A high-yield pholiotus cervinus cultivation method

Abstract

The application relates to a high-yield Dictyophora rubrovolvata planting and cultivation method and belongs to the technical field of edible fungus planting. Mycelium is selected and activated for 5-10 days, and then is inoculated into glucose-agar recovery culture medium for dark culture. The culture medium is prepared by proportionally mixing sawdust, pretreated bamboo chips, auxiliary materials, wood mold inhibitor and eucommia leaf extract, adjusting the water content, sterilizing and bagging. Fungus is inoculated at an inoculation amount of 2-2.5% and is cultured until the bag is full. Temperature, light, humidity and CO2 are controlled in stages during mushroom management, and the mushrooms are harvested when the gill is open, the mushroom cup is red, and the stem is 15-20 cm long. The technology of the application improves the stress resistance of the fungus, reduces the fungus pollution rate, has good water retention and air permeability, has high carbon source utilization rate, reduces the mushroom malformation rate, and improves the high-quality mushroom rate. Compared with the traditional method, the fungus pollution rate is significantly reduced, the total yield of the Dictyophora rubrovolvata is improved, and the proportion of high-quality mushrooms with intact gills and thick stems is greatly improved.

Description

Technical Field

[0001] This invention belongs to the field of edible fungi cultivation technology, specifically, it relates to a cultivation method for high-yield red-topped bamboo fungus. Background Technology

[0002] Red-topped bamboo fungus is a rare edible and medicinal fungus. Due to its rich active ingredients and unique flavor, it has a strong market demand. However, its large-scale cultivation is currently constrained by multiple technical bottlenecks, limiting the development of the industry.

[0003] In existing technologies, most cultures are cultivated at room temperature (20-25℃) without undergoing stress acclimatization. This results in weak strain resistance, making them prone to colonization stagnation when faced with residual tannins in the bamboo shavings culture medium or fluctuating humidity. Contamination rates by Trichoderma and other microorganisms often exceed 10% during the mycelial growth period. Rejuvenation media utilize nutritionally balanced PDA media without inducing stress-resistance gene expression through high-carbon, low-nitrogen nutrient restriction, leading to low carbon source utilization and subsequent poor fruiting quality due to insufficient nutrition. In preparing the culture medium for *Dictyophora indicum*, the bamboo shavings are merely crushed without fermentation. Tannins inhibit mycelial enzyme activity, cellulose is difficult to decompose, and carbon source supply efficiency is low. Trichoderma inhibitors are mostly chemical biological agents such as carbendazim and imazalil, which can leave residues affecting food safety and potentially poisoning the mycelium. Growth promoters are mostly general-purpose and cannot specifically activate the enzyme system that decomposes woody materials, resulting in insufficient carbon source utilization and hindering mycelial growth.

[0004] Based on this, this application provides a cultivation method for high-yield red-topped bamboo fungus. Summary of the Invention

[0005] The purpose of this invention is to provide a high-yield cultivation method for red-topped bamboo fungus, which solves the problems existing in the prior art.

[0006] The objective of this invention can be achieved through the following technical solutions:

[0007] A method for cultivating high-yield red-topped bamboo fungus includes the following steps:

[0008] S1. Strain selection and rejuvenation: The selected *Dictyophora indica* strain was transferred to PDA medium and cultured at 5-8℃ for 5-10 days to obtain an activated strain; then the activated strain was inoculated into a rejuvenation medium and cultured at 5-8℃ in the dark for 15-20 days to obtain a rejuvenated strain.

[0009] S2. Preparation of culture medium: Stir sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor and mycelial growth promoter at room temperature for 30-60 minutes, adjust the water content of the system to 60-70%, keep the pH value natural, sterilize and bag to obtain the culture medium.

[0010] S3. Inoculation and Mycelial Growth: Inoculate the culture medium from step S2 with the revitalized strain from step S1. Maintain an ambient temperature of 22-25℃, an air humidity of 70-75%, and a CO2 concentration of <1600ppm. Ventilate the bag 2-3 times a day until the mycelium has fully colonized the bag.

[0011] S4. Mushroom Management: After the mycelium has filled the bag, treat it at a low temperature of 16-18℃ for 2-3 days, then raise the temperature to 20-22℃ and cultivate for 5-6 days to induce maturity; then transfer it to the fruiting room to induce bud formation for 5-9 days, entering the bud stage; cultivate in the bud stage for 4-5 days to the elongation stage; the elongation stage lasts for 3-5 days, and harvest when the veil is fully open, the volva is red and the stipe is 15-20cm long.

[0012] Furthermore, a cultivation method for high-yield red-topped bamboo fungus includes the following steps:

[0013] S1. Strain selection and rejuvenation: Select strains with white, robust mycelia and fast growth rate; transfer the selected *Dictyophora indica* strain to PDA medium and culture at 5-8℃ for 5-10 days to obtain activated strains; then inoculate the activated strains into rejuvenation medium and culture at 5-8℃ in the dark for 15-20 days to obtain rejuvenated strains.

[0014] S2. Preparation of culture medium: Stir sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor and mycelial growth promoter at room temperature for 30-60 minutes, adjust the water content of the system to 60-70%, keep the pH value natural, sterilize and bag to obtain the culture medium.

[0015] S3. Inoculation and mycelial growth: After sterilizing the culture medium from step S2, cool it to room temperature, then inoculate with the rejuvenated culture from step S1. Maintain an ambient temperature of 22-25℃, an air humidity of 70-75%, and a CO2 concentration of <1600ppm. Ventilate the bag 2-3 times a day until the mycelium fills the bag.

[0016] S4. Fruiting Management: After the mycelium has fully colonized the bag, maintain an ambient temperature of 16-18℃ for 2-3 days, then raise the temperature to 20-22℃ and maintain it for 5-6 days to induce maturity. Next, transfer the bags to a fruiting induction room for bud induction, setting the ambient temperature to 19-20℃, air humidity to 88-90%, light intensity to 200 lux with 10-11 hours of light per day, and CO2 concentration to 800-1200 ppm for 5-9 days to induce the bud stage. Set the ambient temperature to 20-22℃, the ambient humidity to 84-88℃, the light intensity to 300 lux, and the CO2 concentration to 700-900 ppm. Cultivate for 4-5 days to enter the elongation phase. Set the elongation phase temperature to 21-23℃, the air humidity to 94-95%, the light intensity to 350-450 lux, and the CO2 concentration to 650-750 ppm. Maintain this for 3-5 days. Harvest when the veil is fully open, the volva is red, and the stipe is 15-20 cm long.

[0017] Furthermore, in step S1, the PDA culture medium comprises the following raw materials in parts by weight: 200-220g potato, 20-25g glucose, 20-25g agar and 1000mL sterile deionized water.

[0018] Furthermore, in step S1, the rejuvenation culture medium comprises the following raw materials in parts by weight: 15-20g glucose, 15-20g agar and 1000mL sterile deionized water.

[0019] Furthermore, in step S1, the rejuvenation culture medium is specifically prepared by the following steps: sterile deionized water is heated to 60-70°C, glucose is added and stirred until completely dissolved, then agar is added and heated and stirred until the agar is completely dissolved, water is added to 1000mL, sterilized at 121°C for 2-3 hours, and cooled to room temperature to obtain the rejuvenation culture medium.

[0020] Furthermore, in step S2, the pretreated bamboo chips are specifically prepared by the following steps:

[0021] Select fresh, mold-free, and insect-free bamboo shavings, and crush them into pieces less than 2cm in length and less than 0.4cm in diameter using a pulverizer. Then, adjust the moisture content of the system to 55-65% using a 0.5-1wt% urea solution to obtain pre-moistened bamboo shavings. Pile the pre-moistened bamboo shavings into a trapezoidal pile 1.5-2m wide and 1-1.5m high. After 3-5 days of piling, turn the outer layer of bamboo shavings into the inner layer and adjust the moisture content of the bamboo shavings on the surface of the pile to 55-65%. Turn the pile every 7-15 days (7-10 days / time in summer, 10-15 days / time in winter) for a total of 3-4 times. Fermentation is completed when the pile temperature drops to within 5℃ of the ambient temperature, and the pre-treated bamboo shavings are obtained.

[0022] Furthermore, in step S2, the mass ratio of sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter is 40-50:20-30:30-35:0.1-0.3:0.01-0.2.

[0023] Furthermore, in step S2, the auxiliary materials include the following raw materials in parts by weight: 15-20 parts wheat bran, 8-12 parts corn flour, 3-5 parts soybean meal, 0.5-1.5 parts light calcium carbonate, and 0.5-1.5 parts gypsum powder.

[0024] Furthermore, in step S2, the Trichoderma inhibitor is Trichoderma harzianum, 6-pentyl-2H-pyran-2-one, antimicrobial peptide products metabolized by Trichoderma harzianum, and extracts of Trichoderma harzianum metabolites.

[0025] In the above technical solution, the metabolites of Trichoderma harzianum are used as Trichoderma inhibitors in the culture medium of the fungus.

[0026] Furthermore, in step S2, the sterilization process is either high-pressure steam sterilization or atmospheric pressure sterilization.

[0027] Furthermore, in step S2, the sterilization process specifically involves sterilization at 123°C for 2-3 hours.

[0028] Furthermore, in step S2, the sterilization process specifically involves sterilization at 100°C for 12-14 hours.

[0029] Furthermore, in step S2, the mycelial growth promoter is an extract of Eucommia ulmoides leaves.

[0030] Furthermore, the Eucommia ulmoides leaf extract is specifically prepared by the following steps:

[0031] Select dried Eucommia ulmoides leaves, pulverize them, and extract them with ethanol solution for 2-3 hours. Then, reflux the system at 70-75℃ for 2-3 times, with each extraction time being 1.5-2 hours. Collect the filtrate and concentrate it by rotary evaporation to obtain Eucommia ulmoides leaf extract.

[0032] Furthermore, the Eucommia ulmoides leaf extract is specifically prepared by the following steps:

[0033] Select dried Eucommia ulmoides leaves, pulverize them to 20-40 mesh, and extract them with 50-60wt% ethanol solution at a mass-volume ratio of 1g:8-10mL for 2-3 hours. Then, reflux the system at 70-75℃ for 2-3 times, with each extraction time being 1.5-2 hours. Collect the filtrate and concentrate it by rotary evaporation to obtain Eucommia ulmoides leaf extract.

[0034] Furthermore, in step S3, the inoculation amount is 2-2.5% of the weight of the culture medium.

[0035] The beneficial effects of this invention are:

[0036] In this invention, the strain is subjected to low-temperature stress and nutrient restriction during the selection and rejuvenation stage. This nutrient restriction simulates adverse conditions, forcing the strain to activate stress-resistance gene expression and increasing the accumulation of polysaccharide protective substances, thereby reducing contamination rates during the mycelial growth period and increasing fruiting yield. Under low-temperature stress, the strain synthesizes more cold shock proteins and amino acid-based osmotic protectants such as proline, enhancing its tolerance to slight tannin residues in the culture medium and fluctuations in environmental parameters such as humidity during subsequent growth, thus reducing the growth stagnation period in the early stages of colonization. After adapting to a high-carbon, low-nitrogen environment, the rejuvenated strain exhibits significantly improved carbon source utilization. The combination of glucose-based readily available carbon and pretreated bamboo chips in the culture medium quickly meets the carbon requirements of the rejuvenated strain in the early stages of colonization. Furthermore, the enzyme activity activated by Eucommia ulmoides leaf extract further enhances the strain's decomposition efficiency of bamboo and wood chips.

[0037] This invention adds pre-wetted fermented bamboo chips and metabolites of *Trichoderma harzianum* antagonistic bacteria, along with mycelial growth promoters, to the culture medium for fungal inoculum. The bamboo chips undergo pre-wetting with urea and composting fermentation, which not only removes tannins from the bamboo chips through microbial degradation to prevent inhibition of mycelial growth, but also converts recalcitrant cellulose and hemicellulose into small-molecule sugars such as xylose and glucose. Simultaneously, the nitrogen source provided by urea promotes the reproduction of fermenting microorganisms. Combined with readily available carbon source wood chips, this creates a carbon source supply mode suitable for the mycelial growth of *Dictyophora indicum*, meeting the different needs of reproduction and fruiting. The culture medium also exhibits improved water and fertilizer retention and good aeration. The use of ethanol-extracted chlorogenic acid and *Eucommia ulmoides* leaf extracts, such as polysaccharides, activates the cellulase and ligninase activities of the mycelium, improving carbon source decomposition efficiency while enhancing the toughness of the mycelial cell membrane. In addition, the use of natural Trichoderma inhibitors such as harzianum can specifically inhibit the spore germination and mycelial growth of Trichoderma, and is non-toxic to the mycelium of Dictyophora indicum. Compared with traditional carbendazim, the contamination rate of miscellaneous bacteria is reduced, and there is no risk of chemical residues.

[0038] This invention's technical solution, in terms of fruiting management, involves phased parameter control to induce primordia differentiation and veil opening. After the mycelium has filled the bag, low-temperature treatment promotes primordia initiation. Nutrients are accumulated during warming, and the temperature is matched to the corresponding requirements during the budding stage to reduce deformed mushrooms with bent stipes and inability to open veils. During the elongation stage, light exposure is gradually increased to guide upright stipe growth; simultaneously, the CO2 concentration is reduced from 800-1200 ppm during the budding stage to 650-750 ppm during the elongation stage to avoid weak stipes caused by high CO2 levels. Attached Figure Description

[0039] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0040] Figure 1 and Figure 2 The images show the harvesting results of *Dictyophora indicum* in Examples 1 and 2 of this invention, respectively.

[0041] Figure 3 This is a diagram showing the measurement results of the sample length in Embodiment 2 of the present invention. Detailed Implementation

[0042] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0043] The specific embodiments of this application involve:

[0044] The strain used was Hongtuo No. 1.

[0045] The culture medium is packaged in polypropylene corner bags, with dimensions of 17cm×33cm×0.005cm and a filling weight of 1.2kg / bag.

[0046] The 6-pentyl-2H-pyran-2-one used has CAS number 27593-23-3 and a purity ≥96%.

[0047] The CAS number of the imazalil used is 67747-09-5, and the purity is 99%.

[0048] Example 1

[0049] A method for cultivating high-yield red-topped bamboo fungus includes the following steps:

[0050] S1. Strain selection and rejuvenation: Select strains with white, robust mycelia and fast growth rate; transfer the selected *Dictyophora indica* strain to PDA medium and culture at 5℃ for 5 days to obtain activated strains; then inoculate the activated strains into rejuvenation medium and culture at 5℃ in the dark for 15 days to obtain rejuvenated strains.

[0051] S2. Preparation of culture medium: Wood chips, pretreated bamboo chips, auxiliary materials, Trichoderma inhibitor and mycelial growth promoter are stirred at room temperature for 30 minutes. The water content of the system is adjusted to 60%, the pH value is natural, and after sterilization, it is packaged to obtain the culture medium.

[0052] S3. Inoculation and mycelial growth: After sterilizing the culture medium from step S2, cool it to room temperature, then inoculate with the rejuvenated culture from step S1. Maintain an ambient temperature of 22°C, an air humidity of 70%, and a CO2 concentration of <1600ppm. Ventilate twice a day until the mycelium fills the bag.

[0053] S4. Mushroom Management: After the mycelium has fully covered the bag, set the ambient temperature to 16℃ and maintain it for 2 days. Then, raise the temperature to 20℃ and maintain it for 5 days to induce maturity. Afterward, transfer the bags to the fruiting induction room to induce bud formation. Set the ambient temperature to 19℃, the air humidity to 88%, the light intensity to 200 lux and provide 10 hours of light per day, and the CO2 concentration to 800 ppm, and maintain this for 5 days to enter the bud formation stage. During the bud formation stage, set the ambient temperature to 20℃, the ambient humidity to 84℃, the light intensity to 300 lux, and the CO2 concentration to 700 ppm, and cultivate for 4 days to enter the elongation stage. Set the elongation stage temperature to 21℃, the air humidity to 94%, the light intensity to 350 lux, and the CO2 concentration to 650 ppm, and maintain this for 3 days. Harvest when the veil is fully open, the volva is red, and the stipe is 15-20 cm long.

[0054] In step S1, the PDA culture medium comprises the following raw materials in parts by weight: 200g potato, 20g glucose, 20g agar, and 1000mL sterile deionized water; the rejuvenation culture medium comprises the following raw materials in parts by weight: 15g glucose, 15g agar, and 1000mL sterile deionized water. The rejuvenation culture medium is prepared by the following steps: sterile deionized water is heated to 60°C, glucose is added, and stirred until completely dissolved. Then agar is added and heating and stirring continues until the agar is completely dissolved. Water is added to a final volume of 1000mL, and the mixture is sterilized at 121°C for 2 hours. After cooling to room temperature, the rejuvenation culture medium is obtained.

[0055] In step S2, the mass ratio of sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter is 40:20:30:0.1:0.01.

[0056] The pre-treated bamboo chips are prepared by the following steps:

[0057] Fresh, mold-free, and insect-free bamboo shavings are selected and crushed into pieces less than 2cm in length and less than 0.4cm in diameter using a shredder. Then, the moisture content of the system is adjusted to 55% using a 0.5wt% urea solution to obtain pre-wetted bamboo shavings. The pre-wetted bamboo shavings are piled into a trapezoidal pile 2m wide and 1.5m high. After 3 days of pile building, the outer layer of bamboo shavings is turned into the inside, and the moisture content of the bamboo shavings on the surface of the pile is adjusted to 55%. The pile is turned over once every 10 days, for a total of 3 times. When the pile temperature drops to the ambient temperature, fermentation is completed, and the pre-treated bamboo shavings are obtained.

[0058] The auxiliary materials include the following raw materials in parts by weight: 15 parts wheat bran, 8 parts corn flour, 3 parts soybean meal powder, 0.5 parts light calcium carbonate and 0.5 parts gypsum powder; the Trichoderma inhibitor is 6-pentyl-2H-pyran-2-one;

[0059] The mycelial growth promoter is Eucommia ulmoides leaf extract, which is prepared by the following steps: dried Eucommia ulmoides leaves with a moisture content of less than 12% are selected, sieved using a 10-mesh vibrating sieve to remove dead branches, and then pulverized to 40 mesh using a pulverizer. Extraction is performed for 3 hours using a 50wt% ethanol solution at a mass-to-volume ratio of 1g:8mL. The system is then refluxed at 75℃ for 3 extractions, each extraction lasting 2 hours. The filtrate is collected and concentrated by rotary evaporation until the volume of the filtrate is reduced to 1 / 10 of its original volume. During the process, a vacuum of 0.09MPa and a water bath temperature of 55℃ are set to obtain the Eucommia ulmoides leaf extract.

[0060] In step S3, the inoculation amount is 2.5% of the weight of the culture medium.

[0061] Example 2

[0062] A method for cultivating high-yield red-topped bamboo fungus includes the following steps:

[0063] S1. Strain selection and rejuvenation: Select strains with white, robust mycelia and fast growth rate; transfer the selected *Dictyophora indica* strain to PDA medium and culture at 6℃ for 8 days to obtain activated strains; then inoculate the activated strains into rejuvenation medium and culture at 6℃ in the dark for 18 days to obtain rejuvenated strains.

[0064] S2. Preparation of culture medium: The sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor and mycelial growth promoter are stirred at room temperature for 45 minutes. The water content of the system is adjusted to 65%, the pH value is natural, and after sterilization, it is packaged to obtain the culture medium.

[0065] S3. Inoculation and mycelial growth: After sterilizing the culture medium from step S2, cool it to room temperature, then inoculate with the rejuvenated strain from step S1. Maintain an ambient temperature of 23°C, an air humidity of 70%, and a CO2 concentration of <1600ppm. Ventilate the bag three times a day until the mycelium fills the bag.

[0066] S4. Mushroom Management: After the mycelium has fully covered the bag, maintain an ambient temperature of 17℃ for 3 days, then raise the temperature to 22℃ and maintain it for 6 days to induce maturity. Then, transfer the bags to the fruiting induction room to induce bud formation. Set the ambient temperature to 20℃, air humidity to 89%, light intensity to 200 lux with 11 hours of light per day, and CO2 concentration to 1000 ppm, and maintain this for 7 days to enter the bud stage. During the bud stage, set the ambient temperature to 21℃, ambient humidity to 86℃, light intensity to 300 lux, and CO2 concentration to 800 ppm, and cultivate for 5 days to enter the elongation stage. Set the elongation stage temperature to 22℃, air humidity to 95%, light intensity to 400 lux, and CO2 concentration to 700 ppm, and maintain this for 4 days. Harvest when the veil is fully open, the volva is red, and the stipe is 15-20 cm long.

[0067] In step S1, the PDA culture medium comprises the following raw materials in parts by weight: 210g potato, 23g glucose, 23g agar, and 1000mL sterile deionized water; the rejuvenation culture medium comprises the following raw materials in parts by weight: 18g glucose, 18g agar, and 1000mL sterile deionized water. The rejuvenation culture medium is prepared by the following steps: sterile deionized water is heated to 65°C, glucose is added and stirred until completely dissolved, then agar is added and heating and stirring continues until the agar is completely dissolved, water is added to a final volume of 1000mL, sterilized at 121°C for 3 hours, and cooled to room temperature to obtain the rejuvenation culture medium.

[0068] In step S2, the mass ratio of sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter is 45:25:33:0.2:0.1.

[0069] The pre-treated bamboo chips are prepared by the following steps:

[0070] Fresh, mold-free, and insect-free bamboo shavings are selected and crushed into pieces less than 2cm in length and less than 0.4cm in diameter using a shredder. Then, the moisture content of the system is adjusted to 60% using a 0.8wt% urea solution to obtain pre-wetted bamboo shavings. The pre-wetted bamboo shavings are piled into a trapezoidal pile 2m wide and 1.5m high. After 5 days of pile building, the outer layer of bamboo shavings is turned into the inside, and the moisture content of the bamboo shavings on the surface of the pile is adjusted to 60%. The pile is turned over once every 10 days, for a total of 4 times. When the pile temperature drops to the ambient temperature, fermentation is complete, and the pre-treated bamboo shavings are obtained.

[0071] The auxiliary materials include the following raw materials in parts by weight: 18 parts wheat bran, 10 parts corn flour, 4 parts soybean meal powder, 1.0 part light calcium carbonate, and 1.0 part gypsum powder; the Trichoderma inhibitor is 6-pentyl-2H-pyran-2-one;

[0072] The mycelial growth promoter is Eucommia ulmoides leaf extract, which is prepared by the following steps: dried Eucommia ulmoides leaves with a moisture content of less than 12% are selected, sieved using a 10-mesh vibrating sieve to remove dead branches, and then pulverized to 40 mesh using a pulverizer. Extraction is performed for 3 hours using a 50wt% ethanol solution at a mass-to-volume ratio of 1g:8mL. The system is then refluxed at 75℃ for 3 extractions, each extraction lasting 2 hours. The filtrate is collected and concentrated by rotary evaporation until the volume of the filtrate is reduced to 1 / 10 of its original volume. During the process, a vacuum of 0.09MPa and a water bath temperature of 55℃ are set to obtain the Eucommia ulmoides leaf extract.

[0073] In step S3, the inoculation amount is 2.5% of the weight of the culture medium.

[0074] Example 3

[0075] A method for cultivating high-yield red-topped bamboo fungus includes the following steps:

[0076] S1. Strain selection and rejuvenation: Select strains with white, robust mycelia and fast growth rate; transfer the selected *Dictyophora indica* strain to PDA medium and culture at 8℃ for 10 days to obtain activated strains; then inoculate the activated strains into rejuvenation medium and culture at 8℃ in the dark for 20 days to obtain rejuvenated strains.

[0077] S2. Preparation of culture medium: Wood chips, pretreated bamboo chips, auxiliary materials, Trichoderma inhibitor and mycelial growth promoter are stirred at room temperature for 60 minutes. The water content of the system is adjusted to 70%, the pH value is natural, and after sterilization, it is packaged to obtain the culture medium.

[0078] S3. Inoculation and mycelial growth: After sterilizing the culture medium from step S2, cool it to room temperature, then inoculate with the rejuvenated strain from step S1. Maintain an ambient temperature of 25°C, an air humidity of 75%, and a CO2 concentration of <1600ppm. Ventilate the bag three times a day until the mycelium fills the bag.

[0079] S4. Mushroom Management: After the mycelium has fully covered the bag, set the ambient temperature to 18℃ and maintain it for 3 days. Then, raise the temperature to 22℃ and maintain it for 6 days to induce maturity. Afterward, transfer the bags to the fruiting induction room to induce bud formation. Set the ambient temperature to 20℃, the air humidity to 90%, the light intensity to 200 lux and provide 11 hours of light per day, and the CO2 concentration to 1200 ppm, and maintain this for 9 days to enter the bud formation stage. During the bud formation stage, set the ambient temperature to 22℃, the ambient humidity to 88℃, the light intensity to 300 lux, and the CO2 concentration to 900 ppm, and cultivate for 5 days to enter the elongation stage. Set the elongation stage temperature to 23℃, the air humidity to 95%, the light intensity to 450 lux, and the CO2 concentration to 750 ppm, and maintain this for 5 days. Harvest when the veil is fully open, the volva is red, and the stipe is 15-20 cm long.

[0080] In step S1, the PDA culture medium comprises the following raw materials in parts by weight: 220g potato, 25g glucose, 25g agar, and 1000mL sterile deionized water; the rejuvenation culture medium comprises the following raw materials in parts by weight: 20g glucose, 20g agar, and 1000mL sterile deionized water. The rejuvenation culture medium is prepared by the following steps: sterile deionized water is heated to 70°C, glucose is added, and stirred until completely dissolved. Then agar is added and heating and stirring continues until the agar is completely dissolved. Water is added to a final volume of 1000mL, and the mixture is sterilized at 121°C for 3 hours. After cooling to room temperature, the rejuvenation culture medium is obtained.

[0081] In step S2, the mass ratio of sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter is 50:30:35:0.3:2.

[0082] The pre-treated bamboo chips are prepared by the following steps:

[0083] Fresh, mold-free, and insect-free bamboo shavings are selected and crushed into pieces less than 2cm in length and less than 0.4cm in diameter using a shredder. Then, the moisture content of the system is adjusted to 65% using a 1wt% urea solution to obtain pre-wetted bamboo shavings. The pre-wetted bamboo shavings are piled into a trapezoidal pile 2m wide and 1.5m high. After 5 days of pile building, the outer layer of bamboo shavings is turned into the inside, and the moisture content of the bamboo shavings on the surface of the pile is adjusted to 65%. The pile is turned over once every 10 days, for a total of 4 times. When the pile temperature drops to the ambient temperature, fermentation is completed, and the pre-treated bamboo shavings are obtained.

[0084] The auxiliary materials include the following raw materials in parts by weight: 20 parts wheat bran, 12 parts corn flour, 5 parts soybean meal powder, 1.5 parts light calcium carbonate and 1.5 parts gypsum powder; the Trichoderma inhibitor is 6-pentyl-2H-pyran-2-one;

[0085] The mycelial growth promoter is Eucommia ulmoides leaf extract, which is prepared by the following steps: dried Eucommia ulmoides leaves with a moisture content of less than 12% are selected, sieved using a 10-mesh vibrating sieve to remove dead branches, and then pulverized to 40 mesh using a pulverizer. Extraction is performed for 3 hours using a 50wt% ethanol solution at a mass-to-volume ratio of 1g:8mL. The system is then refluxed at 75℃ for 3 extractions, each extraction lasting 2 hours. The filtrate is collected and concentrated by rotary evaporation until the volume of the filtrate is reduced to 1 / 10 of its original volume. During the process, a vacuum of 0.09MPa and a water bath temperature of 55℃ are set to obtain the Eucommia ulmoides leaf extract.

[0086] In step S3, the inoculation amount is 2.5% of the weight of the culture medium.

[0087] Comparative Example 1

[0088] The difference between this comparative example and Example 2 is that step S1 in this comparative example is as follows: Select the strain with white, robust, and fast-growing mycelium; transfer the selected *Dictyophora indica* strain to PDA medium and culture it at 20°C for 8 days to obtain an activated strain; then inoculate the activated strain into a rejuvenation medium and culture it at 20°C in the dark for 18 days to obtain a rejuvenated strain.

[0089] Comparative Example 2

[0090] The difference between this comparative example and Example 2 is that the rejuvenation medium used in step S1 of this comparative example, namely PDA medium, includes the following raw materials by weight: 210g potato, 23g glucose, 23g agar and 1000mL sterile deionized water.

[0091] Comparative Example 3

[0092] The difference between this comparative example and comparative example 2 is that the Trichoderma inhibitor used in step S2 is imazalil.

[0093] Comparative Example 4

[0094] The difference between this comparative example and Example 2 is that the mycelial growth promoter used in step S2 is an algae extract, which is specifically prepared by the following steps:

[0095] Specifically, the seaweed extract is prepared by the following steps: Select brown algae powder, pass it through a 40-mesh sieve, and extract it for 3 hours using a 50wt% ethanol solution at a mass-to-volume ratio of 1g:8mL. Then, reflux the system at 75℃ for 3 times, with each extraction lasting 2 hours. Collect the filtrate and concentrate it by rotary evaporation until the volume of the filtrate is reduced to 1 / 10 of the original volume. During the process, the vacuum degree is set at 0.09MPa and the water bath temperature is set at 55℃ to obtain the seaweed extract.

[0096] The *Dictyophora indicum* obtained in Examples 1-3 and Comparative Examples 1-4 were subjected to performance tests. The *Dictyophora indicum* obtained in Examples 1-3 and Comparative Examples 1-4 were harvested. During harvesting, the volva was cut off at the bottom with a knife, retaining the stipe and veil. After each flush of mushrooms was harvested, residual mushrooms and debris on the surface of the planting bed were promptly cleaned, and a 1cm thick layer of fresh, well-rotted sawdust was added. The substrate moisture content was adjusted to 65%, and steps S3 and S4 for each group were repeated for management before harvesting continued.

[0097] Yield (fresh weight) and quality tests were conducted on the *Dictyophora indicum* from Examples 1-3 and Comparative Examples 1-4. *Dictyophora indicum* was classified into four grades based on morphological integrity, veil opening, stipe thickness, and impurity contamination rate. The specific evaluation criteria are as follows:

[0098] Grade 1: The volva is uniformly dark red and undamaged; the stipe is larger than 1.5cm in diameter and 15cm in length, without bending or hollowing; the veil is fully open, white and transparent, without tearing or yellow spots; free from contamination by other fungi and insect infestation; the moisture content is 85-90%, feels firm to the touch, and is free from soft rot.

[0099] Grade II: The volva is light red with no obvious damage; the stipe is larger than 1.2cm in diameter and 13cm in length, slightly curved with a bending angle of less than 10°; the veil is more than 70° open, with a few small tears, less than 2 tears in length, each less than 1cm; free from contamination by other fungi and insect infestation; moisture content is 90-92%, slightly soft, and free from rot.

[0100] Grade III: The volva is light red or partially whitish; the stipe is larger than 1 cm in diameter and longer than 10 cm, with a curvature between 10-20° and a hollow rate of less than 10%; the veil opening is 50-70%, with fewer than 3 tears, each less than 2 cm, and yellow spots covering less than 5% of the surface area; the contamination rate of miscellaneous bacteria is less than 3%, and there is no insect infestation; the moisture content is 92-94%, the texture is noticeably soft, and there is no large-area rot.

[0101] Grade IV: The volva is white or damaged; the stipe diameter is <1cm, length is <10cm, curvature is >20°, and hollow rate is >10%; the veil opening is <50%, there are more than 3 tears or a single tear >2cm, and yellow spots cover more than 5% of the surface area; the contamination rate of miscellaneous bacteria is >3%, or there are insect infestations; the moisture content is >94%, and there is soft rot.

[0102] The test results are shown in Tables 1 and 2 below.

[0103] Table 1

[0104]

[0105] Table 2

[0106]

[0107] The results in Tables 1 and 2 show that the *Dictyophora indicum* obtained in Example 2 exhibits the best quality and yield. Compared to Example 2, Comparative Example 1 removed the limitations of low-temperature stress in the strain selection and rejuvenation process, while Comparative Example 2 removed the limitations of nutritional stress in the strain selection and rejuvenation process. The results in Comparative Examples 1 and 2 demonstrate that low-temperature stress can induce the strain to synthesize stress-resistant substances and enhance its resistance to contaminating microorganisms; nutritional restriction can cultivate the strain's efficient utilization of carbon sources, accumulating sufficient nutrients for subsequent fruiting.

[0108] The results of Comparative Example 3 show that the natural inhibitor is non-toxic to the mycelium of *Dictyophora indicum* and can inhibit other fungi such as *Trichoderma*. However, replacing it with imazalil has the side effect of inhibiting both bacteria and mycelium, and the residue affects the quality. The results of Comparative Example 4 show that chlorogenic acid in *Eucommia ulmoides* leaf extract can more efficiently activate cellulase and ligninase, further improving the decomposition efficiency of bamboo chips. *Eucommia ulmoides* polysaccharides can enhance the toughness of mycelial cell membranes. However, replacing it with seaweed extract has a weaker activating effect on enzyme activity, resulting in slightly poorer mycelial vitality and mushroom morphology, and a reduced yield.

[0109] In the description of this specification, the references to terms such as "an embodiment," "example," "specific example," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0110] The above description is merely an example and illustration of the concept of the present invention. Those skilled in the art can make various modifications or additions to the specific embodiments described or use similar methods to replace them, as long as they do not deviate from the concept of the invention or exceed the scope defined in the claims, they should all fall within the protection scope of the present invention.

Claims

1. A method for cultivating high-yield red-topped bamboo fungus, characterized in that, Includes the following steps: S1. Strain selection and rejuvenation: The selected *Dictyophora indica* strain was transferred to PDA medium and cultured at 5-8℃ for 5-10 days to obtain an activated strain; then the activated strain was inoculated into a rejuvenation medium and cultured at 5-8℃ in the dark for 15-20 days to obtain a rejuvenated strain; the rejuvenation medium was specifically prepared by the following steps: sterile deionized water was heated to 60-70℃, glucose was added and stirred until completely dissolved, then agar was added and heated and stirred until the agar was completely dissolved, sterile deionized water was added to 1000mL, sterilized and cooled to room temperature to obtain the rejuvenation medium; S2. Preparation of culture medium for fungal inoculum: Wood chips, pretreated bamboo chips, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter are stirred at room temperature for 30-60 minutes. The water content of the system is adjusted to 60-70%, the pH is left to stand naturally, and after sterilization, it is packaged to obtain the culture medium for fungal inoculum. The Trichoderma inhibitor is at least one of Trichoderma harzianum, 6-pentyl-2H-pyran-2-one, antimicrobial peptide products metabolized by Trichoderma harzianum, and extracts of Trichoderma harzianum metabolites. The mycelial growth promoter is Eucommia ulmoides leaf extract. S3. Inoculation and Mycelial Growth: Inoculate the culture medium from step S2 with the revitalized strain from step S1. Maintain an ambient temperature of 22-25℃, an air humidity of 70-75%, and a CO2 concentration of <1600ppm. Ventilate the bag 2-3 times a day until the mycelium has fully colonized the bag. S4. Mushroom Management: After the mycelium fills the bag, treat it at a low temperature of 16-18℃ for 2-3 days, then raise the temperature to 20-22℃ and cultivate for 5-6 days to induce maturity; then induce bud formation for 5-9 days to enter the bud stage; cultivate the bud stage for 4-5 days to the elongation stage; the elongation stage lasts for 3-5 days, and harvest when the veil matures.

2. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, In step S1, the rejuvenation culture medium comprises the following raw materials in parts by weight: 15-20g glucose, 15-20g agar and 1000mL sterile deionized water.

3. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, In step S2, the pretreated bamboo chips are specifically prepared by the following steps: Select bamboo shavings, crush them using a pulverizer, and adjust the moisture content of the system to 55-65% to obtain pre-moistened bamboo shavings. Pile the pre-moistened bamboo shavings into a trapezoidal pile. After 3-5 days of piling, turn the outer layer of bamboo shavings into the inside and adjust the moisture content of the bamboo shavings on the surface of the pile to 55-65%. Turn the pile over once every 7-15 days, for a total of 3-4 times. After fermentation is complete, the pre-treated bamboo shavings are obtained.

4. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, In step S2, the auxiliary materials include the following raw materials in parts by weight: 15-20 parts wheat bran, 8-12 parts corn flour, 3-5 parts soybean meal, 0.5-1.5 parts light calcium carbonate, and 0.5-1.5 parts gypsum powder.

5. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, The Eucommia ulmoides leaf extract is prepared by the following steps: Select dried Eucommia ulmoides leaves, pulverize them, and extract them with ethanol solution for 2-3 hours. Then, reflux the system at 70-75℃ for 2-3 times, with each extraction time being 1.5-2 hours. Collect the filtrate and concentrate it by rotary evaporation to obtain Eucommia ulmoides leaf extract.

6. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, In step S2, the mass ratio of sawdust, pretreated bamboo shavings, auxiliary materials, Trichoderma inhibitor, and mycelial growth promoter is 40-50:20-30:30-35:0.1-0.3:0.01-0.

2.

7. The cultivation method for high-yield red-topped bamboo fungus according to claim 1, characterized in that, In step S3, the inoculation amount is 2-2.5% of the weight of the culture medium.

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