Method for preventing and treating pecan root rot by using chlorine dioxide to disinfect soil and applying microbial agent

Abstract

The invention relates to the technical field of hickory planting, in particular to a hickory root rot prevention and control technology for disinfecting soil by using chlorine dioxide and applying a microbial agent. According to the technical scheme, the method comprises the steps of sample preparation and treatment, sampling and measurement, physiological and biochemical data analysis and microbial community analysis, sample preparation comprises the steps of sampling soil and selecting pecan seedlings, and sample treatment comprises the steps of dividing the sampled soil into six groups, the treated soil in each group is put into a potted plant, a hickory nut seedling is planted, after six weeks, the rhizosphere soil of each pot of plant is sampled, the physicochemical property of the soil, the enzyme activity of the soil and the enzyme activity of a root system are measured, and all rhizosphere soil is collected for amplicon sequencing. The microbial agent is applied after the chlorine dioxide soil with proper concentration is disinfected, so that the growth speed and quality of the hickory nut can be improved, the physical and chemical properties and micro-ecological environment of the soil are improved, and the disease resistance of the hickory nut is enhanced.

Description

Technical Field

[0001] The invention relates to the technical field of pecan planting, and in particular to a method for preventing and controlling pecan root rot by using chlorine dioxide to disinfect the soil and applying a microbial agent. Background Art

[0002] Pecan has high economic and nutritional value. The planting area of ​​pecan is expanding year by year, but its growth process is often affected by soil diseases and insect pests, resulting in a decrease in yield and quality. In order to increase the growth rate and yield of pecan plants, researchers have been looking for effective soil disinfection methods. Traditional soil disinfection methods mainly include chemical pesticides, high-temperature steam, radiation, etc., but these methods have certain limitations. Chemical pesticides can easily cause environmental pollution and damage the ecological balance of the soil; high-temperature steam and radiation treatment costs are high and have strict requirements on equipment. Therefore, seeking an environmentally friendly, efficient and low-cost soil disinfection method has become a hot topic in current research.

[0003] Chlorine dioxide is a highly oxidizing disinfectant widely used in water treatment, food processing and other fields. Studies have found that chlorine dioxide also has significant effects in agricultural soil disinfection. However, the current research on chlorine dioxide in pecan soil disinfection is still insufficient. Although soil disinfection can kill most pathogens in the soil, the stability of the microbial community is destroyed, which can easily lead to the aggravation of the disease. Adding microbial agents during the community recovery process can make the soil community develop in a positive direction. Therefore, this study aims to explore a method for the prevention and control of pecan root rot using chlorine dioxide soil disinfection combined with microbial agents.

[0004] Therefore, we propose a method of using chlorine dioxide soil disinfection combined with the application of microbial agents to prevent and control pecan root rot to solve the existing problems. Summary of the invention

[0005] The purpose of the present invention is to solve the problems existing in the background technology and to provide a method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with the application of microbial agents.

[0006] To achieve the above object, the present invention provides the following technical scheme: a method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents, comprising sample preparation and processing, sampling and measurement, soil and root analysis and microbial community analysis, wherein the sample preparation comprises sampling the soil and selecting pecan seedlings, and the sample processing comprises dividing the sampled soil into six groups, each group corresponding to a different treatment: a clear water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, 900mg / L chlorine dioxide + microbial agent group, the soil treated in each group was placed in pots, with three replicates in each group and nine pots in each replicate. A pecan seedling was planted in each pot for routine maintenance management. After six weeks, the root circle soil of each pot of plant was sampled by the sampling and measurement, the physical and chemical properties of the soil, the soil enzyme activity and the root system were determined by the soil and root system analysis, the enzyme activity in the root system was determined, and all rhizosphere soils were collected for amplicon sequencing through the microbial community analysis.

[0007] Preferably, the sample preparation includes selecting a pecan orchard as a sampling site, in which the pecan trees have typical root rot symptoms such as root rot and blackening, and treetop withering, and randomly selecting a plurality of diseased trees in the orchard as sampling points, using a soil drill or a shovel to dig the soil around the roots from the surface to a depth of 20 cm, and mixing the soil collected from different points evenly.

[0008] Preferably, the sample treatment includes dividing the sampled soil into six groups, each group corresponding to a different treatment: a clean water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, and a 900 mg / L chlorine dioxide + microbial agent group.

[0009] Preferably, the corresponding soil treatment includes: adding only clean water, adding only microbial agents, adding 600 mg / L and 900 mg / L chlorine dioxide solutions respectively, and adding 600 mg / L and 900 mg / L chlorine dioxide solutions respectively and then adding microbial agents ten days later.

[0010] Preferably, the measurement includes measuring the plant height, ground diameter, and fresh weight growth indicators of pecan seedlings with similar growth conditions before the experiment and the plant height, ground diameter, and fresh weight growth indicators of pecan plants six weeks after the experiment, evaluating the disease of the plants, and calculating the disease index.

[0011] Preferably, the disease assessment includes determining disease grading standards, dividing the disease into several levels according to the characteristics of the disease, setting corresponding disease descriptions and scoring standards for each level, and investigating and recording the number of plants with each level of disease.

[0012] Preferably, the condition of each sample plant is observed, and the number of plants with each disease level is recorded according to the disease grading standard. The disease index is calculated according to the disease index calculation formula. The lower the disease index, the lighter the disease of the plant; the higher the disease index, the more serious the disease of the plant, thereby determining the impact on the growth and incidence of pecans.

[0013] Preferably, the soil physicochemical properties to be measured include pH value, organic matter, available nitrogen, available phosphorus, and available potassium; the soil enzyme activities to be measured include urease, sucrase, acid phosphatase, and catalase, thereby measuring the effects on the physicochemical properties of pecan soil; and the root enzyme activities to be measured include propionaldehyde, peroxidase, catalase, and superoxide dismutase, thereby measuring the effects on pecan root enzyme activities.

[0014] Preferably, the amplicon sequencing includes processing the soil sample to extract total DNA from the soil sample, selecting the target region, performing polymerase chain reaction using universal primers, and performing data analysis to better understand the effect of soil disinfection treatment on the microbial community structure.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] The method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with application of microbial agents is used. According to experimental results, in the soil disinfection treatment with chlorine dioxide at a suitable concentration, the number of pathogenic bacteria communities is significantly reduced, and the community structure tends to be healthy and stable by applying the microbial agents. By optimizing the use concentration and treatment method of chlorine dioxide, the growth of pecan can be effectively promoted, the occurrence of root rot can be prevented, the cultivation period can be shortened, and the economic benefits can be improved.

[0017] Improve soil physical and chemical properties: Chlorine dioxide can effectively kill harmful microorganisms in the soil, and microbial agents can increase the abundance of beneficial microorganisms in the soil community, reduce the incidence of soil-borne diseases, improve soil structure, and increase fertility levels.

[0018] Promote healthy root development: Studies have shown that appropriate amounts of chlorine dioxide treatment can help stimulate the growth and development of pecan roots, enhance their ability to absorb nutrients, and provide sufficient nutrient support for the plant.

[0019] Reduce the risk of pesticide residues: Compared with traditional chemical pesticide control measures, chlorine dioxide, as an environmentally friendly disinfectant, can effectively control pests and diseases without developing drug resistance, reducing environmental pollution and food safety risks.

[0020] Save costs and improve efficiency: The implementation of this method not only improves the quality and yield of pecans, but also reduces the input cost in the production process, and realizes the rational allocation and efficient utilization of resources.

[0021] In addition, after chlorine dioxide treatment, the application of microbial agents can increase the number of beneficial microorganisms in the soil and reduce pathogenic bacteria, thereby improving the soil microecological environment and enhancing the disease resistance of pecans. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is a schematic diagram of the technical route of the present invention. DETAILED DESCRIPTION

[0023] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0024] Embodiment 1

[0025] like Figure 1 As shown, the method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection and applying microbial agents proposed by the present invention includes sample preparation and processing, sampling and measurement, soil and root analysis and microbial community analysis. The sample preparation includes sampling the soil and selecting pecan seedlings. The sample processing includes dividing the sampled soil into six groups, each group corresponding to a different treatment: a clear water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, and a 900 mg / L chlorine dioxide + microbial agent group. The soil treated in each group is respectively placed in pots, with three replicates in each group and nine pots in each replicate. A pecan seedling is planted in each pot for routine maintenance management. After six weeks, the root circle soil of each pot plant is sampled by sampling and measurement, the physical and chemical properties of the soil, the soil enzyme activity and the root system are determined by soil and root system analysis, the enzyme activity in the root system is determined, and all rhizosphere soils are collected for amplicon sequencing through microbial community analysis.

[0026] Embodiment 2

[0027] like Figure 1As shown, the present invention proposes a method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with the application of microbial agents. Compared with Example 1, this embodiment also includes: sample preparation includes selecting pecan trees in the park with typical root rot symptoms such as root rot and blackening, and treetop withering, randomly selecting multiple diseased trees in the park as sampling points, using a soil drill or a shovel to dig the soil around the roots to a depth of 20 cm from the surface, and mixing the soil collected from different points evenly to reduce spatial variability.

[0028] Sample treatment includes dividing the sampled soil into six groups, each group corresponding to a different treatment: a water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, and a 900 mg / L chlorine dioxide + microbial agent group. The corresponding soil treatments include: adding only water, adding only microbial agents, adding 600 mg / L and 900 mg / L chlorine dioxide solutions, and adding microbial agents ten days after adding 600 mg / L and 900 mg / L chlorine dioxide solutions. The treated soils of each group were potted separately, with three replicates in each group, and each weight was 1. Nine pots were repeated, and pecan seedlings with similar growth conditions were selected. One seedling was planted in each pot. Before the experiment, the plant height, ground diameter, and fresh weight growth indicators of each seedling were measured and recorded. The pecan seedlings were planted in the treated soil and routine maintenance management was carried out, including regular watering to keep the soil moist and avoid drought or waterlogging. The growth conditions of the plants were observed and recorded, such as whether there were any diseases. After 6 weeks, the root circle soil of each pot of plants was sampled, and the plant height, ground diameter, and fresh weight data were recorded at the same time. The plant height measurement included the preparation of tools: a tape measure or a height meter was placed vertically, and the tape measure or height meter was measured from the base of the plant (at the ground) to the top of the crown to accurately record the height of each plant;

[0029] The ground diameter measurement includes wrapping a ruler around the trunk about 5 cm above the base of the plant and reading the ground diameter value; or using a digital caliper to directly measure the trunk diameter and record the ground diameter size;

[0030] Fresh weight measurement involves preparing an electronic scale, uprooting the plant from the soil, removing the soil from the roots, weighing it using an electronic scale, recording the fresh weight of the plant, and recording the effects of different treatments on the amount of plant growth.

[0031] Embodiment 3

[0032] like Figure 1As shown, the method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents proposed by the present invention, compared with the second embodiment, this embodiment also includes: disease assessment includes determining the disease classification standard, the above 135 plant samples have a sufficient number to reflect the disease conditions of the plants under different conditions, according to the characteristics of the disease, the disease is divided into several levels, and corresponding disease descriptions and scoring standards are set for each level, for example:

[0033] Level 0: No symptoms, no browning of roots

[0034] Level 1: Less than 25% of leaves turn yellow, and less than 1 / 4 of the roots turn brown

[0035] Level 3: Less than 50% of leaves turn yellow, and less than 1 / 3 of the roots turn brown

[0036] Level 5: Less than 75% of leaves turn yellow, more than 1 / 2 of the roots turn brown, and the plant wilts

[0037] Level 7: All leaves of the plant turn yellow, the roots rot or turn brown, and the plant dies.

[0038] Investigate and record the number of plants with different levels of disease, observe the disease condition of each sample plant, record the number of plants with different levels of disease condition according to the disease classification standard, and calculate the disease index according to the disease index calculation formula, where the disease index calculation formula is as follows:

[0039] DI = [(the sum of the representative values ​​of each disease level × the number of plants with the corresponding disease level) / (the total number of plants surveyed × the representative value of the highest disease level)] × 100

[0040] The specific steps are as follows:

[0041] Calculate the product of the representative value of each disease level and the number of plants of the corresponding disease level, and sum them up.

[0042] Calculate the product of the total number of plants surveyed and the representative value of the highest disease level.

[0043] (3) Divide the result of step 1 by the result of step 2, and then multiply by 100 to obtain the disease index.

[0044] For example:

[0045] Assume that 100 plants were surveyed and the number of plants with different disease levels is as follows:

[0046] Level 0: 70 plants

[0047] Level 1: 20 plants

[0048] Level 3: 5 plants

[0049] Level 5: 3 plants

[0050] Level 7: 2 plants

[0051] According to the disease index calculation formula:

[0052] DI=[(0×70+1×20+3×5+5×3+7×2) / (100×7)]×100=[(0+20+15+15+14) / 700]×100=64 / 700×100=0.091×100=9.1

[0053] Therefore, the disease index of the field is 9.1. The lower the disease index, the lighter the disease of the plant; the higher the disease index, the more serious the disease of the plant, so as to determine the effect of soil disinfection with different concentrations of chlorine dioxide on the growth and incidence of pecan.

[0054] Embodiment 4

[0055] like Figure 1 As shown, the method for controlling pecan root rot using chlorine dioxide soil disinfection combined with microbial agents proposed by the present invention, compared with Example 1, this embodiment also includes: the soil physical and chemical properties to be measured include pH value, organic matter, effective nitrogen, fast-acting phosphorus, and fast-acting potassium, the soil enzyme activities to be measured include urease, sucrase, acid phosphatase, and catalase, and the rhizosphere soil collected is the sample;

[0056] Where pH value:

[0057] After some samples were air-dried to remove impurities, 10 g of soil sample was weighed, 50 ml of deionized water was added, and the mixture was shaken for 5 minutes, and then allowed to stand for 30 minutes. The pH value of the supernatant was measured using a pH meter to complete the pH determination;

[0058] Organic matter:

[0059] Weigh 0.5g of soil sample and put it into a digestion tube. Add 10ml of concentrated sulfuric acid and a catalyst (such as copper sulfate). Digest it in a digester for several hours. After cooling, use a Kjeldahl nitrogen analyzer or colorimetry to determine the organic carbon content, and then convert it into organic matter to complete the determination of organic matter.

[0060] Available nitrogen:

[0061] Weigh 5 g of soil sample, extract it with 1% sodium chloride solution, determine the total nitrogen content of the extract by Kjeldahl method, and calculate the effective nitrogen content.

[0062] Fast-acting phosphorus:

[0063] Weigh 5 g of soil sample, extract it with 0.5 M sodium bicarbonate solution, and measure the extract with molybdenum antimony colorimetric method.

[0064] Fast-Acting Potassium:

[0065] Weigh 5 g of soil sample, extract it with 1 M ammonium acetate solution, and measure the extract by atomic absorption spectrophotometry.

[0066] This is to determine the effects of soil disinfection with different concentrations of chlorine dioxide on the physical and chemical properties of pecan soil.

[0067] The root enzyme activities to be measured include malondialdehyde, peroxidase, catalase, and superoxide dismutase. The steps for measuring soil enzyme activities are as follows:

[0068] Urease:

[0069] Weigh 5 g of soil sample, add 10 ml of 10% urea solution, incubate at a certain temperature for 24 hours, and after extraction, determine the released ammonia using Nessler's reagent or colorimetry.

[0070] Sucrase:

[0071] Weigh 5 g of soil sample, add 10 ml of sucrose solution, incubate at a certain temperature for 24 hours, and after extraction, determine the released reducing sugar using 3,5-dinitrosalicylic acid (DNS) colorimetry.

[0072] Acid Phosphatase:

[0073] Weigh 5 g of soil sample, add appropriate amount of phosphate buffer and substrate (such as p-nitrophenyl phosphate), incubate at a certain pH and temperature for a certain period of time, and after extraction, determine the released p-nitrophenol by colorimetry.

[0074] Catalase:

[0075] Weigh 5g of soil sample, add 10ml of hydrogen peroxide solution, and incubate at a certain temperature for a period of time. Calculate the enzyme activity by measuring the reduction of hydrogen peroxide. Usually use colorimetry or titration to determine the effect of different concentrations of chlorine dioxide soil disinfection treatment on the enzyme activity of pecan roots.

[0076] During the measurement process, the experimental conditions, including temperature, time, concentration of reagents, etc., need to be strictly controlled to ensure the accuracy and repeatability of the data. At the same time, in order to reduce errors, each sample should be set up at least three times and the average value should be calculated. During the operation, laboratory safety regulations must be followed to ensure safety.

[0077] Embodiment 5

[0078] like Figure 1As shown, the method for controlling pecan root rot using chlorine dioxide soil disinfection combined with microbial inoculants proposed by the present invention, compared with the first embodiment, this embodiment also includes: amplicon sequencing, which includes filtering and centrifuging all rhizosphere soil samples collected to remove impurities, and then using a commercial DNA extraction kit or a laboratory's own extraction method to extract total DNA from the soil samples;

[0079] Select the appropriate target region according to the research purpose, such as 16S rRNA gene for bacterial diversity analysis and ITS region for fungal diversity analysis, and use universal primers to amplify the target DNA region.

[0080] Purify the amplified DNA fragment from the PCR reaction mixture using a purification kit or gel electrophoresis;

[0081] The purified PCR products are fragmented to suit the requirements of different sequencing platforms, and sequencing platform-specific adapters are attached to both ends of the DNA fragments to add unique molecular tags (indexes) to each sample so that different samples can be distinguished in subsequent analysis;

[0082] The constructed library was quality controlled and quantified, and according to the guidelines of the sequencing platform (Illumina MiSeq), the library was loaded onto the sequencer for sequencing;

[0083] The quality control software (FastQC) was used to evaluate the quality of the raw data obtained by sequencing, and low-quality reads, adapter sequences, and possible contamination sequences were removed. The filtered reads were spliced ​​to obtain complete amplicon sequences. The sequences were clustered using the OTU clustering method (UPARSE) to form OTUs. The OTU representative sequences were compared with the database (Greengenes) for species annotation. The diversity index (Shannon-Wiener index) was used to evaluate the diversity of the samples. Statistical analysis was performed, such as α diversity analysis, β diversity analysis, difference analysis, and co-occurrence network analysis. For α diversity analysis, the following α diversity index was calculated for each sample:

[0084] Shannon-Wiener index (reflects community richness)

[0085] Simpson index (reflects community uniformity)

[0086] Chao1 Index (estimates the total number of species in a community)

[0087] Faith's PD index (reflecting community phylogenetic diversity);

[0088] The differences in alpha diversity index among different treatment groups were compared using a nonparametric test (Kruskal-Wallis test);

[0089] Beta diversity analysis:

[0090] Use OTU table to compare samples, calculate Bray-Curtis distance and Unifrac distance to evaluate the differences between samples, convert distance matrix into two-dimensional or three-dimensional space to visualize the differences between samples, and use PERMANOVA or ANOSIM test to evaluate the significant differences in β diversity between different treatment groups;

[0091] Difference analysis:

[0092] The LEfSe tool was used to perform a significant difference test on OTUs to determine which OTUs were significantly different between different treatment groups, and to perform taxonomic annotations on them. The functional changes of the microbial community were predicted based on the differential OTUs.

[0093] Co-occurrence network analysis:

[0094] The SparCC network analysis tool was used to construct a co-occurrence network based on OTU abundance data, and the network topology parameters such as degree, betweenness, and clustering coefficient were calculated. The MCL method was used to detect modules in the network (interrelated microbial groups in the community) and identify key species in the network (nodes with high degree or betweenness), which may play an important role in community function.

[0095] To explore the differences between different samples, through the above steps, researchers can obtain detailed information on the microbial community in the rhizosphere soil of pecan, so as to better understand the effects of different treatments on the structure of the microbial community.

[0096] The above-mentioned specific embodiments are only several preferred embodiments of the present invention. Based on the technical solutions of the present invention and the relevant inspirations of the above-mentioned embodiments, those skilled in the art can make various alternative improvements and combinations to the above-mentioned specific embodiments.

[0097] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above and that the present invention can be implemented in other specific forms without departing from the spirit or essential features of the present invention. Therefore, the embodiments should be considered exemplary and non-restrictive in all respects, and the scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims be included in the present invention.

Claims

1. A pecan root rot prevention and control technology using chlorine dioxide soil disinfection combined with microbial agents, including sample preparation and processing, sampling and measurement, soil and root system analysis and microbial community analysis, characterized in that: The sample preparation includes sampling the soil and selecting pecan seedlings. The sample treatment includes dividing the sampled soil into six groups, each group corresponding to a different treatment: a clear water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, and a 900 mg / L chlorine dioxide + microbial agent group. The soil treated in each group is respectively placed in pots, with three replicates in each group and nine pots in each replicate. A pecan seedling is planted in each pot for routine maintenance management. After six weeks, the root circle soil of each pot of plant is sampled through the sampling and measurement. The physical and chemical properties of the soil and the soil enzyme activity are determined through the soil and root system analysis, and the enzyme activity in the root system is determined by extracting the root system. All rhizosphere soils are collected and the microbial community analysis is performed through amplicon sequencing.

2. The method for controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents according to claim 1 is characterized in that: The sample preparation includes selecting a pecan orchard with severe root rot as the sampling site. The pecan trees in the orchard have typical root rot symptoms such as root rot and blackening, and treetop withering. Several diseased trees are randomly selected in the orchard as sampling points, and the soil around the roots to a depth of 20 cm is dug using a soil drill or shovel, and the soil collected from different points is mixed evenly.

3. The method for controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents according to claim 1 is characterized in that: The sample treatment includes dividing the sampled soil into six groups, each group corresponding to a different treatment: a clean water control group, a microbial agent control group, a 600 mg / L chlorine dioxide group, a 900 mg / L chlorine dioxide group, a 600 mg / L chlorine dioxide + microbial agent group, and a 900 mg / L chlorine dioxide + microbial agent group.

4. A method for controlling pecan root rot using chlorine dioxide soil disinfection combined with microbial agents according to claim 3, characterized in that: Only clean water was added; only microbial agents were added; 600 mg / L and 900 mg / L of chlorine dioxide solution were added respectively; and microbial agents were added ten days after 600 mg / L and 900 mg / L of chlorine dioxide solution were added respectively.

5. The method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents according to claim 1, characterized in that: The measurement includes measuring the plant height, ground diameter, and fresh weight growth indexes of pecan seedlings with similar growth conditions before the experiment, and the plant height, ground diameter, and fresh weight growth indexes of pecan plants six weeks after the experiment, so as to reflect the growth conditions of pecans under different treatments.

6. A method for controlling pecan root rot using chlorine dioxide soil disinfection combined with microbial agents according to claim 5, characterized in that: The disease assessment includes determining disease grading standards, classifying the disease into several grades according to the characteristics of the disease, setting corresponding disease descriptions and scoring standards for each grade, and investigating and recording the number of plants with each grade of disease.

7. A method for controlling pecan root rot using chlorine dioxide soil disinfection combined with microbial agents according to claim 6, characterized in that: The disease condition of each sample plant is observed, and the number of plants with each disease condition is recorded according to the disease grading standard. The disease index is calculated according to the disease index calculation formula. The lower the disease index, the lighter the disease of the plant; the higher the disease index, the more serious the disease of the plant, thereby determining the impact on the incidence of pecan.

8. The method for preventing and controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents according to claim 1, characterized in that: The soil physicochemical properties to be measured include pH value, organic matter, available nitrogen, available phosphorus, and available potassium; the soil enzyme activities to be measured include urease, sucrase, acid phosphatase, and catalase, so as to measure the effects on the physicochemical properties of pecan soil; the root enzyme activities to be measured include malondialdehyde, peroxidase, catalase, and superoxide dismutase, so as to measure the effects on the root enzyme activities of pecan.

9. The method for controlling pecan root rot by using chlorine dioxide soil disinfection combined with microbial agents according to claim 1, characterized in that: The amplicon sequencing includes processing the soil sample to extract total DNA from the soil sample, selecting the target area, using universal primers for fungi and bacteria to perform polymerase chain reaction, performing high-throughput sequencing, and performing data analysis on the sequencing data, so as to better understand the impact of soil disinfection treatment on the structure of the microbial community.

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