CN108893421B

CN108893421B - Bacillus fusiformis and application thereof in reclamation ecological reconstruction of mining area

Info

Publication number: CN108893421B
Application number: CN201810614833.5A
Authority: CN
Inventors: 李建华; 郜春花; 张强; 李华; 靳东升; 卢晋晶; 李娓
Original assignee: INSTITUTE OF AGRICULTURAL ENVIRONMENT AND RESOURCE SHANXI ACADEMY OF AGRICULTURAL SCIENCES; Shanxi University
Current assignee: INSTITUTE OF AGRICULTURAL ENVIRONMENT AND RESOURCE SHANXI ACADEMY OF AGRICULTURAL SCIENCES; Shanxi University
Priority date: 2018-06-14
Filing date: 2018-06-14
Publication date: 2021-06-08
Anticipated expiration: 2038-06-14
Also published as: CN108893421A

Abstract

The invention discloses a fusiform lysine bacillus and application thereof in reclamation ecological reconstruction of a mining area, wherein the fusiform lysine bacillus is the fusiform lysine bacillus (Lysinibacillus fusiformis), and the preservation number is CGMCC No. 9799. The strain can improve the nutrient content of the reclamation soil in the mining area, and has remarkable effects on improving the contents of organic matters, alkaline hydrolysis nitrogen, quick-acting potassium and available phosphorus in the reclamation soil in the mining area; further experiments show that the bacillus fusiformis can also improve crop biomass, and both can improve the crop biomass in reclamation soil of mining areas for the culture of wheat and rape; more importantly, the application of the lysine bacillus fusiformis can effectively and obviously change the community structure of soil microorganisms, improve the diversity of the soil microorganisms and effectively increase the number of the microorganisms, thereby accelerating the restoration of the soil microorganism community and being widely applied to the restoration of the reclaimed soil in a coal mining area.

Description

Bacillus fusiformis and application thereof in reclamation ecological reconstruction of mining area

Technical Field

The invention belongs to the technical field of ecological reconstruction, and relates to bacillus fusiformis and application thereof in reclamation ecological reconstruction in mining areas.

Background

Coal plays a significant role in energy production and consumption in China, and the specific gravity of the coal is always kept at about 70%. Coal resource mining has lasting and serious negative effects on land resources and ecological environment while supporting social and economic development. The 2007 Chinese geological environment bulletin indicates that the land area occupied and damaged by the mining development in China is 165.8 million hectares, wherein 90.9 million hectares are stacked in tailings, 52.2 million hectares in open pit mining, 20.3 million hectares in mining subsidence, and about 400 million acres of newly-increased damaged land are added every year, wherein more than 60% of the damaged land is cultivated land or other agricultural land, so that the human mole in the coal mining area is continuously aggravated, and the grain safety and the sustainable development of social economy in China are seriously threatened.

The active development of the land reclamation and ecological reconstruction in the mining area is a system engineering with strategic significance. China promulgates a land reclamation regulation in 1988, and definitely regulates responsibility and obligation of land reclamation in the resource development process, the implementation of the regulation can reduce land loss and protect ecological environment, the method is a system engineering benefiting the nation and people, has important economic and social significance, but the land reclamation rate of the mining area in China is less than 10 percent at present. Land reclamation is not only simple reclamation of damaged or disturbed land, but more importantly ecological reclamation, which is the ultimate goal of land reclamation in mining areas. The key of ecological restoration is the restoration of the functions of an ecological system and the construction of a reasonable structure, and the restoration of vegetation and the construction of animal and plant communities and microbial communities are carried out.

At present, in the research of land reclamation in coal mining subsidence areas in China, the traditional fertilization measures are mainly used in various places generally. The main problems of the method are that no living organism substances exist, microorganisms in soil are deficient, the substance energy conversion is slow, the improvement and recovery of soil ecosystems such as physical properties, biological properties and the like of the soil are slow, and the reclamation period is long. Currently, many countries adopt advanced microbial reclamation techniques to reclaim land. The microbial reclamation technology is a biotechnological measure for comprehensively treating and improving the soil in a reclamation area by utilizing the inoculation advantage of microbes. The method comprises the steps of inoculating beneficial and growth-promoting microorganisms to newly-built plants, digging potential fertility of a reclamation matrix by using the microorganisms and life activities of plant rhizosphere microorganisms, recovering and establishing a symbiotic system of the reclamation matrix, promoting plant growth, accelerating matrix improvement of reclamation land soil and continuous development of an ecosystem, and improving the quality of mining area ecological management, so that the reclamation period is shortened. However, the shortage of microbial species with significant efficacy in production practice has been a bottleneck problem in microbial remediation technology.

Disclosure of Invention

The invention aims to overcome the defects of long reclamation period and poor reclamation soil structure in the conventional mining area, and provides lysine bacillus fusiformis with the functions of phosphate solubilizing, potassium solubilizing and nitrogen fixation and application thereof in reclamation ecological reconstruction in the mining area, so that the nutrient level and microbial community of the reclamation soil in the mining area can be effectively improved, the growth of crops is promoted, and the ecological reconstruction speed is accelerated.

The invention is realized by the following technical scheme:

a Bacillus fusiformis with a preservation number of CGMCCNo.9799 is provided.

The lysine bacillus fusiformis can improve the contents of organic matters, alkaline hydrolysis nitrogen, available phosphorus and quick-acting potassium in the reclamation soil in the mine high region.

The lysinibacillus fusiformis can increase the biomass of crops growing in mine reclamation soil.

The lysine bacillus fusiformis can improve the diversity of microbial community structures in reclamation soil of mining areas.

A bacillus fusiformis microbial preparation is prepared by carrying out amplification culture on bacillus fusiformis, then fully mixing active ingredients obtained by culture according to a mass ratio of 2: 10-20 with a carrier, and storing at normal temperature;

the carrier is a solid carrier or a liquid carrier, wherein the solid carrier is a mineral material, a plant material, a high molecular compound or an organic material; the mineral material is one or more of clay, kaolin, montmorillonite, zeolite and fly ash; the plant material is one or more of corn flour, starch, bean flour and peanut flour; the high molecular compound is one or two of polyvinyl alcohol and polyglycol; the organic material is one or more of grass carbon, humic acid and organic fertilizer;

the active ingredients are one or more of living cells obtained by enlarged culture, fermentation liquor containing cells and filtrate of cell culture;

the microbial preparation is prepared into the following dosage forms: liquid, emulsion, suspending agent, granule, wettable powder or water dispersible granule.

The lysine bacillus fusiformis is applied to improving reclamation soil of mining areas.

The lysine bacillus fusiformis or the microbial preparation prepared from the lysine bacillus fusiformis can improve the contents of organic matters, alkaline hydrolysis nitrogen, available phosphorus and available potassium in the reclamation soil of mining areas; and the structural diversity of the microbial community of the reclamation soil in the mining area is improved.

The lysine bacillus fusiformis is applied to preparation of a microbial preparation for improving reclamation soil of mining areas.

The lysine bacillus fusiformis is applied to preparation of a biological fertilizer for promoting growth of reclaimed soil plants in a mining area.

Compared with the prior art, the invention has the following beneficial technical effects:

the lysine bacillus fusiformis provided by the invention is separated and purified from soil collected from a reclamation mining area, then sequentially screened by an inorganic phosphorus selective culture medium, a selective culture medium of solid silicate bacteria and a solid nitrogen-free culture medium, and determined to be a new microorganism through biological identification, 16SrDNA series analysis and phosphorus and potassium dissolving capacity determination: lysine bacillus fusiformis (lysinibacillus fusiformis). The fusiform lysine bacillus shows extremely strong soil adaptability, and is a high-efficiency microorganism with the functions of dissolving phosphorus, potassium and fixing nitrogen; experimental detection shows that the compound fertilizer can improve the nutrient content of the reclaimed soil in a mining area, and has remarkable effects on improving the contents of organic matters, alkaline-hydrolyzable nitrogen, quick-acting potassium and available phosphorus in the reclaimed soil in the mining area; further experiments show that the bacillus fusiformis can also improve crop biomass, and both can improve the crop biomass in reclamation soil of mining areas for the culture of wheat and rape; more importantly, the application of the lysine bacillus fusiformis can effectively and obviously change the community structure of soil microorganisms, improve the diversity of the soil microorganisms and effectively increase the number of the microorganisms, thereby accelerating the restoration of the soil microorganism community, and the lysine bacillus fusiformis can be applied to the improvement of the reclamation soil of a mining area and the promotion of the plant growth of the reclamation soil of the mining area.

Drawings

FIG. 1 shows the form of a Bacillus fusiformis strain of the present invention cultured on a solid beef extract peptone medium for 48 hours;

FIG. 2 shows the morphology of Bacillus fusiformis (Lysinibacillus fusiformis) under an oil microscope (10 x 100 times) after gram staining after 24 hours of culture on solid nutrient agar medium.

Deposit description

The lysine bacillus fusiformis provided by the invention has been preserved in China general microbiological culture Collection center (CGMCC) in 10 and 21 months in 2014, the preservation number is CGMCC No.9799, the preservation unit address is as follows: xilu No. 1 Hospital No. 3, Beijing, Chaoyang, North.

Detailed Description

The present invention will now be described in further detail with reference to specific examples, which are intended to be illustrative, but not limiting, of the invention.

The invention provides a bacillus fusiformis (Lysinibacillus fusiformis), wherein the preservation number of the bacillus fusiformis (Lysinibacillus fusiformis) is CGMCCNo.9799.

The invention provides application of the lysinibacillus fusiformis or the microbial agent in improving reclamation soil of mining areas and promoting plant growth.

The present invention will be further described with reference to the following specific examples.

Example 1

This example illustrates the screening and identification process of lysine bacillus fusiformis (CGMCC No.9799) according to the present invention

1. Sample collection

Collecting soil from typical reclamation mining areas in Shanxi province, respectively collecting 0-20cm soil samples from the weed clumps (N36 degrees 28 '17.846' and E113 degrees 00 '50.383') of mining subsidence reclamation areas in the Shanbei village in Shanxi Changzhi Xiyan county, and preserving at 4 ℃; a0-20 cm soil sample is collected from a weed plexus (N37 degrees 08 '25.485' and E111 degrees 30 '65.453') of a drainage farm of Shanxi Xiaozake Rusmura village and is preserved at 4 ℃.

2. Preliminary screening of bacterial strains

Weighing 10g of the collected reclamation soil of the mining area, adding the reclamation soil into 90mL of sterilized 250mL triangular flask filled with glass beads and sterile water, shaking the flask for 20min at a constant temperature of 30 ℃ and 180r/min, standing the flask for 5min, absorbing 1mL of bacterial liquid into 9mL of sterile water, and fully and uniformly mixing the bacterial liquid (the dilution is 10 at the moment)^-2) Then sucking 1mL of bacteria liquid from the test tube to another sterile water containing 9mL, mixing well, and so on to obtain 10^-3、10^-4、10^-5、10^-6Soil suspensions of various dilutions. Finally, the diluted concentration is 10^-4、10^-5、 10^-60.1mL of the bacterial suspension was applied to a solid inorganic phosphorus selection medium (containing 10g/L, Ca of glucose)₃(PO₄)₂5g/L, 5g/L magnesium chloride, 0.25g/L magnesium sulfate, 0.25g/L potassium chloride, 0.15g/L ammonium sulfate and 18g/L agar), setting three times for each dilution gradient soil suspension, culturing at 30 ℃ for 10D, screening to obtain 8 strains capable of forming large transparent circles, wherein the ratio of the transparent circle diameter (D) to the colony diameter (D) of the 8 strains>1.5。

Inoculating the 8 strains obtained by screening to selective culture medium (Alexander Murfov culture medium containing sucrose 5g/L, magnesium sulfate 0.5g/L, and Na) of solid silicate bacteria by streaking₂HPO₄ 2g/L，CaCO₃0.1g/L，FeCL₃0.005g/L, 1.0g/L of soil mineral and 18g/L of agar) at 28 ℃, after culturing for 4 days, selecting bacterial colonies with large bacterial colonies, high transparent bulges and elastic adhesion, repeatedly drawing lines on an Alexander macroroff culture medium, and performing microscopic examination until obtaining a pure culture, thereby obtaining 4 pure culture bacterial strains in total.

Respectively inoculating the 4 strains obtained by screening to a solid nitrogen-free culture medium (an Asplenium culture medium containing 10G/L of glucose, 0.2G/L of potassium dihydrogen phosphate, 0.2G/L of magnesium sulfate, 0.2G/L of sodium chloride, 0.1G/L of calcium sulfate, 5G/L of calcium carbonate, 18G/L of agar, pH 7.0 and 18G/L of agar) by adopting a streaking method, culturing for 7 days at the temperature of 30 ℃ to obtain 1 strain G-1207 with good growth, and storing the purified strain on a beef extract protein aging culture medium inclined plane.

3. Double sieve

And (4) carrying out phosphorus and potassium dissolving capacity measurement on the screened strain G-1207.

3.1 determination of phosphate solubilizing ability:

strain G-1207 was inoculated into an inorganic phosphorus medium (containing glucose 10G/L, Ca)₃(PO₄)₂5G/L magnesium chloride 5G/L magnesium sulfate 0.25G/L potassium chloride 0.25G/L and ammonium sulfate 0.15G/L) to a G-1207 content of 10⁹cfu/mL, placing on a shaking bed, shaking and culturing at 150r/mim and 30 ℃ for 5 d; simultaneously inoculating Bacillus megaterium (ACCC11107) purchased from agricultural microorganism collection center of Chinese agrology institute of agriculture in the same way as positive control; after inactivation of the strain G-1207, inoculation is used as a negative control, and each treatment is repeated for three times; the obtained fermentation liquor is centrifuged at 5000r/mim for 30min, the content of available phosphorus in the supernatant is measured by molybdenum-antimony colorimetric method, the phosphorus dissolution rate is calculated, and the pH value of the supernatant is measured by an acidimeter (the detection result is shown in Table 1).

TABLE 1 phosphate solubilizing ratios of G-1207

Note: lower case letters on the same column indicate significant differences in p < 0.05.

The detection result shows that the strain has a remarkable phosphate solubilizing effect, and the phosphate solubilizing rate reaches 23.18%.

3.2 determination of Potassium-solubilizing Capacity:

1) preparing a seed solution:

selecting a ring of activated G-1207 slant lawn, inoculating into 30mL seed culture solution (containing starch 5.0G/L, yeast extract 1.0G/L, K)₂HPO₄2.0g/L，MgSO4·7H2O 0.5g/L，CaCO₃0.1 g/L，FeCl₃·6H₂O5 m g/L, pH 7.5-8.5), shaking culture at 180r/min, shaking culture at 28 deg.C for 24 hr to obtain thallus content of 2 × 10⁸cfu/mL. Silicate bacteria (ACCC10013) purchased from the agricultural microorganism Collection of the Chinese agrology institute of agriculture was inoculated in the same manner as a positive control.

2) Shake flask culture of silicate bacteria:

mixing the prepared potassium-dissolving culture medium (containing sucrose 10.0g/L, MgSO)₄·7H₂O 0.5g/L，(NH₄)₂·SO₄ 0.2g/L，NaCl 0.1g/L，CaCO₃0.1G/L, 0.1G/50mL of culture solution of soil minerals, pH 7.2-7.5) 50mL are added into a 250mL conical flask, the flask is sterilized and cooled, 3mL of prepared seed solution is inoculated into each flask, silicate bacteria (ACCC10013) are used as positive control, simultaneously equivalent inactivated G-1207 seed solution is added as negative control, all treatments are repeated for three times, and after inoculation, the flask is subjected to shaking culture for 5 days at the temperature of 28 ℃ at 180 r/min.

3) Research on potassium-dissolving rate of silicate bacteria

Pouring the whole fermentation liquid into an evaporating dish, heating and concentrating in a water bath to about 10ml, adding 2.0ml H₂O₂The solution was evaporated and stirred continuously and 20% H was added repeatedly₂O₂The solution was completely digested to viscous material. 3500r/min centrifugation is carried out for 10min, the supernatant is collected into a 50ml volumetric flask, then distilled water is used for constant volume, the content of water-soluble potassium in the fermentation liquor is measured on a flame photometer, and the detection result is shown in table 2.

TABLE 2 potassium salt removal rates of G-1207

The result shows that the bacterial strain has obvious potassium-dissolving effect, and the increase rate of water-soluble potassium reaches 74.26%.

4. Identification of strains

According to the experimental contents and experimental methods recorded in the handbook of identifying common bacteria systems and the principle, method and practice of bacteria systematics, the strain morphology, physiology and biochemistry and 16SrDNA analysis and identification are carried out on the G-1207 strain obtained by screening.

The morphological characteristics of the strain are as follows: the colony is round, colorless and transparent, has neat edges, and has stick shape and spore shape under electron microscope observation (as shown in FIG. 1).

The physiological and biochemical characteristics of the test pieces are shown in Table 3.

TABLE 3 physiological and biochemical characteristics of G-1207

Contact enzyme	+	b-galactosidase enzyme	－	D-sorbitol	+
						Oxidase enzyme	+	Urease	－	D-mannitol	+
V.P test	－	Arginine double hydrolase	－	D-arabitol	+
						M.R test	－	Lysine dehydroxylase	－	Inositol	+
Nitrate reduction	－	Ornithine dehydroxygenases	+	L-arginine	－
						Liquefaction of gelatin	－	Glycerol	+	L-alanine	－
Starch hydrolysis	－	D-glucose	+	L-aspartic acid	+
						H₂S generation	－	D-galactose	+	L-glutamic acid	+
Indole production	－	D-fructose	+	Growth at 50 deg.C	－
						Citric acid growth	+	D-mannose	+
Growth with 7% NaCl	+	D-maltose	+

The physiological and biochemical determination results of the strain are as follows: gram staining positive (staining results are shown in figure 2), catalase positive, oxidase positive, acetyl methyl alcohol (V.P) negative, methyl red (M.R) negative, nitrate reduction negative, gelatin liquefaction negative, starch hydrolysis negative, hydrogen sulfide production negative, indole test negative, citric acid growth positive, 7% NaCl growth positive, b-galactosidase, urease, arginine double hydrolase, lysine dehydrogenase negative, ornithine dehydrogenase positive, glycerol, D-glucose, D-galactose, D-fructose, the fermentation of D-mannose and D-maltose produces positive acid, the fermentation of D-sorbitol, D-mannitol, D-arabitol and inositol produces positive acid, the fermentation of L-arginine and L-alanine produces negative acid, and the fermentation of L-aspartic acid and L-glutamic acid produces negative acid.

Further performing biological 16SrDNA series analysis, extracting bacterial genome DNA, and performing PCR amplification: 16SrDNA was PCR amplified using 27F/1492R primers with the following sequences:

27F:5'-AGAGTTTGATCCTGGCTCAG-3',

1492R：5'-GGTTACCTTGTTACGACTT-3'

PCR reaction (25. mu.l): template < 1. mu.g, Primer1 (10. mu.M) 1. mu.l, Primer2 (10. mu.M) 1. mu.l, 2 XPfuPCRMastermix 12.5. mu.l, sterilized distilled water to make up to 25. mu.l;

and (3) PCR reaction conditions: pre-denaturation at 93 deg.C for 5min, denaturation at 94 deg.C for 18s, annealing at 56 deg.C for 15s, extension at 72 deg.C for 78s, and circulation for 30 times, and extension at 72 deg.C for 7 min. After the reaction, 5. mu.l of the reaction product was collected and subjected to agarose gel electrophoresis to detect the amplified product. PCR sequencing was performed by the institute for microorganisms of the Chinese academy of sciences.

Sequencing to obtain a 16S rDNA sequence, carrying out BLAST homology comparison by NCBI, and finally determining the sequence to be the Bacillus fusiformis (Lysinibacillus fusiformis) by combining morphological observation and physiological and biochemical test results.

The strain has been preserved in China general microbiological culture Collection center (address: No. 3, Xilu No. 1, Beijing, Chaoyang, Beijing) at 11 months and 5 days in 2014, and the preservation number is CGMCC No. 9799.

Example 2

This example illustrates the cultivation of lysine bacillus fusiformis and the preparation of a microbial agent according to the invention

the carrier is a solid carrier or a liquid carrier, wherein the solid carrier is a mineral material, a plant material, a high molecular compound or an organic material; the mineral material is one or more of clay, kaolin, montmorillonite, zeolite and fly ash; the plant material is one or more of corn flour, starch, bean flour and peanut flour; the high molecular compound is one or two of polyvinyl alcohol and polyglycol; the organic material is one or two of grass carbon, humic acid and organic fertilizer;

Specifically, lysine bacillus fusiformis (Lysinibacillus fusiformis) with the preservation number of CGMCC NO.9799 is inoculated into a broth culture medium (containing 5g/L of beef extract, 10g/L of peptone and 5g/L of NaCl) and cultured under the conditions of 30 ℃ and 200r/min shaking until the bacterial density OD is reached₆₀₀The value is 0.6-0.8, and seed bacterial liquid is obtained.

Adding the seed bacterial liquid into a 100L fermentation tank (fermentation medium contains 10-30 mL/yellow serofluid, 10-50 g/L peanut cake powder, 40-80 g/L sucrose, 2-4 g/L sodium chloride, 2-4 g/L calcium carbonate and 3-6 g/L ferrous sulfate) according to the inoculation amount of 5-10%, culturing at the temperature of 30 ℃, pH 7.2 and the ventilation amount of 0.5vvm, sampling in the culture process, and observing by a microscope direct counting method until the viable count of the fusiform lysine bacillus in each gram of culture solution is 10⁹CFU。

After fermentation is completed, adding turf (carrier) into fermentation liquor (active ingredients) according to the ratio of 2: 10-20, and storing at normal temperature to obtain the microbial agent.

Example 3

This example illustrates the effect of the lysine bacillus fusiformis agent of the present invention on increasing the nutrient content of the reclaimed soil in mining areas

The rape and the spring wheat are planted by a pot culture method, and the planting time is respectively (60 days and 100 days). The matrix soil used for potting is from the reclaimed land of a coal mining subsidence area of southeast, Taihang foot and Luan group in Shanxi province.

The organic matter content of the rape soil is 5.97g/kg, the total nitrogen is 0.50g/kg, the total phosphorus is 0.05g/kg, the total potassium is 20.00g/kg, the alkaline hydrolysis nitrogen is 42.82mg/kg, the available phosphorus is 3.39mg/kg, the quick-acting potassium is 98.60mg/kg, and the pH value is 8.29.

The soil organic matter content of spring wheat is 6.40g/kg, total nitrogen is 0.40g/kg, total phosphorus is 0.4g/kg, total potassium is 19.80g/kg, alkaline hydrolysis nitrogen is 48.76mg/kg, available phosphorus is 2.09mg/kg, quick-acting potassium is 98.5mg/kg, and pH is 8.37.

In the experiment, the microbial inoculum in the example 2 is adopted, and the commercially available ecological restoration microbial inoculum 1 (comparative example 1, gurry dipi biological restoration agent), the ecological restoration microbial inoculum 2 (comparative example 2, proud-aury root microbial inoculum) and the inactivated microbial inoculum of the microbial inoculum in the example 2 (comparative example 3) are selected as controls, and are respectively added into the rape soil and the spring wheat soil for effect comparison. The usage amount of the microbial inoculum is 10 g/kg soil, the application mode is to uniformly mix the microbial inoculum with the soil, then pot the mixture, and then plant crops.

During planting, the soil moisture keeps 70% of the field moisture capacity of the soil. The effect on soil nutrient content after planting is shown in table 4.

TABLE 4 nutrient content to soil after planting

As can be seen from the results in table 4, the lysine bacillus fusiformis microbial agent provided in example 2 has significant effects on increasing the contents of organic matter, alkaline-hydrolyzable nitrogen, quick-acting potassium and available phosphorus in the reclaimed soil in the mining area.

Example 4

This example illustrates the effect of the lysine bacillus fusiformis agent of the present invention on increasing crop biomass.

The soil organic matter content of spring wheat soil is 6.40g/kg, total nitrogen is 0.40g/kg, total phosphorus is 0.4g/kg, total potassium is 19.80g/kg, alkaline hydrolysis nitrogen is 48.76mg/kg, available phosphorus is 2.09mg/kg, quick-acting potassium is 98.5mg/kg, and the pH value is 8.37.

In the experiment, the embodiment is the microbial inoculum in the embodiment 2, and meanwhile, a commercially available ecological restoration microbial inoculum is selected as a comparison; comparative example 1, comparative example 2, comparative example 3 are the same as example 3.

The usage amount of the microbial inoculum is 10 g/kg soil, the application mode is to uniformly mix the microbial inoculum with the soil, then pot the mixture, and then plant crops. Soil moisture during planting maintained 70% of the field capacity of the soil (results for crop biomass are shown in table 5).

TABLE 5 increase in crop Biomass

As can be seen from the results of table 5, the examples have significant effects on increasing the biomass of the reclaimed soil crops in the mining area.

Example 5

This example illustrates the effect of the lysine bacillus fusiformis agent of the present invention on increasing the structural diversity of the microbial community of reclamation soil in mines.

The organic matter content of the rape soil is 5.97g/kg, the total nitrogen is 0.50g/kg, the total phosphorus is 0.05g/kg, the total potassium is 20.00g/kg, the alkaline hydrolysis nitrogen is 42.82mg/kg, the available phosphorus is 3.39mg/kg, the quick-acting potassium is 98.60mg/kg, and the pH value is 8.29. The soil organic matter content of spring wheat soil is 6.40g/kg, total nitrogen is 0.40g/kg, total phosphorus is 0.4g/kg, total potassium is 19.80g/kg, alkaline hydrolysis nitrogen is 48.76mg/kg, available phosphorus is 2.09mg/kg, quick-acting potassium is 98.5mg/kg, and the pH value is 8.37.

In the experiment, the embodiment is the lysine bacillus fusiformis microbial agent in the embodiment 2, and meanwhile, a commercially available ecological restoration microbial agent is selected as a comparison; comparative example 1, comparative example 2, comparative example 3 are the same as example 3.

The usage amount of the microbial inoculum is 10 g/kg soil, the application mode is to uniformly mix the microbial inoculum with the soil, then pot the mixture, and then plant crops. During planting, the soil moisture keeps 70% of the field moisture capacity of the soil.

The soil microbial biomass was characterized by phospholipid fatty acid method (PLFA) and the change in the soil microbial community structure after planting the rape in pots was analyzed (see Table 6).

TABLE 6 alteration of soil microbial community structure after planting oilseed rape in pots

The results in table 6 show that the application of lysenin bacillus fusiformis can effectively and obviously change the community structure of soil microorganisms, improve the diversity of soil microorganisms, obviously increase the number of microorganisms and accelerate the restoration of the soil microorganism community.

Based on the actions of the bacillus fusiformis and the microbial preparation thereof, the following applications are proposed:

application of lysine bacillus fusiformis in improving reclamation soil in mining areas.

Application of lysine bacillus fusiformis in preparing a microbial preparation for improving reclamation soil of mining areas.

The preferred embodiments of the present invention have been described in detail, however, the present invention is not limited to the specific details of the above embodiments, and various simple modifications may be made to the technical solution of the present invention within the technical idea of the present invention, and these simple modifications are within the protective scope of the present invention.

Claims

1. The application of the bacillus fusiformis (Lysinibacillus fusiformis) in improving the contents of organic matters, alkaline hydrolysis nitrogen, available phosphorus and available potassium in the reclaimed soil of a mining area is characterized in that the bacillus fusiformis has a preservation number of CGMCCNo.9799.

2. The application of the lysine bacillus fusiformis microbial preparation according to claim 1, wherein the lysine bacillus fusiformis microbial preparation can improve the contents of organic matters, alkaline-hydrolyzed nitrogen, available phosphorus and available potassium in reclaimed soil of a mining area, and the lysine bacillus fusiformis microbial preparation is prepared by carrying out expanded culture on the lysine bacillus fusiformis, fully mixing active ingredients obtained by culture according to the mass ratio of 2: 10-20 with a carrier, and storing at normal temperature;