CN110041139B - Biological bacterium multilayer tape agent for continuous cropping soil remediation and use method - Google Patents
Biological bacterium multilayer tape agent for continuous cropping soil remediation and use method Download PDFInfo
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Abstract
The invention relates to a biological bacterium multilayer tape agent for continuous cropping soil restoration and a using method thereof, wherein the biological bacterium multilayer tape agent for the continuous cropping soil restoration is sequentially provided with a degradable matrix layer, an adhesive layer, a carrier and a biological bacterium mixing layer from bottom to top; the degradable substrate layer is degradable paper, the adhesive layer is a starch adhesive layer with trace elements, and the carrier and biological bacteria mixing layer comprises the following raw material components in parts by weight, 90-100 parts of rice hull carbon and 0-10 parts of plant ash; the biological bacteria are pseudomonas stutzeri (A)Pseudomonas oryzihabitans) The using method comprises two methods of continuous cropping soil restoration and hole application before cultivation, the multi-layer biological bacterium tape agent can simplify the application operation steps, enlarge the restoration area of the pseudomonas oryzae habitat on the continuous cropping soil, prolong the validity period of the pseudomonas oryzae habitat, and is easy to store.
Description
Technical Field
The invention belongs to the field of biological fertilizer and soil remediation, and particularly relates to a biological bacterium multilayer tape agent for continuous cropping soil remediation and a using method thereof.
Background
Facility agriculture is one of the basic agricultural production modes in China at present, and plays an important role in ensuring that people meet life needs under limited land resources. However, in the facility agriculture, due to the fact that a certain variety is planted in continuous cropping, the physical and chemical properties of soil are reduced, pathogenic microorganisms in the soil are propagated in a large quantity, autotoxic substances in root exudates of crops are accumulated, the normal growth of the crops is further influenced, and the continuous cropping obstacle symptom is obvious.
Soil remediation is one of effective methods for solving continuous cropping obstacles at present, and chemical agents and biological bacteria are mostly adopted for treating continuous cropping obstacles at present. When the greenhouse soil of cucumbers and melon crops is fumigated by chemical agents such as dazomet, the incidence rate of blight of the afterreap cucumbers or melon crops is obviously reduced, but the chemical agents do not degrade phenolic acid substances in the soil and aggravate the deterioration of the physicochemical property of the soil. The application of biological bacteria to treatment of facility continuous cropping soil is also greatly developed, but the biological bacteria has higher requirement on the surrounding environment and lower utilization rate, and particularly the prior application method can not ensure that the biological bacteria can not keep activity for a long time, so that the biological bacteria can not be widely applied to the actual production of repairing the continuous cropping soil.
Pseudomonas oryzae (III)Pseudomonas oryzihabitans) Is a crop rhizosphere autotoxic substance degrading bacterium, can reduce the content of the autotoxic substance in soil and improve the growth of crops by planting in the soil. Patent No. 201810303948.2 discloses the action and application method of the bacterium, the bacterium agent is prepared, and hole application and root irrigation application are adopted in the transplanting period of melon crops, and the method has the beneficial effect that the root irrigation of the bacterium solution can effectively repair the root soil of the transplanted crop plants. However, the optimal use time of the bacteria after fermentation is 5-7 days, the early stage work of the bacteria liquid preparation must be done before the field planting, the root irrigation operation is complicated in the field planting process, and the application area is limited, namely the preparation and use methods of the microbial inoculum limit the application of the pseudomonas oryzae to the continuous cropping soil remediation.
Disclosure of Invention
In order to make up for the defects of the prior art, the invention provides the biological bacterium multilayer tape agent for repairing the continuous cropping soil and the use method thereof, which can simplify the application operation steps, enlarge the repairing area of the pseudomonas oryzae benthic for the continuous cropping soil, improve the degradation rate of the soil phenolic acid, prolong the effective period of the pseudomonas oryzae benthic, and are easy to store.
The invention aims to solve the problems by the following technical schemeThe biological bacterium multilayer carrying agent for continuous cropping soil remediation and the using method thereof are realized, and the technical key points are as follows: the biological bacterium multilayer belt agent for repairing continuous cropping soil is sequentially provided with a degradable matrix layer, an adhesive layer, a carrier and a biological bacterium mixing layer from bottom to top; the degradable substrate layer is degradable paper, the adhesive layer is a starch adhesive layer with trace elements, and the carrier and biological bacteria mixing layer comprises the following raw material components in parts by weight, 90-100 parts of rice hull carbon and 0-10 parts of plant ash; the biological bacteria are pseudomonas stutzeri (A)Pseudomonas oryzihabitans) And the preservation unit: china general microbiological culture Collection center, having the address of No. 3 Xilu No. 1 of the Kyoho district, Beijing, China academy of sciences microbial research institute, with the preservation number of CGMCC number 15325 and the preservation date of 2018, 29 months, are classified and named as Pseudomonas oryzae (Pseudomonas sp.) (Pseudomonas oryzihabitans）；
The preparation method of the biological bacterium multilayer tape agent for repairing continuous cropping soil comprises the following steps:
(1) preparation of starch adhesive with trace elements
The starch adhesive with trace elements comprises the following raw material components in percentage by weight: 15-30 parts of starch, 60-85 parts of cold water, 0.5-1.5 parts of hydrogen peroxide (30%), 0.3-0.5 part of ferrous sulfate, 0.01-0.015 part of magnesium sulfate, 0.5-1.5 part of sodium hydroxide, 1-1.5 parts of calcium sulfate, 0.04-0.06 part of manganese sulfate, 0.04-0.05 part of zinc sulfate and 0.3-0.5 part of sodium tetraborate;
the starch is any one of corn starch, potato starch and sweet potato starch;
the preparation method of the starch adhesive comprises the following steps:
preparing hydrogen peroxide, ferrous sulfate, magnesium sulfate, sodium hydroxide, calcium sulfate, manganese sulfate, zinc sulfate and sodium tetraborate solution;
cold water and starch are fully and uniformly stirred to form starch solution;
③ in sequence: sequentially pouring hydrogen peroxide solution, sodium hydroxide solution, ferrous sulfate, magnesium sulfate, calcium sulfate, manganese sulfate and zinc sulfate into the starch solution, and fully stirring;
fourthly, the solution obtained from the third step is heated to 40 to 60 ℃ by a heating interlayer water-soluble pot, and the heating is stopped;
after gelatinization reaction for 20-30 minutes, adding sodium tetraborate solution to obtain starch adhesive;
(2) glue spreading
Spreading degradable paper, and coating with microelement starch adhesive to form an adhesive layer;
(3) making the carrier layer
Pulverizing rice hull carbon to particle size of 0.5-1.5mm, mixing with plant ash according to weight ratio, uniformly spreading on the glue layer, pressing the carrier layer on the glue layer with roller or pressing plate, and drying under aseptic condition;
(4) preparation of biological agent
The biological agent is fermentation liquor of pseudomonas oryzae, the preparation method of the fermentation liquor of the pseudomonas oryzae comprises the steps of 10g/L tryptone, 5g/L yeast extract and 10g/L sodium chloride (NaCl), wherein the inoculation amount of agar is 2 percent of the weight of a culture medium, the fermentation temperature is 28 ℃, the stirring speed is 120rpm, the fermentation time is 5 days, viscous bacterial suspension is obtained, and the concentration of the bacterial suspension is adjusted to be 5.0-8.0 multiplied by 10 by using the fermentation liquor culture medium8cfu/ml；
(5) Load biological bacteria
And (3) uniformly spraying the bacterial suspension on the carrier layer in the step (3), covering with a film, and hermetically placing the multilayer tape agent loaded with the biological bacteria at the temperature of 25-28 ℃ for 10-180 days.
The invention also provides a using method of the biological bacterium multilayer carrying agent for repairing continuous cropping soil, which comprises the following steps:
(1) and (3) continuous cropping soil remediation before cultivation: during land preparation before crop cultivation, burying the biological bacterium multilayer zone agent for repairing continuous cropping soil 20-30cm below soil surface after ploughing, controlling soil humidity at 40-65%, and allowing the treatment period to be 40-90 d, and then performing crop cultivation normally;
(2) hole application: when crops are transplanted, multilayer tape agents matched with the area of a transplanting hole are cut, the hole bottom is placed before seedlings are transplanted, the distance between the multilayer tape agents and the roots is ensured to be 2-5cm, and then the transplanting is normally carried out.
The invention has the beneficial effects that:
(1) the effective period of the activity of the pseudomonas oryzae is prolonged, the effective period of the active bacteria in the fermentation liquor of the pseudomonas oryzae in the prior art is relatively short, and the inactivation is caused after long-term storage, so the preparation is required before planting crops, and the requirements on the preparation and the use time of the microbial inoculum are high. The multi-layer tape agent of the invention loads the pseudomonas oryzae to provide nutrition required for propagation, and can prolong the effective period of the pseudomonas oryzae.
(2) The use operation is simplified, the use method of the multilayer tape agent comprises direct soil penetration (continuous cropping soil restoration before cultivation) and field planting soil penetration (hole application), and the multilayer tape agent is decomposed after direct soil penetration, so that the soil nutrient is improved, and the phenolic acid substance of the continuous cropping soil is reduced; the damage of phenolic acid substances in the continuous cropping soil to the root system can be reduced by planting and burying, and the soil microenvironment of the root system of the transplanted crop can be improved, so that the survival rate of the transplanted crop can be improved.
Drawings
FIG. 1 is a schematic structural diagram of the multi-layer biological bacterium carrying agent for repairing continuous cropping soil.
The specific names of the structures in the figure are: the degradable substrate layer 1, the glue layer 2, the carrier and biological bacteria mixing layer 3.
Detailed Description
A multi-layer biological bacterium carrying agent for repairing continuous cropping soil and a using method thereof are provided, wherein the multi-layer biological bacterium carrying agent for repairing continuous cropping soil is sequentially provided with a degradable matrix layer, an adhesive layer, a carrier and a biological bacterium mixing layer from bottom to top; the degradable substrate layer is degradable paper, the adhesive layer is a starch adhesive layer with trace elements, and the carrier and biological bacteria mixing layer comprises the following raw material components in parts by weight, namely rice hull carbon 90 and plant ash 10; the biological bacteria are Pseudomonas oryzae (Pseudomonas oryzihabitans), and the preservation unit comprises: the general microbiological center of China Committee for culture Collection of microorganisms, the address is No. 3 of Xilu No. 1 of Beijing, Chaoyang, and the microbiological research institute of China academy of sciences, the collection number is CGMCC No.15325 and the collection date of 2018, 29.01.8, and is classified and named as Pseudomonas oryzae (A)Pseudomonas oryzihabitans）；
The preparation method of the biological bacterium multilayer tape agent for repairing continuous cropping soil comprises the following steps:
(1) preparation of starch adhesive with trace elements
The starch adhesive with trace elements comprises the following raw material components in percentage by weight: 30 parts of corn starch, 80 parts of cold water, 1.3 parts of hydrogen peroxide solution (30%), 0.3 part of ferrous sulfate, 0.012 part of magnesium sulfate, 1 part of sodium hydroxide, 1.2 parts of calcium sulfate, 0.05 part of manganese sulfate, 0.05 part of zinc sulfate and 0.5 part of sodium tetraborate;
the preparation method of the starch adhesive comprises the following steps:
preparing ferrous sulfate, magnesium sulfate, sodium hydroxide, calcium sulfate, manganese sulfate, zinc sulfate and sodium tetraborate solution;
cold water and starch are fully and uniformly stirred to form starch solution;
③ in sequence: sequentially pouring hydrogen peroxide solution, sodium hydroxide solution, ferrous sulfate, magnesium sulfate, calcium sulfate, manganese sulfate and zinc sulfate into the starch solution, and fully stirring;
fourthly, the solution obtained from the third step is heated to 40 to 60 ℃ by a heating interlayer water-soluble pot, and the heating is stopped;
after gelatinization reaction for 20-30 minutes, adding sodium tetraborate solution to complete the preparation of the starch adhesive;
(2) glue spreading
Spreading degradable paper, and coating with microelement starch adhesive to form an adhesive layer;
(3) making the carrier layer
Pulverizing rice hull carbon to particle size of 0.5-1.5mm, mixing with plant ash according to weight ratio, uniformly spreading on the glue layer, pressing the carrier layer on the glue layer with roller or pressing plate, and drying on aseptic operation table;
(4) preparation of biological agent
The biological agent is fermentation liquor of pseudomonas oryzae and the preparation method of the fermentation liquor of the pseudomonas oryzae is trypsin10g/L peptone, 5g/L yeast extract, 10g/L sodium chloride (NaCl), 4-8 g/L agar inoculum size of 2% of culture medium weight, fermenting at 28 deg.C under stirring speed of 120rpm for 5 days to obtain viscous bacterial suspension, and adjusting the concentration of the bacterial suspension to 7.5 × 10 using the above fermentation liquid culture medium8cfu/ml；
(5) Load biological bacteria
And (3) uniformly spraying the bacterial suspension on the carrier layer in the step (3), covering with a film, and sealing and placing the multilayer tape agent loaded with the biological bacteria at the temperature of 25-28 ℃ for 10 days to obtain the application.
The field test of the repairing effect of the biological bacterium multilayer tape agent for repairing the continuous cropping soil comprises the following steps:
(1) test time 3 in mid-ten years in 2018 to 5 months in 2018.
(2) The test site is a muskmelon greenhouse in field test base of Shenyang agricultural university, wherein the muskmelon is continuously cultivated for 3 years, and the muskmelon is a crop on the upper crop.
(3) Test method
The method for testing the influence of the multi-layer biological bacterium carrying agent for repairing the continuous cropping soil on the survival rate of continuous cropping plants comprises the following steps: treatment 1: and (3) continuous cropping soil remediation before cultivation: during soil preparation before crop cultivation, burying the biological bacterium multilayer tape agent for repairing continuous cropping soil 20cm below the soil surface of a previous cropping planting platform, controlling the soil humidity at 50%, controlling the treatment period to 45d, and normally transplanting melon seedlings in the early 5 months; and (3) treatment 2: hole application: when melon seedlings are transplanted in the beginning of 5 months, shearing multiple layers of tape agents with the same area of transplanting holes, placing the tape agents at the bottoms of the holes before transplanting the seedlings, ensuring the distance between the multiple layers of tape agents and roots to be 2-5cm, and then normally transplanting; and (3) treatment: let's not control the above treatment.
② a method for detecting the degradation of phenolic acid substances in soil by using multilayer biological bacterium carrying agents for repairing continuous cropping soil, which comprises the following steps:
25 g of rhizosphere soil of the plants treated 1 and 2 was collected, the soil to be tested was placed in a 75 mL centrifuge tube, and 25 mL of 1 mol. L was added to the soil-1Mixing the NaOH solution with a vortex oscillator, standing for 24 h, mixing with a turnover shaking apparatus for 1 h the next day, oscillating with an ultrasonic oscillator for 30 min,centrifuging, separating supernatant with 12 mol/L-1Adjusting the pH value of the soil solution to 2.5 by hydrochloric acid, standing for 2 h, centrifuging to remove humic acid, separating supernatant, storing at 4 ℃, and preparing for analysis and determination.
The HPLC separation and identification method, HPLC chromatographic condition, and determination of different phenolic acids in soil are described in patent No. 201810303948.2, middle section 0030-0037, and data processing and statistical analysis method are described in the specification.
(4) And 6, surveying the survival rate of transplanted melon plants in the middle ten days of the month.
The plant survival rate calculation method comprises the following steps: plant survival = total number of surviving plants/total number investigated × 100%
The results of the influence of the biological bacterium multilayer tape agent for repairing the continuous cropping soil on the survival rate of continuous cropping crop plants are shown in table 1, and the results show that the survival rate of melons treated by the biological bacterium multilayer tape agent is obviously improved compared with a control.
TABLE 1 Effect of the Multi-layer agent of biological bacteria for repairing continuous cropping soil on the survival Rate of plants of continuous cropping crops
| Treatment of | Survival rate of |
| Process | |
| 1 | 88.24 |
| Treatment | |
| 2 | 92.25% |
| Control | 52.10% |
(5) Degradation effect result of biological bacterium multilayer belt agent for repairing continuous cropping soil on phenolic acid substances in soil
The result of the degradation effect of the biological bacterium multilayer belt agent for repairing the continuous cropping soil on phenolic acid substances in the melon continuous cropping soil shows that both the two using methods have excellent degradation effect on main phenolic acid substances in melon secretions (see table 1), wherein the degradation effect on p-hydroxybenzoic acid, vanillin, coumaric acid, benzoic acid and cinnamic acid is more than 50%, and the excellent degradation effect is shown.
As can be seen from the degradation effect, the degradation rate of the tested phenolic acid is equivalent to that of the phenolic acid treated by the treatment 1 and the treatment 2, and the effect of the multi-layer carrying agent of the treatment 1 is 45 days earlier than that of the treatment 2, but the effect is equivalent to that of the treatment 2, which shows that the multi-layer carrying agent has long effective period and is suitable for continuous cropping soil treatment.
TABLE 1 degradation of phenolic acid substances in melon continuous cropping soil by microbial inoculum
| Autotoxic substances in soil | Degradation rate of |
Degradation rate of treatment 2 |
| P-hydroxybenzoic acid | 51.25% | 55.31% |
| Vanillin | 62.10% | 65.18% |
| Syringic acid | 45.38% | 44.22% |
| Vanillin | 49.07% | 51.37% |
| Coumaric acid | 55.89% | 54.16% |
| Dibutyl phthalate | 42.68% | 48.71% |
| Benzoic acid | 58.64% | 60.28% |
| Cinnamic acid | 71.12% | 72.52% |
The invention principle is as follows:
the invention provides a method for degrading soil phenolic acid by using Pseudomonas oryzae, which comprises the steps of setting a three-layer carrier layer of degradable paper, a starch adhesive, rice hull carbon and plant ash mixture as a substrate of Pseudomonas oryzae (pseudomonias oryzihibitanes) fermentation liquor, covering the sticky Pseudomonas oryzae fermentation liquor by a film, gradually permeating the sticky Pseudomonas oryzae fermentation liquor into the rice hull carbon, forming aggregates in pores of the rice hull carbon, adsorbing and loading the aggregates in the rice hull carbon, wherein the rice hull carbon is carbonized rice hulls, and the porosity and nutrition of the carbonized rice hull carbon are suitable for growth and propagation of the Pseudomonas oryzae.
After the multi-layer biological bacterium tape agent enters soil, the degradable paper, the starch adhesive and the rice husk charcoal delay the release speed of the rice-inhabiting pseudomonas in the soil, reduce the loss after release, starch granules and the degradable paper in the adhesive layer are eroded and decomposed by microorganisms in the soil, the starch adhesive layer and the degradable paper are decomposed in the soil to provide nutrients for the rice-inhabiting pseudomonas to promote the propagation of the rice-inhabiting pseudomonas, and trace elements in the starch adhesive layer are gradually dissolved in the soil to improve the nutrients of the rhizosphere soil of crops. The degradable paper adopted by the invention can be purchased from the market, and can also be prepared by the preparation method in the patent number 201710138527.4 or 201510744895.4.
Claims (1)
1. A biological bacterium multilayer tape agent for continuous cropping soil remediation is characterized in that: the biological bacterium multilayer belt agent for repairing continuous cropping soil is sequentially provided with a degradable matrix layer, an adhesive layer, a carrier and a biological bacterium mixing layer from bottom to top; the degradable substrate layer is degradable paper, the adhesive layer is a starch adhesive layer with trace elements, and the carrier and biological bacteria mixing layer comprises the following raw material components in parts by weight, 90-100 parts of rice hull carbon and 0-10 parts of plant ash; the biological bacteria are pseudomonas stutzeri (A)Pseudomonas oryzihabitans) And the preservation unit: china general microbiological culture Collection center, having the address of No. 3 Xilu No. 1 of the Kyoho district, Beijing, China academy of sciences microbial research institute, with the preservation number of CGMCC number 15325 and the preservation date of 2018, 29 months, are classified and named as Pseudomonas oryzae (Pseudomonas sp.) (Pseudomonas oryzihabitans）；
The preparation method of the biological bacterium multilayer tape agent for repairing continuous cropping soil comprises the following steps:
(1) preparation of starch adhesive with trace elements
The starch adhesive with trace elements comprises the following raw material components in percentage by weight: 15-30 parts of starch, 60-85 parts of cold water, 0.5-1.5 parts of hydrogen peroxide (30%), 0.3-0.5 part of ferrous sulfate, 0.01-0.015 part of magnesium sulfate, 0.5-1.5 part of sodium hydroxide, 1-1.5 parts of calcium sulfate, 0.04-0.06 part of manganese sulfate, 0.04-0.05 part of zinc sulfate and 0.3-0.5 part of sodium tetraborate;
the starch is any one of corn starch, potato starch and sweet potato starch;
the preparation method of the starch adhesive comprises the following steps:
preparing hydrogen peroxide, ferrous sulfate, magnesium sulfate, sodium hydroxide, calcium sulfate, manganese sulfate, zinc sulfate and sodium tetraborate solution;
cold water and starch are fully and uniformly stirred to form starch solution;
③ in sequence: sequentially pouring hydrogen peroxide solution, sodium hydroxide solution, ferrous sulfate, magnesium sulfate, calcium sulfate, manganese sulfate and zinc sulfate into the starch solution, and fully stirring;
fourthly, heating the solution obtained from the third step to 40-60 ℃ by using a heating interlayer water-soluble pot, and stopping heating;
after gelatinization reaction for 20-30 minutes, adding sodium tetraborate solution to obtain starch adhesive;
(2) glue spreading
Spreading degradable paper, and coating with microelement starch adhesive to form an adhesive layer;
(3) making the carrier layer
Pulverizing rice hull carbon to particle size of 0.5-1.5mm, mixing with plant ash according to weight ratio, uniformly spreading on the glue layer, pressing the carrier layer on the glue layer with roller or pressing plate, and drying under aseptic condition;
(4) preparation of biological agent
The biological agent is fermentation liquor of pseudomonas oryzae, the preparation method of the fermentation liquor of the pseudomonas oryzae comprises the steps of 10g/L tryptone, 5g/L yeast extract and 10g/L sodium chloride (NaCl), wherein the inoculation amount of agar is 2 percent of the weight of a culture medium, the fermentation temperature is 28 ℃, the stirring speed is 120rpm, the fermentation time is 5 days, viscous bacterial suspension is obtained, and the concentration of the bacterial suspension is adjusted to be 5.0-8.0 multiplied by 10 by using the fermentation liquor culture medium8cfu/ml；
(5) Load biological bacteria
Uniformly spraying the bacterial suspension on the carrier layer in the step (3), covering the surface with a film, and hermetically placing the multilayer tape agent loaded with the biological bacteria at the temperature of 25-28 ℃ for 10-180 days;
the biological bacterium multilayer tape agent is used for repairing continuous cropping soil of melons, and the using method comprises the following steps of repairing the continuous cropping soil before cultivation or performing hole application:
and (3) repairing continuous cropping soil before tillage: during land preparation before crop cultivation, burying the biological bacterium multilayer zone agent for repairing continuous cropping soil 20-30cm below soil surface after ploughing, controlling soil humidity at 40-65% and treating period of 40-90 d, and then normally performing crop cultivation;
the hole application: when crops are transplanted, multilayer tape agents matched with the area of a transplanting hole are cut, the bottom of the hole is placed before seedlings are transplanted, the distance between the multilayer tape agents and the roots is guaranteed to be 2-5cm, and then transplanting is normally carried out.
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| CN201910461448.6A CN110041139B (en) | 2019-05-30 | 2019-05-30 | Biological bacterium multilayer tape agent for continuous cropping soil remediation and use method |
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| CN201910461448.6A CN110041139B (en) | 2019-05-30 | 2019-05-30 | Biological bacterium multilayer tape agent for continuous cropping soil remediation and use method |
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| CN110041139A CN110041139A (en) | 2019-07-23 |
| CN110041139B true CN110041139B (en) | 2021-11-30 |
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