CN111849828A - Compound microbial inoculum for decomposed compost of livestock and poultry manure as well as preparation method and application thereof - Google Patents
Compound microbial inoculum for decomposed compost of livestock and poultry manure as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a livestock and poultry manure decomposed compost composite microbial inoculum and a preparation method and application thereof, wherein the composite microbial inoculum consists of a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; the amylolytic strain is one or more of bacillus acidophilus, bacillus thuringiensis, high-temperature resistant bacillus X1 and high-temperature resistant bacillus X2; the cellulose decomposition strain is one or more of high temperature resistant bacillus X3, Klebsiella and Klebsiella subspecies; the protein decomposition strain is one or more of Aspergillus niger, monad fungus, Bacillus licheniformis and Fusarium equiseti. The complex microbial inoculum can be heated to a high-temperature period in a short time, has sufficient duration of the high-temperature period, and has the characteristics of strong cellulose decomposition capability and short composting period.
Description
Technical Field
The invention belongs to the technical field of microorganisms, relates to the field of a complex microbial inoculum for decomposed compost of livestock and poultry excrement, and particularly relates to a complex microbial inoculum for decomposed compost of livestock and poultry excrement as well as a preparation method and application thereof.
Background
The continuous improvement of scale, intensification and industrialization of the breeding industry makes the related treatment requirements of livestock and poultry manure generated by the breeding industry increasingly urgent. Because of environmental protection neglect in the early stage, the livestock manure is discharged randomly without being treated, so that serious environmental pollution is caused, and the livestock manure pollution is parallel to industrial wastewater and domestic sewage and becomes one of three major sources of water environment pollution; moreover, the pollution of the breeding industry and the industrial pollution are combined as two pollution sources in China. Therefore, proper treatment of the livestock manure not only has a great support effect on the healthy development of the breeding industry, but also is necessary requirement for environmental protection at present.
The excrement of the livestock contains elements such as nitrogen, phosphorus, potassium and the like which are necessary for the growth of crops, and in addition, trace elements such as calcium, magnesium, iron, boron and the like, and is a valuable resource. High-temperature composting is a most common harmless and recycling technology for treating livestock manure, and refers to a method for converting livestock manure into a stable organic fertilizer under the action of microorganisms. The high temperature generated in the composting process can kill most pathogenic microorganisms, harmful insect eggs, weed seeds and the like, so that the organic matters in the excrement can be utilized again in a harmless and recycling way.
The traditional natural composting has a long composting period (45-60 days), and the fermentation temperature is low or the high-temperature period is too short, so that a large amount of entomogenous germs in composting raw materials cannot be killed, the composting harmless degree is low, and the fertilizer efficiency is poor. The key to the good effect of composting is how to maintain the temperature of the compost at a high temperature for a certain period of time. This determines the degree of harmlessness of the composting and the length of the composting period.
Currently, most of the existing compost fermentation inoculants have the following two problems:
(1) if the medium-temperature microorganisms are selected more, the fermentation starting time of the compost can be shortened, but the subsequent high-temperature period can basically kill the fermentation microorganisms, so that the high-temperature period is too short, the composting is not thorough, and the composting period is still longer;
(2) neglecting the selection of high-efficiency microorganisms such as cellulose and other refractory substances, the microorganisms in the composting environment can only be decomposed at high temperature under the long-term anaerobic condition, and the composting period is prolonged.
In summary, a new compost complex microbial inoculum of decomposed livestock and poultry feces and a preparation method thereof are needed.
Disclosure of Invention
The invention aims to provide a complex microbial inoculum for decomposing and composting livestock and poultry manure as well as a preparation method and application thereof, so as to solve one or more technical problems. The complex microbial inoculum can be heated to a high-temperature period in a short time, has sufficient duration of the high-temperature period, and has the characteristics of strong cellulose decomposition capability and short composting period.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention relates to a livestock and poultry manure decomposed compost composite microbial inoculum, which consists of a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; the amylolytic strain is one or more of bacillus acidophilus, bacillus thuringiensis, high-temperature resistant bacillus X1 and high-temperature resistant bacillus X2; the cellulose decomposition strain is one or more of high temperature resistant bacillus X3, Klebsiella and Klebsiella subspecies; the protein decomposition strain is one or more of Aspergillus niger, monad fungus, Bacillus licheniformis and Fusarium equiseti.
The composite microbial inoculum is used for composting the livestock manure.
The invention discloses an application of a complex microbial inoculum for decomposing manure and compost of livestock, which comprises the following steps:
mixing the livestock and poultry manure with the crushed crop straw particles according to a predetermined proportion to obtain a pile body;
the liquid complex microbial inoculum is inoculated into the pile body according to 2 per mill to 5 per mill of the mass of the pile body; or the solid compound microbial inoculum is inoculated into the pile body according to 6 per mill to 10 per mill of the mass of the pile body;
uniformly mixing the pile raw material and a microbial inoculum, and periodically measuring the internal temperature of the pile; if the internal temperature is more than or equal to 50 ℃, turning the pile to facilitate ventilation and heat dissipation;
and finishing composting treatment of the livestock manure after 25-30 days.
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: fermenting starch decomposing strains, cellulose decomposing strains and protein decomposing strains to obtain bacterial liquid, wherein the bacterial liquid is prepared from the following raw materials in a volume ratio of (1-3): (4-6): (1-3) uniformly mixing to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
The invention further improves the method and also comprises the following steps:
(4) the preparation of the solid complex microbial inoculum comprises the following steps: adding 500g of sterilized and dried zeolite into every 1L of the liquid composite microbial inoculum, uniformly mixing, and airing to obtain a solid composite microbial inoculum; wherein the viable count is more than 1.0 × 109CFU/g。
The invention is further improved in that, in the step (1), the solid culture medium is LB solid culture medium or PDA solid culture medium; the LB solid culture medium formula is: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 15-20 g of agar are added into each 1000mL of deionized water, and the pH value is 7.0; the PDA solid culture medium formula is as follows: 200g of potatoes, 20g of glucose and 15-20 g of agar are put into each 1000mL of water, and the pH value is natural.
In a further improvement of the present invention, in the step (1), the culturing specifically comprises the following steps: culturing the amylolytic strain and the cellulolytic strain at 30-37 ℃ for 1-2 days; culturing the protein decomposing strain at 20-28 ℃ for 3-4 days.
In a further improvement of the present invention, in the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 30-37 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL; in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 20-28 ℃ until the number of viable bacteria capable of being cultured of each strain is more than or equal to 109And the fermentation is completed at CFU/mL.
The invention is further improved in that in the step (2.1), the LB liquid culture medium has the formula as follows: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0; in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating original strains of acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti on corresponding solid culture media, and culturing to activate the strains;
(2) an expanded culture comprising:
(2.1) respectively inoculating activated strains of acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies and bacillus licheniformis to an LB liquid culture medium, and fermenting to preset requirements;
(2.2) inoculating the activated strains of aspergillus niger, monad fungus and fusarium equiseti to a PDA liquid culture medium respectively, and fermenting to the preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: uniformly mixing bacteria liquid fermented by acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti according to the volume ratio of 1: 1 to obtain a liquid composite microbial agent; wherein the viable count is more than 1.5 × 109CFU/mL。
Compared with the prior art, the invention has the following beneficial effects:
aiming at the defects of the traditional compost and the defects of most of the existing compost fermentation inoculants, the invention provides the high-temperature fast-decomposing compost composite inoculant which can quickly start compost fermentation, can be heated to a high-temperature period in a short time, has sufficient duration of the high-temperature period, has stronger cellulose decomposition capacity and also has the function of shortening the composting period. Specifically, the strains used in the invention are screened from a soil humus layer, have no side effect on an ecosystem, and accord with biological safety regulations. The combination of multiple functional strains can quickly start fermentation, quickly raise temperature, effectively prolong the high-temperature stage and shorten the composting period. The use of high-temperature resistant starch degrading strains, protein degrading strains and cellulose degrading strains and the continuous high temperature in the compost can effectively kill germ, worm eggs and weed seeds in the compost and fully degrade complex organic matters in the compost, so that the compost achieves the aims of harmlessness and reduction, and the decomposed finished product has no peculiar smell.
Detailed Description
The livestock and poultry manure decomposed compost composite microbial inoculum provided by the embodiment of the invention comprises a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; wherein the amylolytic strain comprises: bacillus acidophilus (Bacillus acidola strain), Bacillus thuringiensis (Bacillus thuringiensis), Bacillus thermophilic Bacillus X1(Bacillus sp.X1), Bacillus thermophilic Bacillus X2(Bacillus sp.X2); cellulolytic strains include: bacillus thermophilic X3(Bacillus sp. X3), Klebsiella (Klebsiella), Klebsiella subspecies (Klebsiella pneumoniae); proteolytic strains: aspergillus niger, Phomopsis fungi (Phialomoniumsp), Bacillus licheniformis (Bacillus licheniformis), Fusarium equiseti (Fusarium equiseti). The above strains can be purchased from Yulin desert King Biotech Co., Ltd.
The preparation method of the composite microbial inoculum for the decomposed compost of the livestock and poultry manure comprises the following steps:
(1) activating strains: respectively inoculating original strains of 11 bacteria including acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungi, bacillus licheniformis and fusarium equiseti on corresponding solid culture media (LB solid culture media and PDA solid culture media), then culturing 8 bacteria including acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella and bacillus licheniformis at 37 ℃ for 1-2 days, and culturing three bacteria including aspergillus niger, monad fungi and fusarium equiseti at 25 ℃ for 3-4 days to activate the strains;
(2) and (3) amplification culture: activating 8 activated bacteria including Bacillus acidophilus, Bacillus thuringiensis, high temperature resistant Bacillus X1, high temperature resistant Bacillus X2, high temperature resistant Bacillus X3, Klebsiella subspecies, and Bacillus licheniformisRespectively inoculating the strains in LB liquid culture medium, fermenting at 37 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL; respectively inoculating 3 activated strains of Aspergillus niger, Phomopsis fungus and Fusarium equiseti into PDA liquid culture medium, culturing at 25 deg.C for 72-80 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL;
(3) preparing a liquid composite microbial inoculum: uniformly mixing fermented bacteria liquid of 11 bacteria of acidophilic bacillus, bacillus thuringiensis, high-temperature resistant bacillus X1, high-temperature resistant bacillus X2, high-temperature resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti according to the volume ratio of 1: 1 to obtain the liquid composite microbial inoculum, wherein the viable count of the liquid composite microbial inoculum is more than 1.5 multiplied by 109CFU/mL；
(4) Preparing a solid compound microbial inoculum: mixing the liquid composite microbial inoculum and sterilized and dried zeolite at a ratio of 1 (volume) to 2 (mass), and air drying to obtain solid composite microbial inoculum with viable count of more than 1.0 × 109CFU/g。
In the embodiment of the invention, the LB solid culture medium for activation has the formula as follows: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride, 15-20 g of agar, 1000mL of deionized water and 7.0 of pH, wherein the formula of the PDA solid culture medium for activation is as follows: 200g of potatoes, 20g of glucose, 15-20 g of agar, 1000mL of water and natural pH value.
In the embodiment of the invention, the LB liquid culture medium has the formula as follows: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride, 1000mL of deionized water and pH of 7.0, wherein the PDA liquid culture medium comprises the following components in percentage by weight: 200g of potatoes, 20g of glucose and 1000mL of water, and the pH value is natural.
In the embodiment of the invention, the application of the high-temperature fast-decomposing composting composite microbial inoculum for livestock excrement in livestock excrement composting is characterized in that the livestock excrement is mixed with crushed crop straw particles according to a certain proportion (the carbon nitrogen ratio is about 28), a liquid composite microbial inoculum is inoculated into a pile body according to 2-5 per mill of the mass of the pile body (a solid composite microbial inoculum is inoculated into the pile body according to 6-10 per mill of the mass of the pile body), the internal temperature of the pile body is measured periodically after a pile body raw material and the microbial inoculum are mixed uniformly, pile turning is carried out at the temperature of more than 50 ℃ so as to be beneficial to ventilation and heat dissipation, and composting treatment of the livestock excrement can be completed after 20-25 days.
The invention discloses a compost composite microbial inoculum for quickly decomposing livestock and poultry manure at a high temperature and a preparation method thereof, belonging to the technical field of microorganisms. The purpose is to realize the high-temperature fermentation and the rapid decomposition of the livestock manure compost. The microbial agent consists of a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; wherein the amylolytic strain comprises: bacillus acidophilus (Bacillus acidola strain), Bacillus thuringiensis (Bacillus thuringiensis), Bacillus thermophilic Bacillus X1(Bacillus sp.X1), Bacillus thermophilic Bacillus X2(Bacillus sp.X2); cellulolytic strains include: bacillus thermophilic X3(Bacillus sp. x3), Klebsiella (Klebsiella), Klebsiella subspecies (Klebsiella pneumoniae); proteolytic strains: aspergillus niger, Phomopsis fungi (Phialomoniumsp), Bacillus licheniformis (Bacillus licheniformis), Fusarium equiseti (Fusarium equiseti). The invention is used for composting the feces of the livestock and the poultry, can quickly start fermentation, quickly raise temperature, effectively prolong the high-temperature stage, shorten the composting period, realize harmless and quantitative reduction of the compost and produce decomposed finished products without peculiar smell.
Example 1
The preparation method of the composite microbial inoculum comprises the following steps:
(1) activating strains: the method comprises the steps of respectively inoculating original strains of 11 bacteria including acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungi, bacillus licheniformis and fusarium equiseti on corresponding solid culture media (LB solid culture media and PDA solid culture media), then culturing 8 bacteria including acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies and bacillus licheniformis at 37 ℃ for 1-2 days, and culturing three bacteria including aspergillus niger, monad fungi and fusarium equiseti at 25 ℃ for 3-4 days to activate the strains.
(2) And (3) amplification culture: respectively inoculating 8 activated strains of bacillus acidophilus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies and bacillus licheniformis to an LB liquid culture medium, fermenting at 37 ℃ for 20-24 h until the number of culturable live bacteria of each strain is more than or equal to 109Completing fermentation when CFU/mL; respectively inoculating 3 activated strains of Aspergillus niger, Phomopsis fungus and Fusarium equiseti into PDA liquid culture medium, culturing at 25 deg.C for 72-80 hr until the number of viable bacteria of each strain is greater than or equal to 109And the fermentation is completed at CFU/mL.
(3) Preparing a liquid composite microbial inoculum: uniformly mixing fermented bacteria liquid of 11 bacteria of acidophilic bacillus, bacillus thuringiensis, high-temperature resistant bacillus X1, high-temperature resistant bacillus X2, high-temperature resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti according to the volume ratio of 1: 1 to obtain the liquid composite microbial inoculum, wherein the viable count of the liquid composite microbial inoculum is more than 1.5 multiplied by 109CFU/mL。
(4) Preparing a solid compound microbial inoculum: mixing the liquid composite microbial inoculum and sterilized and dried zeolite at a ratio of 1 (volume) to 2 (mass), air drying to obtain solid composite microbial inoculum with viable count of 1.0 × 10 or more9CFU/g。
Example 2
The application of the composite microbial inoculum in the livestock manure compost provided by the embodiment of the invention comprises the following steps:
(1) mixing fresh excrement of livestock (such as cow dung and pig dung) and crushed crop straw particles (such as corn straw with the length of 2-3 cm) according to a certain proportion, so that the carbon-nitrogen ratio of the mixture is about 26-30, and the water content is 60-65%. The solid complex microbial inoculum is prepared according to 6-10 per mill of the mass of the heap raw materials, and according to the method of scattering a layer of solid complex microbial inoculum on a layer of heap raw materials, the heap raw materials and the microbial inoculum are uniformly mixed to build a strip-stack compost fermentation body with the length of 3.5 meters, the width of 1.8 meters and the height of 1 meter, and meanwhile, a control group is built to carry out natural composting. And measuring the internal temperature of the compost, starting turning the compost if the internal temperature of the compost is higher than 50 ℃, turning the compost once every three days, and turning the compost once every five days when the temperature is reduced to 35 ℃ until the compost fermentation is finished when the internal temperature of the compost is stabilized to the ambient temperature.
(2) After the windrow type compost fermentation body is formed, measuring the internal temperature of the compost body every day, and measuring indexes such as nutrient content of the compost body and seed Germination Index (GI) when the compost fermentation is finished.
The test results of the embodiment of the invention are as follows:
TABLE 1 temperature Change during composting
The temperature of the compost in the compost fermentation is an important influence factor for determining the quality of the compost, influences the microbial activity in the fermentation and the degradation degree of related substances, and is one of the determination parameters for judging whether the compost reaches the harmless degree. The occurrence of high temperature and sufficient duration guarantee that the composting treatment can kill germ and insect eggs, weed seeds and degrade organic matters. In the embodiment, the days of maintaining the compost group inoculated with the microbial inoculum at the temperature of more than 50 ℃ reach 16 days, the requirements that the compost temperature specified by feces innocent health standard (GB7959-87) in China is maintained at the temperature of more than 50-55 ℃ for 5-7 days are completely met, the aim that the compost is used for killing germ and insect eggs in the compost raw materials is achieved, the days of maintaining the compost group at the temperature of more than 50 ℃ are only 5 days in comparison, the standard formulated by the regulation is barely reached, and the rotten degree is obviously poor. The compost group inoculated with the invention is heated to 60 ℃ on the next day because the protein and starch degrading strains in the microbial inoculum quickly utilize related substances, the vital activities are vigorous, and the metabolic products 'detonate' the vital activities of other microorganisms, so that the compost temperature is rapidly increased, and in addition, the high-temperature resistant strains are contained in the invention, so that the microbial activities in the high-temperature period are still normally carried out, the maintenance days of more than 50 ℃ reach 14 days, wherein the days of the highest temperature reaching 80 ℃ also have 4 days, the complete decomposition of compost fermentation is fully ensured, the composting period is greatly shortened, the whole process is only 27 days, and the period of a control group of natural compost reaches 54 days, and the difference between the two can fully show the superiority of the invention.
TABLE 2 index parameters of compost products
According to the regulations of compost quality requirements and excrement innocent health standards (GB7959-87) and the like, the mark that the compost is thoroughly decomposed can be taken as the mark that the carbon-nitrogen ratio is less than 20. From the above table, the carbon-nitrogen ratio of the compost group inoculated with the microbial inoculum at the end of fermentation is 14.69, the comparison group is 19.89, compared with the comparison group of natural fermentation, the compost inoculated with the microbial inoculum has higher rotten degree, and the contents of total nitrogen, total phosphorus and total potassium are higher than those of the comparison group, which further indicates that the organic fertilizer with higher quality can be obtained when the organic fertilizer is used for compost fermentation.
The germination index of the seeds is an index for characterizing the composting degree and is a parameter for reflecting the biological toxicity of the compost. When GI is more than or equal to 80 percent, the organic fertilizer is completely decomposed and has no toxicity to plants. As can be seen from Table 2, the GI of the compost group inoculated with the microbial inoculum is far more than 80 percent, while the GI of the control group is only 81 percent, which fully shows that the inoculation of the microbial inoculum can promote the decomposition of compost fermentation with high quality, and achieves the purposes of harmless and resource treatment of the compost.
Example 3
The livestock and poultry manure decomposed compost composite microbial inoculum provided by the embodiment of the invention comprises a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; the amylolytic strain is bacillus acidophilus; the cellulose decomposition strain is high-temperature resistant bacillus X3; the protein decomposition strain is Aspergillus niger and monad fungus.
Example 4
The livestock and poultry manure decomposed compost composite microbial inoculum provided by the embodiment of the invention comprises a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; the amylolytic strain is bacillus acidophilus and bacillus thuringiensis; the cellulose decomposition strain is high-temperature resistant bacillus X3 and Klebsiella; the protein decomposing strain is aspergillus niger.
Example 5
The livestock and poultry manure decomposed compost composite microbial inoculum provided by the embodiment of the invention comprises a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain; the amylolytic strain is bacillus thuringiensis; the cellulose decomposition strain is Klebsiella; the protein decomposition bacterial strain is fusarium equiseti.
Example 6
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: fermenting starch decomposing strains, cellulose decomposing strains and protein decomposing strains to obtain bacterial liquid, wherein the bacterial liquid is prepared by mixing the following components in a volume ratio of 1: 4: 1 to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
In the step (1), the solid culture medium is an LB solid culture medium or a PDA solid culture medium; the LB solid culture medium formula is: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 15-20 g of agar are added into each 1000mL of deionized water, and the pH value is 7.0; the PDA solid culture medium formula is as follows: 200g of potatoes, 20g of glucose and 15-20 g of agar are put into each 1000mL of water, and the pH value is natural.
In the step (1), the culturing specifically comprises the following steps: culturing the amylolytic strain and the cellulolytic strain at 37 ℃ for 1-2 days; the proteolytic strain is cultured at 28 ℃ for 3-4 days.
In the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 37 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL; in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 28 ℃, and keeping the number of viable bacteria capable of being cultured of each strain to be more than or equal to 109And the fermentation is completed at CFU/mL.
In the step (2.1), the LB liquid culture medium has the formula: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0; in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
Example 7
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: fermenting starch decomposing strains, cellulose decomposing strains and protein decomposing strains to obtain bacterial liquid, wherein the bacterial liquid is prepared by mixing the following components in a volume ratio of 1: 2: 1 to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
(4) The preparation of the solid complex microbial inoculum comprises the following steps: adding 500g of sterilized and dried zeolite into every 1L of the liquid composite microbial inoculum, uniformly mixing, and airing to obtain a solid composite microbial inoculum; wherein the viable count is more than 1.0 × 109CFU/g。
In a further improvement of the present invention, in the step (1), the culturing specifically comprises the following steps: culturing the amylolytic strain and the cellulolytic strain at 30 ℃ for 1-2 days; the proteolytic strain is cultured at 20 ℃ for 3-4 days.
In a further improvement of the present invention, in the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 30 ℃ for 20-24 h until the number of culturable live bacteria of each strain is more than or equal to 109Completing fermentation when CFU/mL; in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 20-28 ℃ until the number of viable bacteria capable of being cultured of each strain is more than or equal to 109And the fermentation is completed at CFU/mL.
The invention is further improved in that in the step (2.1), the LB liquid culture medium has the formula as follows: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0; in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
Example 8
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: dividing the starch intoBacterial liquid obtained by fermenting the zymolytic strain, the cellulose decomposition strain and the protein decomposition strain according to the volume ratio of 2: 5: 2, uniformly mixing to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
In the step (1), the solid culture medium is an LB solid culture medium or a PDA solid culture medium; the LB solid culture medium formula is: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 15-20 g of agar are added into each 1000mL of deionized water, and the pH value is 7.0; the PDA solid culture medium formula is as follows: 200g of potatoes, 20g of glucose and 15-20 g of agar are put into each 1000mL of water, and the pH value is natural.
In the step (1), the culturing specifically comprises the following steps: culturing the amylolytic strain and the cellulolytic strain at 35 ℃ for 1-2 days; the proteolytic strain is cultured at 25 ℃ for 3-4 days.
In the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 35 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL; in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 25 ℃, and keeping the number of viable bacteria capable of being cultured of each strain to be more than or equal to 109And the fermentation is completed at CFU/mL.
In the step (2.1), the LB liquid culture medium has the formula: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0; in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
Example 9
The preparation method of the composite microbial inoculum comprises the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: fermenting starch decomposing strains, cellulose decomposing strains and protein decomposing strains to obtain bacterial liquid, wherein the bacterial liquid is prepared from the following raw materials in a volume ratio of (1-3): (4-6): (1-3) uniformly mixing to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
The invention further improves the method and also comprises the following steps:
(4) the preparation of the solid complex microbial inoculum comprises the following steps: adding 500g of sterilized and dried zeolite into every 1L of the liquid composite microbial inoculum, uniformly mixing, and airing to obtain a solid composite microbial inoculum; wherein the viable count is more than 1.0 × 109CFU/g。
The invention is further improved in that, in the step (1), the solid culture medium is LB solid culture medium or PDA solid culture medium; the LB solid culture medium formula is: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 15-20 g of agar are added into each 1000mL of deionized water, and the pH value is 7.0; the PDA solid culture medium formula is as follows: 200g of potatoes, 20g of glucose and 15-20 g of agar are put into each 1000mL of water, and the pH value is natural.
In a further improvement of the present invention, in the step (1), the culturing specifically comprises the following steps: culturing the amylolytic strain and the cellulolytic strain at 30-37 ℃ for 1-2 days; culturing the protein decomposing strain at 20-28 ℃ for 3-4 days.
In a further improvement of the present invention, in the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 30-37 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL; in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 20-28 ℃ until the number of viable bacteria capable of being cultured of each strain is more than or equal to 109And the fermentation is completed at CFU/mL.
The invention is further improved in that in the step (2.1), the LB liquid culture medium has the formula as follows: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0; in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art can make modifications and equivalents to the embodiments of the present invention without departing from the spirit and scope of the present invention, which is set forth in the claims of the present application.
Claims (10)
1. A compost complex microbial inoculum is characterized by comprising a starch decomposition strain, a cellulose decomposition strain and a protein decomposition strain;
the amylolytic strain is one or more of bacillus acidophilus, bacillus thuringiensis, high-temperature resistant bacillus X1 and high-temperature resistant bacillus X2;
the cellulose decomposition strain is one or more of high temperature resistant bacillus X3, Klebsiella and Klebsiella subspecies;
the protein decomposition strain is one or more of Aspergillus niger, monad fungus, Bacillus licheniformis and Fusarium equiseti.
2. The complex microbial inoculum of claim 1 is used for composting livestock manure.
3. The application of the livestock manure decomposed compost composite microbial inoculum of claim 1, which is characterized by comprising the following steps:
mixing the livestock and poultry manure with the crushed crop straw particles according to a predetermined proportion to obtain a pile body;
the liquid complex microbial inoculum is inoculated into the pile body according to 2 per mill to 5 per mill of the mass of the pile body; or the solid compound microbial inoculum is inoculated into the pile body according to 6 per mill to 10 per mill of the mass of the pile body;
uniformly mixing the pile raw material and a microbial inoculum, and periodically measuring the internal temperature of the pile; if the internal temperature is more than or equal to 50 ℃, turning the pile to facilitate ventilation and heat dissipation;
and finishing composting treatment of the livestock manure after 25-30 days.
4. The preparation method of the complex microbial inoculum of claim 1, which is characterized by comprising the following steps:
(1) strain activation, comprising: respectively inoculating starch decomposing strain, cellulose decomposing strain and protein decomposing strain on corresponding solid culture medium, and culturing to activate strain;
(2) an expanded culture comprising:
(2.1) respectively inoculating the amylolytic strain and the cellulolytic strain activated in the step (1) into an LB liquid culture medium, and fermenting to meet the preset requirement;
(2.2) inoculating the activated protein decomposing strain in the step (1) into a PDA liquid culture medium, and fermenting to a preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: fermenting starch decomposing strains, cellulose decomposing strains and protein decomposing strains to obtain bacterial liquid, wherein the bacterial liquid is prepared from the following raw materials in a volume ratio of (1-3): (4-6): (1-3) uniformly mixing to obtain a liquid composite microbial inoculum; wherein the viable count is more than 1.5 × 109CFU/mL。
5. The method for preparing a complex microbial inoculum according to claim 4, which further comprises:
(4) the preparation of the solid complex microbial inoculum comprises the following steps: adding 500g of sterilized and dried zeolite into every 1L of the liquid composite microbial inoculum, uniformly mixing, and airing to obtain a solid composite microbial inoculum; wherein the viable count is more than 1.0 × 109CFU/g。
6. The method for preparing a complex microbial inoculum according to claim 4, wherein in the step (1), the solid culture medium is LB solid culture medium or PDA solid culture medium;
the LB solid culture medium formula is: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 15-20 g of agar are added into each 1000mL of deionized water, and the pH value is 7.0;
the PDA solid culture medium formula is as follows: 200g of potatoes, 20g of glucose and 15-20 g of agar are put into each 1000mL of water, and the pH value is natural.
7. The preparation method of the complex microbial inoculum according to claim 4, wherein in the step (1), the culture specifically comprises the following steps:
culturing the amylolytic strain and the cellulolytic strain at 30-37 ℃ for 1-2 days; culturing the protein decomposing strain at 20-28 ℃ for 3-4 days.
8. The method for preparing a complex microbial inoculum according to claim 4,
in the step (2.1), the fermentation to the preset requirement specifically comprises: fermenting at 30-37 deg.C for 20-24 hr until the number of viable bacteria of each strain is greater than or equal to 109Completing fermentation when CFU/mL;
in the step (2.2), the fermentation to the preset requirement specifically comprises: culturing for 72-80 h at 20-28 ℃ until the number of viable bacteria capable of being cultured of each strain is more than or equal to 109And the fermentation is completed at CFU/mL.
9. The method for preparing a complex microbial inoculum according to claim 4,
in the step (2.1), the LB liquid culture medium has the formula: 10g of tryptone, 5g of yeast extract powder and 10g of sodium chloride are added into each 1000mL of deionized water, and the pH value is 7.0;
in the step (2.2), the PDA liquid culture medium formula is as follows: 200g of potato and 20g of glucose are put into each 1000mL of water, and the pH value is natural.
10. The preparation method of the complex microbial inoculum of claim 1, which is characterized by comprising the following steps:
(1) strain activation, comprising: respectively inoculating original strains of acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti on corresponding solid culture media, and culturing to activate the strains;
(2) an expanded culture comprising:
(2.1) respectively inoculating activated strains of acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies and bacillus licheniformis to an LB liquid culture medium, and fermenting to preset requirements;
(2.2) inoculating the activated strains of aspergillus niger, monad fungus and fusarium equiseti to a PDA liquid culture medium respectively, and fermenting to the preset requirement;
(3) the preparation of the liquid complex microbial inoculum comprises the following steps: uniformly mixing bacteria liquid fermented by acidophilic bacillus, bacillus thuringiensis, high-temperature-resistant bacillus X1, high-temperature-resistant bacillus X2, high-temperature-resistant bacillus X3, klebsiella subspecies, aspergillus niger, monad fungus, bacillus licheniformis and fusarium equiseti according to the volume ratio of 1: 1 to obtain a liquid composite microbial agent; wherein the viable count is more than 1.5 × 109CFU/mL。
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