A method for identifying resistance to macadamia anthracnose

Abstract

This invention discloses a method for identifying anthracnose resistance in macadamia nuts, belonging to the field of macadamia nut resistance breeding technology. The method includes steps such as: isolation of the anthracnose pathogen, preparation of mycelial cakes, inoculation of healthy macadamia nut leaves by scrambling, disease index classification, and evaluation of resistance and susceptibility characteristics. This application involves culturing macadamia anthracnose to obtain purified anthracnose mycelia, then inoculating healthy leaves by scrambling; classifying the disease index by the diameter of leaf lesions, and evaluating the anthracnose resistance and susceptibility characteristics of macadamia nuts based on the disease index; providing a convenient method for identifying anthracnose-resistant macadamia nut varieties.

Description

Technical Field

[0001] This invention belongs to the field of macadamia nut resistance breeding technology, specifically relating to a method for identifying macadamia nut anthracnose resistance. Background Technology

[0002] Anthracnose is one of the common diseases of macadamia nuts, mainly affecting the leaves, young shoots and fruits. Symptoms include the death of young shoots, yellowing of young leaves with black patches on the surface, and dark brown coloration of young fruits and shells, which gradually develop into black conidiophores, affecting the yield and quality of macadamia nuts.

[0003] Chemical agents are a common means of controlling anthracnose, but long-term use of a single chemical agent can lead to drug resistance in anthracnose. Currently, anthracnose has developed varying degrees of resistance to existing chemical agents. In order to improve the control effect, fruit growers usually increase the dosage of chemical agents, which will accelerate the development of resistance and also lead to pesticide residue problems.

[0004] Therefore, breeding disease-resistant varieties can avoid the resistance problems caused by the use of chemical agents. Breeding anthracnose-resistant varieties of macadamia nuts requires precise resistance identification. Currently, there is no readily available method for identifying macadamia nut resistance. Summary of the Invention

[0005] In view of the above, it is necessary to provide a method for identifying resistance to macadamia anthracnose, so as to identify resistant varieties quickly and accurately.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: A method for identifying resistance to macadamia anthracnose, comprising the following steps: S1. Material Preparation Prepare macadamia nut anthracnose fungal cakes, 6-10 mm in size; Harvest healthy, fresh, and uniformly growing macadamia nut leaves, disinfect them, and then dry them. S2, Inoculation via laceration The fungal cake was fixed to the scratched area of ​​the macadamia leaf and placed in a light incubator with a temperature of 30℃-32℃ and a humidity of 70%, with 12 hours of light and 12 hours of darkness. S3, Resistance Evaluation Seven days after inoculation, the diameter of the lesions was measured, and the percentage of the lesion area covering the entire leaf was calculated to confirm the disease severity of the macadamia nut. Grade 0: The area of ​​the lesion accounts for 0% of the total area of ​​the sample tissue; Grade 1: 0% < lesion area accounts for < 10% of the total sample tissue area; Grade 2: 10% ≤ lesion area accounts for < 20% of the total sample tissue area; Grade 3: 20% ≤ lesion area accounts for < 50% of the total sample tissue area; Grade 4: 50% ≤ lesion area accounts for < 75% of the total sample tissue area; Grade 5: 75% ≤ lesion area as a percentage of the total sample tissue area; The formula for calculating the disease index is: Incidence rate I (%) = (Number of cases investigated / Total number of cases investigated) × 100 Disease Index (DI) = [(∑leaf size for each disease level × representative value for each disease level) / (total number of leaves surveyed × highest disease level value)] × 100; Based on the disease index, the resistance level of macadamia nut anthrax is divided into 5 categories: High resistance (HR): 0 ≤ DI < 10; Disease resistance (R): 10 ≤ DI < 25; Mid-range antibody (MR): 25 ≤ DI < 45; Sickness (S): 45 ≤ DI < 75; High sensitivity (HS): DI≥75.

[0007] Furthermore, the method for preparing purified mycelial cake is as follows: collect diseased specimens from macadamia leaves, rinse them with sterile water, cut off the diseased tissue at the junction of diseased and healthy tissue, disinfect and rinse with sterile water again, air dry, and then invert them on the culture medium for cultivation; when the anthracnose pathogen has covered the slant culture medium, take a single mycelium from the edge of the colony for purification and cultivation, repeat the substitution purification three times to obtain purified mycelial cake.

[0008] Further purification and culture were carried out for 4-7 days at a temperature of 25°C.

[0009] Furthermore, the inoculated macadamia leaves were placed in a light incubator and first treated in the dark at 30℃-32℃ for 24 hours.

[0010] Furthermore, the nutritional components of the culture medium include the following: sweet potato flour, glucose, yeast powder, maifanite, sucrose, peptone, calcium nitrate, hydrogel, and vitamins.

[0011] Furthermore, the hydrogel is a chitosan-hyaluronic acid hydrogel, and the weight percentage of chitosan is 15-20%; the molecular weight of the chitosan is ≤2.5×10⁻⁶. 5 Its weight percentage is 8-12%.

[0012] Furthermore, the method for preparing the nutrients in the culture medium is as follows: S1, sweet potato powder is added to water, heated and stirred to dissolve, and stirred to obtain substance A; S2, maifan stone is ground and treated, added to alkaline solution and ultrasonically treated for 3-5 minutes, precipitated, filtered, and dried to obtain maifan stone powder; S3, glucose, yeast powder, and vitamins are taken, added to water and stirred to dissolve, then maifan stone powder is added and stirred evenly to obtain substance B; S4, glucose, sucrose, yeast powder, peptone, and calcium nitrate are taken, added to water and stirred to dissolve, then chitosan-hyaluronic acid hydrogel is added and stirred evenly to obtain substance C; S5, substance C is added to substance B and stirred evenly, then substance A is added, and stirring is continued until uniform, dried, and pulverized to obtain the nutrients.

[0013] Furthermore, in step S2, the maifan stone is ground to a particle size of 100-150 mesh; before adding the alkaline solution, the ground maifan stone is first added to the acid solution and stirred for 1-2 minutes, then filtered and washed until neutral.

[0014] Furthermore, the weight ratio of glucose in step S3 to glucose in step S4 is 1:2-5; the weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:0.4-1.

[0015] Furthermore, the nutrient particles obtained by pulverizing in step S4 have a particle size of 0.5-2 mm.

[0016] The present invention has the following beneficial effects: This application describes a method for identifying anthracnose-resistant macadamia varieties by culturing the fungus, obtaining purified anthracnose mycelium, and then inoculating healthy leaves with the fungus through scrambling. The application also describes the classification of disease index by statistically analyzing the diameter of leaf lesions, and the evaluation of anthracnose resistance characteristics of macadamia nuts based on the disease index.

[0017] The nutrients used in the culture medium of this invention conform to the growth characteristics of anthrax bacteria, which can accelerate the growth and reproduction of the bacteria and provide a basis for subsequent identification. In the early stage of inoculation, the bacteria grow slowly and consume less nutrients. The carbon source at this stage is sweet potato starch. As the bacteria grow and reproduce, their demand for nutrients increases, and the demand for nitrogen, carbon, vitamins, etc., gradually increases. At this time, the nutrients adsorbed by maifanite are slowly absorbed by the bacteria. Moreover, the loose and porous nature of maifanite also helps the mycelium grow and facilitates the absorption of nutrients. As the bacteria multiply and their numbers increase, the nutrients in the culture medium are consumed, and the chitosan-hyaluronic acid hydrogel is exposed. It swells under alkaline conditions, allowing the nutrients encapsulated in it to flow out, which helps the bacteria absorb them.

[0018] Maifan stone is first acid-treated to make it loose and porous, and then alkali-treated. This allows the metal ions on the maifan stone to adsorb some hydroxide ions. During the mixing process with carbon and nitrogen sources, some complexation reactions occur. When the pH of the bacterial growth medium decreases, the pH can be adjusted by hydroxide ions, and nutrients can be released for the bacteria to absorb. Attached Figure Description

[0019] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.

[0020] Figure 1 Leaf of macadamia nut plant with anthracnose.

[0021] Figure 2 The mycelial growth is shown after one day of cultivation in a light incubator.

[0022] Figure 3 The mycelial growth is shown after 7 days of cultivation in a light incubator. Detailed Implementation

[0023] To facilitate a better understanding of the present invention, the following examples are provided in conjunction with the accompanying drawings. These examples fall within the scope of protection of the present invention, but do not limit the scope of protection of the present invention.

[0024] Example A method for identifying resistance to macadamia anthracnose, comprising the following steps: S1. Material Preparation Prepare macadamia nut anthracnose fungal cakes, 6-10 mm in size; Harvest healthy, fresh, and uniformly growing macadamia nut leaves, disinfect them, and then dry them. S2, Inoculation via laceration The fungal cake was fixed to the scratched area of ​​the macadamia leaf and placed in a light incubator with a temperature of 30℃-32℃ and a humidity of 70%, with 12 hours of light and 12 hours of darkness. S3, Resistance Evaluation Seven days after inoculation, the diameter of lesions was measured, and the percentage of the lesion covering the entire leaf area was calculated to determine the disease resistance index of macadamia nuts. Grade 0: The area of ​​the lesion accounts for 0% of the total area of ​​the sample tissue; Grade 1: 0% < lesion area accounts for < 10% of the total sample tissue area; Grade 2: 10% ≤ lesion area accounts for < 20% of the total sample tissue area; Grade 3: 20% ≤ lesion area accounts for < 50% of the total sample tissue area; Grade 4: 50% ≤ lesion area accounts for < 75% of the total sample tissue area; Grade 5: 75% ≤ lesion area as a percentage of the total sample tissue area; The formula for calculating the disease index is: Incidence rate I (%) = (Number of cases investigated / Total number of cases investigated) × 100 Disease Index (DI) = [(∑ number of leaves at each disease level × representative value of each disease level) / (total number of leaves surveyed × highest disease level value)] × 100; Based on the disease index, the resistance level of macadamia nut anthrax is divided into 5 categories: High resistance (HR): 0 ≤ DI < 10; Disease resistance (R): 10 ≤ DI < 25; Mid-range antibody (MR): 25 ≤ DI < 45; Sickness (S): 45 ≤ DI < 75; High sensitivity (HS): DI≥75.

[0025] The method for preparing purified mycelial cake is as follows: collect diseased specimens from macadamia leaves, rinse them with sterile water, cut off the diseased tissue at the junction of diseased and healthy tissue, disinfect and rinse with sterile water again, air dry, and then invert them on the culture medium for cultivation; when the anthracnose pathogen has covered the slant culture medium, take a single mycelium from the edge of the colony for purification and cultivation, repeat the substitution purification three times to obtain purified mycelial cake.

[0026] Purification culture for 4-7 days at a temperature of 25℃.

[0027] Macadamia nut leaves were placed in a light incubator and first treated in darkness at 30℃-32℃ for 24 hours.

[0028] This application also provides a culture medium for anthrax cultivation, the nutritional components of which include the following components in the following weight ratios: sweet potato flour 1-2%, glucose 1.5-2%, yeast powder 0.6-1.2%, maifanite, sucrose 0.8-1.5%, peptone 0.2-0.8%, calcium nitrate 0.1-0.4%, hydrogel 1.5-2.5%, and vitamins 0.02-0.08%.

[0029] The hydrogel is a chitosan-transparent acid hydrogel, with chitosan accounting for 15-20% by weight; the molecular weight of the chitosan is ≤2.5×10⁻⁶. 5 Its weight percentage is 8-12%.

[0030] The method for preparing the nutrients in the culture medium is as follows: S1, sweet potato powder is added to water, heated and stirred to dissolve, and stirred to obtain substance A; S2, maifan stone is ground and treated, added to alkaline solution and ultrasonically treated for 3-5 minutes, precipitated, filtered, and dried to obtain maifan stone powder; S3, glucose, yeast powder, and vitamins are taken, added to water and stirred to dissolve, then maifan stone powder is added and stirred evenly to obtain substance B; S4, glucose, sucrose, yeast powder, peptone, and calcium nitrate are taken, added to water and stirred to dissolve, then chitosan-hyaluronic acid hydrogel is added and stirred evenly to obtain substance C; S5, substance C is added to substance B and stirred evenly, then substance A is added, and stirring is continued until uniform, dried, and pulverized to obtain the nutrients.

[0031] Step S2: After grinding the maifan stone, the particle size is 100-150 mesh. Before adding the alkali solution, add the ground maifan stone to the acid solution and stir for 1-2 minutes. Filter and wash until neutral.

[0032] The weight ratio of glucose in step S3 to glucose in step S4 is 1:2-5; the weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:0.4-1.

[0033] The nutrient particles obtained by pulverizing in step S4 have a particle size of 0.5-2 mm.

[0034] Example 1 This application also provides a culture medium for anthrax cultivation, the nutritional components of which include the following components in the following weight ratios: 1% sweet potato flour, 1.5% glucose, 0.6% yeast powder, 1% maifanite, 0.8% sucrose, 0.2% peptone, 0.1% calcium nitrate, 1.5% hydrogel, and 0.02% vitamins.

[0035] The hydrogel is a chitosan-transparent acid hydrogel, and the chitosan content is 15% by weight; the molecular weight of the chitosan is ≤2.5×10⁻⁶. 5 Its weight percentage is 8%.

[0036] The method for preparing the nutrients in the culture medium is as follows: S1, sweet potato powder is added to water, heated and stirred to dissolve, and stirred to obtain substance A; S2, maifan stone is ground and treated with alkali solution, ultrasonicated for 3 minutes, precipitated, filtered, and dried to obtain maifan stone powder; S3, glucose, yeast powder, and vitamins are taken, added to water and stirred to dissolve, then maifan stone powder is added and stirred evenly to obtain substance B; S4, glucose, sucrose, yeast powder, peptone, and calcium nitrate are taken, added to water and stirred to dissolve, then chitosan-hyaluronic acid hydrogel is added and stirred evenly to obtain substance C; S5, substance C is added to substance B and stirred evenly, then substance A is added, and stirring is continued until uniform, dried, and pulverized to obtain the nutrients.

[0037] Step S2: After grinding the maifan stone, the particle size is 100 mesh. Before adding the alkaline solution, add the ground maifan stone to the acid solution and stir for 1 minute. Filter and wash until neutral.

[0038] The weight ratio of glucose in step S3 to glucose in step S4 is 1:2; the weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:0.4.

[0039] The nutrient particles obtained by pulverizing in step S4 have a particle size of 0.5 mm.

[0040] Example 2 This application also provides a culture medium for anthrax cultivation, the nutritional components of which include the following components in the following weight ratios: sweet potato flour 1.5%, glucose 1.8%, yeast powder 0.9%, maifanite 1.5%, sucrose 1.2%, peptone 0.5%, calcium nitrate 0.3%, hydrogel 2%, and vitamin 0.05%.

[0041] The hydrogel is a chitosan-transparent acid hydrogel, and the chitosan content is 17% by weight; the molecular weight of the chitosan is ≤2.5×10⁻⁶. 5 Its weight percentage is 10%.

[0042] The method for preparing the nutrients in the culture medium is as follows: S1, sweet potato powder is added to water, heated and stirred to dissolve, and stirred to obtain substance A; S2, maifan stone is ground and treated with alkali solution, ultrasonicated for 4 minutes, precipitated, filtered, and dried to obtain maifan stone powder; S3, glucose, yeast powder, and vitamins are taken, added to water and stirred to dissolve, then maifan stone powder is added and stirred evenly to obtain substance B; S4, glucose, sucrose, yeast powder, peptone, and calcium nitrate are taken, added to water and stirred to dissolve, then chitosan-hyaluronic acid hydrogel is added and stirred evenly to obtain substance C; S5, substance C is added to substance B and stirred evenly, then substance A is added, and stirring is continued until even, dried, and pulverized to obtain the nutrients.

[0043] After grinding the maifan stone in step S2, the particle size is 150 mesh. Before adding the alkaline solution, the ground maifan stone is added to the acid solution and stirred for 1.5 minutes, then filtered and washed until neutral.

[0044] The weight ratio of glucose in step S3 to glucose in step S4 is 1:3; the weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:0.7.

[0045] The nutrient particles obtained by pulverizing in step S4 have a particle size of 1.5 mm.

[0046] Example 3 This application also provides a culture medium for anthrax cultivation, the nutritional components of which include the following components in the following weight ratios: sweet potato flour 2%, glucose 2%, yeast powder 1.2%, maifanite 2%, sucrose 1.5%, peptone 0.8%, calcium nitrate 0.4%, hydrogel 2.5%, and vitamin 0.08%.

[0047] The hydrogel is a chitosan-transparent acid hydrogel, and the chitosan content is 20% by weight; the molecular weight of the chitosan is ≤2.5×10⁻⁶. 5 Its weight percentage is 12%.

[0048] The method for preparing the nutrients in the culture medium is as follows: S1, sweet potato powder is added to water, heated and stirred to dissolve, and stirred to obtain substance A; S2, maifan stone is ground and treated with alkali solution, ultrasonicated for 5 minutes, precipitated, filtered, and dried to obtain maifan stone powder; S3, glucose, yeast powder, and vitamins are taken, added to water and stirred to dissolve, then maifan stone powder is added and stirred evenly to obtain substance B; S4, glucose, sucrose, yeast powder, peptone, and calcium nitrate are taken, added to water and stirred to dissolve, then chitosan-hyaluronic acid hydrogel is added and stirred evenly to obtain substance C; S5, substance C is added to substance B and stirred evenly, then substance A is added, and stirring is continued until uniform, dried, and pulverized to obtain the nutrients.

[0049] After grinding the maifan stone in step S2, the particle size is 150 mesh. Before adding the alkaline solution, the ground maifan stone is added to the acid solution and stirred for 2 minutes, then filtered and washed until neutral.

[0050] The weight ratio of glucose in step S3 to glucose in step S4 is 1:5; the weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:1.

[0051] The nutrient particles obtained by pulverizing in step S4 have a particle size of 2 mm.

[0052] test The macadamia trees used in the experiment were grown in Nanning, with trees aged 5-8 years. The experimental groups were numbered 1-9, and the control group was numbered 10. The resistant macadamia variety was HAES 660, and the susceptible variety was numbered 11. All macadamia trees used in the experiment were growing well and free from pests and diseases.

[0053] The names of the macadamia varieties and their ages used in the experiment are detailed in Table 1.

[0054] Table 1. List of Macadamia Nut Varieties

[0055] Healthy, fresh, mature, and uniformly growing macadamia leaves were collected from each of the 11 varieties, with 11 leaves per variety. After disinfection, the leaves were dried. The epidermis of the macadamia leaves was gently punctured with a needle. Ten leaves were inoculated with the pathogen cultured in Example 3 (the mycelial cake cultured in Example 3 was fixed at the puncture site of the macadamia leaf), and one leaf was inoculated with culture medium as a negative control. After inoculation, the leaves were placed in a light incubator at a temperature of 30°C and a humidity of 70%, with 12 hours of light and 12 hours of darkness. Seven days after inoculation, the lesion area and relative fungal biomass of the leaves were measured.

[0056] The percentage of lesion area was determined by measuring the lesion area using a grid method and then calculating its ratio to the leaf area.

[0057] The relative biomass of fungi was measured using real-time quantitative PCR. Specifically, DNA was extracted from leaves, diluted 5-10 times until the concentration was consistent, and then used for real-time quantitative PCR. The SYBR Super Mix from Vazyme was used as the real-time quantitative PCR reagent, and amplification was performed using a Bio-RAD real-time quantitative PCR instrument. Gene expression levels were calculated using a relative quantitative method, with the expression level of the Ubiquitin gene as a reference. The formula for calculating the relative expression level of the target gene was 2 - (Δtarget gene Ct - ΔUbiquitin Ct). The experimental results were taken as the average of three replicates, and the error was expressed as standard error (SE). The identification results are shown in Table 2.

[0058] Table 2 Results of macadamia nut experiments in each group

[0059] The disease index for each group was calculated based on Table 2, and the results are shown in Table 3.

[0060] Table 3. Results of macadamia nut disease index in each experimental group.

[0061] As shown in Tables 2 and 3, using the identification method of this invention, with varieties HAES 660 and A16 as control groups, the results are as follows: HAES 660 is a highly resistant variety, Guire 1, Guire 5, and Guire 7 are disease-resistant varieties, OC is a moderately resistant variety, JW, 900, Guire Yin 2, and Guire Yin 3 are disease-susceptible varieties, and 695 is a susceptible variety.

[0062] The leaf samples collected in Example 3 are shown below. Figure 1 The cultured mycelia are seen Figure 2 , Figure 3 .

[0063] It should be noted that, in this document, relational terms such as "first" and "second" are used merely to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, the phrase "comprising an element defined as..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0064] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for identifying resistance to macadamia anthracnose, characterized in that, Includes the following steps: S1. Material Preparation Prepare macadamia nut anthracnose fungal cakes, 6-10 mm in size; Harvest healthy, fresh, and uniformly growing macadamia nut leaves, disinfect them, and then dry them. S2, Inoculation via laceration The fungal cake was fixed to the scratched area of ​​the macadamia leaf and placed in a light incubator with a temperature of 30℃-32℃ and a humidity of 70%, with 12 hours of light and 12 hours of darkness. S3, Resistance Evaluation Seven days after inoculation, the diameter of the lesions was measured, and the percentage of the lesion area covering the entire leaf was calculated to confirm the disease severity of the macadamia nut. Grade 0: The area of ​​the lesion accounts for 0% of the total area of ​​the sample tissue; Grade 1: 0% < lesion area accounts for < 10% of the total sample tissue area; Grade 2: 10% ≤ lesion area accounts for < 20% of the total sample tissue area; Grade 3: 20% ≤ lesion area accounts for < 50% of the total sample tissue area; Grade 4: 50% ≤ lesion area accounts for < 75% of the total sample tissue area; Grade 5: 75% ≤ lesion area as a percentage of the total sample tissue area; The formula for calculating the disease index is: Incidence rate I (%) = (Number of cases investigated / Total number of cases investigated) × 100 Disease Index (DI) = [(∑ number of leaves at each disease level × representative value of each disease level) / (total number of leaves surveyed × highest representative value of the disease level)] × 100; Based on the disease index, the resistance level of macadamia nut anthrax is divided into 5 categories: High resistance (HR): 0 ≤ DI < 10; Disease resistance (R): 10 ≤ DI < 25; Mid-range antibody (MR): 25 ≤ DI < 45; Sickness (S): 45 ≤ DI < 75; High sensitivity (HS): DI≥75.

2. The method for identifying macadamia anthracnose according to claim 1, characterized in that, The method for preparing purified mycelia is as follows: Diseased specimens were collected from macadamia leaves, rinsed with sterile water, and diseased tissue at the junction of diseased and healthy tissue was cut off. After disinfection, the tissue was rinsed with sterile water again, dried, and then placed upside down on a culture medium for incubation. When the anthracnose pathogen covered the slant culture medium, a single hyphae from the edge of the colony was taken for purification and culture. This process was repeated three times to obtain purified hyphae.

3. The method for identifying macadamia anthracnose according to claim 2, characterized in that, Purification culture for 4-7 days at a temperature of 25℃.

4. The method for identifying macadamia anthracnose according to claim 3, characterized in that, Macadamia nut leaves were placed in a light incubator and first treated in darkness at 30℃-32℃ for 24 hours.

5. The method for identifying macadamia anthracnose according to claim 4, characterized in that, The nutritional components of the culture medium include the following: sweet potato flour, glucose, yeast powder, maifanite, sucrose, peptone, calcium nitrate, hydrogel, and vitamins.

6. The method for identifying macadamia anthracnose according to claim 5, characterized in that, The hydrogel is a chitosan-transparent acid hydrogel, and the weight percentage of chitosan is 15-20%. The chitosan contained therein has a molecular weight ≤2.5×10 5 Its weight percentage is 8-12%.

7. A method for identifying macadamia anthracnose according to any one of claims 1-6, characterized in that, The method for preparing the nutrients in the culture medium is as follows: S1. Sweet potato starch, add water, heat and stir to dissolve, and stir to obtain substance A; S2. Grind the maifan stone, add it to the alkaline solution and sonicate for 3-5 minutes, precipitate and filter, dry to obtain maifan stone powder; S3. Take glucose, yeast powder, and vitamins, add water and stir to dissolve, then add maifanite powder and stir evenly to obtain substance B. S4. Take glucose, sucrose, yeast powder, peptone, and calcium nitrate, add water and stir to dissolve, then add chitosan-hyaluronic acid hydrogel and stir evenly to obtain substance C; S5. Add substance C to substance B, stir well, then add substance A, continue stirring until uniform, dry, and pulverize to obtain the nutritional components.

8. The method for identifying macadamia anthracnose according to claim 7, characterized in that, After grinding in step S2, the maifan stone has a particle size of 100-150 mesh. Before adding the alkaline solution, add the ground maifan stone to the acid solution and stir for 1-2 minutes, then filter and wash until neutral.

9. The method for identifying macadamia anthracnose according to claim 8, characterized in that, The weight ratio of glucose in step S3 to glucose in step S4 is 1:2-5; The weight ratio of yeast powder in step S3 to yeast powder in step S4 is 1:0.4-1.

10. The method for identifying macadamia anthracnose according to claim 9, characterized in that, The nutrient particles obtained by pulverizing in step S4 have a particle size of 0.5-2 mm.

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