CN111423998A - Compound microbial agent for mushroom residue basification and preparation method and application thereof - Google Patents
Compound microbial agent for mushroom residue basification and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a compound microbial agent for mushroom residue basification and a preparation method and application thereof. The compound microbial agent special for mushroom residue matrixing utilization fermentation consists of bacillus urealyticum, acetobacter pasteurianus, aspergillus fumigatus, thermomyces, brevibacillus thermoerythraeus and bacillus safensis; the compound microbial agent special for mushroom residue fermentation, which is prepared by the invention, provides technical support and method means for promoting mushroom residue fermentation process, mushroom residue fermentation decomposition degree and mushroom residue matrixing utilization.
Description
Technical Field
The invention belongs to the technical field of microbial fermentation of organic solid wastes, and relates to a compound microbial agent for mushroom residue basification, and a preparation method and application thereof.
Background
The mushroom residue is also called mushroom residue, is a culture medium with low nutritive value after the edible mushrooms are harvested, and is a main byproduct of the edible mushroom industry. China is the biggest edible fungus producing country in the world, the annual output exceeds 3500t and accounts for more than 80 percent of the total edible fungus production in the world. According to related researches, 5kg of waste mushroom residues are generated on average when 1kg of edible mushrooms are produced, so that the quantity of the actually generated mushroom residues is extremely large, and if the mushroom residues cannot be reasonably utilized, huge resource waste and environmental pollution are caused. With the increase of the production amount of the mushroom residues, the problem of environmental pollution is increasingly highlighted by randomly disposing the mushroom residues, and how to reasonably and effectively utilize the mushroom residues becomes a key point of research. At present, the utilization approaches of mushroom residues mainly comprise base fertilizer, feed, ingredients for edible mushroom production, fuel and soilless culture medium, wherein, when the mushroom residues are used as the culture medium, specific process treatment is required, and the adopted method is composting fermentation.
The edible fungus culture medium is prepared from wood chips, corncobs, straws and other biomass as raw materials, and is rich in organic matters, calcium, magnesium, nitrogen, phosphorus and other nutritional ingredients. The components of the mushroom residues have certain difference along with different factors such as the cultivation of mushroom seeds, raw material formulas and mushroom growing times, but the main components of most of the mushroom residues are cellulose, hemicellulose and lignin. Microorganisms capable of metabolizing to produce cellulase widely exist in the nature, but part of cellulase has generally low specific activity, and the efficient decomposition and utilization of cellulose are difficult to realize really, while thermophilic cellulose degrading bacteria have the characteristics of fast metabolism, short generation time, high thermal stability of enzyme and the like, and the high-temperature period of composting fermentation is the fastest time for degrading organic matters and other substances in the whole composting fermentation process, so the development and utilization of thermophilic bacteria in the high-temperature environment are widely regarded. But currently, a special microbial agent and a product for mushroom residue fermentation using thermophilic bacteria as main strains are still lacked, and the development and utilization of the efficient and safe mushroom residue fermentation microbial agent has important practical significance for improving the resource utilization of organic solid wastes.
Disclosure of Invention
The present invention has been made in view of the above circumstances, and an object of the present invention is to provide a complex microbial agent for the basification of mushroom dregs.
The second purpose of the invention is to provide a preparation method of the compound microbial agent.
The third purpose of the invention is to provide the application of the compound microbial agent.
The technical scheme adopted by the invention for solving the technical problems is as follows:
a compound microbial agent for mushroom residue basification mainly comprises thermophilic cellulose degradation bacteria, thermophilic cellulose degradation fungi and thermophilic cellulose degradation actinomycetes; wherein the thermophilic cellulose-degrading bacteria are Bacillus urealyticus (Ureibacillus subsinus) and Acetobacter pasteurianus (Acetobacter pasteurianus); the thermophilic cellulose-degrading fungi are Aspergillus fumigatus (Aspergillus fumigatus) and Thermomyces thermophilus (Thermomyces sp.Strain); the thermophilic cellulose-degrading actinomycetes are Brevibacillus thermovirens (Brevibacillus thermoruber) and Bacillus safensis (Bacillus safensis).
The sources of the 6 microorganisms are not particularly required, the 6 microorganisms can be used for preparing the microbial inoculum of the invention as long as the taxonomy meets the species classification, and the 6 microorganisms can be screened by themselves or purchased through a commercial way.
As a preferred technical scheme of the application, the compound microbial agent mainly comprises bacterial liquid of Bacillus urealyticum (Ureibacillus subswerensis), Acetobacter pasteurianus (Acetobacter pasteurianus), Aspergillus fumigatus (Aspergillus fumigatus), Thermomyces sp.
As a preferred technical scheme of the application, the compound microbial agent is mainly prepared by mixing bacterial liquids of Bacillus ureafaciens (Ureibacillus subsynensis), Acetobacter pasteurianus (Acetobacter pasteurianus), aspergillus fumigatus (aspergillus fumigatus), Thermomyces sp.
The preparation method of the compound microbial agent for mushroom residue basification comprises the steps of respectively and independently culturing and propagating 6 types of microorganisms as claimed in any one of claims 1 to 4, and mixing the microorganisms in equal volume to form the compound microbial agent after inoculation conditions are reached.
As a preferred technical scheme of the application, the culture and propagation of the thermophilic cellulose degradation bacteria uses a beef extract peptone liquid medium; culturing and propagating thermophilic cellulose degradation fungi by using a potato glucose liquid culture medium; the culture and propagation of the thermophilic cellulose degradation actinomycetes uses an improved Gao's No. I liquid culture medium.
The culture medium preferably comprises beef extract peptone liquid culture medium comprising beef extract 5.0 g/L, peptone 10.0 g/L5.0.0 g/L, distilled water 1000m L, and pH7.4-7.6, and improved Gao I liquid culture medium comprising KNO31 g/L, soluble starch 20 g/L, K2HPO40.5g/L，MgSO4.7H2O 0.5g/L，NaCl 0.5g/L，FeSO40.01 g/L, 1000m L water, 3% potassium dichromate 3.3m L per 1000m L culture medium, potato glucose agar culture medium including potato 200.0 g/L, glucose 20.0 g/L, agar powder 18.0 g/L, distilled water 1000m L, and natural pH.
As a preferred technical scheme of the application, the inoculation conditions are that the OD value of the bacterial liquid of the bacterial microorganisms cannot be lower than 1, and the spore concentration of the bacterial liquid of actinomycetes and fungal microorganisms is not lower than 1 × 10-7And each m L.
As a preferred technical scheme of the application, the preparation method comprises the following steps:
step 1, respectively carrying out shake culture on thermophilic bacteria, thermophilic actinomycetes and thermophilic fungi in a beef extract peptone liquid culture medium, an improved Gao's I liquid culture medium and a potato glucose liquid culture medium at 50 ℃ at 200 rpm for 2-3 d;
(2) measuring OD value of bacteria at 600nm, measuring spore concentration of actinomycetes and fungi, diluting OD value of bacteria to 1.0 with sterile water, mixing to obtain bacteria solution, and respectively diluting spore concentration of actinomycetes and fungi to 1 × 107Actinomycete and fungus bacteria liquid of L per m, and mixing all the bacteria in equal volume to obtain bacteria liquid.
Advantageous effects
The invention provides a compound microbial agent for mushroom residue basification and a preparation method and application thereof, and the prepared microbial agent special for mushroom residue fermentation provides technical support and method means for promoting mushroom residue fermentation process and mushroom residue fermentation decomposition degree. The method provided by the invention has important scientific significance and application value for matrix utilization of mushroom residues.
The composite microbial agent prepared by the invention has thermophilic bacterial strains and strong cellulose decomposition capacity, can realize good symbiosis among the bacterial strains, has no antagonistic action among the microbial strains, is prepared by fermenting the mushroom residues with the liquid microbial agent, is simple and easy to operate, has strong practicability, reduces the production cost, accelerates the fermentation process of the mushroom residues and improves the fermentation quality of the mushroom residues.
Drawings
Fig. 1 shows a technical scheme diagram of a complex microbial inoculant for mushroom residue basification and a preparation method and application thereof.
FIG. 2 shows the isolation of thermophilic bacteria from mushroom dregs.
FIG. 3 shows the growth of fermenting thermophiles on cellulose Congo red medium.
FIG. 4 shows the filter paper disintegration of fermented thermophilic bacteria in the filter paper disintegration test.
FIG. 5 is a graph showing the antagonism test between different species.
FIG. 6 is a temperature change diagram of a mushroom residue composting fermentation test using the composite microbial inoculum prepared by the invention and specially used for mushroom residue fermentation.
FIG. 7 is a lignocellulose change diagram of a mushroom residue composting fermentation test by using the compound microbial inoculum special for mushroom residue fermentation prepared by the invention.
FIG. 8 is a graph showing the change of germination index of seeds in a mushroom residue composting fermentation test by using the composite microbial agent specially used for mushroom residue fermentation prepared by the present invention.
Detailed Description
The present invention will be further described with reference to the following examples, which are intended to illustrate the present invention and not to limit the scope of the present invention, and all simple modifications of the preparation method of the present invention based on the idea of the present invention are within the scope of the present invention. The following examples are experimental methods without specifying specific conditions, and generally follow the methods known in the art. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified.
Test materials
Fresh needle mushroom residues come from Jiangsu Xingnong substrate science and technology limited of Zhenjiang city, Jiangsu province. The basic physicochemical properties of the fresh mushroom residues are shown in the table, and the table shows that the water content of the fresh mushroom residues is 62 percent, is neutral and can be directly subjected to composting fermentation treatment; the mushroom dregs mainly comprise cellulose, hemicellulose and lignin, wherein the cellulose is the highest and reaches 25 percent. In addition, the mushroom residues are rich in nutrients, the C/N is 22.4, and the EC value is higher and reaches 4.76 mS/cm.
Beef extract peptone solid culture medium comprises beef extract 5.0 g/L, peptone 10.0 g/L5.0.0 g/L, distilled water 1000m L, agar powder 18.0 g/L, and pH 7.4-7.6.
Beef extract peptone liquid culture medium comprises beef extract 5.0 g/L, peptone 10.0 g/L5.0.0 g/L, distilled water 1000m L, and pH 7.4-7.6.
The Martin's medium comprises 5.0 g/L of peptone, 10.0 g/L of glucose, 1.0 g/L of monopotassium phosphate, 0.5 g/L of magnesium sulfate, 0.1 g/L of streptomycin, 0.033 g/L of Bengal, 18.0 g/L of agar powder, 1000m L of distilled water and natural pH.
Composition of modified solid culture medium No. one of gao: KNO31g/L，Soluble starch 20 g/L, K2HPO40.5g/L，MgSO4.7H2O 0.5g/L，NaCl 0.5g/L，FeSO40.01 g/L, agar 18 g/L, water 1000m L, 3% potassium dichromate per 1000m L of culture medium 3.m L.
Composition of modified Gao's No. one liquid medium: KNO31 g/L, soluble starch 20 g/L, K2HPO40.5g/L，MgSO4.7H2O 0.5g/L，NaCl 0.5g/L，FeSO40.01 g/L, water 1000m L, 3% potassium dichromate per 1000m L medium 3.m L.
Composition of solid culture medium No. gao shi: KNO31 g/L, soluble starch 20 g/L, K2HPO40.5g/L，MgSO4.7H2O 0.5g/L，NaCl 0.5g/L，FeSO40.01 g/L, agar 18 g/L, water 1000m L, pH 7.4-7.6.
Potato glucose agar culture medium comprising potato 200.0 g/L, glucose 20.0 g/L, agar powder 18.0 g/L, distilled water 1000m L, and natural pH.
The potato glucose liquid culture medium comprises 200.0 g/L of potatoes, 20.0 g/L of glucose and 1000m L of distilled water, and has natural pH.
The cellulose Congo red culture medium comprises 2.0 g/L of carboxymethyl cellulose, 2.0 g/L of ammonium sulfate, 0.5 g/L of magnesium sulfate heptahydrate, 1.0 g/L of dipotassium hydrogen phosphate, 0.5 g/L of sodium chloride, 18 g/L of agar powder, 0.2 g/L of Congo red, 1000m L of tap water and pH 7.0.
Paper strip disintegration culture medium: (NH)4)2SO41.0g/L，MgSO4·7H2O 0.5g/L，KH2PO41.0 g/L, yeast extract 0.1 g/L, filter paper strip (1cm × 6cm)1 strip/triangular flask, pH 7.0.
Example 1 preparation of Complex microbial Agents
(1) Respectively culturing thermophilic cellulose degrading bacteria, thermophilic cellulose degrading actinomycetes and thermophilic cellulose degrading fungi in a beef extract peptone liquid culture medium, a Gao's first liquid culture medium and a potato glucose liquid culture medium at 50 ℃ and 200 rpm for 2-3 days in a shaking way.
(2) Thermophilic cellulose-degrading bacteria measure OD value at 600nm, addicted toMeasuring spore concentration in bacteria liquid by using thermocellulose-degrading actinomycetes and thermophilic cellulose-degrading actinomycetes, diluting OD value of thermophilic cellulose-degrading bacteria to 1.0 by using sterile water, mixing to obtain bacteria liquid, respectively diluting spore concentrations of bacteria liquid of thermophilic cellulose-degrading actinomycetes and thermophilic cellulose-degrading actinomycetes to 1 × 107Actinomycetes and fungi bacteria liquid of per m L.
(3) All strains are mixed into bacterial liquid according to the equal volume of 1:1:1(V: V: V).
Example 2
1. High temperature culture of strain
(1) Preparing bacterial suspension of different samples, taking 10g of fresh sample, placing into a conical flask containing 90m L sterile water, shaking for 30min at 200 r, filtering, and diluting the filtrate in gradient manner to obtain 10-1-10-7A concentration gradient of the bacterial suspension. (2) coating: bacterial selection 10-5、10-6、10-7Three concentration gradients, fungal selection 10-2、10-3、10-4Three concentration gradients, actinomycete selection 10-4、10-5、10-6The three concentration gradients are respectively coated on a beef extract peptone solid culture medium, a martin culture medium and an improved Gao's I solid culture medium.
(3) High-temperature culture: standing and culturing in a biochemical incubator at 50 deg.C.
2. Thermophilic bacterial strain separation and purification culture
Separating and purifying the dominant strain on beef extract peptone solid medium, martin medium and modified Gao's No. one solid medium by streaking, and continuously purifying for 2-3 times.
3. Primary screening for cellulose degradation capability of thermophilic bacterial strain
The primary screening of the cellulose degradation capability of the strain adopts a cellulose Congo red culture medium screening method, and the strain with stronger cellulose degradation capability is screened out mainly according to the ratio of the diameter of a hydrolysis ring to the diameter of a bacterial colony.
(1) The inoculating needle picks up the microbial colony to be inoculated into a cellulose Congo red culture medium, 2d (bacteria), 3d (fungi) and 5d (actinomycetes) are inversely cultured in a biochemical incubator, and the culture temperature is set to be 50 ℃.
(2) After culturing for corresponding days, dripping 0.2% Congo red solution into a culture dish with bacterial colonies, shaking uniformly, standing and dyeing for 15min, then flushing the Congo red culture medium with 1M NaCl solution, and decolorizing for 15 min.
(3) And screening out the bacterial strain with stronger cellulose decomposition capacity according to the ratio of the diameter of the hydrolysis ring to the diameter of the bacterial colony.
4. Thermophilic bacterial strain cellulose degradation ability re-screening
The bacterial strain cellulose degradation capability is re-screened, a filter paper disintegration experiment screening method is adopted, and the strength of the bacterial strain cellulose degradation capability is judged mainly according to the disintegration condition of the filter paper.
(1) Placing 100m L filter paper disintegration culture medium in conical flask, placing a filter paper strip (3 times) of 1cm × 6cm at the bottom of each conical flask, autoclaving at 121 deg.C for 20min, and cooling the liquid culture medium for use.
(2) The inoculating needle picks thermophilic bacteria, thermophilic fungi and thermophilic actinomycetes, and inoculates the thermophilic bacteria, thermophilic fungi and thermophilic actinomycetes into a centrifugal tube containing sterilized beef extract peptone liquid culture medium, potato glucose liquid culture medium and Gao's No. one liquid culture medium 2m L respectively, and cultures for 24h in a shaking table at 50 ℃ and 180 turns.
(3) In an ultra-clean workbench, a pipette is used for sucking 1m L microbial strains to be inoculated into a filter paper disintegration culture medium, shaking culture is carried out for 7 days at 50 ℃ and 180rpm, and strains with stronger cellulose decomposition capacity are screened according to the disintegration condition of the filter paper.
5. Thermophilic cellulose degradation strain identification
(1) Extracting the genome DNA of the screened strains.
(2) And carrying out PCR amplification on the extracted strain genome DNA by adopting a 16S universal primer.
(3) Sending the PCR product to a biological company for sequencing, performing Blast on the sequencing result in an NCBI database, and selecting a result with the comparison similarity of more than 98%; the identified and finished strains with stronger cellulose decomposition capacity comprise: bacteria (Bacillus urealyticum, Acetobacter pasteurianus), actinomycetes (Bacillus caldovelox, Bacillus salofumei), and fungi (Aspergillus fumigatus, Thermomyces thermophilus).
6. Thermophilic cellulose degrading bacterial strain antagonistic verification
(1) Antagonism verification between fungi and bacteria, fungi and actinomycetes
1) Respectively inoculating fungus strains to a potato liquid culture medium, culturing at 37 ℃ and 180r/min for 24h, and then using the potato liquid culture medium for an antagonistic test, 2) adding each fungus suspension to a potato glucose solid culture medium flat plate at 0.2m L/dish, uniformly coating, inoculating the tested fungus, bacterium or actinomycetes to the center part of the cross, then inoculating two other fungi or bacteria or actinomycetes at the symmetrical 25 mm position of the cross, culturing at 50 ℃ for 1-2 days, then observing the growth condition, finally judging whether the compound is suitable according to whether the bacteriostatic ring exists, if so, not, then, suitably compounding.
(2) Antagonism between bacteria and actinomycetes
1) Preparing a potato glucose agar solid culture medium, carrying out autoclaving at 121 ℃ for 20min, pouring the culture medium into a flat plate, and cooling the culture medium for later use. 2) And (4) streaking and culturing the activated strains on a PDA culture medium plate respectively, and culturing at constant temperature of 50 ℃ for 3 d. 3) And observing the growth conditions of two strains at the crossed streaks. If the two strains at the cross point grow relatively weakly or do not grow, the antagonism between the two strains is shown; if the bacteria at the cross points grow well, the antagonism between the two strains is proved, and the method can be used for mixed culture.
7. According to the preparation method of the compound microbial agent in the embodiment 1, the bacillus urealyticum (laboratory separation), the acetobacter pasteurianus (laboratory separation), the brevibacillus thermoerythraeus (laboratory separation), the bacillus saffron (laboratory separation), the aspergillus fumigatus (laboratory separation) and the myceliophthora thermophila (laboratory separation) are respectively cultured in corresponding culture media, and all bacterial liquids are mixed into the compound microbial agent according to the equal volume of 1:1:1(V: V: V).
Example 3
According to the preparation method of the compound microbial inoculum in the embodiment 1, the ureabacterium (CGMCC1.15355), the acetobacter pasteurianus (CGMCC1.2269), the brevibacillus thermoerythraeus (CGMCC1.8649), the bacillus safensis (CGMCC1.9052), the aspergillus fumigatus (CGMCC3.15720) and the myceliophthora thermophila (CGMCC3.17993) are respectively cultured in corresponding culture media, and all the bacterial liquids are mixed into the compound microbial inoculum according to the equal volume of 1:1:1(V: V: V). The strains are all purchased from the China general microbiological culture Collection center.
Example 4
According to the preparation method of the compound microbial inoculum in the embodiment 1, the ureabacterium (CGMCC1.15301), the acetobacter pasteurianus (CGMCC1.1810), the brevibacillus thermoerythraeus (CGMCC1.3483), the bacillus safensis (CGMCC1.8112), the aspergillus fumigatus (CGMCC3.14869) and the myceliophthora thermophila (CGMCC3.9539) are respectively cultured in corresponding culture media, and all the bacterial liquids are mixed into the compound microbial inoculum according to the equal volume of 1:1:1(V: V: V). The strains are all purchased from the China general microbiological culture Collection center.
Example 5 Effect evaluation of fermentation application of Complex microbial inoculum
1. The test was carried out in 2019, 8 months, by Jiangsu Xingnong substrate science and technology Limited, Jiangsu province, wherein the raw material is fresh needle mushroom residues, and the water content is 61.51%. The complex microbial inoculum prepared in the invention in the example 2(T2), the example 3(T3) and the example 4(T4) is inoculated in a pile with the length of 2m, the width of 1.5m and the height of 1m according to the mass volume ratio of 5 per mill, and a blank (T1) without adding the inoculum and a purchased commercial inoculum (T5) are set as controls.
2. Temperature measurement: during the composting fermentation period, 10 a.m. every 2 days: 00 the four positions of the front, the back, the left and the right of the pile body are measured by a probe thermometer, and the average value is taken to record the ambient temperature at that time.
3. Turning: and (4) on the day of pile building, recording as day 0, starting natural ventilation, turning the pile once every three days, and stopping turning the pile until the temperature of the pile body is reduced to be below 40 ℃ and does not rise any more.
4. The lignocellulose content is measured by placing a fresh sample in an electrothermal blowing dry box for drying at 105 ℃ for 1h, crushing the sample by a crusher, sieving the crushed sample (100 meshes), and then respectively measuring by adopting a 72% concentrated sulfuric acid hydrolysis method, a 2 mol/L hydrochloric acid hydrolysis method and a concentrated sulfuric acid method.
5. The germination index is measured by weighing 20.0g of air-dried matrix, uniformly mixing with 100.0m L ultrapure water, placing the mixture in a shaking table for shaking extraction for 2-3h, filtering by using qualitative filter paper, taking 6.0m L filtrate, dripping the filtrate into culture dishes (the diameter is 9cm) paved with sterile filter paper, placing 20 radish seeds in each culture dish, taking the ultrapure water as a control, setting 3 times of treatment for each treatment, placing the culture dishes in a constant-temperature incubator at 26 ℃ for 48h, calculating the germination rate and the root length, and calculating the germination index by using the following formula:
6. the experimental results are as follows: the results are shown in fig. 6, 7 and 8, all the treatment temperatures show the trend of increasing first and then decreasing, the temperature of the compost can be increased by inoculating the mushroom residue fermentation compound microbial inoculum, and the temperature is advanced to a high temperature period (>55 ℃) 2d compared with the treatment of T1 and a commercial microbial inoculum T5; the high-temperature period can be prolonged, and the inoculated mushroom residue fermentation compound microbial agents T2, T3 and T4 are maintained for 26 days at the high temperature of more than 55 ℃ when being treated, and the treatment time is more than 4 days when being treated by T1 and T5; the inoculated mushroom residue fermentation compound microbial agent can also accelerate the degradation speed of lignocellulose of the stacked materials, and the degradation rates of the lignocellulose of T2, T3 and T4 are all higher than that of the lignocellulose of contrast T1 and the lignocellulose of commercial bacteria agent T5; in addition, the compost can be decomposed more quickly, the GI value is improved, the basic decomposition state is reached by the treatment of T2, T3 and T4 10-15 days earlier than the treatment of the control T1 and T5, and the final germination index of the seeds is obviously higher than that of the treatment of the control T1 and T5. As shown above, the microbial agent special for mushroom residue matrixing fermentation has excellent composting, fermenting and decomposing effects, and can be applied to the matrix utilization and fermentation production of needle mushroom residues.
Claims (9)
1. The compound microbial agent for mushroom residue basification is characterized by mainly comprising thermophilic cellulose degrading bacteria, thermophilic cellulose degrading fungi and thermophilic cellulose degrading actinomycetes; wherein the thermophilic cellulose degrading bacteria are Bacillus urealyticum and Acetobacter pasteurianus; the thermophilic cellulose degradation fungi are aspergillus fumigatus and thermomyces thermophilus; the thermophilic cellulose degradation actinomycetes are Brevibacillus thermoerythraeus and Bacillus saffron.
2. The compound microbial inoculant for mushroom residue basification according to claim 1, wherein the compound microbial inoculant mainly consists of a bacterial solution of bacillus ureafaciens, acetobacter pasteurianus, aspergillus fumigatus, myceliophthora thermophila, brevibacillus thermoerythraeus and bacillus safensis.
3. The compound microbial inoculant for mushroom residue basification according to claim 1, wherein the compound microbial inoculant is mainly prepared by equal-volume mixing of bacterial solutions of bacillus uredoides, acetobacter pasteurianus, aspergillus fumigatus, thermomyces, brevibacillus thermoerythraeus and bacillus safensis.
4. The complex microbial inoculant according to claim 1, wherein the bacillus uredoides, acetobacter pasteurianus, aspergillus fumigatus, thermomyces thermophilus, brevibacillus thermoerythraea and bacillus saffron are self-screened or commercially available.
5. The method for preparing a complex microbial inoculant for the basification of mushroom dregs according to any one of claims 1 to 4, wherein the 6 types of microorganisms described in any one of claims 1 to 4 are cultured and propagated separately, respectively, and after reaching the inoculation condition, the microorganisms are mixed into the complex microbial inoculant in equal volume.
6. The method according to claim 5, wherein the thermophilic cellulose-degrading bacterium is cultured in a beef extract peptone liquid medium; culturing and propagating thermophilic cellulose degradation fungi by using a potato glucose liquid culture medium; the culture and propagation of the thermophilic cellulose degradation actinomycetes uses an improved Gao's No. I liquid culture medium.
7. The method according to claim 5, wherein the inoculation condition is that the OD value of the bacterial solution of the thermophilic cellulose-degrading bacterium is not higher than the OD valueLess than 1, the spore concentration of the bacteria liquid of the thermophilic cellulose degradation fungi and the thermophilic cellulose degradation actinomycetes is not less than 1 × 10-7And each m L.
8. The method of claim 5, comprising the steps of:
step 1, respectively carrying out shake culture on thermophilic cellulose degrading bacteria, thermophilic cellulose degrading fungi and thermophilic cellulose degrading actinomycetes in a beef extract peptone liquid culture medium, a potato glucose liquid culture medium and an improved Gao's I liquid culture medium at 50 ℃ and 200 r/min for 2-3 d;
(2) measuring OD value of thermophilic cellulose degradation bacteria under 600nm, measuring spore concentration in bacterial liquid by thermophilic cellulose degradation fungi and thermophilic cellulose degradation actinomycetes, diluting OD value of bacteria to 1.0 by aseptic water, mixing to obtain bacterial liquid of thermophilic cellulose degradation bacteria, respectively diluting spore concentrations of bacterial liquid of thermophilic cellulose degradation fungi and thermophilic cellulose degradation actinomycetes to 1 × 107Actinomycete and fungus bacteria liquid of L per m, and mixing all the bacteria in equal volume to obtain bacteria liquid.
9. The use of the complex microbial inoculant defined in any one of claims 1 to 4 for the basification of mushroom dregs.
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Application publication date: 20200717 |