CN110218666A - A kind of compound bacteria preparation method and its application for improveing feldspathic sandstone soil - Google Patents
A kind of compound bacteria preparation method and its application for improveing feldspathic sandstone soil Download PDFInfo
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- C—CHEMISTRY; METALLURGY
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Abstract
The invention belongs to agricultural technology, environmental protection and ecological improved application fields; in particular to a kind of compound bacteria preparation method and its application for improveing feldspathic sandstone soil, compound bacteria are made of bacillus P75, rhizobium D10, phosphate solubilizing bacteria H3: the compound bacteria can be used for improveing feldspathic sandstone soil fertility, promotion Growth of Alfalfa, the increase feldspathic sandstone soil organism, increase feldspathic sandstone soil ammonium and nitrate nitrogen, increase feldspathic sandstone soil quick-effective phosphor and available potassium by bacteria agent prepared by the compound bacteria, improve feldspathic sandstone soil saccharase and urease activity.
Description
Technical field
The invention belongs to agricultural technology, environmental protection and ecological improved application field, in particular to a kind of compound bacteria preparation
Method and its application for improveing feldspathic sandstone soil.
Background technique
Triassic period that feldspathic sandstone was specifically referred to by the Permian Period in the Paleozoic Era, Mesozoic Era, Jurassic Period and cretaceous thick-layer sandstone, sand
The rock alternating layers of shale and argillaceous sandstone composition, main integrated distribution is in northern Loess Plateau Shanxi, Shan Inner Mongol contiguous areas
Ordos Plateau.Feldspathic sandstone belongs to continental clastic system, and diagenetic grade is low, degree of consolidation is poor between the grains of sand, structural strength is low,
Have the characteristics that it is anhydrous it is as hard as flint, meet soft such as the mud of yew and to the wind if at sand.The serious weathering in Stands in Arsenic Sandstone Area rock stratum, distribution ground
Area's plant growth is difficult, and ecological environment is severe, and soil erosion situation is extremely serious, has seriously affected local life production, when
The ground masses regard its harm poison such as arsenic, therefore are called " feldspathic sandstone ".Meanwhile the difficulty of governance of feldspathic sandstone is very big, by China and foreign countries expert
Referred to as " world's soil erosion most " and " earth environment cancer ".
It is well known that soil erosion is becoming arid in the world or a common serious problems of semiarid zone,
And plant or biological control measure and other ecological engineerings related with vegetation are the conservation of soil and water and the good ecological environment of realization
Most basic method.Currently, the measure for administering semifinished product and glaze soil erosion is mainly engineering measure and biological control measure, but all deposit
In limitation, can not popularize and large-area construction operation.Both at home and abroad have using microorganism improve barren property soil case and
Research.Liu Lijun administers the aeolian sandy soil of the Northeast using microbe soil conditioner by long term experiment, has with conventional
The plantation of machine fertilizer and traditional planting patterns compare, and primary study microbe soil conditioner is to the Northeast's aeolian sandy soil retain water and nutrients
The influence of effect inquires into microbe soil conditioner to aeolian sandy soil modifying mechanism；Du Hui equality microorganisms are in mine reclamation
Effect and its effect during soil base material improvement can be effectively improved plant nutrient item using microorganism modification method
While part, promotion plant growth and development, using the vital movement of plant rhizosphere microbe, make to lose reclaiming for microbial activity
Area's soil re-establishes and restores edaphon system, increases biological activity of soil, accelerates the base material improvement of reclaimed land soil,
Accelerate natural soils to agrological conversion process, improves immature soil into cultivated land, soil fertility.But microbe application is in feldspathic sandstone
The report of improvement is less, yet there are no the application of composite bacteria agent improvement feldspathic sandstone characteristic.
Chinese patent CN107488618A discloses a kind of bacillus megaterium (Bacillus megaterium) H3, protects
It is hidden in China typical culture collection center, the deposit date is on October 10th, 2016, culture presevation number was CCTCC NO:M
2016551；It can be used for promoting plant growth and reducing crop absorption heavy metal；Chinese patent CN107937302A discloses one
Kind Sinorhizobium meliloti D10 (Sinorhizobiummeliloti D10), is preserved in China typical culture collection center,
The deposit date is on October 10th, 2016, culture presevation number was CCTCC NO:M 2016554；It can be used for promoting energy-source plant raw
Long and joint energy-source plant improves heavy-metal contaminated soil；Above two microorganism is applied individually to any the comprehensive effect of feldspathic sandstone improvement
Fruit is bad.
Summary of the invention
The present invention solves the above-mentioned technical problems in the prior art, provides a kind of compound bacteria preparation method and its improvement
The application of feldspathic sandstone soil.
To solve the above problems, technical scheme is as follows:
A kind of compound bacteria is made of bacillus P75, rhizobium D10, phosphate solubilizing bacteria H3:
The bacillus P75 classification naming is Bacillus halotolerans, is preserved in Chinese Typical Representative culture guarantor
Hiding center, preservation address are Wuhan, China Wuhan University, and the deposit date is on October 17th, 2018, culture presevation number was CCTCC
NO:M 2018690；
The rhizobium D10 classification naming is Sinorhizobium meliloti, is preserved in Chinese Typical Representative culture guarantor
Hiding center, preservation address are Wuhan, China Wuhan University, and the deposit date is on October 27th, 2016, culture presevation number was CCTCC
NO:M 2016554；Sinorhizobium meliloti disclosed in as Chinese patent CN107937302A
(Sinorhizobiummeliloti D10)
The phosphate solubilizing bacteria H3 classification naming is that Bacillus megaterium. is preserved in China typical culture collection
The heart, preservation address are Wuhan, China Wuhan University, and the deposit date is on October 27th, 2016, culture presevation number was CCTCC NO:M
2016551；Bacillus megaterium disclosed in as Chinese patent CN107488618A (Bacillus megaterium H3).
Preferably, in the compound bacteria the ratio between biomass of bacillus P75, rhizobium D10 and phosphate solubilizing bacteria H3 be 4~
6:0.5~1.5:4~6.
A kind of bacteria agent, including the compound bacteria.
Preferably, in the bacteria agent total number of cells of compound bacteria in 500,000,000 CFU/mL or more.
The preparation method of the bacteria agent, comprising the following steps:
Bacillus P75 and phosphate solubilizing bacteria H3 slant strains are inoculated in LB solid medium and are cultivated；Then it chooses full
Single colonie be inoculated in LB liquid medium culture, obtain P75 bacterium solution and H3 bacterium solution；
Rhizobium D10 slant strains are inoculated in TY solid medium and are cultivated, full, sticky single colonie is then chosen
It is inoculated in TY fluid nutrient medium culture, obtains D10 bacterium solution；
By the P75 bacterium solution, H3 bacterium solution, D10 bacterium solution, 1:1:1 is mixed by volume, obtains composite bacteria liquid.
The compound bacteria or bacteria agent can be used for improveing feldspathic sandstone soil fertility.
The compound bacteria or bacteria agent can be used for promoting Growth of Alfalfa.
The compound bacteria or bacteria agent can be used for increasing the feldspathic sandstone soil organism, increase feldspathic sandstone soil ammonium and
Nitrate nitrogen increases feldspathic sandstone soil quick-effective phosphor and available potassium, improves feldspathic sandstone soil saccharase and urease activity.
Compared with the existing technology, advantages of the present invention is as follows,
Compound bacteria of the invention can improve feldspathic sandstone soil fertility, promote rye grass and alfalfa growing, and it is raw to increase plant
Object amount:
(1) the feldspathic sandstone soil pH value for being inoculated with compound bacteria has been remarkably decreased 0.45 unit (P < 0.05).
(2) the feldspathic sandstone soil organic matter content for being inoculated with compound bacteria dramatically increases 38.46% (P < 0.05).
(3) the feldspathic sandstone soil ammonium content for being inoculated with compound bacteria dramatically increases 1.60 times (P < 0.05).
(4) the feldspathic sandstone content of soil nitrate-N for being inoculated with compound bacteria dramatically increases 55.60% (P < 0.05).
(5) sucrase active for being inoculated with the feldspathic sandstone soil of compound bacteria dramatically increases 22.55% (P < 0.05).
(6) urease activity for being inoculated with the feldspathic sandstone soil of compound bacteria dramatically increases 1.36 times (P < 0.05).
(7) available phosphorus contents for being inoculated with the feldspathic sandstone soil of compound bacteria dramatically increase 76.32% (P < 0.05).
(8) quick-acting potassium content for being inoculated with the feldspathic sandstone soil of compound bacteria dramatically increases 41.38% (P < 0.05).
(9) biomass of the alfalfa of inoculation compound bacteria processing dramatically increases 44.52% (P < 0.05).
Specific embodiment
Embodiment 1: the activation of bacterial strain P75, D10 and H3 and the preparation of compound bacteria
It is solid that P75 (CCTCC NO:M 2018690) and H3 (CCTCC NO:M 2016551) slant strains are inoculated in LB
In body culture medium (tryptone 10.0g, yeast extract 5.0g, NaCl 10.0g, distilled water 1000mL, agar 20g, pH 7.0),
30 DEG C of culture 1d.Then choose full single colonie be inoculated in LB liquid medium (tryptone 10.0g, yeast extract 5.0g,
NaCl 10.0g, distilled water 1000mL, pH 7.0), 30 DEG C, 160rpm shaken cultivation 32h, it is ensured that the formation of gemma obtains P75 bacterium
Liquid and H3 bacterium solution.
D10 (CCTCC NO:M 2016554) slant strains are inoculated in TY solid medium (peptone 5.0g, yeast extract
3.0g, anhydrous CaCl20.647g, distilled water 1L, pH 7.0) in, 30 DEG C of culture 1d.Then full, sticky single colonie is chosen
It is inoculated in TY fluid nutrient medium (peptone 5.0g, yeast extract 3.0g, anhydrous CaCl20.647g, distilled water 1 L, pH 7.0),
30 DEG C, 160rpm shaken cultivation 18h obtains D10 bacterium solution；
Finally 1:1:1 mixes bacterium solution P75, D10 and H3 by volume, obtains composite bacteria liquid.In composite bacteria liquid, bacillus
The ratio between biomass of P75, rhizobium D10 and phosphate solubilizing bacteria H3 is 4~6:0.5~1.5:4~6.
Embodiment 2: influence of the composite bacteria agent to feldspathic sandstone soil pH
The P75 bacterium solution of embodiment 1, D10 bacterium solution, H3 bacterium solution and composite bacteria liquid are inoculated in by 5% inoculum concentration respectively
In feldspathic sandstone soil, using alfalfa as test plant, scientific management one month.The feldspathic sandstone soil that will be air-dried and be ground up, sieved
Earth is mixed with deionized water by the mass ratio of 1:2.5, is placed in 200rmin-1Room temperature shakes 2h on shaking table, and centrifugation keeps soil insoluble
After object precipitating, pH meter measures supernatant pH value, as soil pH value.As shown in Table 1, it compared with not connecing bacterium control group, connects at bacterium
0.43~0.45 unit of reason group pH decreased, compound bacteria processing feldspathic sandstone soil pH reduce 0.45 unit (P <
0.05)。
Influence of 1 composite bacteria agent of table to feldspathic sandstone soil pH
| Processing | Soil pH |
| CK | 8.21±0.05 |
| P75 | 7.78±0.12 |
| D10 | 7.77±0.06 |
| H3 | 7.76±0.05 |
| Composite bacteria agent | 7.76±0.03* |
Embodiment 3: the effect of composite bacteria agent raising feldspathic sandstone soil organic matter content
The P75 bacterium solution of embodiment 1, D10 bacterium solution, H3 bacterium solution and composite bacteria liquid are inoculated in by 5% inoculum concentration respectively
In feldspathic sandstone soil, using alfalfa as test plant, scientific management one month.Using potassium bichromate titrimetric method-heat of dilution
The content of method measurement soil with organic matter.As shown in Table 2, compared with not connecing bacterium control group, the feldspathic sandstone soil for connecing bacterium processing has
Machine matter content increases by 6%~38.46%, the feldspathic sandstone soil with organic matter content of compound bacteria processing dramatically increase 38.46% (P <
0.05)。
Influence of 2 composite bacteria agent of table to feldspathic sandstone soil organic matter content
| Processing | Content of organic matter g/kg |
| CK | 14.51±2.07 |
| P75 | 15.62±1.30 |
| D10 | 15.38±1.55 |
| H3 | 18.63±0.97 |
| Composite bacteria agent | 20.10±2.18* |
Embodiment 4: the effect of composite bacteria agent raising feldspathic sandstone soil nitrogen phosphorus potassium content
The P75 bacterium solution of embodiment 1, D10 bacterium solution, H3 bacterium solution and composite bacteria liquid are inoculated in by 5% inoculum concentration respectively
In feldspathic sandstone soil, using alfalfa as test plant, scientific management one month.Using 2molL-1KCl extraction-indophenols
The content of blue colorimetric method measurement soil ammonium-state, ultraviolet spectrophotometry measure the content of nitrified nitrogen in soil, 0.5mol/L
NaHCO3Method measures the content of rapid available phosphorus in soil, the content of flame spectrophotometric determination soil effective K.As shown in Table 3, with
It does not connect bacterium control group to compare, the feldspathic sandstone soil ammonium-state content amplification for connecing bacterium processing is -10%~160%, and nitrate nitrogen contains
Measuring amplification is 16.28%~55.60%, and available phosphorus contents amplification is 18.10%~76.32%, and quick-acting potassium content amplification is
24.56~72.04%, wherein the feldspathic sandstone soil ammonium-state content of compound bacteria processing dramatically increases 1.60 times, and nitrate nitrogen contains
Amount obviously increases 55.60%, and available phosphorus contents obviously increase 76.32%, quick-acting potassium content obviously increase 41.38% (P <
0.05)。
Influence of 3 composite bacteria agent of table to feldspathic sandstone soil nitrogen phosphorus potassium content
Embodiment 5: the effect of composite bacteria agent raising feldspathic sandstone soil enzyme activities
The P75 bacterium solution of embodiment 1, D10 bacterium solution, H3 bacterium solution and composite bacteria liquid are inoculated in by 5% inoculum concentration respectively
In feldspathic sandstone soil, using alfalfa as test plant, scientific management one month.Using 3,5- dinitrosalicylic acid (DNS)
Colorimetric method measures soil invertase activity, sodium phenate-sodium hypochlorite colorimetric method for determining urease activity.As shown in Table 4, with do not connect bacterium
Control group is compared, and the feldspathic sandstone soil invertase activity amplification for connecing bacterium processing is 22.55~36.77%, and urease activity amplification is
104%~148%, wherein the feldspathic sandstone soil invertase activity of compound bacteria processing obviously increases 22.55%, and urease activity is significant
Increase by 1.38 times (P < 0.05).
Influence of 4 composite bacteria agent of table to feldspathic sandstone soil enzyme activities
| Processing | Sucrase active mg/g | Urease activity mg/g |
| CK | 5.63±0.82 | 0.21±0.02 |
| P75 | 7.70±0.66 | 0.49±0.03 |
| D10 | 7.29±0.75 | 0.43±0.02 |
| H3 | 7.30±0.08 | 0.52±0.01 |
| Composite bacteria agent | 6.90±0.42* | 0.50±0.01* |
Embodiment 6: composite bacteria agent promotes the effect of Growth of Alfalfa in feldspathic sandstone soil
Soil is derived from the soil of Inner Mongol Stands in Arsenic Sandstone Area.Potted plant experiment is taken in test, each hole tray dress 300g soil, and to cave
Deionized water 30g is added in disk, and entire soil environment is made to keep certain degree of drought.By test plant rye grass and clover kind
After son carries out surface sterilization 20min with 5% liquor natrii hypochloritis respectively, seed is placed on sterile gauze, is placed on 30 DEG C
Sterile culture case, keep humidity 60%~80% carry out vernalization, picking is in the same size show money or valuables one carries unintentionally after seed be placed in water planting alms bowl
On cultivated.When seed bud grows to 2cm or so, growing way consistent alfalfa seedling uniform in size is selected into hole tray,
Be inoculated with embodiment 1 P75 bacterium solution, D10 bacterium solution, H3 bacterium solution and composite bacteria liquid 20ml, three repetitions of each processing, every two weeks again
Secondary inoculation 20ml bacterium solution intensive treatment, carries out one month by a definite date scientific management.Alfalfa dry weight is measured after one month, with
Growth-promoting effect of the judge composite bacteria agent to it.
As shown in Table 5, with do not connect bacterium control compared with, connect bacterium processing biomass of alfalfa amplification be 18.58%~
44.52%, treated that biomass of alfalfa dramatically increases 44.52% for compound bacteria.
Growth-promoting functions of 5 composite bacteria agent of table to alfalfa
| Processing | Alfalfa dry weight g/ basin |
| CK | 0.103±0.005 |
| P75 | 0.130±0.011 |
| D10 | 0.122±0.020 |
| H3 | 0.146±0.012 |
| Composite bacteria agent | 0.149±0.017* |
It should be noted that above-described embodiment is only presently preferred embodiments of the present invention, there is no for the purpose of limiting the invention
Protection scope, the equivalent substitution or substitution made on the basis of the above all belong to the scope of protection of the present invention.
Claims (9)
1. a kind of compound bacteria, which is characterized in that be made of bacillus P75, rhizobium D10, phosphate solubilizing bacteria H3:
The bacillus P75 classification naming is Bacillus halotolerans, is preserved in China typical culture collection
The heart, preservation address are Wuhan, China Wuhan University, and the deposit date is on October 17th, 2018, culture presevation number was CCTCC NO:M
2018690；
The rhizobium D10 classification naming is Sinorhizobium meliloti, is preserved in China typical culture collection
The heart, preservation address are Wuhan, China Wuhan University, and the deposit date is on October 27th, 2016, culture presevation number was CCTCC NO:M
2016554；
The phosphate solubilizing bacteria H3 classification naming is that Bacillus sp. is preserved in China typical culture collection center, and preservation address is
Wuhan, China Wuhan University, the deposit date is on October 27th, 2016, culture presevation number was CCTCC NO:M 2016551.
2. compound bacteria as described in claim 1, which is characterized in that in the compound bacteria bacillus P75, rhizobium D10 and
The ratio between biomass of phosphate solubilizing bacteria H3 is 4~6:0.5~1.5:4~6.
3. a kind of bacteria agent, which is characterized in that including compound bacteria as claimed in claim 1 or 2.
4. bacteria agent as claimed in claim 3, which is characterized in that the total number of cells of compound bacteria are 5 in the bacteria agent
Hundred million CFU/mL or more.
5. the preparation method of bacteria agent as claimed in claim 3, which comprises the following steps:
Bacillus P75 and phosphate solubilizing bacteria H3 slant strains are inoculated in LB solid medium and are cultivated；Then full list is chosen
Colony inoculation obtains P75 bacterium solution and H3 bacterium solution in LB liquid medium culture；
Rhizobium D10 slant strains are inoculated in TY solid medium and are cultivated, full, sticky single colonie inoculation is then chosen
In TY fluid nutrient medium culture, D10 bacterium solution is obtained；
By the P75 bacterium solution, H3 bacterium solution, D10 bacterium solution, 1:1:1 is mixed by volume, obtains composite bacteria liquid.
6. compound bacteria as claimed in claim 1 or 2 is in improvement feldspathic sandstone soil fertility or promotes answering in Growth of Alfalfa
With.
7. compound bacteria as claimed in claim 1 or 2 increases the feldspathic sandstone soil organism, increases feldspathic sandstone soil ammonium, increases
Add feldspathic sandstone soil nitrate-N, increases feldspathic sandstone soil quick-effective phosphor, increase feldspathic sandstone soil available nitrogen, improve feldspathic sandstone soil sugarcane
Application in carbohydrase or raising feldspathic sandstone soil urease liveness.
8. bacteria agent as described in claim 3 or 4 is in improvement feldspathic sandstone soil fertility or promotes in Growth of Alfalfa
Using.
9. bacteria agent as described in claim 3 or 4 is increasing the feldspathic sandstone soil organism, is increasing feldspathic sandstone soil ammonium state
Nitrogen increases feldspathic sandstone soil nitrate-N, increases feldspathic sandstone soil quick-effective phosphor, increase feldspathic sandstone soil available nitrogen, improve feldspathic sandstone
Application in soil saccharase or raising feldspathic sandstone soil urease liveness.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111919533A (en) * | 2020-07-09 | 2020-11-13 | 亿利资源集团有限公司 | Comprehensive improvement method for desertification land |
| CN112553100A (en) * | 2019-09-26 | 2021-03-26 | 有研工程技术研究院有限公司 | Composite microbial agent and method for soil fertility improvement and ecological restoration of heavy metal-containing field by using same |
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| CA2845408A1 (en) * | 2012-11-30 | 2014-05-30 | Manas Ranjan Banerjee | Phosphate solubilizing rhizobacteria bacillus firmus as biofertilizer to increase canola yield |
| CN105861420A (en) * | 2016-05-20 | 2016-08-17 | 广州赛莱拉干细胞科技股份有限公司 | Culture medium for inducing inner ear stem cells to be differentiated into hair cells and method of culture medium |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2845408A1 (en) * | 2012-11-30 | 2014-05-30 | Manas Ranjan Banerjee | Phosphate solubilizing rhizobacteria bacillus firmus as biofertilizer to increase canola yield |
| CN105861420A (en) * | 2016-05-20 | 2016-08-17 | 广州赛莱拉干细胞科技股份有限公司 | Culture medium for inducing inner ear stem cells to be differentiated into hair cells and method of culture medium |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112553100A (en) * | 2019-09-26 | 2021-03-26 | 有研工程技术研究院有限公司 | Composite microbial agent and method for soil fertility improvement and ecological restoration of heavy metal-containing field by using same |
| CN111919533A (en) * | 2020-07-09 | 2020-11-13 | 亿利资源集团有限公司 | Comprehensive improvement method for desertification land |
| CN111919533B (en) * | 2020-07-09 | 2022-12-02 | 亿利资源集团有限公司 | Comprehensive improvement method for desertification land |
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