A greenhouse gas emission reduction inhibitor and its application in livestock and poultry breeding litter

Abstract

The invention belongs to the field of microorganisms and discloses a greenhouse gas emission reduction inhibitor and its application in livestock and poultry breeding bedding. The greenhouse gas emission reduction inhibitor provided by the invention couples a methane oxidizing strain with a nitrous oxide inhibitor. The strain G-5 with methane oxidizing activity is Bacillus velezinoffe. Bacillus from Velez , G-5 can grow with methane as the only carbon source and oxidize methane; the chemical reagents with nitrous oxide inhibition are superphosphate, n-butylthiophosphoric triamide, and dicyandiamide, which can effectively reduce the loss of N in the breeding litter. The greenhouse gas emission reduction inhibitor of the present invention has a methane emission reduction rate of 34.18% and a nitrous oxide emission reduction rate of 37.26%, which can effectively reduce greenhouse gas emissions.

Description

Technical Field

[0001] The invention relates to a greenhouse gas emission reduction inhibitor and application thereof in livestock and poultry breeding bedding, belonging to the technical field of microorganisms. Background Art

[0002] With the rapid development of livestock and poultry farming in my country, the greenhouse gases generated in the farming process have also increased year by year. As the second largest greenhouse gas, CH4 contributes about 16% to the greenhouse effect. The N2O content in the atmosphere is lower than that of CO2 and CH4, but its warming potential is 298 times that of CO2. The large amount of CH4 and N2O emissions in the livestock and poultry farming process seriously restricts the green development of animal husbandry. However, there are currently no good treatment measures for greenhouse gases in the livestock and poultry farming industry. Generally, they are directly discharged into the air without treatment. Therefore, achieving carbon and nitrogen emission reduction in the livestock and poultry farming process is a necessary way for the development of animal husbandry. Therefore, the screening of microbial coupling nitrous oxide inhibitors that efficiently oxidize methane is applied to breeding litter to reduce the emission of methane and nitrous oxide from the source of livestock and poultry farming, providing a theoretical basis and technical support for green and low-carbon carbon and nitrogen emission reduction in animal husbandry. Summary of the invention

[0003] The problem to be solved by the present invention is to achieve carbon and nitrogen emission reduction in the process of livestock and poultry breeding, and to provide a group of greenhouse gas emission reduction inhibitors and their application in breeding bedding. A bacterial agent with methane oxidation ability is coupled with a nitrous oxide inhibitor. Adding it to breeding bedding can achieve carbon and nitrogen emission reduction in the process of livestock and poultry manure management.

[0004] The technical solution provided by the present invention is as follows:

[0005] A greenhouse gas emission reduction inhibitor, wherein the greenhouse gas emission reduction inhibitor is a bacterial agent containing Bacillus Velez and a nitrous oxide inhibitor coupled, wherein the Bacillus Velez was deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on January 22, 2024, with a deposit number of CGMCC NO. 29723, and is classified as Bacillus Velez Bacillus velezensis , the storage place is Beijing; the nitrous oxide inhibitors include superphosphate, n-butylthiophosphoric triamide and dicyandiamide.

[0006] Furthermore, the mass ratio of the nitrous oxide inhibitor to the bacterial agent containing Bacillus Velezii is 1:900-1500; the mass ratio of superphosphate, n-butylthiophosphoric triamide and dicyandiamide is 1:1:1.

[0007] Furthermore, the bacterial agent containing Bacillus Velez subtilis is a mixture of the above-mentioned Bacillus Velez subtilis and rice husks.

[0008] Furthermore, the preparation method of the bacterial agent containing Bacillus Velez subtilis includes: inoculating the above-mentioned Bacillus Velez subtilis subtilis into a culture medium, preparing a seed solution in a constant temperature shaking incubator; and mixing the cultured seed solution with rice husks.

[0009] Furthermore, the preparation method of the bacterial agent containing Bacillus Velezii comprises the following steps:

[0010] (1) Inoculate the G-5 bacterial solution into the culture medium at a 3% inoculum volume, and culture it in a constant temperature shaking incubator at a temperature of 28°C and a rotation speed of 180 r / min for 48 h to prepare the seed solution;

[0011] (2) The cultured seed solution was mixed with rice husks and the bacterial content reached 20×10 8 CFU / g.

[0012] Furthermore, the culture medium is: 10g of peptone, 5g of yeast powder, 10g of sodium chloride, 1000ml of distilled water, the pH is adjusted to neutral with sodium hydroxide, and the medium is sterilized at 121°C and autoclaved for 15min.

[0013] Furthermore, the culture conditions of the seed solution are a temperature of 25-40° C. and a rotation speed of 150-200 r / min.

[0014] Furthermore, the culturing time of the seed solution is 40 to 50 hours.

[0015] The present invention also provides application of the greenhouse gas emission reduction inhibitor in livestock and poultry breeding bedding.

[0016] Furthermore, the greenhouse gas emission reduction inhibitor is used to reduce greenhouse gas emissions in livestock and poultry farming.

[0017] Furthermore, the greenhouse gases include methane and nitrous oxide.

[0018] Furthermore, greenhouse gases generated in the management of livestock and poultry manure can be reduced by adjusting the thickness, moisture content and turning depth of the breeding litter.

[0019] Furthermore, the greenhouse gas emission reduction inhibitor is added to livestock and poultry breeding litter at an inoculation rate of 0.05-0.2% by mass.

[0020] Furthermore, the greenhouse gas emission reduction inhibitor is added to the breeding litter at an inoculation rate of 0.1% by mass.

[0021] Furthermore, the thickness of livestock and poultry breeding litter is 40 cm; the moisture content is 41%; and the turning depth is 24 cm. Beneficial Effects

[0022] The greenhouse gas emission reduction inhibitor prepared by the present invention can play a role in greenhouse gas emission reduction in aquaculture bedding, with a methane emission reduction rate of 34.18% and a nitrous oxide emission reduction rate of 37.26%, and has the characteristics of good emission reduction effect and no secondary pollution. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The Bacillus Velez Bacillus velezensis Photo of the G5 strain;

[0024] Figure 2 The Bacillus Velez Bacillus velezensis Phylogenetic tree of G5 strain;

[0025] Figure 3 is the effect of different litter culture treatments on methane emissions;

[0026] Figure 4 is the effect of different litter culture treatments on the cumulative methane emissions;

[0027] Figure 5 is the effect of different litter culture treatments on nitrous oxide emissions;

[0028] Figure 6 It is the effect of different litter culture treatments on the cumulative nitrous oxide emissions. DETAILED DESCRIPTION

[0029] The present invention is further described below. The following examples are only used to more clearly illustrate the technical solution of the present invention, and are not intended to limit the protection scope of the present invention.

[0030] Samples were collected from farms, sludge, and rice fields. The collected samples were screened, including continuous enrichment, separation and purification, and the strains were screened again, including methane oxidation capacity, adaptability screening under different temperatures, pH, and oligotrophic concentrations, to obtain methane-efficient oxidizing strain G-5. Its bacterial photos and phylogenetic tree are shown in the figure below. Figure 1 and Figure 2 shown.

[0031] Biomaterial sample storage:

[0032] G-5 has been deposited in the "China General Microbiological Culture Collection Management Center" in Beijing, China. The deposit address is: Culture Collection Center of the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. The deposit number is 29723 and the deposit date is January 22, 2024.

[0033] The 16S rRNA sequence is shown in SEQ ID No. 1:

[0034]

[0035] Example 1: Greenhouse gas emission reduction inhibitor effect verification test

[0036] (1) The validation experiment was conducted in a cowshed and divided into three groups: Group A+B, Group A, and Control. Each group used cows of similar growth stages and traits. The validation experiment lasted for 15 days.

[0037] (2) The experiment was designed with three different treatments, as follows: Group A+B: The inoculum containing Bacillus Velez subtilis was mixed with a nitrous oxide inhibitor and stirred evenly to obtain a greenhouse gas emission reduction inhibitor, which was then sprinkled on the breeding litter at an inoculation rate of 0.1% by mass (the greenhouse gas emission reduction inhibitor contained 2 billion bacteria / g and was added once every 5 days). The inoculum containing Bacillus Velez subtilis contained Bacillus Velez subtilis and rice husks; Group A: The inoculum containing Bacillus Velez subtilis was sprinkled on the breeding litter at an inoculation rate of 0.1% by mass (the inoculum containing Bacillus Velez subtilis contained 2 billion bacteria / g and was added once every 5 days. No nitrous oxide inhibitor was added to this group); Control group (CK): The same breeding litter was laid in the pens.

[0038] Preparation method of litter: The main components of the litter are various organic matter such as straw powder, sawdust, mushroom residue, feces residue, and sludge. After evenly mixing, the bulk density is ≤0.45 and the moisture content is ≤60%. High-temperature composting and fermentation are carried out for 7-10 days. When the moisture content drops below 45% and meets the requirements for harmless treatment, it can be spread out to cool and set aside.

[0039] (3) The static box-gas chromatograph method was used to collect and analyze the methane produced in the sty. Gas was collected on the 0th, 3rd, 6th, 9th, 12th, and 15th day of the experiment. Each collection time was 7:00-10:00 am. The gas at each point was collected at 0, 5, 10, and 15 minutes, respectively. After being placed in a 0.5L aluminum foil gas sampling bag, it was measured using a gas chromatograph.

[0040] The result is as follows:

[0041] Depend on Figure 3 As shown in Figure 2, the methane emission fluxes of the three treatments showed a sharp downward trend in the first 9 days of the experiment, which was related to the sharp drop in the external ambient temperature. The methane emission flux of the CK group increased from an initial 2.06 mg·m -2 ·h -1 The lowest value was 0.14 mg·m -2 ·h -1 , Group A from 1.58 mg·m -2 ·h -1 Decreased to 0.56 mg·m -2 ·h -1, A+B group from 1.71mg·m -2 ·h -1 The minimum value was 0.08 mg·m -2 ·h -1 After the 9th day, the methane emission fluxes of the three treatments fluctuated slightly until the end of the experiment. During the experiment, the methane emission fluxes of group A and group A+B were significantly lower than that of CK.

[0042] Depend on Figure 4 It can be seen that the cumulative methane emissions under the three treatments were quite different. The cumulative methane emission in the CK group reached 1933 mg·m -2 The cumulative methane emissions of Group A are 1584 mg·m -2 The cumulative methane emissions of group A+B are 1273 mg·m -2 , compared with the CK group, the methane emission reduction was 18.05% and 34.18% respectively. The test results show that under the same environmental conditions, the methane emission flux of the test group with the addition of methane oxidizing bacteria and greenhouse gas emission reduction inhibitors was significantly lower than that of the control group, and the combined use of bacteria and nitrous oxide chemical inhibitors was more effective than the use of bacteria alone.

[0043] Depend on Figure 5 It can be seen that the nitrous oxide emission fluxes under the three treatments showed a trend of first decreasing and then increasing sharply. This was due to the decrease in ambient temperature in the early stage of the experiment. The nitrous oxide emission fluxes under the three treatments also decreased during the period from 0 d to 9 d. On the 9th day, the nitrous oxide emission fluxes of the three treatments all dropped to the minimum value, which was 0.06 mg·m in the CK group. -2 ·h -1 , Group A 0.09 mg·m -2 ·h -1 , A+B group: 0.04 mg·m -2 ·h -1 In the later stage of the experiment, the rise in ambient temperature caused a sharp increase in the nitrous oxide emission flux. On the 15th day, the nitrous oxide emission flux of the CK group was 0.42 mg·m -2 ·h -1 , group A: 0.13 mg·m -2 ·h -1 , group A+B: 0.31 mg·m -2 ·h -1 During the experiment, the nitrous oxide emission fluxes of group A and group A+B were significantly lower than that of CK.

[0044] Depend on Figure 6 It can be seen that the cumulative emission of nitrous oxide in the test group was significantly lower than that in the control group. The cumulative emission of nitrous oxide in group A for 15 days was 283 mg·m -2The cumulative nitrous oxide emissions of group A+B for 15 days were 263 mg·m -2 The cumulative nitrous oxide emission of the CK group was 420 mg·m -2 , and the nitrous oxide emission reduction was 32.61% and 37.26% respectively. The test results show that under the same environmental conditions, the nitrous oxide emission flux of the test group with the addition of methane oxidizing bacteria and greenhouse gas emission reduction inhibitors was significantly lower than that of the control group, and the combined use of bacteria and nitrous oxide chemical inhibitors was more effective than the use of bacteria alone.

[0045] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the technical principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A greenhouse gas emission reduction inhibitor, characterized in that: The greenhouse gas emission reduction inhibitor is a bacterial agent containing Bacillus Velezii coupled with a nitrous oxide inhibitor. The Bacillus Velezii was deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on January 22, 2024, with a deposit number of CGMCC NO. 29723 and a classification name of Bacillus Velezii. Bacillus velezensis , the storage place is Beijing; the nitrous oxide inhibitor includes superphosphate, n-butylthiophosphoric triamide and dicyandiamide; the greenhouse gases are methane and nitrous oxide.

2. The greenhouse gas emission reduction inhibitor according to claim 1, characterized in that: The mass ratio of the nitrous oxide inhibitor to the bacterial agent containing Bacillus Velezii is 1:900-1500; the mass ratio of superphosphate, n-butylthiophosphoric triamide and dicyandiamide is 1:1:

1.

3. The greenhouse gas emission reduction inhibitor according to claim 1, characterized in that: The bacterial agent containing Bacillus Velez is a mixture of the Bacillus Velez and rice husks as claimed in claim 1.

4. The greenhouse gas emission reduction inhibitor according to claim 3, characterized in that: The preparation method of the bacterial agent containing Bacillus Velezii comprises: inoculating the Bacillus Velezii described in claim 1 into a culture medium, preparing a seed solution in a constant temperature shaking incubator; and mixing the cultured seed solution with rice husks.

5. The greenhouse gas emission reduction inhibitor according to claim 4, characterized in that: The culture conditions of the seed solution are a temperature of 25-40° C. and a rotation speed of 150-200 r / min.

6. The greenhouse gas emission reduction inhibitor according to claim 4, characterized in that: The culturing time of the seed solution is 40 to 50 hours.

7. Use of the greenhouse gas emission reduction inhibitor according to claim 1 in livestock and poultry breeding bedding.

8. The use according to claim 7, characterized in that: The greenhouse gas emission reduction inhibitor is used for reducing greenhouse gas emissions in livestock and poultry farming.

9. The use according to claim 7, characterized in that: The greenhouse gas emission reduction inhibitor is added to livestock and poultry breeding litter at an inoculation rate of 0.05-0.2% by mass.

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