CN109867564A - A method of it eliminating biogas slurry agricultural biological fertilizer flexible package flatulence and improves fertilizer efficiency - Google Patents
A method of it eliminating biogas slurry agricultural biological fertilizer flexible package flatulence and improves fertilizer efficiency Download PDFInfo
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Abstract
The invention discloses a kind of methods eliminated biogas slurry agricultural biological fertilizer flexible package flatulence and improve fertilizer efficiency.The present invention is by by composite bacteria agent and CaO2It is added in biogas slurry agricultural biological fertilizer, which is mainly made of bacillus pumilus (Bacillus pumilus WP8), pseudomonas putida (Pseudomonasputida sp.RBP1), acinetobacter calcoaceticus (Acinetobacter sp.A6).The present invention not only efficiently solves high temperature flatulence problem present in biogas slurry agricultural biological fertilizer flexible package, and the biogas slurry effect of fertilizer that can also effectively improve that treated, improves the effect of edaphon abundance and diversity, seed soaking are to improve plant seed germination rate, disease resistance etc..
Description
Technical field
The invention belongs to agricultural wastes processing technology fields, more particularly to one kind to pass through chemical reagent and composite microbial bacteria
The method that agent is used in combination to eliminate biogas slurry agricultural biological fertilizer flexible package flatulence, and by biogas slurry agricultural biological fertilizer, chemistry
Reagent combines the method for improving fertilizer efficiency with complex micro organism fungicide.
Background technique
In recent years, country supports and promotes energetically the harmless treatment and energy benefit of the organic wastes such as cultivation waste
With.The behave can not only eliminate environmental pollution, produce clean energy resource biogas, and the biogas slurry and biogas residue of generation can produce organic fertilizer
Material adapts to energy-saving and emission-reduction and the renewable growth requirement of the energy, meets cultivation-plantation Agricultural recycling economy and country
The strategy of sustainable development.With the implementation of the behave, biogas project (especially large and medium-sized biogas project) increases year by year, generates
A large amount of biogas slurry and biogas residue.The biogas residue of solid state component is generally used for production organic fertilizer or soil conditioner, the natural pond of liquid parts
Liquid yield is big, and biogas slurry commercialization, direct returning to farmland, discharge are current principal modes after process operation.
Nutrient rich in biogas slurry, the bioactivity such as amino acid, humic acid, auxin, hydrolase, vitamin
Substance, abundant, enormous amount the microorganism containing type, has the double grading of bio-feritlizer and biological pesticide simultaneously.By natural pond
Liquid imposes on farmland or breeding water body as organic fertilizer, can not only improve crops/aquatic products yield, improves soil/water body
Quality, and the application of chemical fertilizer and pesticide can be reduced, improve energy resource base, the level of resources utilization.
With the development of membrane technology, the concentration techniques such as reverse osmosis, nanofiltration membrane of use can realize biogas slurry concentration.Commodity
Change concentration biogas slurry fertilizer uses composite soft membrane bag filling to reduce product packaging cost more, but as the temperature rises, in biogas slurry fertilizer
Fermentation process aggravation, the serious even broken bag of biogas slurry fertilizer mantle bag flatulence phenomenon, this not only causes huge warp
Ji loss, while there are security risks in product storage, transport, sale.Therefore, it is soft how to solve biogas slurry agricultural biological fertilizer
Packaging flatulence is the serious problem that enterprise faces.
Summary of the invention
The primary purpose of the present invention is that providing a kind of elimination biogas slurry agricultural biological fertilizer flexible package flatulence and improving fertilizer efficiency
Method, it is intended to while solving biogas slurry agricultural biological fertilizer soft dress flatulence problem, moreover it is possible to effectively improve the biogas slurry agricultural biological
Effect of the fertilizer in terms of improving soil fertility, promoting crop sprout rate, improve seedling quality, Genes For Plant Tolerance disease.
The invention is realized in this way a kind of side for eliminating biogas slurry agricultural biological fertilizer flexible package flatulence and improving fertilizer efficiency
Method, this method are as follows: by composite bacteria agent and CaO2It is added in biogas slurry agricultural biological fertilizer, the composite bacteria agent includes short and small brood cell
Bacillus (Bacillus pumilus WP8), pseudomonas putida (Pseudomonasputida sp.RBP1), acinetobacter calcoaceticus
(Acinetobacter sp.A6)；Wherein,
The bacillus pumilus (Bacillus pumilus WP8), on September 25th, 2011 in " China Microbiological bacterium
Kind preservation administration committee common micro-organisms center " preservation, deposit number are CGMCC No. 5206；
The pseudomonas putida (Pseudomonasputida sp.RBP1), on August 31st, 2011 " Chinese micro-
Biological inoculum preservation administration committee common micro-organisms center " preservation, deposit number CGMCCNo.5205；
The acinetobacter calcoaceticus (Acinetobacter sp.A6), on July 2nd, 2015 in " Chinese microorganism strain preservation
Administration committee's common micro-organisms center " preservation, deposit number are CGMCC No.11033.
Preferably, in the biogas slurry agricultural biological fertilizer, CaO2Concentration be 0.2~0.5g/L, WP8 bacteria concentration be 106~
109Cfu/L, RBP1 bacteria concentration are 105~108Cfu/L, A6 bacteria concentration are 105~108cfu/L。
Preferably, the composite bacteria agent is mainly by bacillus pumilus (Bacillus pumilus WP8) suspension, stench
Pseudomonad (Pseudomonasputida sp.RBP1) suspension, acinetobacter calcoaceticus (Acinetobacter sp. A6) suspension are mixed
It is obtained after supernatant is removed in centrifugation plus sterile water is uniform after conjunction；Wherein,
Bacillus pumilus (Bacillus pumilus WP8) the suspension OD600For 0.8~1.3, concentration 107~
1011cfu/mL；
Pseudomonas putida (Pseudomonasputida sp.RBP1) the suspension OD600It is for 0.8~1.3, concentration
106~1010cfu/mL；
Acinetobacter calcoaceticus (Acinetobacter sp.A6) the suspension OD600For 0.6~1.2, concentration 105~109cfu/
mL。
Preferably, the centrifugation is to be centrifuged 10min under 8000rpm.
Preferably, CaO in the biogas slurry agricultural biological fertilizer2Concentration be 0.2~0.5g/L.
The present invention overcomes the deficiencies of the prior art and provide a kind of elimination biogas slurry agricultural biological fertilizer flexible package flatulence and improves
The method of fertilizer efficiency.The present invention is used in combination by chemical reagent calcium oxide with microbial bacterial agent, to solve biogas slurry agricultural biological fertilizer
Expect flexible package flatulence problem, and effectively improves the biogas slurry agricultural biological fertilizer simultaneously and improving soil fertility, promoting crop sprout
Effect in terms of rate, raising seedling quality, Genes For Plant Tolerance disease.
In the present invention, calcium oxide (CaO2) a kind of environmentally friendly material for having both oxygen release and oxidisability, odorless, nothing
Poison, the property with oxygen release and neutralization acid soil, improves soil physical and chemical property and soil nutrient status, while promoting microorganism
Activity, is widely used to soil improvement, aquatic products oxygenation, sewage treatment, fruit and vegetable is fresh-keeping etc..
In addition, microbial bacterial agent by Bacillus pumilus WP8, Pseudomonasputida sp.RBP1,
Acinetobacter sp.A6 is constituted.Wherein, Bacillus pumilus WP8 be obtained from rhizosphere soil it is 1 plant really both wide
Compose it is disease-resistant, and efficiently growth-promoting plant growth-promoting rhizobacteria (plant growth promoting rhiz obacteria,
PGPR), and the application success on raising rice seedlings, cowpea, tomato, semen viciae fabae etc. plant.Bacillus pumilus WP8 is
Bacillus pumilus, August in 2011 31 days in " China Committee for Culture Collection of Microorganisms's common micro-organisms center " preservation,
Preservation address is the institute 3 of BeiChen West Road, Chaoyang District, BeiJing City 1, and deposit number is CGMCC No.5206.
Pseudomonasputida sp.RBP1 be from the reed rhizosphere soil of Jiangsu Province, China Dafeng Milu Nature Reserve separation,
The plant growth-promoting rhizobacteria (plant growth promoting rhizobacteria, PGPR) filtered out promotes rice seedling
Seedling quality improves the pseudomonas pseudomonas putida of plant biological and ecological methods to prevent plant disease, pests, and erosion effect, and on August 31st, 2011 in " China Microbiological bacterium
Kind preservation administration committee common micro-organisms center " preservation, preservation address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, is protected
Hiding number is CGMCCNo.5205.Acinetobacter sp.A6 bacterial strain is to produce culture pond from the big abundance of water in Jiangsu Province, China Yancheng
It separated in bed mud, screen gained, have purification function to freshwater aquiculture eutrophication water, on July 2nd, 2015 in " China
Microbiological Culture Collection administration committee common micro-organisms center " preservation, preservation address are BeiChen West Road, Chaoyang District, BeiJing City 1
Institute 3, deposit number are CGMCC No.11033.
Compared with the prior art the shortcomings that and deficiency, the invention has the following advantages:
(1) present invention effectively eliminates high temperature flatulence problem present in biogas slurry agricultural biological fertilizer flexible package；
(2) soil fertility after the application of biogas slurry agricultural biological fertilizer, edaphon abundance and more can be improved in the present invention
Sample；
(3) seed germination rate after the seed soaking of biogas slurry agricultural biological fertilizer can be improved in the present invention；
(4) the disease resistance ability of industrial crops after the application of biogas slurry agricultural biological fertilizer can be improved in the present invention.
Detailed description of the invention
Fig. 1 is pressure differential method experimental provision schematic diagram；Wherein, 1 is U-shaped differential manometer, and 2 be emptying folder, and 3 be surge flask, and 4 be taper
Bottle, 5 be thermostat water bath；
Fig. 2 is different formulations to the effect for eliminating biogas slurry agricultural biological fertilizer (commodity A) flexible package production gas；Wherein, column
Line shows three duplicate standard errors；Ck: control group；CaO2: add certain density CaO2；Composite bacteria agent: Bacillus
pumilus WP8,Pseudomonasputida sp.RBP1,Acinet obacter sp.A6；CaO2+ composite bacteria agent: CaO2
It is mixed with composite bacteria agent；
Fig. 3 is different formulations to the effect for eliminating biogas slurry agricultural biological fertilizer (commodity B) flexible package production gas；Wherein, column
Line shows three duplicate standard errors, ck: control group, CaO2: add certain density C aO2, composite bacteria agent: Bacillus
pumilus WP8,Pseudomonasputida sp.RBP1,Acineto bacter sp.A6；CaO2+ composite bacteria agent: CaO2
It is mixed with composite bacteria agent；
Fig. 4 is the soil 16s rRNA abundance for applying the biogas slurry fertilizer of different formulations；Wherein, columnar alignment show three it is duplicate
Standard error；Ck: fertilizer is not applied；A: commercialization biogas slurry agricultural biological fertilizer A；B: commercialization biogas slurry agricultural biological fertilizer B；Natural pond
Liquid fertilizer (CaO2): addition CaO2Biogas slurry agricultural biological fertilizer；Composite bacteria agent: addition Bacillus pumilus WP8,
The biogas slurry agricultural biological fertilizer of Pseudomonasputida sp.RBP1, Ac inetobacter sp.A6；CaO2+ compound bacteria
Agent: addition CaO2With the biogas slurry agricultural biological fertilizer of composite bacteria agent；
Fig. 5 is the influence for applying different formulations biogas slurry fertilizer to rice seed germination rate；Wherein, 0 is control group, does not add natural pond
Liquid；10%, the lettuce of various concentration (is added to composite bacteria agent and CaO after 20%, 40%, 60%, 80% dilution2)；Figure
Middle letter is different to indicate significant difference (p ﹤ 0.05)；
Fig. 6 is the influence for applying different formulations biogas slurry fertilizer to tomato seedling resistance to bacterial wilt；Wherein, Rs1115 is addition natural pond
The pathogen experimental comparison group of liquid fertilizer；Biogas slurry+composite bacteria agent is the pathogen experiment after adding biogas slurry fertile (containing composite bacteria agent)
Group；Biogas slurry+CaO2(contain CaO for addition biogas slurry fertilizer2) after pathogen experimental group；Biogas slurry+composite bacteria agent+CaO2To add natural pond
Liquid fertilizer (contains composite bacteria agent and CaO2) after pathogen experimental group.
Specific embodiment
In order to make the objectives, technical solutions, and advantages of the present invention clearer, with reference to the accompanying drawings and embodiments, right
The present invention is further elaborated.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, and
It is not used in the restriction present invention.
Embodiment
Bacterial strain activation medium (LB culture medium): being added tryptone 10g, yeast extract 5g in 950ml deionized water,
NaCl 10g shakes container up to solute dissolution, is settled to 1L with deionized water to 7.0 with 1mol/L NaOH tune pH,
Steam sterilizing 20min under 15psi high pressure.
The activation of Bacillus pumilus WP8 bacterial strain: bacterial strain is gone on LB solid plate, and 30 DEG C of inversion cultures 18~
After for 24 hours, picking single colonie goes to LB liquid medium, and 30 DEG C, 120~150rpm shaken cultivation 1 8~prepare seed afterwards for 24 hours
Liquid.It is gone in LB liquid medium with 1% inoculum concentration, after 30 DEG C of shaken cultivations 1 8~for 24 hours, goes to 4 DEG C of environment and stand
Bacteria suspension is prepared after 30min.
The activation of Pseudomonasputida sp.RBP1 bacterial strain: bacterial strain is gone on LB solid plate, 30 DEG C of inversion cultures
18~for 24 hours after, picking single colonie goes to LB liquid medium, 30 DEG C, 120~150rpm shaken cultivation 18~prepare kind afterwards for 24 hours
Sub- liquid.It is gone in LB liquid medium with 1% inoculum concentration, 30 DEG C of shaken cultivations 18~prepare bacteria suspension afterwards for 24 hours.
The activation of Acinetobacter sp.A6 bacterial strain: bacterial strain is gone on LB solid plate, and 25 DEG C of inversion cultures 18~
After for 24 hours, picking single colonie goes to LB liquid medium, and 25 DEG C, 120~150rpm shaken cultivation 18~prepare seed afterwards for 24 hours
Liquid.It is gone in LB liquid medium with 1% inoculum concentration, 25 DEG C of shaken cultivations 18~prepare bacteria suspension afterwards for 24 hours.
Bacteria suspension preparation: taking above-mentioned bacterium solution to go to centrifuge tube, and 8000rpm is centrifuged 10min, abandons supernatant, adds a small amount of sterile
Water mixes, adjustment WP8 bacteria suspension to OD600It is 0.8~1.3, adjustment RBP1 bacteria suspension to OD600It is 0.8~1.3, adjusts A6 bacterium
Suspension is to OD600It is spare after=0.6~1.2.
The preparation of composite bacteria agent: above-mentioned WP8, RBP1 and A6 bacteria suspension is that 2:11 ratio is mixed with volume ratio by sterile working
It closes, plastc ring 4mL, 8000rpm is taken to be centrifuged 10min, abandon supernatant, a small amount of sterile water goes to the sterile XiLin 1mL after mixing
Bottle, freeze-drying obtain composite bacteria agent afterwards for 24 hours.
Chemical reagent and microbial bacterial agent system putting mode: CaO is added in biogas slurry agricultural biological fertilizer2, additive amount is
0.2~0.5g/L；It takes suitable quantity of water or biogas slurry to dissolve above-mentioned composite bacteria agent, is added in 1L biogas slurry agricultural biological fertilizer and mixes, obtain
Into agricultural biological fertilizer, WP8 bacteria concentration is 106~109Cfu/L, RBP1 bacteria concentration are 105~108Cfu/L, A6 bacteria concentration is
105~108cfu/L。
Effect example
One, experimentation
200mL biogas slurry agricultural biological fertilizer is added in 500mL triangular flask, adds the CaO of 0.4g/L thereto2It is mixed
It is even；WP8, RBP1, A6 composite powder are added, with being added in system after the dissolution of a small amount of sterile water, bacterium initial concentration divides in system
It Wei 109cfu/L、108cfu/L、107cfu/L.After system mixes, the closed bottleneck of rubber stopper simultaneously smears vaseline, is sequentially connected
Surge flask U-tube manometer (device such as Fig. 1).Triangular flask is placed in 35 DEG C of water-baths, each experimental group is recorded and produces atmospheric pressure.
3 samplings are randomly selected in the sample ground of biogas slurry agricultural biological fertilizer (commodity biogas slurry fertilizer A, commodity biogas slurry fertilizer B)
Area selects 3 sampled points in each sample region.Each sampled point acquires soil sample 3 of 10cm depth respectively, and soil sample is vertical after acquisition
It is fitted into sterile envelope, car refrigerator takes back -4 DEG C of laboratory cryo-conservation.Removing stone, plant residue waited 2mm to sieve, fast
Speed extracts soil microbial DNA to do microbiological analysis.Using the DNA rapidly extracting kit (Power of Mo BIO company
Soil edaphon total DNA) is extracted, step is carried out by operating instruction.The PCR amplification of microbe genome DNA: bacterium amplification
Primer uses primer pair:
Positive F338:5'-ACTCCTACGGGAGGCAGCAG-3'；
Reversed R518:5'-ATTACCGCGGCTGCTGG-3'.
95 DEG C of initial denaturation 3min before this headed by 16s rRNA amplification condition, 40 circulations are 95 DEG C of denaturation 1m in, and 56 DEG C are moved back
Fiery 1min, 72 DEG C are prolonged 1min, and last 72 DEG C of extensions 5min is stored in 4 DEG C of refrigerators.Each sample is repeated 3 times.
PCR product after amplification detects quality with 1.2% agarose gel electrophoresis.Denaturing gradient gel electrophoresis (DGGE):
The preparation of denaturing gradient gel uses 475 type gradient glue system (475 Gradient of Mo del of Bio-RAD company
DeliverySystem).Denatured gradient is 35%~60% from top to bottom, and polyacrylamide concentration is 8%.90V's
Under voltage, 60 DEG C of electrophoresis 9h.After electrophoresis, gel is fixed into 3min in fixer, dyes 10min with silver staining liquid, then
It is washed with distilled water 2 times, then plus developing liquid developing 3min；By the Gel Doc-2000 gel of the Bio-RAD of the gel after dyeing
Image analysis system is taken pictures；The quantity of sample electrophoresis band, luminance peaks are determined with Imange lab analysis software (Bio-RAD)
Value, carries out clustering with SPSS 13.0.
Concentration biogas slurry is diluted with deionized water, obtains the biogas slurry of 10%, 20%, 40%, 60% and 80% concentration gradient.?
The concentration biogas slurry 50mL after each concentration gradient dilutes, CaO in above-mentioned biogas slurry are added in beaker2Concentration is 0.2g/L, WP8 bacteria concentration
It is 108Cfu/L, RBP1 bacteria concentration are 106Cfu/L, A6 bacteria concentration are 106cfu/L.It is put into 200 seeds in each beaker, makes
Seed is completely immersed in biogas slurry, and each processing is repeated 3 times.For rice paddy seed anoxic during anti-seed soaking, constant temperature is placed the beaker
Oscillator culture, temperature is 30 DEG C, revolving speed 80rmin, after seed soaking for 24 hours, after biogas slurry is planted away in deionized water cleaning, by seed
It is transferred to illumination box.Seed is placed in glass culture dish, spreads 1 layer of filter paper, with deionized water in 2 times daily morning and evening wet filters
Paper is subject to without obvious ponding, and each culture dish places 20 seeds.Rice seed germination rate is recorded after vernalization 3d.
After Ralstonia solanacearum Rs1115 strain transfer is checked its pathogenicity to TTC culture medium, picking
Pink is spare in the bacterium colony expansion culture of flow regime.Picking has the Rs1115 of pathogenicity to be seeded to NB fluid nutrient medium, and 30
DEG C, 180r.min-1Shaken cultivation for 24 hours after, go to sterile 50mL centrifuge tube, 5 000r.min-1It is centrifuged 10min, abandons supernatant,
Cell is resuspended in sterile water, mixes, is adjusted to required concentration with sterile water using preceding.
Add composite bacteria agent, CaO2Into concentration biogas slurry, CaO is adjusted2Concentration be 0.2~0.5g/L, WP8 bacteria concentration is
106~109Cfu/L, RBP1 bacteria concentration are 105~108Cfu/L, A6 bacteria concentration are 105~108cfu/L.Before inoculation, in tomato children
3~5cm zanjon is drawn with knife in the side of seedling, and with the above-mentioned biogas slurry of 50mL centrifuge tube quantitative, in lateral irrigation note biogas slurry of being completely cured, every plant is filled
5~10mL of biogas slurry or so is infused, control group equivalent sterile water replaces.Additional 1 above-mentioned biogas slurry every 3 days is inoculated with 3 times altogether.After
Watering keeps ground moistening daily, cannot neglect dry suddenly wet.
Ralstonia solanacearum Rs1115 pathogen inoculation method: after inoculation biogas slurry the 12nd day, by spare blueness
Withered bacterium Rs1115 bacteria suspension is configured to 3 × 10 with sterile water6Cfu/mL suspension.With same method, being completely cured, lateral irrigation infuses bacterium
Liquid, 100 plants of perfusion 500mL (every plant of perfusion bacterium solution 5mL or so), control group is replaced with equivalent sterile water.Watering is protected daily later
Ground moistening is held, cannot be neglected dry suddenly wet.
Biocontrol effect group seedling tomato diseased plant number is observed and recorded in the same time daily from idiopathy, the state of an illness is calculated and refers to
Number, biocontrol effect of the analysis biogas slurry to bacterial wilt of tomato.Disease scale method are as follows: 0 grade asymptomatic for blade face, and 1 grade is on plant
1/4 blade face below shows wilting symptom, and 2 grades show wilting symptom for 1/4~1/2 blade face on plant, and 3 grades are on plant 1/2
The above blade face shows wilting symptom, and 4 grades dead for complete stool wilting.
Disease index calculation method: disease index (%)=∑ (diseased plant numbers at different levels × typical value at different levels)/(investigation total strain number
× superlative degree typical value) × 100.
Biocontrol effect calculation method: biocontrol effect (%)=(biogas slurry+Rs1115 experimental group disease index-CK+Rs1115
Experimental group disease index)/CK+Rs1115 experimental group disease index × 100.
Two, experimental result
As a result as shown in Fig. 2~4, table 1:
The commercialization biogas slurry agricultural biological fertilizer A (addition microbial bacterial agent) of different component is added it can be seen from Fig. 2
Pressure change under 35 DEG C of environment of high temperature in closed system.Wherein, as time goes by, biogas slurry agricultural biological fertilizer is commercialized
A enclosed system pressure reaches 2800Pa, and biogas slurry flatulence phenomenon can be reduced by respectively handling in 20h；After 23h, compared with the control group
Compared with addition CaO2It is remarkably decreased with the experimental group pressure of composite bacteria agent.
It can be seen from figure 3, the commercialization biogas slurry agricultural biological fertilizer B for adding different component is sealed under 35 DEG C of environment of high temperature
Pressure change in closure system.Wherein, under high temperature, commercialization biogas slurry agricultural biological fertilizer B reaches 1500Pa in 5h, adds CaO2
Experimental group with composite bacteria agent is only 400Pa；After 21h, the pressure for adding the biogas slurry business fertilizer B of different formulations declines to a great extent；25h
Afterwards, control group pressure still rises, and the pressure of each processing group is remarkably decreased, and adds CaO2With the experimental group most final pressure of composite bacteria agent
Power maintains -300Pa or so.
As seen from Figure 4, the edaphon abundance of commercialization biogas slurry agricultural biological fertilizer A, B of different formulations are applied
Variation.Compared with the control group, after biogas slurry agricultural biological fertilizer A, B is commercialized in application, 1 6S rRNA abundance is with composite bacteria agent
Addition rise, the rich value of highest appear in adds CaO2It is 1.6 × 10 in the soil of composite bacteria agent11Copy g-1.In addition,
Biodiversity index based on 16S rRNA map is as shown in table 1 below:
Biodiversity index of the table 1 based on 16S rRNA map
In table 1, ck is not apply fertilizer；A: commercialization biogas slurry agricultural biological fertilizer A；B: commercialization biogas slurry agricultural biological fertilizer
Expect B；Biogas slurry fertilizer (CaO2): addition CaO2Biogas slurry agricultural biological fertilizer；Composite bacteria agent: addition Bacillus pumilus WP8,
The biogas slurry agricultural biological fertilizer of Pseudomonasputida sp.RBP1, Acinetobacter sp.A6；CaO2+ compound bacteria
Agent: addition CaO2With the biogas slurry agricultural biological fertilizer (agricultural biological fertilizer i.e. of the present invention) of composite bacteria agent.
As can be seen from Table 1, composite bacteria agent, CaO are added2In the experimental group of composite bacteria agent, abundance index is high, this table
Bright, composite bacteria agent has positive influence for edaphon；Add CaO2Most with the Shannon index of composite bacteria agent experimental group
Height illustrates the diversity highest of bacterium in soil sample；For the coverage index Coverage of sample all 99% or more, this illustrates institute
Have the sampling of sample all very rationally, sequencing result can in representative sample Multiple drug resistance truth, composite bacteria agent with
CaO2The fertilizer efficiency that soil can be improved is used in combination, improves edaphon abundance and diversity.
Concentration biogas slurry is diluted with deionized water, obtains the biogas slurry of 10%, 20%, 40%, 60% and 80% concentration gradient.?
Concentration biogas slurry 50mL after each concentration gradient dilution is added in beaker, every glass is put into 200 seeds, and seed is made to be completely immersed in biogas slurry
In, each processing is repeated 3 times.For rice paddy seed anoxic during anti-seed soaking, constant temperature oscillator culture is placed the beaker, temperature is
30 DEG C, revolving speed 80rmin-1, after seed soaking for 24 hours, after biogas slurry is planted away in deionized water cleaning, seed is transferred to illumination cultivation
Case.Seed is placed in glass culture dish, spreads 1 layer of filter paper, with deionized water in 2 wet filter paper daily morning and evening, without obvious product
Subject to water, each culture dish places 20 seeds.Rice seed germination rate is recorded after vernalization 3d.
As seen from Figure 5, with the increase of lettuce (0~20%), rice seed germination rate is gradually increased；Work as natural pond
When liquid concentration is 20%, experimental group germination rate reaches peak 81%, adds CaO2Experimental group germination rate with composite bacteria agent is
85%；With further increasing (40~100%) for lettuce, seed germination rate decline, 100% is concentrated biogas slurry treatment group
Seed germination rate is only 9.8%.As a result illustrate, the biogas slurry of 20% or less concentration, which sprouts seed, has facilitation, concentration mistake
Height will inhibit the sprouting of seed, and the best lettuce for promoting seed to sprout is 20%.
As can be seen from Figure 6, Rs1115 group and biogas slurry+CaO2Group plant all starts to present bacterial wilt illness in 11d, and biogas slurry+
CaO2Group bacterial wilt disease index is higher than Rs1115 group；After 12d, the disease index of each processing group is below control group Rs1115
Experiment process group；17d~21d, Rs1115 group bacterial wilt disease index rise without obvious, remaining each experimental group disease index is
Downward trend；Disease index is begun to decline after 21d, and maximum value 77.5% occurs in 21d disease index.In whole experiment process
In, it is added to composite bacteria agent+CaO2Biogas slurry it is best always for the control efficiency of bacterial wilt.
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all in essence of the invention
Made any modifications, equivalent replacements, and improvements etc., should all be included in the protection scope of the present invention within mind and principle.
Claims (5)
1. a kind of method eliminated biogas slurry agricultural biological fertilizer flexible package flatulence and improve fertilizer efficiency, which is characterized in that this method are as follows:
By composite bacteria agent and CaO2It is added in biogas slurry agricultural biological fertilizer, the composite bacteria agent includes bacillus pumilus
(Bacillus pumilus WP8), pseudomonas putida (Pseudomonasputida sp.RBP1), acinetobacter calcoaceticus
(Acinetobacter sp.A6)；Wherein,
The bacillus pumilus (Bacillus pumilus WP8), on August 31st, 2011 in " Chinese microorganism strain guarantor
Hide administration committee's common micro-organisms center " preservation, deposit number is CGMCC No.5206；
The pseudomonas putida (Pseudomonasputida sp.RBP1), on August 31st, 2011 in " China Microbiological
Culture presevation administration committee common micro-organisms center " preservation, deposit number CGMCCNo.5205；
The acinetobacter calcoaceticus (Acinetobacter sp.A6), on July 2nd, 2015 in " Chinese microorganism strain preservation management
Committee's common micro-organisms center " preservation, deposit number are CGMCC No.11033.
2. the method eliminated biogas slurry agricultural biological fertilizer flexible package flatulence as described in claim 1 and improve fertilizer efficiency, feature
It is, in the biogas slurry agricultural biological fertilizer, CaO2Concentration be 0.2~0.5g/L, WP8 bacteria concentration be 106~109Cfu/L,
RBP1 bacteria concentration is 105~108Cfu/L, A6 bacteria concentration are 105~108cfu/L。
3. the method eliminated biogas slurry agricultural biological fertilizer flexible package flatulence as described in claim 1 and improve fertilizer efficiency, feature
It is, the composite bacteria agent is mainly by bacillus pumilus (Bacillus pumilus WP8) suspension, pseudomonas putida
Through being centrifuged after (Pseudomonasputida sp.RBP1) suspension, the mixing of acinetobacter calcoaceticus (Acinetobacter sp.A6) suspension
It goes to obtain after supernatant plus sterile water are uniform；Wherein,
Bacillus pumilus (Bacillus pumilus WP8) the suspension OD600For 0.8~1.3, concentration 107~
1011cfu/mL；
Pseudomonas putida (Pseudomonasputida sp.RBP1) the suspension OD600For 0.8~1.3, concentration 106~
1010cfu/mL；
Acinetobacter calcoaceticus (Acinetobacter sp.A6) the suspension OD600For 0.6~1.2, concentration 105~109cfu/mL。
4. the method eliminated biogas slurry agricultural biological fertilizer flexible package flatulence as claimed in claim 3 and improve fertilizer efficiency, feature
It is, the centrifugation is to be centrifuged 10min under 8000rpm.
5. the method eliminated biogas slurry agricultural biological fertilizer flexible package flatulence as described in claim 1 and improve fertilizer efficiency, feature
It is, CaO in the biogas slurry agricultural biological fertilizer2Concentration be 0.2~0.5g/L.
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