻¹, respectively. However, the concentration of Ci does not show different patterns. Under the rainfall duration of 3 and 6 days, the concentration of Ci shows a significant downward trend with the deepening of karst cracks, and under the rainfall duration of 9 days, it shows a trend of first decreasing and then increasing. This phenomenon indicates that P. yunnanensis seedlings consume the most CO₂ concentration during photosynthesis, resulting in the highest Pn. As for P. elliottii seedlings, Tr, Pn, Ci, and Gs showed extremely significant differences (P < 0.01) under different karst fissures and rainfall duration treatments, with only Tr and Pn showing extremely significant differences in their interaction (P < 0.01). Under the same rainfall duration, the values of Tr, Pn, and Gs decrease with the increase of karst fractures, while the performance of Ci is exactly the opposite. In the whole soil (S₀) habitat, the concentrations of Tr, Pn, and Gs in P. elliottii seedlings were higher in 3d rainfall than in 6d by 0.136 mol·m⁻²·s⁻¹, 0.439µmol·m⁻²·s⁻¹, and 0.110 mol·m⁻²·s⁻¹. Under mild rocky desertification (S_1/4), the concentrations of Tr, Pn, and Gs were 167.925%, 35.636%, and 53.731% higher under 3d rainfall than 6d. Under moderate rocky desertification (S_1/2), the concentrations of Tr, Pn, and Gs were the lowest 6 days of rainfall, with values of 0.032 mol·m⁻²·s⁻¹, 2.942µmol·m⁻²·s⁻¹, and 0.024 mol·m⁻²·s⁻¹. Research has shown that increasing the consumption of Ci concentration in P. yunnanensis seedlings through karst cracks can enhance the photosynthetic capacity of plants and promote the synthesis of Pn, Tr, and Gs. Reducing rainfall duration promotes the synthesis of Pn, Tr, and Gs in P. elliottii seedlings, thereby improving the photosynthetic capacity of plants, increasing karst cracks, inhibiting photosynthesis in P. elliottii seedlings, and promoting the accumulation of Ci concentration.

Fig. 8

Effects of different karst fissures and rainfall distribution on photosynthetic gas exchange parameters of two coniferous seedlings

Pn-PAR and Pn-CO2 curves

From Fig. 9; Table 4, it can be seen that there are differences in the Pn-PAR response curves of two types of coniferous seedlings under different treatments. For P. yunnanensis seedlings, the fitting degree of Pn-PAR response curves varies between 0.9136 and 0.9949 under each treatment. The fitting degrees of I_6dS_1/4 and I_6dS_1/2 treatments are the highest, both reaching above 0.99. Under 9 treatments, with the increase of PAR, the Pn of P. yunnanensis seedlings showed a trend of first increasing and then stabilizing, with PAR ranging from 0 to 200µmol·m^− 2·s^− 1, the Pn values of the 9 treatments showed a rapid increase trend with the increase of PAR. When PAR > 200µmol·m^− 2·s^− 1, the Pn of P. yunnanensis seedlings gradually increased and tended to plateau under each treatment (Fig. 9-A). In addition, under the I_6dS_1/2 treatment, the maximum net photosynthetic rate of P. yunnanensis seedlings reached its maximum value (14.951µmol·m^− 2·s^− 1), and under this treatment, the LSP value is the highest (14.951µmol·m^− 2·s^− 1), Rd and LCP have the lowest values of 0.590 and 21.395µmol·m^− 2·s^− 1, respectively, indicates that P. yunnanensis seedlings have the strongest photosynthetic ability under this treatment, with a shorter dark response time, and can undergo light response under lower light conditions (Table 4). For P. elliottii seedlings, the fitting degree of Pn-PAR response curves under each treatment varied between 0.9027 and 0.9952, and the fitting degree was good. When PAR is between 0 and 400µmol·m^− 2·s^− 1, the Pn value of P.elliottii seedlings under 9 treatments showed a sharp increasing trend with the increase of PAR. When PAR > 400µmol·m^− 2·s^− 1, the Pn of P. elliottii seedlings gradually increased and tended to stabilize under each treatment (Fig. 9-B). However, under the treatment of I_3dS₀ and I_6dS₀, the LSP of P. elliottii seedlings was the highest, with values of 1081.100 and 1019.310µmol·m^− 2·s^− 1, respectively, indicating that the maximum saturated light intensity of Pinus elliottii E. seedlings in this environment is 1081.100µmol·m^− 2·s^− 1, exceeding this light intensity will inhibit plant photosynthesis, thereby affecting plant growth and development (Table 4). Moreover, under I_3dS₀ treatment, the net photosynthetic rate of P. elliottii seedlings reached its maximum (17.771µmol·m^− 2·s^− 1).

Fig. 9

Effects of different karst habitats and rainfall distribution on the Pn-PAR curves of two coniferous seedlings

Table 4 Effects of different karst habitats and rainfall distribution on the characteristic parameters of Pn-PAR curves in two coniferous seedlings

According to Fig. 10; Table 5, there are differences in the effects of different treatments on the Pn-CO₂ response curves of two coniferous seedlings. For P. yunnanensis, the fitting degree of all 9 treatments is higher than 0.94, with a maximum fitting degree of 0.9876, indicating good fitting effects. Ci between 0 and 400µmol·m^− 1, the Pn values of the 9 treatments showed an accelerated increase trend with the increase of Ci. When Ci > 400µmol·m^− 1, the Pn of P. yunnanensis seedlings gradually increased and tended to plateau under various treatments (Fig. 10-A). In addition, the I_6dS_1/2 treatment CSP was significantly higher than I_6dS₀ by 1086.610µmol·m^− 1, while under I₉S_1/2 treatment, CSP and Rp were the lowest, with values of 690.507µmol·m^− 1 and 12.007µmol·m^− 2·s^− 1(Table 5), respectively, indicates that under this treatment, P. yunnanensis seedlings have a lower ability to utilize CO₂ and weak photosynthesis. For P. elliottii seedlings, the fitting degree of all 9 treatments was higher than 0.9338, with a maximum fitting degree of 0.9874, indicating good fitting effects. Under each treatment, the Pn value of P. elliottii seedlings showed a significant trend of first increasing sharply and then decreasing with the increase of Ci (Fig. 10-B). However, under the treatment of I₃S₀ and I₃S_1/4, the P. elliottii seedlings showed the best utilization efficiency of CO₂, indicating that the photosynthetic capacity of the treated plants was the best.

Fig. 10

Effects of different karst habitats and rainfall distribution on Pn-CO₂ curves of two coniferous seedlings

Table 5 Effects of different karst habitats and rainfall distribution on the characteristic parameters of Pn-CO₂ curves of two conifer seedlings

Structural equation modeling

The structural equation model (Fig. 11) shows that under different treatments, P.yunnanensis G, Pro, and RV all have a direct effect on SOD, with path coefficients of 0.46, 0.78 (P < 0.05), and 0.74 (P < 0.001), respectively. Pro and POD have indirect effects on RB, while SOD, H, and SP all have direct effects on RB, with path coefficients of 0.46 (P < 0.05), 1.51 (P > 0.05), and − 0.92 (P > 0.05), respectively. G and Pro have a direct effect on H, with path coefficients of 0.97 and 0.40, respectively (P < 0.001). LB, RB, and RD all have a direct effect on RV, with path coefficients of 0.68 (P < 0.05), -0.90 (P < 0.01), and 1.18 (P < 0.001), respectively. This fully demonstrates the high correlation between the physiological growth of P. yunnanensis seedlings, and the plant’s seedling height, ground diameter, biomass, and root characteristics have a significant impact on antioxidant enzymes. For example, as the diameter of the ground increases, the SOD activity of plants will be enhanced, and the increase in root biomass promotes the increase in root volume, which in turn promotes the accumulation of stem and leaf biomass; However, the enhancement of antioxidant activity can, in turn, promote the accumulation of plant biomass and growth (Fig. 11-A). From Fig. 10-B, it can be seen that there are certain direct or indirect effects between the photosynthetic pigments, fluorescence parameters, and photosynthetic gas exchange parameters of P. yunnanensis seedlings. Car, Chlb, Fv, and Fo all have a direct effect on Fm, with path coefficients of 0.40 (P < 0.01), 0.11 (P < 0.01), 0.82 (P < 0.001), and − 0.29 (P < 0.05), respectively. Tr and qP have a direct effect on Chla, with path coefficients of 0.34 (P < 0.01) and 0.40 (P < 0.01), respectively. Pn and Ci have a positive direct effect on GS and Ci has a positive effect on Pn. Although Fm has no direct effect on Pn, it indirectly affects Pn through Ci. These fully demonstrate that the increase in photosynthetic pigment content and fluorescence parameters in P. yunnanensis seedlings can indirectly or directly reflect the net photosynthetic rate of the plant, thereby reflecting the photosynthetic capacity of the plant.

However, for P. elliottii seedlings, H and SOD have a direct effect on G, with path coefficients of 0.94 (P < 0.01) and 0.09 (P > 0.05), respectively. MDA, Pro, and H all have a negative direct effect on RS, indicating that an increase in MDA, Pro, and H reduces RS. RV and LB have a positive direct effect on RD, while RV has a positive direct effect on SOD and SP has a positive direct effect on LB, with path coefficients of 0.36 (P > 0.05) and 0.39 (P < 0.05), respectively. RB has no direct effect on RD, but it can indirectly affect RD through RV. Therefore, under different treatments, the physiological growth of P. elliottii seedlings will adapt to changes in different environments through the adjustment of their physiological indicators, to maintain their growth and development (Fig. 11-C). In addition, from Fig. 10-D, it can be seen that Chla has a direct effect on Fo, Pn, and Chlb, with path coefficients of -1.22 (P < 0.01), 0.31 (P < 0.05), and 0.61 (P < 0.001), respectively. Pn has a direct effect on Chlb, Tr, and Ci, indicating that as the net photosynthetic rate of plants increases, Chlb, Tr, and Ci also increase. Ci can indirectly affect Tr through Gs, and Chla can also indirectly affect Car. These studies can all demonstrate that P. elliottii seedlings can directly or indirectly affect the photosynthetic capacity of plants through the conditions of photosynthetic pigment content or fluorescence parameters. The relationship between photosynthetic pigment, fluorescence parameters, and gas exchange parameters of P. elliottii seedlings can also be intuitively seen through this graph.

Fig. 11

Structural equation model of physiological growth and photosynthetic characteristics of two coniferous seedlings under different karst fissures and rainfall distribution. Note The orange path represents a positive effect, while the green path represents a negative effect. The width of the arrow indicates the strength of the causal effect* P < 0.05, * * P < 0.01, * * * P < 0.001

Fuzzy comprehensive analysis

According to fuzzy membership analysis (Table 6). The average membership function values of P. yunnanensis seedlings under different karst fissures and rainfall distribution treatments are I_6dS_1/4> I_6dS_1/2> I_3dS_1/4> I_3dS_1/2> I_6dS₀> I_9dS_1/4> I_3dS₀> I_9dS₀> I_9dS_1/2. From this, it can be seen that the optimal treatment group is the I_6dS_1/4 treatment, indicating that under natural rainfall duration, increasing karst cracks can promote the growth of P. yunnanensis seedlings. Combined with previous analysis, it can also be seen that the antioxidant system, photosynthetic pigments, and root characteristics of P. yunnanensis seedlings in this treatment are the best; Secondly, the I_6dS_1/2 treatment indicates that under moderate rocky desertification, P. yunnanensis seedlings can also grow and develop normally. However, the average membership function values of P. elliottii seedlings under different treatments were I_3dS₀ > I_3dS_1/4> I_6dS₀> I_3dS_1/2> I_6dS_1/4> I_9dS₀> I_6dS_1/2> I_9dS_1/4> I_9dS_1/2. From this, it can be seen that reducing rainfall duration significantly promotes the growth of P. elliottii seedlings, and I_3dS₀ treatment has the best effect, followed by I_3dS_1/4.

From Fig. 12-A, it can be seen that under different karst fissures and rainfall distribution treatments, the physiological growth indicators of P. yunnanensis seedlings exhibit different trends, but overall, the I_6dS_1/2 treatment has the best effect, with multiple indicators such as Fv, Fm, Chlb, qP, ETR reaching their maximum values under this treatment. The second-best treatment is I_6dS_1/4, which is consistent with the analysis results in Table 6. However, P. elliottii seedlings showed different trends of change (Fig. 12-B). Under different treatments, the overall physiological growth indicators of P. elliottii seedlings showed the best effect under the I_3dS₀ treatment, followed by the I_3dS_1/4 treatment. These analyses further validate the fuzzy membership analysis.

Table 6 Membership analysis of physiological growth indicators of two coniferous seedlings under different karst fissures and rainfall distributions

Fig. 12

Radar analysis of various indicators of two types of coniferous seedlings under different karst fissures and rainfall distribution

Discussion

Plant biomass and root morphology characteristics

The ecosystem in KRD areas is unstable, prone to degradation, nutrient loss, and decreased water-holding capacity, ultimately leading to soil erosion. Biomass is an important indicator of plant energy accumulation, and the distribution differences of biomass in various organs can reflect the growth strategy of plants [47]. In this study, with the deepening of karst cracks, the seedling height, ground diameter, and biomass of various organs of P. yunnanensis seedlings showed a significant trend of first increasing and then decreasing (P < 0.05). Properly increasing karst cracks promoted the growth of P. yunnanensis seedlings, and P. yunnanensis seedlings did not have a good effect on mild rocky desertification in the whole soil habitat. This is similar to the research findings of Zong and Shi [48]. This may be because under full soil treatment, although plants can obtain the most nutrients and living space, the permeability of karst soil is poor, and the soil in this habitat is prone to compaction, resulting in certain deep-water stress [49]; The soil permeability in habitats with fewer rocks and more soil allows plants to have better contact with oxygen, which is beneficial for plant growth. However, P. elliottii seedlings showed a significant downward trend with the deepening of karst cracks, and increasing karst cracks significantly inhibited the growth of P. elliottii seedlings, consistent with Wu et al. [50] study on the effect of drought stress on Sophora davidii K. seedlings. This may be because increasing karst fissures improves soil permeability, but it also leads to rapid soil water loss, causing drought problems. In addition, as P. elliottii is an exotic tree species, its physiological mechanisms have not yet adapted to changes in environmental conditions. Therefore, under the whole soil treatment, P. elliottii has the best biomass accumulation.

In this study, the root characteristics of P. yunnanensis seedlings increased in thickness with the deepening of karst cracks, and prolonged rainfall duration (I_9d) inhibited root growth, which is similar to previous research results [51, 52]. Our research indicates that by increasing the thickness of karst cracks, the root activity of P. yunnanensis seedlings is higher, which also indicates the adaptability of P. yunnanensis seedlings to karst habitat stress. However, when the duration of rainfall, the root activity of most seedlings will gradually lose vitality, thereby inhibiting root growth and ultimately leading to death. However, the root growth of P. elliottii seedlings decreases with the increase of karst cracks. This may be determined by the physiological characteristics of P. elliottii seedlings themselves, which increase karst cracks, cause rapid soil water loss, and result in less water absorption by the roots, which is not conducive to the development of the roots. The results of this study also indicate that reducing rainfall duration (3d) has a promoting effect on the growth of the root structure of P. elliottii seedlings, which fully indicates that P. elliottii seedlings have a high water demand.

Physiological growth and antioxidant system

This study found that P. yunnanensis seedlings and P. elliottii seedlings had the lowest K⁺ and Mg²⁺ content and less root biomass accumulation in karst habitats of S_1/2 with a rainfall duration of 9 days. The main reason for this phenomenon may be related to karst stress and longer rainfall intervals [53]. The environment where plants with large karst cracks are located has less water, and the plants are affected by water deficiency, which leads to the loss of K⁺ and thus affects the accumulation of plant root biomass [54]. In addition, the mineral element content of the two conifer seedlings showed that Ca²⁺> Mg²⁺> K⁺> Na⁺, This is likely related to KRD habitats, where more highly weathered carbonate bedrock is exposed to the surface, and carbonates are already rich in high Ca²⁺ [55]. Moreover, the accumulation of element content in various organs of P.yunnanensis showed that leaves > roots > stems, while P. elliottii showed that roots > stems > leaves. The nutrient accumulation of P. elliottii mainly tends to be in the roots, indicating that to grow in rocky desertification areas, P. elliottii distributes more nutrients to the roots, thereby ensuring their growth and development. This is because karst fissures can cause certain drought stress, and P. elliottii will adjust its root structure (root length, root volume, and root surface area) to increase the absorption of water and nutrients from the soil, thereby responding to the drought stress caused by karst fissures [56]. From the root characteristics, it can also be seen that the root length, root volume, and root surface area of P. elliottii seedlings grow better than those of P.yunnanensis This further proves that P. elliottii has made adjustments to its root structure to adapt to drought caused by karst fissures, such as increasing root length, root width, and root surface area. However, the root length, root width, and root surface area of P. yunnanensis are not as good as those of P. elliottii. This is due to the genetic characteristics of P. yunnanensis itself. P. yunnanensis is better able to adapt to drought caused by karst fissures than P. elliottii, or the negative effects of this drought stress on P. yunnanensis are relatively small. Therefore, P. yunnanensis does not need to adjust its root structure to adapt to the karst fissure environment in this environment. In this study, the content of SOD, POD, and SP in P. yunnanensis seedlings gradually increased with the increase of karst cracks under the rainfall duration of 3 and 6 days. The extension of rainfall interval (9 days) first increased and then decreased, which is consistent with previous research results [57, 58]. Their report showed that after 21 days of pretreatment, compared with the negative control, the CAT activity of rice seedlings significantly increased in the supernatant of PNSB diluted with 1 µM commercial ALA or 10X under NaCl stress. This means that commercial ALA and 10X diluted PNSB supernatant containing ALA can decompose H₂O₂ under NaCl stress conditions. On the other hand, for all treatment types, CAT activity under normal conditions was significantly higher than that under NaCl stress. However, this study found that P. yunnanensis seedlings actively regulate their protective enzyme activity to eliminate the damage caused by rocky desertification and karst fissures. Secondly, P. yunnanensis seedlings reduce their cell osmotic potential by increasing SP content, thereby increasing the activity of protective enzymes and promoting normal plant growth. Mild rocky desertification (S_1/4) alleviated the damage of P. yunnanensis seedlings to karst cracks under 9 days of rainfall, which is consistent with previous results on nitrogen fertilizer alleviating stress-induced membrane damage [59]. On the contrary, the POD, SOD, and SP of P. elliottii seedlings gradually decrease with the increase of karst cracks. P. elliottii seedlings did not increase the activity of antioxidant enzymes to alleviate the damage caused by stress under this environmental stress, which was different from the previous studies [57, 58]. This situation is likely due to the inability of P. elliottii seedlings to adapt to environmental changes through antioxidant enzymes due to their genetic factors under karst and long-term rainfall stress. Secondly, insufficient water leads to redox imbalance within plant leaf cells, thereby increasing the likelihood of oxidative damage. Moreover, the increase of karst fissures accelerates soil moisture loss, resulting in low soil moisture content. Therefore, the antioxidant enzymes of P. elliottii in karst fissure habitats are lower than those in whole soil habitats. This indicates that karst fissures affect the ability of plant antioxidant systems to remove reactive oxygen species, which may be due to the membrane lipid peroxidation triggered by karst stress, which destroys the active centers of enzymes, alters enzyme structure, or inhibits enzyme expression [60]. Finally, it is also possible that karst stress inhibits the transcription or genetic mutations of genes related to P. elliottii seedlings, thereby affecting the incomplete expression of antioxidant enzyme activity [61]. Of course, this reason still needs further verification from us. In addition, this study also found that the MDA and Pro levels of two coniferous seedlings were significantly higher under the I_9dS_1/2 treatment than other treatments (Fig. 5), indicating that prolonged rainfall duration and S_1/2 karst habitat led to an increase in MDA content in the bodies of the two coniferous seedlings. Karst fissure stress has already caused damage to the membrane system, exacerbating membrane lipid peroxidation, which is consistent with the results of Li et al. [62]. In summary, an appropriate increase of karst fissures could increase the antioxidant enzyme activities of P. yunnanensis seedlings, thereby reducing the content of MDA and Pro; The P. elliottii seedlings have the highest antioxidant enzyme activity in the whole soil habitat.

Photosynthetic pigments

In this study, the photosynthetic pigment content (Chla, Chlb, Car) of P. elliottii seedlings showed a trend of first increasing and then decreasing with the increase of karst habitat, and gradually decreasing with the extension of rainfall time, which is consistent with the research results of Ahmadizadeh [63]. It is possible that low-frequency heavy rainfall (9d) in a semi-soil and semi-stone habitat can lead to leaching, causing nutrient loss in the soil with the loss of rainfall. It is also possible that the increase of karst cracks causes plants to experience certain droughts, leading to excessively high temperatures and inhibiting the formation of chlorophyll esters in the plant body, resulting in a decrease in chlorophyll formation. The content of photosynthetic pigments in P. yunnanensis seedlings gradually increases with the increase of karst cracks (except for 9 days of rainfall). This may be due to the stress of karst fissures in P. yunnanensis needles, where cell fluid tends to move to the leaf epidermis, increasing pressure and leading to an increase in surface active substance concentration, thereby promoting the synthesis and accumulation of photosynthetic pigments. In addition, under drought conditions caused by karst fissures, plant roots also release organic matter, providing nutrients for the synthesis of photosynthetic pigments, which can promote the synthesis of photosynthetic pigments in plants. However, in this study, moderate rocky desertification inhibited the synthesis of photosynthetic pigments in P. elliottii, but promoted the accumulation of photosynthetic pigments in P. yunnanensis. This may be due to differences in genetic factors and genes regulating photosynthetic pigment synthesis in the genomes of the two coniferous seedlings. P. yunnanensis may have a genotype with photosynthetic pigment content, while P. elliottii. does not, and this phenomenon needs further verification. In addition, from the photosynthetic intensity, it can be concluded that the light saturation point of P. yunnanensis is higher than that of P. elliottii (Fig. 9), which also indicates that the accumulation of photosynthetic pigments in P. yunnanensis is higher than that of P. elliottii.

Fluorescence and gas exchange parameters

In this study, with the thickening of karst cracks, the Fo/Fv, Fv/Fm, Fv’/Fm’, qP, ETR, and NPQ of P, yunnanensis seedlings gradually increased, while Fo gradually decreased. Under half-soil and half-stone habitats with extended rainfall duration, these indicators all decreased. The fluorescence indicators of P. elliottii seedlings gradually decrease with the thickness of the karst. The decrease in fluorescence parameters indicates that the deterioration of rocky desertification habitats and the extension of rainfall intervals can affect the light-harvesting protein complexes and activity of enzymes, leading to an impact on the activity of PSII reaction centers, inhibiting the photosynthetic pigments from converting the captured light energy into chemical energy [64]. The decrease in Fo indicates that the PSII reaction center has not been completely deactivated, and more energy is dissipated in the form of heat [65]. NPQ is the excess light energy dissipated in thermal form by the PSII reaction center [66, 67]. The qP and NPQ of P. yunnanensis increase with the deepening of rocky desertification, while those of P. elliottii gradually decrease. This indicates that in habitats with lower levels of rocky desertification, the electron transfer activity of P. elliottii is higher, but with the reverse succession of rocky desertification ecosystems, the habitat further deteriorates. The opening ratio of PSII reaction centers and the energy involved in photochemical reactions will be suppressed, causing plants to dissipate excess light energy in the form of heat, reducing damage to photosynthetic structures. In addition, beyond increasing heat dissipation and other regulatory mechanisms, plants can also consume excess light energy through processes such as photorespiration and nitrogen metabolism, avoiding burns to photosynthetic organs.

The LCP and AQY reflect the plant’s ability to utilize weak light [68]. Stomatal conductance (Gs) also plays an important role in photosynthesis by absorbing CO₂ and controlling water loss [69]. Plant species with higher Gs are expected to have higher maximum CO₂ assimilation [70]. This study found that P. yunnanensis seedlings generally exhibited higher Gs and transpiration rate (Tr) compared to P. elliottii (Table 5). According to other studies, the higher the maximum photosynthetic rate of plants, the higher the Gs value [71], and the results of this study also indicate this pattern (Table 5).

In this study, P. yunnanensis seedlings showed lower LCP and higher AQY under I_6dS_1/4 treatment (Table 4), which is consistent with previous research results [72]. This indicates that P. yunnanensis has a higher sensitivity to light and respiration under this treatment, and cells can carry out photosynthesis under lower light intensity. The lower the LCP, the stronger the plant’s adaptability. This is likely because P. yunnanensis chloroplasts have larger basal grains, and the more layers there are in the basal grains, the higher the content of chloroplasts. This ensures that the photosynthesis and respiration rates of the plant are equal under lower light intensity [73]. P. yunnanensis has a higher AQY under I_6dS_1/4 treatment, which means its photosynthetic rate is higher. This also indicates that the closer the light wavelength in the photosynthetic spectrum is to the absorption peak of the plant, the higher the AQY. In addition, the LPnmax of P. elliottii seedlings showed a trend of increasing and then decreasing with the increase of karst fissure thickness. The semi-soil and semi-stone habitat inhibited the photosynthesis of P. elliottii seedlings, while P. yunnanensis promoted it. This may be due to factors such as increased karst fissures and infiltration, leading to a water deficit in the soil niche. In this situation, the water transported to the leaves is also insufficient, causing stomatal closure and a decrease in CO₂ entering the leaves, both raw materials for photosynthesis. When the leaves are dehydrated, starch hydrolysis is enhanced [74], sugars continue to accumulate, and the output of photosynthetic products becomes slow, inhibiting the rate of photosynthesis. At the same time, due to water deficiency in the leaves, the electron transfer rate in chloroplasts decreases, coupled with the uncoupled photosynthetic phosphate, which affects the formation of the assimilation force [75]. In addition, water deficiency can also affect the growth of leaves, inhibit the expansion of photosynthetic area, and reduce the absorption of light energy by leaves, all of which can lead to a decrease in photosynthetic rate. However, due to factors such as thin soil, mineral nutrients are deficient in the soil niche. For example, when the oxygen content in the soil is insufficient, it can hinder aerobic respiration in the roots, and the roots are one of the mechanisms for absorbing mineral elements, i.e., active uptake is carried out through the energy provided by the metabolism of root cells. The aerobic respiration of roots is inhibited, and the function of absorbing mineral elements is also inhibited. The lack of mineral elements can have an impact on plant photosynthesis in many aspects. Figure 4 also shows a decrease in mineral element content by increasing karst thickness. When lacking certain mineral elements, chlorophyll synthesis is hindered, and the collection, transmission, and conversion of light energy by plants are inhibited, resulting in a decrease in photosynthetic capacity. If severe hypoxia occurs, toxic substances such as alcohol produced by anaerobic respiration in roots can also lead to root rot, preventing it from performing its function of absorbing and transporting water and nutrients. Therefore, hindering the activity of the root system will indirectly hurt photosynthesis, so that, increasing the thickness of the karst will reduce the photosynthetic rate of P. elliottii seedlings. The LCP, Rd, and AQY can all reflect the plant’s ability to utilize weak light. In this study, except for the S_1/4 and S_1/2P. elliottii seedlings under I9d rainfall, the AQY of two coniferous seedlings in the other treatments was relatively low, lower than the general plant level of 0.03 ≤ AQY ≤ 0.05 [76], indicating that the light energy utilization efficiency of two coniferous seedlings is relatively low under weak light. The CO₂ response curve reflects the response characteristics of leaf photosynthetic capacity to different CO₂ concentration changes. The higher the CSP of plants, the stronger their adaptability to high CO₂ concentration environments, while plants with low CCP have the characteristics of high Pn and fast growth [77]. The P. yunnanensis seedlings treated with I_6dS_1/2 and the P. elliottii seedlings treated with I_3dS₀ have larger CSP, lower CCP, and the lowest photorespiration, indicating that this treatment enhances the CO₂ utilization ability of P. yunnanensis and P. elliottii needles.

Conclusion

In summary, the physiological growth and photosynthetic characteristics of two types of coniferous seedlings in different soil niches are affected by the interval time of rainfall, and the photosynthetic physiological growth during the interval time of rainfall is also constrained by the soil niche. Under the same rainfall duration, with the increase of karst thickness, the growth of seedling height and ground diameter, the biomass of various organs, K⁺, Ca²⁺, Na⁺, and Mg²⁺accumulation of P. yunnanensis seedlings showed a significant change pattern of first increasing and then decreasing (P < 0.05), while P. elliottii seedlings showed a gradually decreasing trend (except for Ca²⁺). The K⁺, Ca²⁺, and Mg²⁺ in each organ of P. yunnanensis seedlings showed that leaves > roots > stems, and Na⁺ showed that roots > leaves > stems. The accumulation of mineral elements in various organs of P. elliottii seedlings is as follows: roots > stems > leaves, and the accumulation of mineral elements in two types of coniferous seedlings is as follows: Ca²⁺ > Mg²⁺ > K⁺ > Na⁺. The root characteristics (root length, root volume, root surface area, root diameter), SOD, POD, SP, photosynthetic pigment content, fluorescence parameters, and gas exchange parameters of P. yunnanensis seedlings gradually increase with the increase of karst thickness (except for 9 days of rainfall). The root characteristics, antioxidant enzyme activity, and photosynthetic indicators of P. elliottii seedlings gradually decrease.

Under different treatments, our study showed that the maximum saturated light intensity and minimum light intensity of P. yunnanensis seedlings were 1624.530 and 21.395µmol·m^− 2·s^− 1, respectively, while the values for P. elliottii seedlings were 1081.100 and 27.148µmol·m^− 2·s^− 1, respectively. The CO₂ saturation points of P. yunnanensis and P. elliottii are 2171.070 and 1444.840µmol·m^− 1, respectively. Fuzzy membership analysis showed that under different treatments, P. yunnanensis seedlings showed that I_6dS_1/4> I_6dS_1/2> I_3dS_1/4> I_3dS_1/2> I_6dS₀> I_9dS_1/4> I_3dS₀> I_9dS₀> I_9dS_1/2. The P. elliottii seedlings showed that I_3dS₀ > I_3dS_1/4> I_6dS₀> I_3dS_1/2> I_6dS_1/4> I_9dS₀> I_6dS_1/2> I_9dS_1/4> I_9dS_1/2. In conclusion, prolonging rainfall duration has an inhibitory effect on the growth of two types of coniferous seedlings. Reducing rainfall duration promotes the growth and development of P. elliottii seedlings while increasing karst thickness promotes the growth and development of P. yunnanensis seedlings, which is better than P. elliottii seedlings. However, it has an inhibitory effect on the growth of P. elliottii seedlings. Therefore, P. yunnanensis is preferred as a tree species for vegetation restoration or rocky desertification governance in karst areas at different degrees of desertification.