A method for tracing the spread of invasive alien plants using stable isotopes

Abstract

The present invention discloses a method for tracing the spread of invasive alien plants using stable isotopes, which includes tracing the source of invasive alien plants and identifying the risk of invasive alien plant spread. The tracing the source of invasive alien plants includes S1, plant sample collection, S2, sample pretreatment, S3, stable isotope determination, S4, analysis of the spread source and spread path of invasive alien plants; the risk identification of invasive alien plants includes P1, construction of a spread network of invasive alien plants, and P2, identification of spread risk areas. The present invention measures the stable isotope ratios of plants from newly introduced areas and potential source areas, and combines a multi-dimensional analysis method to trace the source and spread path of invasive alien plants, with high tracing accuracy. Compared with traditional genetic analysis methods, this method has lower costs and higher experimental efficiency, and can significantly reduce tracing costs.

Description

Technical Field

[0001] The present invention relates to the technical field of biological traceability, and in particular to a method for tracing the diffusion of invasive alien plants using stable isotopes. Background Art

[0002] The invasion of alien species has become a major factor in global biodiversity loss and ecosystem degradation. Solanum quinquefolium is a nationally managed invasive alien species and a malignant invasive plant in the natural grasslands of the agricultural-pastoral transition zone in northern my country. It has caused great harm to the grassland and farmland ecosystems in Liaoning, Inner Mongolia, and Hebei. It is still being introduced and spreading, showing an exponential growth trend. The main problem currently faced is that the introduction route and source are unclear, which greatly restricts the efficiency of prevention and control. Therefore, accurately identifying the source of spread of alien invasive plants such as Solanum quinquefolium, especially tracking their spread routes, is crucial for the prevention and control of alien invasive plants.

[0003] Due to the lack of analytical methods to trace the spread of invasive alien plants such as Solanum serrata, current prevention and control of Solanum serrata is extremely passive and inefficient. Existing traceability methods mainly rely on plant morphological characteristics and molecular genetic structure. These methods have many limitations when dealing with plant invasions that spread rapidly and in complex ecological contexts. On the one hand, when an alien invasive plant is introduced only once, the species' gene pool is very limited and the variability of genetic markers is insufficient, resulting in samples collected from multiple locations showing similar genetic characteristics, making it impossible to effectively distinguish samples from different sources. On the other hand, genetic traceability methods are costly and time-consuming, requiring large-scale sample collection and high-precision genetic analysis. At the same time, data processing and interpretation are complex, increasing costs and experimental complexity. In addition, molecular genetic methods can only analyze the differences in the alien invasive plant itself and obtain information on its source of spread, but its spread pathway cannot be identified or determined. Summary of the Invention

[0004] The purpose of the present invention is to provide a method for tracing the spread of alien invasive plants using stable isotopes, which can trace the source and spread path of alien invasive plants through the stable isotope ratio in the sample.

[0005] The present invention is implemented by the following technical solutions:

[0006] A method for tracing the spread of invasive alien plants using stable isotopes, which includes tracing the spread of invasive alien plants and identifying the risk of invasive alien plants spreading; wherein,

[0007] The invasive alien plant source tracing analysis includes the following steps S1-S4;

[0008] S1. Plant Sample Collection: S11. Identify the new introduction site; S12. Collect invasive alien plants and crop products adulterated with these invasive alien plants from the new introduction site. These crop products may contain one or more species, as plant samples from the new introduction site; S13. Collect data to determine the potential source of the plant samples from the new introduction site; S14. Collect the invasive alien plants and crop products from the potential source site as plant samples from the potential source site. The invasive alien plants and crop products collected should be free of insect holes and lesions, and the plant parts collected from the new introduction site and the potential source site must be the same.

[0009] S2, sample pretreatment;

[0010] S3. Stable isotope determination. Since the ratio of stable isotopes in an organism is closely related to its environment, and different geographical regions, ecosystems, and climatic conditions can lead to differences in these isotope ratios, by analyzing the stable isotope composition of the invasive alien plant, its possible habitat or spread path can be inferred. The source and spread path of the alien plant can also be further confirmed by detecting the stable isotopes of the spread medium (referring to the agricultural product itself).

[0011] S4. Analyze the sources and pathways of spread of invasive alien plants;

[0012] The identification of the risk of spread of invasive alien plants includes the following steps P1 and P2;

[0013] P1, constructing a diffusion network for the invasive alien plant; P11, identifying diffusion network nodes; P12, generating road network links between the diffusion network nodes; P13, constructing a trade network for the crop products based on the diffusion network nodes and the road network links;

[0014] P2. Identify diffusion risk areas: Identify risk areas for invasion by invasive alien plants in the trade network of the crop products by analyzing the correlation between the centrality of the diffusion network nodes and the distribution frequency of the invasive alien plants.

[0015] Furthermore, the method for determining the new introduction site in step S11 is: based on the national spatiotemporal distribution data of the alien invasive plants, the areas where the alien invasive plants have been newly recorded to have appeared in the past five years are selected as the new introduction site.

[0016] Furthermore, the data collection in step S13 includes: investigating the sources of the agricultural products purchased in the newly introduced areas, collecting spatiotemporal data of the planting of the agricultural products across the country, and collecting spatiotemporal data of the occurrence of the alien invasive plants across the country.

[0017] Furthermore, the preprocessing of step S2 includes:

[0018] S21. Dust and debris removal: The collected plant parts should be stored at room temperature in a dry and ventilated place to prevent rotting, and then pollutants (such as dust and dirt) on the plant surface should be removed.

[0019] S22, drying: drying the newly introduced plant samples and the potential source plant samples after dust and impurity removal to a constant weight;

[0020] S23, crushing: crushing the dried plant samples of the newly introduced area and the plant samples of the potential source area, and then sieving;

[0021] S24. Sampling: Take 2-3 mg of the sieved sample as a subsample.

[0022] Furthermore, the step S3 is specifically to perform stable isotope determination on the pretreated subsample to calculate the stable isotope ratio of the subsample; the stable isotope ratio includes δ 13 C (carbon isotope ratio reflects the source and environmental characteristics of plant photosynthesis. Plants in different regions will show different carbon isotope values, which can distinguish different ecological regions), δ 15 N (nitrogen isotope ratios help identify the soil type and fertilization history of plant growth; since nitrogen isotopes vary in different soils and environmental conditions, they are important indicators of the spread of invasive alien plants), δ 18 O (oxygen isotope ratios can reflect changes in water sources; plants absorb water through different water sources under different climatic conditions, so oxygen isotope ratios can provide information on the water source of plant growth), δ 2 H (Water in plants is absorbed by the roots and enters the plant tissues through transpiration. The hydrogen isotope ratio of water is affected by the hydrogen isotope composition of local water sources (such as groundwater, precipitation, river water, etc.); water from different sources has different δ 2 H value, so with the help of hydrogen isotopes, the type of water source and the source of water for plants can be determined).

[0023] Furthermore, step S4 is specifically as follows: using statistical methods (such as cluster analysis, principal component analysis, multivariate regression analysis, etc.) to process the stable isotope ratios of the subsamples of the newly introduced plant samples and the stable isotope ratios of the subsamples of the plant samples from the potential source area to analyze the diffusion sources and diffusion paths of the alien invasive plants.

[0024] Furthermore, the step P11 is specifically as follows: according to the diffusion source and diffusion path of the alien invasive plant analyzed in the step S4, the diffusion network nodes of the alien invasive plant are determined, including the production site of the crop product, the logistics transit center, the distribution site of the crop product, the grass market for transporting the crop product, and the market or farmers and herdsmen who receive the crop product.

[0025] Furthermore, in step P2, the centrality measurement indicators include degree centrality, betweenness centrality, and closeness centrality; centrality reflects the importance of a diffusion network node in the diffusion network, and also reflects the invasion risk of alien invasive plants. In the agricultural product trade network constructed in this scheme, the degree centrality of a diffusion network node reflects the number of nodes that trade agricultural products with the node. A high degree of centrality means that the node has trade relations with more regions, which further indicates that the node plays a hub role in the trade network; closeness centrality reflects the total number of times that transshipments are required during trade between nodes; and betweenness centrality reflects the number of times that transshipments need to pass through a certain node during trade between nodes.

[0026] Degree Centrality D(i) The calculation formula is shown in formula (1):

[0027] (1);

[0028] Where, d i is the degree of node i, N is the total number of nodes in the network;

[0029] Betweenness Centrality C(i) The calculation formula is shown in formula (2):

[0030] (2);

[0031] Where, β ij Indicates the starting node i To the end j The number of links contained in the shortest path, N is the total number of nodes in the network;

[0032] closeness centrality B(i) The calculation formula is shown in formula (3):

[0033] (3)

[0034] In the formula α xy For nodes x To Node yThe number of all shortest paths, α xy (i) For nodes x To Node y and passing through the node i The number of all shortest paths.

[0035] Furthermore, when the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node are all significantly correlated with the distribution frequency of the alien invasive plants, the diffusion network node is determined to be a high-risk invasion area; when one or two of the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node are significantly correlated with the distribution frequency of the alien invasive plants, the diffusion network node is determined to be a medium-risk invasion area; when the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node do not show a significant correlation with the distribution frequency of the alien invasive plants, the diffusion network node is determined to be a low-risk invasion area.

[0036] Advantages of the present invention:

[0037] (1) High traceability accuracy: Stable isotope ratios are significantly affected by environmental and geographical factors and show spatial heterogeneity. By measuring the stable isotope ratios of plants from newly introduced areas and potential source areas, and combining geographic information systems (GIS) with multi-dimensional analysis methods such as spatial analysis and cluster analysis, it is possible to trace the sources and diffusion paths of alien invasive plants under variable environmental conditions, and to analyze the invasion risk of alien invasive plants at a certain node. The traceability accuracy is high, and the traceability results are more accurate and reliable.

[0038] (2) Low cost and high efficiency: Compared with traditional genetic analysis methods, the cost of stable isotope single sample detection is low, and only 1-3 samples need to be collected at each location for testing; in addition, the time required for stable isotope detection and analysis is short, and the detection time for a single sample is 10-15 minutes. Therefore, it has low cost and high experimental efficiency, which can significantly reduce the traceability cost.

[0039] (3) Convenient and easy to store: The samples used for isotope determination are not limited to a specific part of the plant. It is only required that the samples collected from the newly introduced area and the potential source area belong to the same part. The branches or seeds of the plant can be selected. The collected samples can be stored in a ventilated and dry place at room temperature. Unlike traditional genetic testing methods, they do not need to be poured into silica gel for preservation, nor do they need to be stored in liquid nitrogen or placed in a refrigerator for low-temperature storage. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0041] Figure 1 Cluster diagram of stable isotope content in Solanum serrata branches

[0042] Figure 2 This is the traceability analysis diagram of the stable isotope content in the branches of Solanum serrata;

[0043] Figure 3 This is the discriminant analysis result of the origin of Solanum truncatum branches;

[0044] Figure 4 This is the cluster diagram of stable isotope content of Solanum serrata seeds;

[0045] Figure 5 This is the traceability analysis diagram of the stable isotope content of Solanum serrata seeds;

[0046] Figure 6 This is the discriminant analysis result of the origin of Solanum serrata seeds;

[0047] Figure 7 is the cluster diagram of stable isotope content of straw;

[0048] Figure 8 This is the traceability analysis diagram of the stable isotope content in straw;

[0049] Figure 9 This is the result of the discriminant analysis of the origin of straw;

[0050] Figure 10 This is the cross-regional diffusion network of Solanum truncatum. DETAILED DESCRIPTION

[0051] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.

[0052] Example 1

[0053] Solanum truncatum Solanum rostratumSolanum spp. (Dund.) is a highly invasive, annual weed in the Solanaceae family. In recent years, newly invasive Solanum spp. (Solanum spp.) has been discovered in natural grasslands across several banners in Xilin Gol League, Inner Mongolia, necessitating the development of effective control measures.

[0054] This example provides a method for tracing the spread of invasive alien plants using stable isotopes. The carbon (δ 13 C), nitrogen (δ 15 N), oxygen (δ 18 O), hydrogen (δ 2 H) Stable isotope ratios are used to track the sources and pathways of Solanum serratum’s spread, and to analyze and determine risk areas where Solanum serratum may invade, thereby providing accurate data support for the management and control of its spread, and to inhibit its further rapid spread.

[0055] This method includes the traceability analysis of Solanum luteum and the identification of the spread risk of Solanum luteum.

[0056] 1. Origin Analysis of Solanum truncatum

[0057] 1. Plant sample collection

[0058] 1) Determine the newly introduced areas: Based on the spatiotemporal distribution data of Solanum serrata across China, select the areas where Solanum serrata has been newly recorded in the past five years as the newly introduced areas.

[0059] 2) It was found that the main crop product adulterated with Solanum serrata was straw. Therefore, straw purchased by farmers in the newly introduced area and the remains and seeds of Solanum serrata plants mixed in it were collected as plant samples from the newly introduced area.

[0060] 3) Conduct field investigations on the sources of straw purchased in newly introduced areas, collect spatiotemporal data on straw planting across the country, collect spatiotemporal data on the occurrence of Solanum serrata across the country, and identify potential sources of plant samples from newly introduced areas.

[0061] 4) Collect Solanum serrata and straw from different areas of the potential source, using the same collection sites as plant samples from the newly introduced areas, as plant samples from the potential source areas.

[0062] When collecting plant samples from newly introduced areas and potential source areas, select plants that are free of insect holes and lesions. After collection, store the samples in a ventilated, dry place at room temperature. Based on actual needs, more dense sampling points can be planned. For example, sampling straw and Solanum serrata plants from adjacent farmland plots less than 1 km apart can help more accurately capture local differences and improve traceability accuracy.

[0063] 2. Sample preprocessing

[0064] 1) Dust and debris removal: Remove surface pollutants (such as dust and dirt) from newly introduced plant samples and plant samples from potential sources.

[0065] 2) Drying: After dust and impurity removal, plant samples from newly introduced areas and potential source areas should be placed in a freeze dryer or a constant temperature dryer and dried to constant weight. If a freeze dryer is used, during the drying process, the temperature is lowered to below freezing (-80~-40°C) during the freezing stage to freeze the water, ensuring that the structure and biological activity of the sample remain as unchanged as possible. During the sublimation stage (primary drying stage), the sample is heated to -50~-20°C under vacuum conditions, and the water sublimates directly from the solid state to the gaseous state. To remove the small amount of remaining water in the sample, a secondary drying process can be performed, gradually increasing the temperature to -10~20°C. If a constant temperature dryer is used, the temperature needs to be controlled between 40~60°C to avoid changes in the isotopic composition of the plant sample due to excessively high temperatures.

[0066] 3) Pulverization: Pulverize the dried plant samples from newly introduced areas and potential source areas using a grinder, and then pass them through a 100-mesh standard sieve for isotope analysis. Pulverize different plant parts separately to prevent cross-contamination.

[0067] 4) Sampling: Take 2-3 mg of the sieved sample as a subsample.

[0068] 3. Stable isotope determination

[0069] The pre-treated sub-samples are subjected to stable isotope determination using a mass spectrometer or an isotope ratio mass spectrometer (IRMS) to calculate the stable isotope ratio of the sub-samples. The stable isotope ratio includes δ 13 C, δ 15 N, δ 18 O, δ 2 H, unit is ‰.

[0070] When performing carbon and nitrogen stable isotope analysis, a 2 mg subsample is first weighed using a high-precision balance with an accuracy of one millionth of a gram to ensure accurate weighing. The subsample is then placed in a tinfoil boat to prevent contamination or loss during analysis. The subsample is then placed in the elemental analyzer's solid sample autosampler, ensuring that one or two reference materials are inserted every 12 subsamples for quality control. The subsample is converted to CO₂ and N₂ gases through a combustion-reduction reaction at high temperature, and then enters the isotope ratio mass spectrometer for precise analysis. Instrument settings include a combustion tube temperature of 950°C, a reduction tube temperature of 600°C, a carrier gas of high-purity helium, a reference gas of CO₂ and N₂, and an oxygen flow rate of 40 mL min⁻¹. -1The oxygen injection time was 70 seconds. The accelerating voltage of the mass spectrometer was 3992 V for carbon isotope analysis and 4230 V for nitrogen isotope analysis, with magnetic field strengths of 3600 mA (C mode) and 2800 mA (N mode), respectively.

[0071] For hydrogen and oxygen stable isotope analysis, a 2 mg subsample was weighed and placed in a silver boat for sample preparation. The sample was then placed in the solid sample autosampler of the elemental analyzer and subjected to quality control with a standard material. The hydrogen and oxygen in the subsample were converted to hydrogen and carbon monoxide gases through a high-temperature cracking reaction. The resulting gases were then detected by an isotope ratio mass spectrometer. The cracking furnace temperatures for hydrogen and oxygen were set at 1080°C and 1180°C, respectively, with a helium purge flow rate of 230 mL / min. -1 The reference gases were CO and H2. The accelerating voltage of the mass spectrometer was 4032 V for hydrogen isotope analysis and 4230 V for oxygen isotope analysis. The magnetic field strengths were 820 mA (H mode) and 2800 mA (O mode), respectively.

[0072] 4. Analyze the diffusion source and diffusion path of Solanum serrata

[0073] After completing the stable isotope ratio measurements of subsamples of Solanum serratum and crop straw collected from newly introduced and potential source areas in the above steps, statistical methods are used to process the data and analyze the sources and pathways of Solanum serratum's spread. In this example, principal component analysis (PCA) was used to process the stable isotope ratio data obtained from subsamples of plant samples from newly introduced and potential source areas. Cluster analysis of these stable isotope ratio data using PCA software revealed the stable isotope signatures of straw and Solanum serratum from different sources. In this way, subsamples from different sources can be classified to identify the sources of Solanum serratum's spread. The pathways between these sources can then be used to determine the pathways of spread.

[0074] II. Identification of the risk of Solanum spicata spreading

[0075] 1. Constructing a diffusion network for Solanum serrata

[0076] Based on the traceability results of stable isotopes of Solanum xanthophyllum, combined with the spatial location of its diffusion network nodes, and according to the minimum cost principle of actual straw transportation, ArcGIS spatial analysis software and Python data analysis tools were used to construct the Solanum xanthophyllum diffusion network. By calculating the centrality of different nodes, high-risk areas in the Solanum xanthophyllum diffusion network were identified. The specific steps include the following.

[0077] 1) Identify diffusion network nodes: Based on the diffusion sources and diffusion paths of Solanum serrata analyzed in the above steps, determine the diffusion network nodes of Solanum serrata, including straw production sites, logistics transit centers, straw distribution sites, straw transfer markets, and straw receiving markets or farmers and herdsmen.

[0078] 2) Generate road network links between diffusion network nodes: Since the transportation cost of straw increases with the increase of transportation distance, roads with shorter distances will be given priority in the actual transportation process. Therefore, this plan uses the nearest neighbor analysis tool of ArcGIS 10.2 to calculate the distance between diffusion network nodes, and uses the nearest facility tool to realize the optimal path analysis between nodes to complete the road network link: ① In order to determine the spatial location of the diffusion network nodes of Solanum serrata, the latitude and longitude of the above nodes were obtained from Google Maps; ② Download the main road data of my country and use the geographic analysis tool of ArcGIS software to cut out the main roads in the study area; ③ Merge the road network vector data. Using the Arctoolbox data management tool, merge the national highway, expressway, provincial highway, county road, and township road data within the study area into a shapefile file; ④ Create a network dataset. In the ArcGIS Catalog, use the road vector data generated in the above steps to create a road network dataset with topological relationships. ⑤ Call the NewClosest function under the ArcGIS Network Analyst module of ArcGIS software. The Facility tool adds a road network dataset, uses road length as impedance, finds the travel cost between nodes, solves the optimal path and path length between nodes, and generates road network links between diffusion network nodes.

[0079] 3) Use the Python software Networkx package to construct a straw trade network based on the diffusion network nodes and road network links.

[0080] 2. Identify areas of risk of spread

[0081] By analyzing the correlation between the centrality of diffusion network nodes and the frequency of Solanum serratum distribution, the risk areas of Solanum serratum invasion in the straw trade network were identified. The centrality measurement indicators include degree centrality, betweenness centrality and closeness centrality.

[0082] Degree centrality D(i) The calculation formula is shown in formula (1):

[0083] (1);

[0084] Where, d i is the degree of node i, N is the total number of nodes in the network;

[0085] Betweenness centrality C(i) The calculation formula is shown in formula (2):

[0086] (2);

[0087] Where, β ij Indicates the starting node i To the end j The number of links contained in the shortest path, N is the total number of nodes in the network;

[0088] Closeness Centrality B(i) The calculation formula is shown in formula (3):

[0089] (3)

[0090] In the formula α xy For nodes x To Node y The number of all shortest paths, α xy (i) For nodes x To Node y and passing through the node i The number of all shortest paths.

[0091] When the degree centrality, betweenness centrality, and closeness centrality of a diffusion network node are significantly correlated with the distribution frequency of invasive alien plants (P < 0.05), the diffusion network node is judged to be a high-risk invasion area; when one or two of the degree centrality, betweenness centrality, and closeness centrality of a diffusion network node are significantly correlated with the distribution frequency of invasive alien plants, the diffusion network node is judged to be a medium-risk invasion area; when the degree centrality, betweenness centrality, and closeness centrality of a diffusion network node do not show a significant correlation with the distribution frequency of invasive alien plants, the diffusion network node is judged to be a low-risk invasion area.

[0092] In addition, since tracing the spread of invasive plants is a long-term task, a database of the spread sources of alien invasive plants can also be constructed to facilitate subsequent work: including (1) continuously collecting plant samples from all over the country, and establishing an isotope database of sub-samples of plant samples from newly introduced areas and an isotope database of sub-samples of plant samples from potential source areas; (2) collecting data on natural environmental factors and human activity factors from newly introduced areas and potential source areas. Since these factors directly affect the isotope composition of alien invasive plants, they must be recorded together with the isotope data to establish an environmental factor database; among them, natural environmental factor data include climate, soil type, and water source data of plant sample sampling points in newly introduced areas and potential source areas; human activity factor data include land use data, road network density, distance from roads, and human activity footprint index of newly introduced areas and potential source areas; (3) collecting the introduction time, distribution area, and spread path of alien invasive plants across the country, and establishing a historical spread information database. The spread path and speed of the alien invasive plants can be inferred through historical records to assist in analyzing the spread source.

[0093] Example 2

[0094] Conduct traceability analysis on Solanum sylvestris and the straw that carries Solanum sylvestris.

[0095] 1) Solanum truncatum branches: A total of 55 subsamples of Solanum truncatum branches of the invasive alien plant were tested for their C, N, O, and H stable isotope contents and ratios. Principal component analysis (PCA) was used to construct an origin discrimination model, such as Figure 1 The results show that the subsamples collected from potential source areas can be divided into 7 groups. Figure 2 As shown in the figure, the branches of Solanum serrata intercepted from the newly introduced goods in Inner Mongolia mainly belong to the same group as the potential source cities of Chifeng and Zhangjiakou. One-way ANOVA and PLS-DA were further used to determine the accuracy of the model traceability, with 70% of the samples as the training set and 30% of the samples as the test set. Figure 3 As shown in the figure, the results show that the accuracy of the model is 88.24%, indicating that the model has strong stability and can accurately trace the branches of Solanum serrata.

[0096] 2) Solanum serratum seeds: A total of 43 subsamples of Solanum serratum seeds, an invasive alien plant, were tested for their C, N, O, and H stable isotope contents and ratios, such as Figure 4 As shown in the figure, the analysis results show that the yellow flower spinulosa seed samples collected from the source area can be divided into 6 groups. Figure 5As shown in the figure, the yellow flower Solanum seeds intercepted from the cargo transported from the newly introduced area in Inner Mongolia belong to the same group as the potential source cities of Chifeng and Zhangjiakou. One-way ANOVA and PLS-DA were further used to judge the accuracy of the model traceability, using 70% of the samples as the training set and 30% of the samples as the test set. Figure 6 As shown in the figure, the discrimination results show that the accuracy of the model is 88.24%, indicating that the model has strong stability and can accurately trace the seeds of Solanum serrata.

[0097] 3) Straw: A total of 92 straw subsamples were tested for their C, N, O, and H stable isotope contents and ratios. The PCA-X model was used to analyze the stable isotope contents. Figure 7 As shown in Figure 2, cluster analysis results show that all subsamples from potential sources can be divided into 7 groups by analyzing stable isotopes. Figure 8 As shown in Figure 2, the traceability analysis results showed that straw collected from the newly introduced area in Inner Mongolia belonged to the same group as the potential source cities of Chifeng, Zhangjiakou, Tongliao, and Ulanqab. One-way ANOVA and PLS-DA were further used to assess the accuracy of the model traceability, using 70% of the subsamples as the training set and 30% of the subsamples as the test set. Figure 9 As shown in the figure, the discrimination results show that the accuracy of the model is 86.84%, indicating that the model has strong stability and can accurately trace the source of straw.

[0098] The traceability results for straw, Solanum serratum branches, and seeds were comprehensively analyzed, and a chi-square test was used to test whether there were significant differences between the traceability results. The results, shown in Table 1, showed no significant differences in the traceability results for the three plant subsamples, indicating that the source of straw purchased in the newly introduced areas is consistent with that of Solanum serratum, further confirming that interregional straw trade is a mechanism for the spread of Solanum serratum.

[0099] Table 1. Significance of differences in the traceability results of branches, seeds and straw of Solanum truncatum

[0100]

[0101] Example 3

[0102] Conduct a spread risk analysis on Solanum truncatum.

[0103] Samples of Solanum serrata branches, seeds, and straw were collected from nine cities. The straw intercepted in the Qianshao area originated only from Zhangjiakou (Source 1), Ulanqab (Source 2), Tongliao (Source 3), and Chifeng (Source 4). Solanum serrata originated only from Zhangjiakou and Chifeng. Because Solanum serrata plants are spread among straw, a straw trade network was constructed based on the principle of minimizing transportation costs, transporting straw from its source to the grass market and then to herders' homes. This is known as the Solanum serrata cross-regional diffusion network.

[0104] like Figure 10 As shown in the figure, the results of analyzing the source areas in the straw trade network showed that there were potential source areas with Solanum serrata distributed in millet fields, such as the source areas in Zhangjiakou City and Chifeng City, and Solanum serrata plants were intercepted in the grass markets or herders in their downstream transshipment; while there were no potential source areas with Solanum serrata distributed in millet fields, such as the source areas in Tongliao City and Ulanqab City, and no Solanum serrata plants were intercepted in the grass markets or herders in their downstream transshipment.

[0105] Further analysis of the correlation between the centrality of diffusion network nodes and the occurrence frequency of Solanum truncatum was conducted, such as Figure 2 As shown, the results show that in the diffusion network nodes from Chifeng and Zhangjiakou, the degree centrality of the diffusion network nodes D(i) , betweenness centrality C(i) and closeness centrality B(i) The results showed a significant positive correlation with the frequency of Solanum oxyphylla distribution (P < 0.05), that is, the higher the centrality of the node, the higher the frequency of Solanum oxyphylla distribution, indicating that the higher the centrality of the node in the diffusion network, the higher the risk of Solanum oxyphylla invasion. The diffusion network nodes from Chifeng and Zhangjiakou were judged to be high-risk areas.

[0106] Table 2 Correlation between node centrality and distribution frequency in the Solanum truncatum diffusion network

[0107]

[0108] In summary, stable isotope analysis can identify the source of the invasive plant Solanum serratum in newly introduced areas. This method yielded high accuracy for tracing straw and its contaminated branches, exceeding 80%. Tracing results were not significantly different among the three plant subsamples, confirming that the straw trade network represents a network of Solanum serratum dispersal pathways. Furthermore, based on isotope tracing, a diffusion network model constructed by combining Solanum serratum occurrence data and straw cultivation data identified high-risk areas for Solanum serratum invasion as key nodes with high centrality in the diffusion network.

[0109] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

[0110] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for tracing the spread of invasive plants using stable isotopes, characterized in that: It includes the analysis of the origin of invasive alien plants and the identification of the risk of their spread; the invasive alien plant is Solanum spicata; among them, The invasive alien plant source tracing analysis includes the following steps S1-S4; S1. Plant sample collection: S11. Determine the newly introduced area; S12. Collect straw purchased by farmers in the newly introduced area and the remains and seeds of Solanum serratum plants mixed therein as plant samples from the newly introduced area; S13. Collect data to determine the potential source of the newly introduced plant samples; wherein the data collection includes: investigating the source of the straw purchased from the newly introduced area, collecting spatiotemporal data on the planting of the straw across the country, and collecting spatiotemporal data on the occurrence of Solanum serratum across the country; S14. Collect Solanum serratum and straw from the potential source as plant samples from the potential source; S2, sample pretreatment; S3. Stable isotope determination: The pretreated subsample is subjected to stable isotope determination to calculate the stable isotope ratio of the subsample; the stable isotope ratio includes δ 13 C, δ 15 N, δ 18 O, δ 2 H; in the detection of carbon and nitrogen stable isotopes: the combustion tube temperature was set at 950°C, the reduction tube temperature was set at 600°C, the carrier gas was high-purity helium, the reference gases were CO2 and N2, and the oxygen flow rate was set at 40 mL min -1 , the oxygen injection time was 70 seconds; in the detection of hydrogen and oxygen stable isotopes: the cracking furnace temperatures of hydrogen and oxygen were set to 1080℃ and 1180℃ respectively, and the helium purge flow rate was 230 mL·min -1 , the reference gases are CO and H2; S4. Analyze the spread sources and spread paths of the alien invasive plants: Compare the stable isotope ratios of the subsamples of the plant samples from the newly introduced area with the stable isotope ratios of the subsamples of the plant samples from the potential source area, and use statistical methods to process the data to analyze the spread sources and spread paths of the alien invasive plants; The identification of the risk of spread of invasive alien plants includes the following steps P1 and P2; P1. Constructing a diffusion network for the invasive alien plant; P11. Identifying diffusion network nodes: Based on the diffusion sources and diffusion paths of the invasive alien plant analyzed in step S4, determining the diffusion network nodes for the invasive alien plant, including the straw production site, logistics transit center, straw distribution site, straw market for transferring the straw, and markets or farmers and herdsmen receiving the straw; P12. Generating road network links between the diffusion network nodes; P13. Constructing a straw trade network based on the diffusion network nodes and the road network links; P2. Identify diffusion risk areas: Identify risk areas for invasion by invasive alien plants in the straw trade network by analyzing the correlation between the centrality of the diffusion network nodes and the distribution frequency of the invasive alien plants.

2. The method for tracing the spread of invasive alien plants using stable isotopes according to claim 1, characterized in that: The method for determining the new introduction site in step S11 is: based on the national spatiotemporal distribution data of the alien invasive plants, the areas where the alien invasive plants have been newly recorded in the past five years are selected as the new introduction sites.

3. The method for tracing the spread of invasive plants using stable isotopes according to claim 1, characterized in that: The pre-processing of step S2 includes: S21, dust and debris removal; S22, drying: drying the newly introduced plant samples and the potential source plant samples after dust and impurity removal to a constant weight; S23, crushing: crushing the dried plant samples of the newly introduced area and the plant samples of the potential source area, and then sieving; S24. Sampling: Take 2-3 mg of the sieved sample as a subsample.

4. The method for tracing the spread of invasive alien plants using stable isotopes according to claim 1, characterized in that: In step P2, the centrality measurement indicators include degree centrality, betweenness centrality and closeness centrality; Degree Centrality D(i) The calculation formula is shown in formula (1): （1）； Where, d i is the degree of node i, N is the total number of nodes in the network; Betweenness Centrality C(i) The calculation formula is shown in formula (2): （2）； Where, β ij Indicates the starting node i To the end j The number of links contained in the shortest path, N is the total number of nodes in the network; closeness centrality B(i) The calculation formula is shown in formula (3): （3） In the formula α xy For nodes x To Node y The number of all shortest paths, α xy (i) For nodes x To Node y and passing through the node i The number of all shortest paths.

5. The method for tracing the spread of invasive alien plants using stable isotopes according to claim 4, characterized in that: When the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node are all significantly correlated with the distribution frequency of the alien invasive plant, the diffusion network node is determined to be a high-risk invasion area; When one or two of the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node are significantly correlated with the distribution frequency of the alien invasive plant, the diffusion network node is determined to be a medium-risk area for invasion; When the degree centrality, the betweenness centrality, and the closeness centrality of the diffusion network node do not show a significant correlation with the distribution frequency of the alien invasive plant, the diffusion network node is determined to be a low-risk invasion area.

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