Paracoccus pantotrophicus, deodorant composed of paracoccus pantotrophicus and application of paracoccus pantotrophicus

Abstract

The invention provides paracoccus pantotrophus, a deodorant composed of the paracoccus pantotrophus and application of the deodorant, and belongs to the technical field of microbial agents. According to the invention, a paracoccus pantotrophus strain is screened from biogas residues of a pig farm, is named as Paracoccus pantotrophus SZZ1 and is preserved in the China Center for Type Culture Collection, the preservation address is Wuhan University, Wuhan, China, and the biological preservation number is CCTCC M20242509. The SZZ1 strain is used for animal manure fermentation, and results show that the microbial inoculum can effectively reduce odor generated during manure fermentation, and especially has a significant inhibition effect on release of hydrogen sulfide. Therefore, the method not only enriches the types of existing deodorization strains, but also has an excellent removal effect on hydrogen sulfide and a stable and lasting deodorization effect, and is suitable for popularization and application.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbial agents, and specifically relates to a Paracoccus pantotrophus, a deodorant composed of the Paracoccus pantotrophus, and its application. Background Art

[0002] With the rapid development of the livestock and poultry breeding industry, the impact of livestock and poultry manure and the generated odor on the surrounding environment has become increasingly serious, which has affected the quality of life and physical and mental health of the surrounding people, and has also attracted great attention from the government. Manure, feed residues, etc. in livestock and poultry farms will all cause odors. The odor has a certain stimulating effect on the gustatory nerves of livestock, which can slow down the growth rate of livestock. In a relatively smelly environment for a long time, the physical quality of animals will decline, the immunity will decrease, and the output of livestock and poultry products will also decrease. As the people's requirements for the ecological environment are increasing day by day, the proportion of breeding odor complaints in the national odor complaints is gradually increasing, and it has become an urgent problem to be solved.

[0003] In order to effectively control the malodorous gases generated from composting, a variety of treatment technologies have been developed at home and abroad. Currently, the commonly used deodorization technologies mainly include biological methods and chemical methods, supplemented by physical methods. The deodorization costs of physical and chemical methods are high, and there are also potential risks of secondary pollution. Biological deodorization mainly uses microorganisms to decompose malodorous substances, including in-situ addition of microbial agents and ex-situ bioreactor treatment (such as biofilters, biotrickling filters, and bioscrubbers). It usually has the advantages of high efficiency, no secondary pollution, simple operation, and low cost, and is an effective method for removing odors. However, biological deodorization is extremely dependent on suitable strains, and has relatively high requirements for the specificity, diversity, heat resistance, adaptability, and deodorization efficiency of the strains.

[0004] The currently discovered types of deodorizing strains mainly include Bacillus subtilis, Bacillus licheniformis, Alcaligenes faecalis, Pseudomonas, etc. The types are few, and antagonistic effects may occur when used in combination, affecting their deodorization effect.

[0005] Currently, regarding Paracoccus pantotrophus, there are relevant reports mentioning its corresponding microbial agents, but mainly concentrated in denitrification and dephosphorization, and are mostly used in fields such as sewage and wastewater treatment. Currently, there is no report on using Paracoccus pantotrophus for deodorization. New deodorizing agents, as well as microbial agents with stable and lasting deodorization effects, remain to be further developed. Summary of the Invention

[0006] The present invention is to solve the above technical problems, and thus provides a Paracoccus pantotrophus, a deodorant composed of the Paracoccus pantotrophus, and its applications. The present invention screens a strain of Paracoccus pantotrophus (Paracoccus pantotrophus SZZ1) from pig farm biogas residues, which has a good deodorizing effect. When this strain of Paracoccus pantotrophus is used in the fermentation of animal feces, it can effectively reduce the odor generated by feces fermentation, especially significantly inhibit the release of hydrogen sulfide, thus broadening the types of microbial deodorant agents and having good application prospects.

[0007] The present invention first provides a strain of Paracoccus pantotrophus, which is named Paracoccus pantotrophus SZZ1, and the preservation unit is: China Center for Type Culture Collection, the preservation address is: Wuhan University, Wuhan, China, the preservation center number is: CCTCC No. M 20242509, and the preservation time is: November 11, 2024.

[0008] Furthermore, the gene sequence of this Paracoccus pantotrophus is as shown in SEQ NO.1.

[0009] The present invention also provides a biocontrol agent, and the biocontrol agent contains the Paracoccus pantotrophus as described above.

[0010] The present invention also provides a deodorant, and the deodorant contains the Paracoccus pantotrophus as described above.

[0011] The present invention also provides the application of the Paracoccus pantotrophus as described above in deodorization.

[0012] The present invention also provides the application of the Paracoccus pantotrophus as described above in the preparation of a deodorant.

[0013] Specifically, it is the application of this Paracoccus pantotrophus in the deodorization of animal feces.

[0014] Specifically, it is the application of this Paracoccus pantotrophus in the preparation of an animal feces deodorant.

[0015] Furthermore, the substance targeted by the deodorization is hydrogen sulfide.

[0016] Furthermore, the bacterial liquid concentration of the Paracoccus pantotrophus is 1×10 11 CFU / m 3 .

[0017] The beneficial effects of the present invention are as follows:

[0018] The currently discovered types of deodorizing strains mainly include Bacillus subtilis, Bacillus licheniformis, Alcaligenes faecalis, Pseudomonas, etc. The types are few, and it is difficult to develop new deodorizing strains. In addition, when these strains are used in combination, there may be antagonistic effects, thus affecting their deodorizing effect and long-term stability. The present invention provides a new strain, Paracoccus pantotrophus, which has not been used in the deodorizing field. When Paracoccus pantotrophus of the present invention is used for animal feces fermentation, it is found that the bacterial agent can effectively reduce the odor generated during feces fermentation, especially significantly inhibit the release of hydrogen sulfide. Therefore, the present invention not only enriches the types of deodorizing strains, but also has an excellent effect on the removal of hydrogen sulfide, and the deodorizing effect can be stable and lasting. Brief Description of the Drawings

[0019] Figure 1 Colony morphology diagram of Paracoccus pantotrophus SZZ1 of the present invention.

[0020] Figure 2 Gram staining diagram of Paracoccus pantotrophus SZZ1 of the present invention.

[0021] Figure 3 Growth curve of Paracoccus pantotrophus SZZ1 of the present invention. Detailed Description of the Invention

[0022] In order to make the objectives, technical solutions and advantages of the present invention clearer, the following describes the present invention in detail with reference to the embodiments. It should be noted that the following embodiments are only used to explain and illustrate the present invention, and are not used to limit the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the above invention content still fall within the protection scope of the present invention.

[0023] Example 1

[0024] 1. Isolation of Strains

[0025] 1.1 Enrichment and Domestication of Strains

[0026] (1) The composition formula of the culture medium used in this application is as follows:

[0027] Basic inorganic salt medium: KH2PO4 2.0 g / L, NH4Cl 0.4 g / L, Na2CO3 0.4 g / L, MgCl2·6H2O 0.2 g / L, pH = 7.0, sterilized at 121 °C for 20 min.

[0028] S1 medium: Sodium sulfide is added to the basic inorganic salt medium to make the S 2- concentration 5 mmol / L.

[0029] S2 medium: Sodium sulfide is added to the basal inorganic salt medium to make the S 2- concentration 10 mmol / L.

[0030] S3 medium: Sodium sulfide is added to the basal inorganic salt medium to make the S 2- concentration 15 mmol / L.

[0031] S4 medium: Sodium sulfide is added to the basal inorganic salt medium to make the S 2- concentration 20 mmol / L.

[0032] TSB medium: Tryptone 17.0 g / L, Phytone 3.0 g / L, Sodium chloride 5.0 g / L, Dipotassium hydrogen phosphate 2.5 g / L, Glucose 2.5 g / L.

[0033] (2) Prepare 100 mL of S1 medium, inoculate 5 g of the sample (the sample is from different environments in the pig farm (feces, soil, compost, biogas slurry)), place it in a shaker at 30 °C and 150 r / min for 72 h, and transfer it successively to S2 medium, S3 medium, and S4 medium for enrichment culture according to 10% (volume percentage).

[0034] 1.2 Isolation and purification of strains

[0035] Dilute and spread the enrichment solution on plates, pick single colonies, and isolate and purify them by the streak plate method.

[0036] 2. Primary screening of deodorizing microorganisms

[0037] Cultivate the purified strains in shake flasks to make a bacterial suspension. Weigh 50 g of air-dried pig manure and put it into a 1000 mL large beaker. Add 10 mL of the bacterial suspension to the large beaker, then add 40 mL of sterile water to the large beaker. Add 50 mL of sterile water to the control group. Seal it with double-layer plastic wrap, and set up 3 replicates for each group. After culturing for 5 days, divide the odor level by sensory evaluation (odorless - smelly are M0 - M4 in turn), and initially screen the strains with better effects for rescreening.

[0038] 3. Screening of hydrogen sulfide-removing microorganisms

[0039] The purified strain was cultured in a shake flask to prepare a bacterial suspension, which was inoculated into a 300 mL large test tube containing 100 mL of basic fermentation medium at an inoculation amount of 10% (volume percentage). Additionally, 2 mL of a hydrogen sulfide standard solution with a mass concentration of 1000 μg / mL was added, and a small container filled with 10 mL of solution was suspended in the large test tube to absorb hydrogen sulfide gas. In the control group, no bacterial solution was added, and an equal amount of sterile water was added. Three replicates were set for each treatment, and they were placed in an incubator at 30 °C for constant temperature culture in the dark. After 48 h, the hydrogen sulfide content in the absorption solution and the remaining hydrogen sulfide content in the large test tube were measured, and through comparison, 1 strain of strain SZZ1 with the highest hydrogen sulfide removal efficiency was selected.

[0040] 4. Morphological characteristics of the strain

[0041] As Figure 1 shown, the colony diameter of strain SZZ1 was 1.5 - 1.8 mm, milky white, round, raised, with a smooth and moist surface and neat edges. As Figure 2 shown, the Gram staining of strain SZZ1 was negative.

[0042] 5. Growth curve

[0043] Fresh single colonies on the solid medium were picked and inoculated into a bacterial bottle containing TSB liquid medium, and placed in a shaker at 30 °C for culture at a rotation speed of 180 rpm / min. Samples were taken every 2 h, and the bacterial growth curve was plotted based on the number of bacterial CFUs.

[0044] 6. Molecular biology identification

[0045] The strain was sent to BGI Genomics Co., Ltd. for molecular biology sequence identification, and the results showed that the strain belonged to Paracoccus pantotrophus SZZ1. At the same time, the strain was sent to the China Center for Type Culture Collection of Wuhan University for biological preservation. The preservation unit was: China Center for Type Culture Collection, and the preservation address was: Wuhan University, Wuhan, China. The preservation center number was: CCTCC No. M 20242509, and the preservation time was: November 11, 2024.

[0046] 7. Gene 16S rRNA sequence

[0047] The gene sequence of Paracoccus pantotrophus SZZ1 of the present invention is as shown in SEQ NO.1.

[0048] Example 2

[0049] Application of SZZ1 in reducing the odor of livestock manure in a mutton sheep farm

[0050] 1. Implementation location: Xilingxue Agriculture Co., Ltd., Dayi County, Chengdu City

[0051] 2. Experimental Methods

[0052] In the harmless treatment area of the sheep farm, composting windrows were established with a length of 3 m, a height of 1 m, and a width of 1.5 m for each treatment group. A total of 2 treatment groups were set up in the experiment, with each group replicated 3 times. No addition of microbial inoculant was used as the blank control. The experimental period was 4 weeks, and the odor value and hydrogen sulfide release were measured weekly using instruments.

[0053] Paracoccus pantotrophus treatment group: The inoculant was evenly sprayed at a dosage of 1×10 11 cfu / m 3 , and the water content was adjusted to be between 60 - 65%. After 14 days, it was supplemented and applied once.

[0054] The corresponding degradation rates (odor value, hydrogen sulfide) were calculated according to the following formula:

[0055] Degradation rate (%) = [(Release amount of blank control - Release amount of treatment group) / Release amount of blank control group] × 100%

[0056] 3. Experimental Results

[0057] 3.1 Growth Curve

[0058] The fresh bacterial liquid SZZ1 began to enter the growth phase after 2 h of cultivation in TSB medium and entered the plateau phase at 10 h. When cultivated for 14 h, the colony count reached 12.7×10 9 CFU, as Figure 3 shown.

[0059] 3.2 Detection Results of Odor Value and Hydrogen Sulfide in the Experiment on Reducing Odor of Farm Manure

[0060] Table 1 Detection Results of Odor Value in the Experiment on Reducing Odor of Farm Manure

[0061]

[0062] Table 2 Detection Results of Hydrogen Sulfide in the Experiment on Reducing Odor of Farm Manure

[0063]

[0064] After detection, the odor value of the control group exceeded the detection range of the instrument and reached 999 OU / m 3 after 1 week, and only decreased to 847.67 OU / m 3 in the 4th week. The odor values of the Paracoccus pantotrophus SZZ1 treatment group decreased to 493 OU / m 3 , 84.67 OU / m 3 , 32.67 OU / m 3 , 8.33 OU / m 3, the odor degradation rates reached 50.65%, 91.52%, 96.73%, and 99.02% successively from the 7th day to the 28th day.

[0065] After detection, the hydrogen sulfide levels in the control group were 26.67 ppm, 38.33 ppm, 20 ppm, and 12 ppm on the 7th day, 14th day, 21st day, and 28th day respectively. In the Paracoccus pantotrophus SZZ1 treatment group, the hydrogen sulfide level dropped to 1 ppm on the 7th day and then to 0 ppm thereafter. The hydrogen sulfide degradation rate reached 96.15% on the 7th day and 100% thereafter.

Claims

1. A strain of Paracoccus pantrophicus, characterized in that The pan-trophic Paracoccus is named ParacoccuspantotrophusSZZ1, the preservation unit is: China Center for Type Culture Collection, the preservation address is: Wuhan University, Wuhan, China, the preservation center number is: CCTCC No.M 20242509, and the preservation time is: November 11, 2024.

2. The pantrophic Paracoccus according to claim 1, characterized in that The gene sequence of the pantrophic Paracoccus is shown in SEQ NO.

1.

3. A biocontrol agent, characterized in that: The biocontrol agent comprises the Paracoccus pantotrophicus according to claim 1 or 2.

4. A deodorant, characterized in that: The deodorant comprises the Paracoccus pantotrophicus according to claim 1 or 2.

5. Use of the Paracoccus pantotrophicus according to claim 1 or 2 in deodorization.

6. Use of the Paracoccus pantotrophicus according to claim 1 or 2 in the preparation of a deodorant.

7. The use according to claim 5, characterized in that: The invention relates to the application of the pantrophic Paracoccus in deodorizing animal excrement.

8. The use according to claim 6, characterized in that: The invention relates to the application of the pantrophic Paracoccus in preparing an animal excrement deodorant.

9. The use according to claim 7 or 8, characterized in that: The substance targeted by the deodorization is hydrogen sulfide.

10. The use according to claim 7 or 8, characterized in that: The concentration of the Paracoccus pantrophicus solution was 1×10 11 CFU / m 3 .

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