CN103710272A - Compound microorganism bacterium agent for aerobic fermentation of biogas residue and preparation method of bacterium agent - Google Patents

Compound microorganism bacterium agent for aerobic fermentation of biogas residue and preparation method of bacterium agent Download PDF

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CN103710272A
CN103710272A CN201310745642.XA CN201310745642A CN103710272A CN 103710272 A CN103710272 A CN 103710272A CN 201310745642 A CN201310745642 A CN 201310745642A CN 103710272 A CN103710272 A CN 103710272A
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natural pond
aerobic fermentation
pond slag
organism fungicide
micro organism
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CN201310745642.XA
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Chinese (zh)
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于芳
吕志敏
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青岛福瑞斯生物能源科技开发有限公司
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Priority to CN201310745642.XA priority Critical patent/CN103710272A/en
Publication of CN103710272A publication Critical patent/CN103710272A/en

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Abstract

The invention provides a compound microorganism bacterium agent for aerobic fermentation of biogas residue and a preparation method of the bacterium agent. The method comprises the steps of I, screening and identifying dominant strains: 1, cultivating strains, 2, screening dominant strains and 3, identifying the dominant strains; and II, preparing a compound solid bacterium agent: inoculating beef extract soup to a triangular flask, cultivating for 48 hours to obtain a seed solution, inoculating to a fermentation flask, implementing amplification cultivation for 48 hours to obtain a fermentation broth, mixing and fermenting a mixture of the fermentation broth and sterilized bran at ratio of 1: 1 for 24-72 hours, and drying to obtain the compound solid bacterium agent which contains 200,000,000 beneficial living bacteria per gram. The preparation method disclosed by the invention is applicable to preparation of the compound solid bacterium agent for aerobic fermentation of biogas residue, so as to realize high added value production, harmless treatment and resource utilization of the biogas residue; therefore, the preparation method can solve problems on rural energy and environment, and can promote rapid development of new rural and urban construction.

Description

A kind of complex micro organism fungicide for aerobic fermentation natural pond slag and preparation method thereof
Technical field
The present invention relates to a kind of complex micro organism fungicide for aerobic fermentation natural pond slag and preparation method thereof.
Background technology
In China rural area, oneself is very general to utilize the direct fermentation producing methanes such as crop stalk, animal excrement, but also rest on as fertilizer and be applied directly on farm crop for the utilization of natural pond slag, this too coarse simple to the utilization of natural pond slag, cannot bring into play its real effect, also environment caused to certain pollution.There are some researches show the further aerobic fermentation of natural pond slag, can form a kind of novel, efficient, safe organic composite fertilizer.It can improve natural pond slag utilization ratio effectively, regulates plant metabolism, strengthens improving activity of root system and nutrient absorption capability.By natural pond slag aerobic fermentation, being the fertilizer that added value is higher, making organic fertilizer production industrialization, product seriation, input commercialization, is the Important Action that improves environment, improves the performance of enterprises, is also the optimal selection of environment protection, the utilization of resources.And microorganism is the key of natural pond slag aerobic fermentation, the variation of its kind and quantity is very large on fermentation impact.And single bacterium, fungi, actinomycetes colony, no matter how high its activity is, and the effect in accelerating Composting Process is all lower than the acting in conjunction of mixing microorganisms flora.At present, the research that natural pond slag aerobic fermentation is produced fertilizer seldom, and not for the research of natural pond slag aerobic fermentation complex bacterial agent special, causes natural pond slag fermentation time long, ferments not thorough, and utilization ratio is low.
Summary of the invention
The invention provides a kind of complex micro organism fungicide for aerobic fermentation natural pond slag and preparation method thereof, by adding described microbiobacterial agent, can greatly improve the speed of natural pond slag aerobic fermentation, and the fertility of biological organic fertilizer, reduce natural pond slag pollution on the environment, promote agriculture development.
For achieving the above object, the present invention is achieved by the following technical solutions:
For a complex micro organism fungicide for aerobic fermentation natural pond slag, it comprises the bacterial strain of bacillus, pseudomonas, Staphylococcus, streptomyces, Penicillium, Eurotium and Trichoderma,
Described bacillus is selected from least one in subtilis BCRC10058, Bacillus licheniformis BCRC15413, bacillus megaterium BAB-2927, subtilis JM4, bacillus megaterium BAB-2761;
Described pseudomonas is Pseudomonas fluorescens PC17;
Described Staphylococcus is selected from least one in staphylococcus xylosus JCM2418, staphylococcus xylosus ATCC29971, Staphylococcus carnosus IP105758, staphylococcus xylosus F11, Staphylococcus carnosus CIP103274;
Described streptomyces is streptomyces microflavus HBUM174047;
Described Penicillium is Paecilomyces lilacinus;
Described Eurotium is selected from least one in aspergillus niger BAB1132, Aspergillus amstelodami NBRC15417, aspergillus niger Ter331, aspergillus niger C3-1;
Described Trichoderma is selected from least one in viride NW537, viride Gr135, koning trichoderma FUE9.
The described complex micro organism fungicide for aerobic fermentation natural pond slag, the preparation method of above-mentioned bacterial strains is as follows:
(1) spawn culture: get multiple-microorganism composite fungus agent and mix with sterile purified water, and by the membrane filtration in 0.25 μ m aperture, the sterile purified water serial dilution of cleaning filter membranes becomes 10 -1-10 -9the diluent of concentration, gets respectively 10 -7-10 -9each l00 μ L of the diluent of concentration is coated with on bacterium, actinomycetes and mould isolation medium, and inoculation is cultivated;
(2) screening of dominant strain: by bacterium colony different on flat board picking cultivation respectively, and select by Starch Hydrolysis test, gelatin liquification test, fat hydrolysis test, cellulose degradation testing sieve the bacterial strain that each degradation capability is the strongest;
(3) evaluation of effective strain: obtain the 16SrRNA fragment of these 20 bacterial strains by the method for 16SrRNA PCR, and check order, go out above-mentioned dominant strain by identify.
Further improvement to technique scheme: in described step (1), spawn culture complex microbial inoculum used is magnificent micro-straw decomposing inoculant D201, green prosperous earthworm power god complex micro organism fungicide, prosperous blue or green cr-j6480 composite fungus agent and Hua Yuangu bio-fertilizer fermenting agent.
Further improvement to technique scheme: described complex micro organism fungicide is containing 200,000,000/g of useful total viable count.
The preparation method of the complex micro organism fungicide for aerobic fermentation natural pond slag described in the present invention also provides, comprise the following steps: the bacteria suspension of making described bacterial strain, be inoculated in seed culture medium, 30 ℃, 220r/min cultivates 48h and obtains seed liquor, by the seed liquor of 10% concentration, be inoculated into enlarged culturing in fermention medium and obtain fermented liquid, after fermented liquid and the mixed fermentation of sterilizing wheat bran, dry, make complex micro organism fungicide.
Further improvement to technique scheme: described seed culture medium and fermention medium are extractum carnis soup, its formula is 0.3% beef extract+1% peptone+0.5%NaCl+ distilled water, regulates pH7.4-7.6 after heating for dissolving.
Further improvement to technique scheme: described fermented liquid and sterilizing wheat bran dry after with the ratio mixed fermentation 48h of 1: 1.
Further improvement to technique scheme: described natural pond slag nutritive ingredient comprises organic matter 36%-49%, humic acid 10.1-24.6%, crude protein 5-9%, nitrogen 0.4-0.6%, potassium 0.6-1.2%, surplus is mineral substance.
The present invention compared with prior art has the following advantages:
1, take organic waste that agriculture production produces is fermented the natural pond slag of generation as composite solid microbial inoculum aerobic fermentation raw material as raw material, preserves the ecological environment, promotes agricultural sustainable development.
2, from various agricultural microbiobacterial agent, filter out the required multiple-microorganism of the most applicable natural pond slag aerobic fermentation, and make composite solid microbial inoculum, effectively improve the fertility of fermentation rate and the biological organic fertilizer of natural pond slag, promote the suitability for industrialized production of natural pond slag fermentation.
3,200,000,000/g of the contained useful total viable count of the made composite solid microbial inoculum of the present invention, surpasses national standard, meets the requirement of biological organic fertilizer suitability for industrialized production.The present invention can prepare the required composite fungus agent of natural pond slag aerobic fermentation, to promote natural pond slag high added value production, harmless treatment and recycling; Solve the villages and small towns energy and environmental problem, promote the fast development of new rural area and city-building.
Read by reference to the accompanying drawings after the specific embodiment of the present invention, it is clearer that the other features and advantages of the invention will become.
Accompanying drawing explanation
Fig. 1 is the DNA extraction result electrophoretogram of bacterial strain in the embodiment of the present invention 1.
Fig. 2 is the 16S PCR result of bacterial strain in the embodiment of the present invention 1.
Embodiment
Below in conjunction with embodiment, technical scheme of the present invention is further described in detail.
Embodiment 1
The present embodiment is to relate to the preparation method that a kind of aerobic fermentation natural pond slag is produced the required complex micro organism fungicide of biological organic fertilizer, comprises the following steps:
One, the screening of dominant strain and evaluation:
A. spawn culture: by multiple-microorganism composite fungus agent (magnificent micro-straw decomposing inoculant D201, green prosperous earthworm power god (complex micro organism fungicide), prosperous blue or green cr-j6480 composite fungus agent, Hua Yuangu bio-fertilizer fermenting agent, all from commercially available composite fungus agent) respectively get 2g and mix with 1L sterile purified water, and by the membrane filtration in 0.25 μ m aperture, with 10ml sterile purified water cleaning filter membranes, this 10ml sterile purified water is mixed, and then serial dilution becomes 10 -1-10 -9the diluent of concentration, gets respectively 10 -7-10 -9each l00 μ L of the diluent of concentration is coated with on bacterium, actinomycetes and mould isolation medium, three repetitions of each concentration gradient, 36 ℃ of inoculation 48h.Spawn culture bacterium used, mould and actinomycetes isolation medium are respectively finished product beef-protein medium, potato sucrose substratum, Gause I substratum.Distilled water used need be used high-pressure sterilizing pot autoclaving 20min under the condition of 121 ℃ of temperature.
Wherein, the bacterial strain mark screening, magnificent micro-straw decomposing inoculant D201 represents with A1-A15; Green prosperous earthworm power god (complex micro organism fungicide) represents with B1-B15; Prosperous blue or green cr-j6480 composite fungus agent represents with C1-C20; Hua Yuangu bio-fertilizer fermenting agent represents with D1-D20.
The screening of b, dominant strain: by bacterium colony different on flat board respectively picking cultivate, and by Starch Hydrolysis test, gelatine liquefication (proteolysis) test, fat hydrolysis are tested, cellulose degradation testing sieve is selected 5 bacterial classifications that each degradation capability is the strongest.
Starch Hydrolysis testing sequence of the present invention is: the single colony inoculation of picking, on starch culture-medium, is cultivated after 48h by 36 ℃, with Lugol's iodine solution uniform fold on substratum, Observe and measure result.Diameter is measured each bacterial strain Starch Hydrolysis ability with transparent circle, to have that it's too late.Bacterial strain hydrolysis ability is with formula Up=(D/d) 2 calculating.In formula, D is transparent circle diameter (mm), and d is colony diameter (mm).The preparation method of described Lugol's iodine solution is: first get 2g liquor kalii iodide in a small amount of distilled water, then get the 1g crystalline flake of iodine and be dissolved in potassium iodide aqueous solution, after all dissolving, adding distil water is mended to 300ml.Starch Hydrolysis test-results is as shown in table 1:
The Up value of table 1 bacterial strain hydrolyzed starch ability
Bacterial strain Up value Bacterial strain Up value Bacterial strain Up value Bacterial strain Up value Bacterial strain Up value
A27.21 A1110.53 B1111.32 C126.31 D1617.81
A37.79 A1413.65 B159.54 C138.72 D1810.32
A48.69 B312.34 C37.43 C145.37 D199.27
A67.91 B68.54 C610.54 C169.18 D207.54
A910.11 B88.21 C99.31 D910.33 Mean value 7.04
Wherein, A11, A14, B3, B11, D16 hydrolyzed starch ability are the strongest.
Gelatine liquefication of the present invention (proteolysis) testing sequence is: the single bacterium colony percutaneous puncture-inoculation of picking is in gelatine liquefication substratum respectively, and 36 ℃ of temperature are cultivated 5d, observe Degree of Liquefaction, before observing, test tube is positioned over to 0 ℃ of cooling 30min.Use formula P=(L 1/ L 2) * 100% represents the power of liquification, and wherein P is liquefaction percentage, L 1the gelatin column length (mm) of liquefaction, L 2gelatin post overall length (mm).Proteolysis test-results is as shown in table 2:
Table 2 bacterial strain is to gelatin (protein) liquification result
Bacterial strain h value Bacterial strain h value Bacterial strain h value Bacterial strain h value Bacterial strain h value
A53.73 B53.32 B154.52 C104.34 D113.13
A63.21 B72.57 C12.54 C163.71 D143.73
A82.04 B82.02 C22.98 C183.63 D154.48
A113.81 B93.83 C34.59 D12.79 D165.21
A123.34 B102.96 C43.89 D22.54 D184.04
A131.51 B122.54 C64.18 D52.31 D194.33
A142.02 B134.03 C72.17 D63.62 D204.68
B32.21 B144.37 C95.91 D72.05 Mean value 1.81
Wherein, B15, C3, C9, D16, D20 liquefy gelatin (protein) ability are the strongest.
Fat hydrolysis testing sequence of the present invention is: the single colony inoculation of picking is in fat hydrolysis substratum respectively, and 36 ℃ of temperature are cultivated 72h, and observation flat-plate bacterial colony and the around colour-change of substratum occur that red person is positive for being hydrolyzed, otherwise negative.Fat hydrolysis test-results is as follows:
In 70 kinds of bacterial strains, the bacterial strain of the grease of degrading has 27 strains (A1, A6, A13, A14, A15, B1, B2, B4, B11, B12, B13, C1, C8, C9, C10, C11, C15, C16, C18, D3, D8, D13, D16, D17, D18, D19, D20).Wherein, A14, B12, C9, C10, the D20 ability that cuts grease is the strongest.
Cellulose degradation testing sequence of the present invention is: the single colony inoculation of picking is in cellulose degradation substratum respectively, and 36 ℃ of temperature are cultivated 30d, and every 3d observed and recorded once.In nutrient solution, filter paper bar (Mierocrystalline cellulose) is as sole carbon source, to Mierocrystalline cellulose, do not have the bacterial strain of Degradation can not grow, can according to whether growing, the speed of growth, filter paper bar attenuation degree and the degradable required time of filter paper judge the power of each bacterial decomposition.
Cellulose degradation test-results is as follows:
In 70 kinds of bacterial strains, the bacterial strain of energy degraded cellulose has 20 strains (A5, A6, A11, A12, A13, B11, B12, B13, C1, C14, C16, C17, C18, D3, D4, D5, D15, D16, D17, D20).Wherein, A6, A11, C16, D5, D16 degraded cellulose ability are the strongest.
Starch culture-medium, gelatine liquefication substratum, fat hydrolysis substratum, cellulose degradation substratum are finished product substratum, without other configuration.
The evaluation of c, effective strain: obtain the 16SrRNA fragment of these 20 bacterial classifications by the method for 16SrRNA PCR, and check order, by identify dominant strain.
The concrete steps of described 16SrRNA PCR are: the single bacterium colony of picking is in 1.5mlEP pipe respectively, add 1ml sterile purified water, mix, place 15min in boiling water, the centrifugal 5min of 10000r, removes distilled water, and precipitation is extracted with DNA rapid extraction test kit, and it is carried out to PCR, result is verified with agarose gel electrophoresis.
Distilled water used need be used high-pressure sterilizing pot autoclaving 20min under the condition of 121 ℃ of temperature.
As shown in Figure 1, the result of the 16S PCR of the bacterial strain filtering out all can adopt electrophoretogram as shown in Figure 2 to represent to DNA extraction result.
Through comparison, the genbank accession number of the dominant strain screening is as shown in table 3:
The genbank accession number of table 3 bacterial strain
Starch based organic matter is had to capacity of decomposition: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride of the degraded organic staphylococcus xylosus of fats and degraded cellulose genus repeats to screen gained.
Two, the preparation of complex micro organism fungicide
Concrete steps are: with the washing of 10ml stroke-physiological saline solution, preserve inclined-plane, to shake up in aseptic triangular flask bacteria suspension, respectively get 1ml and be inoculated in 100ml extractum carnis soup triangular flask, 36 ℃, 220r/min cultivates 48h and obtains seed liquor, is inoculated into enlarged culturing 48h in fermentation flask obtains fermented liquid by the seed liquor of 10% concentration.Fermented liquid and sterilizing wheat bran dry after with the ratio mixed fermentation 48h of 1: 1, make composite solid microbial inoculum, 200,000,000/g of contained useful total viable count.
All substratum and vessel all need with high-pressure sterilizing pot autoclaving 20min under the condition of 121 ℃ of temperature.
Embodiment 2
Present embodiment is different from embodiment one is that the composite fungus agent of multiple-microorganism described in step 1 is respectively got 5g and mixed with 1L sterile purified water, and by the membrane filtration in 0.25 μ m aperture, with 10ml sterile purified water cleaning filter membranes, this 10ml sterile purified water is mixed, and then serial dilution becomes 10 -1-10 -11the diluent of concentration, gets respectively 10 -9-10 -11each l00 μ L of the diluent of concentration is coated with on bacterium, actinomycetes and mould isolation medium.Other step is identical with embodiment one with parameter.
In present embodiment, what identify has capacity of decomposition to starch based organic matter: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride of the degraded organic staphylococcus xylosus of fats and degraded cellulose genus repeats to screen gained.
Embodiment 3
What present embodiment was different from embodiment one or two is that bacterial screening cultivation, enlarged culturing and the fermentation culture temperature used described in step 1 is 45 ℃.Other step is identical with embodiment one or two with parameter.
In present embodiment, what identify has capacity of decomposition to starch based organic matter: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride of the degraded organic staphylococcus xylosus of fats and degraded cellulose genus repeats to screen gained.
Embodiment 4
What present embodiment was different from embodiment one to three is that bacterial screening cultivation, enlarged culturing and the fermentation culture temperature used described in step 1 is 28 ℃.Other step is identical with embodiment one to three with parameter.
What identify has capacity of decomposition to starch based organic matter: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride of the degraded organic staphylococcus xylosus of fats and degraded cellulose genus repeats to screen gained.
Embodiment 5
What present embodiment was different from embodiment one to four is the screening of the dominant strain described in step 2: by bacterium colony different on flat board respectively picking cultivate, and by Starch Hydrolysis test, gelatine liquefication (proteolysis) test, fat hydrolysis are tested, cellulose degradation testing sieve is selected 6 bacterial classifications that each degradation capability is the strongest.Other step is identical with embodiment one to four with parameter.
In present embodiment, its genbank accession number of the dominant strain identifying is as shown in table 4.
Starch based organic matter is had to capacity of decomposition: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride genus of the aspergillus niger of the subtilis of degraded starch class, degrade proteins class, the organic staphylococcus xylosus of degraded fats, degraded cellulose repeats to screen gained.
The genbank accession number of table 4 bacterial strain
Embodiment 6
What present embodiment was different from embodiment one to five is the screening of the dominant strain described in step 2: by bacterium colony different on flat board picking cultivation respectively, and by Starch Hydrolysis test, gelatine liquefication (proteolysis) test, fat hydrolysis test, cellulose degradation testing sieve select 8 bacterial classifications that each degradation capability is the strongest, other step is identical with embodiment one to five with parameter.
In present embodiment, what identify has capacity of decomposition to starch based organic matter: subtilis, bacillus megaterium, streptomyces microflavus, Paecilomyces lilacinus, aspergillus niger; Protein-based organism is had to Decomposition: Bacillus licheniformis, staphylococcus xylosus, Staphylococcus carnosus, Pseudomonas fluorescens, aspergillus niger; Fats organism is had to capacity of decomposition: bacillus megaterium, staphylococcus xylosus, Staphylococcus carnosus, streptomyces microflavus; Mierocrystalline cellulose is had to degradation capability: Aspergillus amstelodami, aspergillus niger, viride, koning trichoderma.
Wherein, the viride of the aspergillus niger of the subtilis of degraded starch class and streptomyces microflavus, degrade proteins class and Staphylococcus carnosus and Bacillus licheniformis, the organic staphylococcus xylosus of degraded fats and bacillus megaterium, degraded cellulose and aspergillus niger genus repeat to screen gained.
The genbank accession number of the bacterial strain screening is as shown in table 5:
The genbank accession number of table 5 bacterial strain
Embodiment 7
By following experimental verification beneficial effect of the present invention:
Composite solid microbial inoculum is linked in the slag muck fertilizer of natural pond, not add the contrast that is treated to of composite fungus agent.Composting conditions is: between waste height 1m-2.5m, and 3d turning 1 time, compost 15d.Envrionment temperature is 25 ℃ of left and right, and compost temperature is controlled at 55 ℃ of left and right, and pH value is controlled between 5.5-8.
Described in this experiment, natural pond slag is that the organic waste that agriculture production produces is the natural pond slag that raw material ferments and produces.It is organic 36%-49% that its nutritive ingredient comprises, humic acid 10.1-24.6%, crude protein 5-9%, nitrogen 0.4-0.6%, potassium 0.6-1.2%, and the mineral substance such as a small amount of copper, iron, manganese, zinc.
This test-results shows, with do not add composite solid microbial inoculum and compare, add appropriate composite solid microbial inoculum greatly to improve the speed of natural pond slag aerobic fermentation, natural pond slag fermentation completely, utilization ratio is high, and the fertilizer after fermentation includes the various crop required nutritive element of growing, and can greatly promote plant growth.
The concrete dominant strain of embodiment of the present invention 1-7 screening all can adopt commercially available bacterial strain, because above-mentioned bacterial strains comes from 4 kinds of described commercially available complex micro organism fungicides (magnificent micro-straw decomposing inoculant D201, green prosperous earthworm power god (complex micro organism fungicide), prosperous blue or green cr-j6480 composite fungus agent, Hua Yuangu bio-fertilizer fermenting agent), commercially available above-mentioned microbial inoculum selects commercially available bacterial strain to be prepared from, the dominant strain of the present invention's screening can come from commercially available bacterial strain, also can adopt screening method screening of the present invention to obtain.
The present invention is by the required composite fungus agent of preparation natural pond slag aerobic fermentation, solved the emission problem of discarded natural pond slag for a long time, the villages and small towns energy and environmental problem have greatly been alleviated, reduce the pollution of agricultural organic waste to environment, realize the recycle of resource, significant to establishing resource saving type, friendly environment society.
Above embodiment is only in order to technical scheme of the present invention to be described, but not is limited; Although the present invention is had been described in detail with reference to previous embodiment, for the person of ordinary skill of the art, the technical scheme that still can record previous embodiment is modified, or part technical characterictic is wherein equal to replacement; And these modifications or replacement do not make the essence of appropriate technical solution depart from the spirit and scope of the present invention's technical scheme required for protection.

Claims (8)

1. for a complex micro organism fungicide for aerobic fermentation natural pond slag, it is characterized in that it comprises the bacterial strain of bacillus, pseudomonas, Staphylococcus, streptomyces, Penicillium, Eurotium and Trichoderma,
Described bacillus is selected from least one in subtilis BCRC 10058, Bacillus licheniformis BCRC 15413, bacillus megaterium BAB-2927, subtilis JM4, bacillus megaterium BAB-2761;
Described pseudomonas is Pseudomonas fluorescens PC17;
Described Staphylococcus is selected from least one in staphylococcus xylosus JCM 2418, staphylococcus xylosus ATCC 29971, Staphylococcus carnosus IP105758, staphylococcus xylosus F11, Staphylococcus carnosus CIP103274;
Described streptomyces is streptomyces microflavus HBUM174047;
Described Penicillium is Paecilomyces lilacinus;
Described Eurotium is selected from least one in aspergillus niger BAB1132, Aspergillus amstelodami NBRC 15417, aspergillus niger Ter331, aspergillus niger C3-1;
Described Trichoderma is selected from least one in viride NW537, viride Gr135, koning trichoderma FUE9.
2. the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 1, is characterized in that: the preparation method of above-mentioned bacterial strains is as follows:
(1) spawn culture: get multiple-microorganism composite fungus agent and mix with sterile purified water, and by the membrane filtration in 0.25 μ m aperture, the sterile purified water serial dilution of cleaning filter membranes becomes 10 -1-10 -9the diluent of concentration, gets respectively 10 -7-10 -9each l00 μ L of the diluent of concentration is coated with on bacterium, actinomycetes and mould isolation medium, and inoculation is cultivated;
(2) screening of dominant strain: by bacterium colony different on flat board picking cultivation respectively, and select by Starch Hydrolysis test, gelatin liquification test, fat hydrolysis test, cellulose degradation testing sieve the bacterial strain that each degradation capability is the strongest;
(3) evaluation of effective strain: obtain the 16SrRNA fragment of these 20 bacterial strains by the method for 16SrRNA PCR, and check order, go out above-mentioned dominant strain by identify.
3. according to the required composite solid microbial inoculum of preparation natural pond slag aerobic fermentation described in claims 1, it is characterized in that in described step (1), spawn culture complex microbial inoculum used is magnificent micro-straw decomposing inoculant D201, green prosperous earthworm power god complex micro organism fungicide, prosperous blue or green cr-j6480 composite fungus agent and Hua Yuangu bio-fertilizer fermenting agent.
4. the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 1, is characterized in that: described complex micro organism fungicide is containing 200,000,000/g of useful total viable count.
5. the preparation method of the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 1, it is characterized in that comprising the following steps: the bacteria suspension of making described bacterial strain, be inoculated in seed culture medium, 30 ℃, 220r/min cultivates 48h and obtains seed liquor, by the seed liquor of 10% concentration, be inoculated into enlarged culturing in fermention medium and obtain fermented liquid, after fermented liquid and the mixed fermentation of sterilizing wheat bran, dry, make complex micro organism fungicide.
6. the preparation method of the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 5, it is characterized in that: described seed culture medium and fermention medium are extractum carnis soup, its formula is 0.3% beef extract+1% peptone+0.5%NaCl+ distilled water, regulates pH 7.4-7.6 after heating for dissolving.
7. the preparation method of the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 5, is characterized in that: described fermented liquid and sterilizing wheat bran dry after with the ratio mixed fermentation 48h of 1: 1.
8. the preparation method of the complex micro organism fungicide for aerobic fermentation natural pond slag according to claim 5, is characterized in that: described natural pond slag nutritive ingredient comprises organic matter 36%-49%, humic acid 10.1-24.6%, crude protein 5-9%, nitrogen 0.4-0.6%, potassium 0.6-1.2%, surplus is mineral substance.
CN201310745642.XA 2013-12-30 2013-12-30 Compound microorganism bacterium agent for aerobic fermentation of biogas residue and preparation method of bacterium agent CN103710272A (en)

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CN110699286A (en) * 2019-10-28 2020-01-17 湖南农业大学 Microbial agent for relieving potato continuous cropping obstacle and preparation method and application thereof

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