Method for preparing organic fertilizer by utilizing defective grapefruit and application of organic fertilizer

Abstract

The invention discloses a method for preparing an organic fertilizer by utilizing defective grapefruit and application of the organic fertilizer, and belongs to the technical field of agricultural waste resource utilization and microbial fertilizers. The microbacterium A5, the bacillus flexus HL-37 and the bacillus cereus TH-35 are used for degrading the defective grapefruit fruits to prepare the organic bacterial fertilizer, the liquid bacterial fertilizer prepared through a staged aerobic fermentation process is mixed with soybean meal / manure according to the ratio of 1: 1 to be returned to a grapefruit orchard, the weight of the grapefruit fruits can be increased by 10.2%, the sugar content can be increased by 5.3%, meanwhile, odor is eliminated, and the pH value of alkaline soil is neutralized; when the solid organic fertilizer is used for vegetable planting, the yield is increased by 23.8% compared with that of a chemical fertilizer, and the microbial structure of soil is remarkably improved. The method realizes resource utilization of agricultural wastes, and has both environmental protection and yield increase benefits.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of agricultural waste resource utilization and microbial fertilizer technology, in particular to a method for preparing organic fertilizer from residual and inferior pomelo fruits and application thereof. BACKGROUND

[0002] With the expansion of the planting scale of pomelo in the main producing areas year by year, the biomass of residual and inferior pomelo fruits produced by thinning every year and the secondary fruits without fresh value is twice that of fresh pomelo fruits. Since the pomelo peel is thick and hard, it is difficult to be naturally degraded by microorganisms in nature, and it has become a new agricultural waste garbage when piled up in the field, causing a huge environmental burden. How to realize the rapid on-site return of residual and inferior pomelo fruit resources and realize the resource recycling of agricultural waste is a problem to be solved.

[0003] Organic fertilizer is a carbon-containing substance extracted from plants and animals, which is applied to soil to provide nutrients for plant growth. Organic fertilizer, including compost, biogas fertilizer, green manure, etc., is the residue extracted from fertilizer after a period of decomposition. In a narrow sense, organic fertilizer refers to a substance that needs to be removed by a certain process. According to the nature and function of organic fertilizer, it can be divided into organic fertilizer, urine fertilizer, straw fertilizer, green fertilizer, soil fertilizer and microbial fertilizer. The application of organic fertilizer in agricultural production not only can improve yield and quality, but also can change the composition, properties and microorganisms of soil. Organic fertilizer can change the porosity of soil particles and form a more suitable soil aggregate structure. Organic fertilizer can also change the composition of soil, including nitrogen, phosphorus, potassium and other elements, as well as organic matter existing in soil as nutrients for soil, providing nutrients required for crop growth and accelerating crop growth. The application of organic fertilizer can change the amount and proportion of humic acid, increase the amount of humic acid, activate the quality of humic acid, and improve the fertility of soil by improving the quality of humic acid. Organic fertilizer not only can improve soil quality, but also can achieve the effect of promoting crop growth and improving quality, and promote agricultural development in many aspects.

[0004] Pomelo fertilizer is a type of green manure, a type of organic fertilizer. This invention utilizes on-site, segmented aerobic fermentation of defective pomelo fruit, using the fermentation product as organic fertilizer for vegetable and pomelo cultivation. Comparisons of soil microbial community composition, soil physical and chemical properties, and crop quality before and after application of this organic fertilizer confirm the effectiveness of the green fertilizer formed by rapid aerobic fermentation of pomelo peel and defective fruit. First, this method addresses the high labor, transportation, and centralized processing costs associated with collecting defective pomelo fruit, enabling the on-site return of agricultural waste to the fields and the recycling of resources. Second, it provides a new type of organic fertilizer for vegetable and fruit cultivation, which is of great significance for the efficient utilization of biomass, the development of a bio-circular economy, and the scientific and technological support of rural revitalization. Third, the organic fertilizer degraded from the pomelo fruit has a distinctive essential oil aroma unique to pomelo. When mixed with organic fertilizers such as poultry manure and peanut bran produced through anaerobic fermentation, it effectively eliminates odors generated by other organic fertilizers, acting as a deodorant. This can promote the sustainable development of agriculture, reduce negative environmental impacts, and advance the development of green agriculture. Summary of the Invention

[0005] The purpose of the present invention is to provide a method for preparing organic fertilizer using defective pomelo fruits and its application, so as to solve the problems existing in the above-mentioned prior art. The organic fertilizer provided by the present invention can be used in vegetable and pomelo planting, and can increase the weight of pomelo fruit by 10.2%, increase the sugar content by 5.3%, and eliminate odor. When the solid organic fertilizer is used in vegetable planting, the yield is increased by 23.8% compared with chemical fertilizer, and the soil microbial structure is significantly improved.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] The present invention provides a method for preparing organic fertilizer by utilizing defective pomelo fruits, comprising any one of the following steps:

[0008] Bacillus flexus HL-37 was inoculated into No. 1 pomelo fruit culture medium for cultivation to obtain HL-37 fermentation liquid;

[0009] Bacillus cereus TH-35 was inoculated into No. 1 pomelo fruit culture medium for cultivation to obtain TH-35 fermentation liquid;

[0010] Inoculating Microbacterium A5 into a No. 1 pomelo fruit culture medium for culture; then inoculating into a No. 2 pomelo fruit culture medium for culture, and then inoculating into a No. 3 pomelo fruit culture medium for culture, to obtain a Microbacterium A5 complete pomelo fruit fermentation liquid;

[0011] Add the HL-37 fermentation liquid and the TH-35 fermentation liquid to the complete pomelo fruit fermentation liquid of Microbacterium A5, and continue fermenting to obtain liquid bacterial fertilizer;

[0012] The liquid bacterial fertilizer is dried to obtain solid organic fertilizer;

[0013] The No. 1 pomelo fruit culture medium: 500 mL of LB medium is added with 250 g of pomelo residual and secondary fruits, and water is added to make up to 1 L.

[0014] The No. 2 pomelo fruit culture medium: 200 mL of LB medium is added with 400 g of pomelo residual and secondary fruits, and water is added to make up to 1 L.

[0015] The No. 3 pomelo fruit culture medium: 500 g of pomelo residual and secondary fruits are added with water to make up to 1 L.

[0016] Optionally, the preservation number of the Curvularia genitana HL-37 is GDMCC No. 60045, the preservation number of the Bacillus cereus TH-35 is GDMCC No. 60044, and the preservation number of the Microbacterium A5 is CCTCC NO: M209174.

[0017] Optionally, the inoculation amount of the Microbacterium A5 is 5-15%.

[0018] Optionally, the inoculation amount of the HL-37 fermentation liquid and the TH-35 fermentation liquid is 10%.

[0019] The application further provides a liquid microbial fertilizer prepared according to the method.

[0020] The application further provides a solid organic fertilizer prepared according to the method.

[0021] The application further provides application of the liquid microbial fertilizer in promoting growth of pomelos.

[0022] Optionally, the promoting growth of pomelos includes increasing single fruit weight and / or sugar content of pomelos.

[0023] The application further provides application of the solid organic fertilizer in promoting growth of vegetables.

[0024] Optionally, the promoting growth of vegetables includes increasing fresh weight and / or plant height of vegetables.

[0025] The vegetables include amaranth.

[0026] The application discloses the following technical effects:

[0027] The application provides a method for preparing organic microbial fertilizer by degrading residual and inferior pomelo fruits by three strains (Microbacterium A5, HL-37 and TH-35). The prepared liquid microbial fertilizer is mixed with soybean meal / fecal fertilizer at a ratio of 1:1 and then applied to pomelo orchards, so that the weight of pomelo fruits is increased by 10.2%, the sugar content is increased by 5.3%, the odor of soybean meal / fecal fertilizer anaerobic fermentation is eliminated, and the alkaline soil pH is neutralized; when the solid organic fertilizer is used for vegetable planting, the yield is increased by 23.8% compared with chemical fertilizer, and the soil microbial structure is significantly improved. The application realizes the resource utilization of agricultural waste, and has the benefits of environmental protection and yield increase. BRIEF DESCRIPTION OF DRAWINGS

[0028] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed in the embodiments will be briefly introduced as follows. Obviously, the drawings in the following description only constitute some embodiments of the present application, and for those skilled in the art, other drawings can also be obtained without creative labor on the basis of these drawings.

[0029] Figure 1 It is the colony morphology of Microbacterium A5;

[0030] Figure 2 It is the growth curve of Microbacterium A5 seed extension culture;

[0031] Figure 3 It is the growth curve of Microbacterium A5 scale-up fermentation;

[0032] Figure 4 It is the rhizosphere soil microbial alpha diversity of different treatment groups;

[0033] Figure 5 It is a PCA analysis diagram;

[0034] Figure 6 It is a sample clustering heat map;

[0035] Figure 7 It is a species distribution diagram, the left side is the door level, and the right side is the class level;

[0036] Figure 8 It is a species abundance clustering heat map;

[0037] Figure 9 It is a network diagram of each species at the genus level. DETAILED DESCRIPTION

[0038] Now, various exemplary embodiments of the present application will be described in detail, which should not be considered as a limitation of the present application, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present application.

[0039] It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the present application. In addition, where a range of values is provided, it is understood that each intervening value, to the upper and lower limit of the range is also specifically disclosed. Each smaller range between any stated value or intervening value in the stated range and any other stated or intervening value in that stated range is encompassed. The upper and lower limits of these smaller ranges can independently be included or excluded in the range.

[0040] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application. All documents mentioned herein are incorporated by reference to disclose and describe in detail the methods and / or materials which are described in them. In the case of conflict between the present specification and any document incorporated herein by reference, the present specification will control.

[0041] Many modifications and variations of the present application described in the specification are possible without departing from the scope or spirit of the application. Other embodiments of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application disclosed herein. The specification and examples are illustrative only.

[0042] As used herein, the terms "comprises", "comprising", "includes", "including", "has", "having", "contains", "containing", or variations thereof, are intended to be open-ended terms that mean including, but not limited to.

[0043] The microbacterium A5 used in the present application has a preservation number of CCTCC NO: M209174, and has been disclosed in patent CN101701198B "Peach gum-decomposing enzyme producing strain and its application in preparing peach gum polysaccharide"; the bacillus flexus HL-37 has a preservation number of GDMCC No. 60045, and has been disclosed in patent CN106591173A "Bacillus flexus HL-37 capable of activating heavy metal cadmium in soil and its application"; the bacillus cereus TH-35 has a preservation number of GDMCC No. 60044, and has been disclosed in patent CN106167776A "Bacillus cereus TH-35 capable of activating heavy metal cadmium in soil and its application".

[0044] Example 1

[0045] 1. Preparation of pomelo culture medium

[0046] Pretreatment of pomelo residual fruits: wash the pomelo with water to remove dirt and impurities on the surface of the pomelo peel. Cut into small pieces of 2-3 cm in size.

[0047] (1) No. 1 nectarine medium, per liter: LB medium 500 mL + 250 g nectarine peel pieces, add water to 1 L, adjust pH = 4 with 0.2 mol / L NaOH. Divide into 250 mL conical flasks, 150 mL / flask, 121 °C, sterilize for 20 min.

[0048] (2) No. 2 nectarine medium, per liter: LB medium 200 mL + 400 g nectarine peel pieces, add water to 1 L, adjust pH = 4 with 0.2 mol / L HCL. Divide into 1 L shake flasks, 500 mL / flask, 121 °C, sterilize for 20 min.

[0049] (3) No. 3 nectarine medium: 500 g nectarine fruit pieces, add water to 1 L, adjust pH = 4 with 1 mol / L NaOH, 121 °C, sterilize for 20 min.

[0050] 2. Domestication and scale-up fermentation of nectarine peel degrading bacteria

[0051] (1) The microbacterium A5 strain stored at 4 °C was streaked on LB solid medium plates and placed in a 30 °C incubator for 24 h. Single colonies were picked and inoculated into 4 mL of LB liquid medium in a test tube for activation, and cultured at 30 °C with 150 rpm / min shaking for 12 h.

[0052] (2) 3 mL of the bacterial solution was inoculated into 150 mL of No. 1 nectarine medium and cultured at 30 °C with 150 rpm shaking for 3 days. After the nectarine particles in the 150 mL medium / 250 mL conical flask were basically degraded and there were no large particles, 100 mL of the culture solution was transferred into 500 mL of No. 2 nectarine medium / 1 L conical flask and cultured at 30 °C with 150 rpm shaking for 5-7 days. During the fermentation process, samples were taken every 4 h and OD 600 was measured. Finally, the growth curve of microbacterium A5 was drawn (vertical coordinate: OD 600 , horizontal coordinate: shaking bed fermentation time).

[0053] (3) After the nectarine particles in the 1 L conical flask disappeared and became a paste, 1 L of the fermentation product was inoculated into 3 No. 3 nectarine medium in a 10 L or 15 L plastic bucket, the top of the bucket was covered with double-layer gauze or 300 mesh gauze, and the room temperature was opened once a day to stir and ventilate until the nectarine particles completely became a paste to obtain the complete nectarine fermentation solution of microbacterium A5.

[0054] 3. Strain combination

[0055] Microbacterium A5, Bacillus flexus HL-37 and Bacillus cereus TH-35 were used for segmented fermentation.

[0056] (1) HL-37 fermentation liquid preparation: 1 mL of liquid seed was inoculated into 4 mL of LB medium and cultured for 12 hours, then inoculated into 100 mL of LB medium and cultured for 12 hours, and then inoculated into No. 1 pomelo medium at a 15% inoculation amount for 12 hours of shaking culture, and the viable cell count was ≥10 9 CFU / mL of fermentation liquid.

[0057] (2) TH-35 fermentation liquid preparation: 1 mL of liquid seed was inoculated into 4 mL of LB medium and cultured for 12 hours, then inoculated into 100 mL of LB medium and cultured for 12 hours, and then inoculated into No. 1 pomelo medium at a 15% inoculation amount for 12 hours of shaking culture, and the viable cell count was ≥10 9 CFU / mL of fermentation liquid.

[0058] (3) Liquid / solid bacterial fertilizer preparation

[0059] After pretreatment of the residual pomelo fruits, the microbacterium A5 was acclimated in No. 1 pomelo medium→No. 2 pomelo medium→No. 3 pomelo medium, and then mixed with HL-37 fermentation liquid and TH-35 fermentation liquid at a volume ratio of 10:1:1 to obtain a liquid bacterial fertilizer.

[0060] The HL-37 fermentation liquid / TH-35 fermentation liquid was mixed with the microbacterium A5 complete pomelo fermentation liquid at a volume ratio of 1:10 to obtain a two-bacterial combination liquid bacterial fertilizer.

[0061] The liquid bacterial fertilizer was dried at 60°C for 5 hours to obtain a liquid organic fertilizer.

[0062] 4. Field scale-up bacterial fertilizer preparation process

[0063] A 200L plastic bucket was cleaned and disinfected with diluted 84 disinfectant, 5cm-sized residual young fruit blocks (residual young fruit weight: 75kg) were added, 75L of water was added to the bucket to a total volume of 150L, 10-15L of microbacterium A5 complete pomelo fermentation liquid was added, and the mixture was stirred evenly. A 200-mesh gauze was placed on the bucket mouth, and the mixture was stirred once a day. After 3-4 days, the fruit blocks became a paste, which was used as a field fermentation medium. The HL-37 fermentation liquid and the TH-35 fermentation liquid prepared above were inoculated at a volume ratio of 10%, and the mixture was stirred once a day. After 2 days, a liquid bacterial fertilizer was formed and could be used.

[0064] The liquid bacterial fertilizer was dried at 60°C for 5 hours to obtain a solid organic fertilizer.

[0065] Application method: liquid bacterial fertilizer application in pomelo orchard: mixed with bean meal / fecal fertilizer (bean meal and fecal fertilizer at a volume ratio of 1.5:4.5) and other anaerobic fermentation organic fertilizers at a volume ratio of 1:1, 20 pounds per tree, and fertilized every two weeks for a total of four times.

[0066] Solid organic fertilizer is used in vegetable cultivation: instead of chemical fertilizer, the application amount is 25g / kg soil (liquid fertilizer is 200mL / kg).

[0067] 5. Results

[0068] 5.1 Colony morphology of Microbacterium A5

[0069] like Figure 1 As shown in the figure, the starting colony of Microbacterium A5 used for grapefruit peel degradation is round, light yellow, thick in the middle and thin at the edge, and relatively moist.

[0070] 5.2 Acclimation of Microbacterium to Grapefruit Peel Degradation Activity and Growth Curves During Scale-up Fermentation

[0071] The step-by-step amplification process of Microbacterium degradation of grapefruit fermentation is, on the one hand, to gradually enhance the degradation activity of the bacteria on grapefruit peel, and on the other hand, to increase the processing capacity and shorten the fermentation cycle. Figure 2 As shown, under the conditions of 30°C and 150rpm shaking culture, the microbacterium enters the logarithmic growth phase at 30h-36h, and the fermentation liquid at the middle and late logarithmic growth phase at 35-36h is determined as the seed liquid for the next 500mL / 1L scale-up fermentation. Figure 3 As shown, in 80% grapefruit peel culture medium, at 30°C and 150 rpm / min, Microbacterium enters the logarithmic growth phase at 20-28 hours. The fermentation liquid at 24 hours is determined as the seed liquid for inoculation into the complete grapefruit peel culture medium.

[0072] 5.3 Degradation rate of defective grapefruit fruit

[0073] The degradation efficiency of the three-bacteria synergistic fermentation and the single / two-bacteria combination on the defective grapefruit fruit is shown in Table 1. The degradation rate of the three-bacteria synergistic fermentation composite agent on the main components of grapefruit peel (cellulose, hemicellulose and lignin) is shown in Table 2.

[0074] Table 1 Comparison of degradation efficiency of defective grapefruit fruit by three-bacteria synergistic fermentation and single / two-bacteria combination

[0075]

[0076] Table 2 Degradation rate of main components of pomelo peel by three-bacteria synergistic fermentation composite agent

[0077] Ingredients Initial content Content after fermentation Degradation rate Cellulose 32.5％ 2.6％ 92.0％ Hemicellulose 28.7％ 3.4％ 88.1％ Lignin 18.3％ 6.4％ 65.0％

[0078] Example 2 Application of liquid bacterial fertilizer in pomelo orchard

[0079] 1. Materials and Methods

[0080] The experiment was conducted using the liquid bacterial fertilizer prepared by the field-amplified bacterial fertilizer preparation process in Example 1.

[0081] In Meizhou Songyuan pomelo orchard, 4 groups of treatments (n = 20 trees) were set: T1: conventional chemical fertilizer; T2: liquid microbial fertilizer + soybean meal / manure (volume ratio of liquid microbial fertilizer to soybean meal / manure is 1:1, volume ratio of soybean meal to manure is 1.5:4.5); T3: soybean meal / manure (anaerobic compost); T4: blank control.

[0082] Fertilization scheme: T2 group applied 20 kg of mixed fertilizer (10 kg of liquid microbial fertilizer + 10 kg of soybean meal / manure) per tree each time, a total of 4 times, with an interval of 14 days; the other groups were applied with corresponding fertilizers according to the same treatment.

[0083] Detection index: single fruit weight, sugar content (handheld sugar density meter) were measured when the pomelo fruits were mature, and soil organic matter, alkali-hydrolyzable nitrogen content, air volatile organic compounds (VOCs) concentration, odor sensory score standard, soil element and organic matter content detection methods are as follows.

[0084] 1.1 Soil element and organic matter content detection

[0085] (1) Soil organic matter content detection

[0086] Potassium dichromate oxidation-external heating method (classic method)

[0087] Standard basis: "Soil Testing Part 6: Determination of Soil Organic Matter" (NY / T 1121.6-2006).

[0088] Detection steps: weigh an appropriate amount of air-dried soil sample, add excess potassium dichromate-sulfuric acid solution, and heat in an oil bath to oxidize organic matter. Titrate the remaining potassium dichromate with ferrous sulfate solution, and calculate the organic matter content (converted to carbon content, unit: g / kg) according to the consumption.

[0089] (2) Soil element detection

[0090] Pre-treatment method: soil digestion: use mixed acid of hydrofluoric acid + nitric acid + perchloric acid to digest the soil, so that the elements are converted into ionic state (standard: HJ 803-2016).

[0091] Element determination: inductively coupled plasma optical emission spectrometry (ICP-OES), using standard: HJ 700-2014.

[0092] 1.2 Determination of volatile organic compounds (VOCs) concentration in air

[0093] Determination was performed using a portable PID detector.

[0094] a. Instrument preparation and calibration

[0095] Preheating: Turn on the power, preheat the instrument for 10-15 minutes until the indicator light is stable (avoid the influence of environmental temperature on the accuracy).

[0096] Zero calibration: Clean odorless air (or high-purity nitrogen) is passed through the sampling tube into the instrument for 1-2 minutes, and the instrument display value is adjusted to 0 ppm (or automatic zero calibration is performed according to the instructions). Span calibration (optional): Connect the isobutene standard gas (known concentration), and pass it into the instrument at a flow rate of 100-200 mL / min. After the reading is stable, adjust the instrument to make the display value consistent with the standard gas concentration (calibration frequency: before each test or after instrument failure).

[0097] b. Field sampling and testing

[0098] Direct sampling: Place the sampling probe at the detection point (about 1.5 m from the ground), turn on the instrument pump suction function, and directly inhale the air sample (the sampling flow rate is usually 200-500 mL / min).

[0099] Adsorption tube sampling (if enrichment is required): After sampling with activated carbon or Tenax tube, the sample is introduced into the PID detector by thermal desorption or solvent analysis (suitable for low concentration scenarios).

[0100] c. Detection operation:

[0101] Insert the sampling probe into the environment to be tested, avoiding contact with water or particulate matter (if necessary, install a filter head).

[0102] Start the detection program, and the instrument displays the VOCs concentration value in real time (usually updated every second), and record the maximum value or the average value after stabilization.

[0103] If multiple points are detected, clean air should be used to purge the sampling tube for 1-2 minutes when switching points to avoid cross contamination.

[0104] d. Data recording and processing

[0105] Real-time recording: Simultaneously record the detection time, location, environmental temperature (PID response is affected by temperature, which needs to be corrected), humidity (humidity > 80% may affect accuracy, and dehumidification is recommended), and instrument display concentration value.

[0106] Unit conversion: Convert ppm to mg / m 3 , concentration (mg / m 3 ) = concentration (ppm) * molecular weight / 24.45;

[0107] Note: 24.45 is the molar volume of gas under standard conditions, unit L / mol; the molecular weight is the average molecular weight of the VOCs mixture, or the molecular weight of isobutene is assumed to be 56.1.

[0108] 1.3 Odor sensory score criteria (three-point comparison odor bag method)

[0109] Standard basis: Determination of air quality odor-three point comparison odor bag method (GB / T 14675-1993).

[0110] Olfactometer requirements: Train 6 olfactometers (no olfactory disorder), aged 18-45 years.

[0111] Sample preparation: The sample gas is diluted with odorless air in stages to prepare gas bags of different dilution ratios.

[0112] Olfactory steps: Provide 3 gas bags to the olfactometer each time, one containing sample dilution gas and two odorless air, and let it identify the bag with odor. Test from low concentration to high concentration until the olfactometer cannot correctly identify it, and record the lowest odor concentration (olfactory threshold).

[0113] The scoring criteria are shown in Table 3.

[0114] Table 3 Odor sensory score criteria

[0115]

[0116] 2. Experimental results

[0117] 2.1 Effect of liquid bacterial fertilizer on average weight of single fruit, sugar content, soil organic matter and alkali-hydrolyzable nitrogen content of grapefruit

[0118] The average weight of single fruit, sugar content, soil organic matter and alkali-hydrolyzable nitrogen content of each group of grapefruit are shown in Table 4.

[0119] Table 4 Average weight of single fruit, sugar content, soil organic matter and alkali-hydrolyzable nitrogen content of each group of grapefruit

[0120] Group Average weight of single fruit / g Sugar content / % Soil organic matter / % Alkaline nitrogen / mg / kg T1 856±32 12.5±0.8 1.82±0.15 156.7±9.2 T2 943±28 17.8±0.5 3.14±0.12 189.3±11.5 T3 835±25 12.7±0.6 1.79±0.13 152.4±8.7 T4 720±20 10.2±0.4 1.35±0.10 112.5±7.3

[0121] Note: Compared with the T4 non-fertilized blank group, both chemical fertilizer and soybean meal / fecal fertilizer can increase the weight of grapefruit, but the average weight of single fruit, sugar content, soil organic matter and alkali-hydrolyzable nitrogen of T2: liquid bacterial fertilizer + soybean meal / fecal fertilizer (1:1) are the most significant.

[0122] 2.2 Deodorization effect

[0123] After fertilization, the concentration of volatile organic compounds (VOCs) in the air of the orchard in the T2 group was reduced by 68.5% compared with the T3 group, and the odor sensory score decreased from 5.2 (strong odor) to 1.8 (no obvious odor).

[0124] 2.3 Effect of liquid bacterial fertilizer on soil element and organic matter content

[0125] Table 5 Soil element and organic matter content of each group

[0126]

[0127]

[0128] As can be seen from Table 5, (1) the organic matter and soil fertility are significantly improved. After applying the pomelo peel organic fertilizer, the soil organic matter content increases from 0.19% to 17.46%, with an increase of 81.4 times, indicating that the soil structure and fertilizer retention capacity are significantly improved; at the same time, the conductivity increases from 57us / cm to 983.1us / cm, indicating that the content of soluble nutrients (such as nitrogen, phosphorus and potassium ions) in the soil is greatly increased, providing more sufficient nutrients for plant growth. (2) The content of nitrogen, phosphorus and potassium elements is greatly improved. The total nitrogen content increases from 205.17mg / kg to 6247.21mg / kg, the alkali-hydrolyzable nitrogen (the form of nitrogen that can be directly absorbed by plants) increases from 24.42mg / kg to 177.5mg / kg, with an increase of 6.3 times, promoting the growth of plant leaves and photosynthesis; the available phosphorus content increases from 75.72mg / kg to 1945.09mg / kg, with an increase of 24.4 times, significantly enhancing the root development and fruit quality of plants; the available potassium increases from 64.00mg / kg to 2185.85mg / kg, with an increase of 33.8 times, which is helpful for plant stress resistance (such as drought resistance and cold resistance) and sugar accumulation in fruits. (3) The trace elements such as calcium, magnesium and iron are improved synergistically. The contents of calcium, magnesium and iron and other trace elements are significantly increased (such as the content of iron increases by 122.7 times), which can prevent plant deficiency and promote fruit development; the content of copper decreases slightly, which may be related to the slow-release characteristics of copper elements in organic fertilizer, but the overall content is still within the suitable absorption range of plants. (4) The pH value is adaptively adjusted. The soil pH value decreases from 6.76 (weak alkaline) to 4.25 (acidic), which is close to the pH value (4.09) of the organic fertilizer itself. Although the acidity is increased, pomelo and other crops are suitable for slightly acidic soil (pH 5.5-6.5), and the increase of organic matter can buffer the change of soil acidity and alkalinity, and does not inhibit the growth of plants in actual use.

[0129] Meanwhile, the liquid microbial fertilizer prepared by the present application is compared with single strain fermentation (completely pomelo fruit degradation liquid microbial fertilizer prepared by using only A5 microbacterium) and natural composting (pomelo residual fruits are buried in the soil for natural composting), and the results are shown in Table 6.

[0130] Table 6 Comparison of liquid microbial fertilizer with prior art

[0131] Index The present invention Single strain fermentation Natural composting Degradation period 3-7 days (completely paste) 10 days 90-180 days Degradation rate of inferior pomelo fruit 95.3% (weight loss rate) 62.7％ <30％ Odor removal rate 68.5% (VOCs concentration reduction) <30％ None Pomelo fruit weight increase 10.2% (20 kg per plant, 4 times) <5％ None Soil organic matter increase times 81.4 times (from 0.19% to 17.46%) <10 fold <5 fold Nitrogen, phosphorus and potassium retention rate Total nitrogen retention rate 82.3% Total nitrogen retention rate 58.6% Total nitrogen loss over 40%

[0132] In summary, the embodiment verifies that the application of liquid bacterial fertilizer significantly improves the content of organic matter and key nutrients such as nitrogen, phosphorus and potassium in the soil, and at the same time improves the supply of trace elements, creating a better soil environment for plant growth. The data show that the organic fertilizer has excellent effect in improving soil fertility and promoting nutrient balance, significantly better than the prior art, and has the effect of promoting growth and quality improvement.

[0133] Example 3 Application of solid organic fertilizer in vegetable (amaranth) planting

[0134] 1. Experimental design and treatment group setting

[0135] The treatment groups are shown in Table 7.

[0136] Among them, compound fertilizer: market purchase of special compound fertilizer for vegetables (Stanley, SDL-FHF01).

[0137] Table 7 Setting of each treatment group

[0138] Treatment group Fertilizer type Application amount Soil type T1 group (blank control) No fertilizer - Black soil, red soil, loess T2 group Liquid pomelo peel microbial fertilizer 200 mL / kg soil Black soil, red soil, loess T3 group Solid pomelo peel microbial fertilizer 25 g / kg soil Black soil, red soil, loess T4 group (commercial compound fertilizer) Compound fertilizer 2.5 g / kg soil Black soil, red soil, loess

[0139] 2. Results

[0140] 2.1 Effect on the physical indicators of amaranth growth and soil physical and chemical properties

[0141] The comparison of the physical indicators of amaranth growth in each treatment group (after 60 days of planting) is shown in Table 8, and the changes in soil physical and chemical properties (after 60 days of planting) are shown in Table 9.

[0142] As shown in Table 8, the fresh weight of GSF was significantly higher than that of Con: the fresh weight of GSF in black soil was 3.8 times that of Con, and in red soil and yellow soil, it was 3.2 times and 2.9 times respectively; and the fresh weight of GSF was better than that of GLF: the fresh weight of GSF in black soil was 15.3% higher than that of GLF, and in red soil and yellow soil, it was 12.1% and 9.7% higher respectively; while the effect of CF on fresh weight was the best: the fresh weight of CF in black soil was 22.4% higher than that of GSF, but the difference between GSF and CF in red soil and yellow soil was reduced (10.8% and 8.3% lower respectively). GSF significantly promoted the growth of plant height: the plant height of GSF in black soil was 3.1 times that of Con, and in red soil and yellow soil, it was 2.9 times and 2.7 times respectively; and the plant height of GSF was better than that of GLF: the plant height of GSF in black soil was 18.7% higher than that of GLF, and in red soil and yellow soil, it was 14.3% and 11.3% higher respectively. The plant height of CF was the highest: the plant height of CF in black soil was 25.2% higher than that of GSF, but the difference between GSF and CF in red soil and yellow soil was smaller (11.6% and 9.1% lower respectively).

[0143] From Table 9, GSF significantly increased soil organic matter: GSF organic matter was 2.1 times higher than Con in black soil, 2.3 times and 2.6 times higher in red soil and loess, respectively. GSF organic matter was superior to GLF: GSF organic matter was 19.5% higher than GLF in black soil, 18.0% and 17.9% higher in red soil and loess, respectively. CF had limited effect on increasing organic matter: CF organic matter was close to GSF (only 5.4% higher) in black soil, and lower than GSF (12.6% and 15.3% lower, respectively) in red soil and loess. GSF reduced soil pH: GSF pH was 2.3 lower than Con in black soil, 1.8 and 1.5 lower in red soil and loess, respectively, which may be related to the acidity of organic fertilizer (pH 4.09). GSF acidity had a more significant effect: GSF pH was 0.4 lower than GLF in black soil, 0.3 and 0.2 lower in red soil and loess, respectively, but did not completely inhibit the growth of amaranth (as the initial pH of black soil was closer to neutral). CF had a smaller effect on pH: CF pH was 0.8 higher than GSF in black soil, 0.6 and 0.5 higher in red soil and loess, respectively.

[0144] Table 8 Comparison of physical indicators of amaranth growth in each treatment group

[0145]

[0146]

[0147] Note: *p<0.05 compared with Con.

[0148] Table 9 Changes in soil physical and chemical properties

[0149]

[0150] Note: *p<0.05 compared with Con.

[0151] In summary, solid organic fertilizer significantly promoted the growth of amaranth in three types of soil, with fresh weight and plant height increasing by 2.7-3.8 times compared to the blank group, and the effect was superior to that of liquid microbial fertilizer. In black soil, the growth indicators of GSF (such as fresh weight 33.9 g / plant, plant height 30.1 cm) were close to those of compound fertilizer (CF), but the gap between GSF and CF narrowed in red soil and loess, indicating that GSF was adaptable in different soils.

[0152] After applying GSF, the organic matter content of the three types of soil increased significantly (black soil 2.57%, red soil 1.51%, loess 1.24%), which was 1.6-2.6 times higher than that of the blank group, and was superior to that of liquid fertilizer, indicating that the soil improvement effect of GSF was durable.

[0153] Potential for partial replacement of chemical fertilizers: Although the growth-promoting effect of GSF was slightly lower than that of CF, its effect on increasing soil organic matter was significantly superior to that of CF, and the growth gap between GSF and CF was smaller in red soil and loess, which could be used as a replacement for chemical fertilizers.

[0154] Acid effect and adaptability: GSF application leads to soil pH decrease, with less effect in black soil (initial pH 6.64) and good growth of amaranth; attention should be paid to acid adjusting measures such as lime application in red and yellow soils to optimize the effect.

[0155] 2.2 Analysis of the effect on soil microorganisms

[0156] 2.2.1 Alpha diversity

[0157] The results of the detection of the alpha diversity of the rhizosphere soil bacteria of amaranth in the blank group and the solid organic fertilizer group showed that (Table 10 and Figure 4 ), the solid fertilizer treatment significantly reduced the abundance of rhizosphere bacteria by 18.0% compared with the blank group, which may be related to the inhibition of part of the bacterial flora by the acidity (pH 4.09) of the organic fertilizer. The bacterial species richness decreased by 12.6%, indicating that the solid fertilizer may change the soil microenvironment, leading to the disappearance of part of the species. The bacterial diversity decreased by 7.0%, indicating that the complexity of the community structure decreased, but the dominant flora may be more concentrated. The species evenness decreased, and the dominance of the dominant bacterial phylum (such as Proteobacteria) increased under the solid fertilizer treatment.

[0158] Table 10 Alpha diversity of rhizosphere soil bacteria of amaranth in different treatment groups

[0159] Index Blank group (Con) Solid fertilizer group (GSF) Significant difference Note ACE index 2100±85 1722±68 p<0.0001 Abundance Chao1 index 1950±72 1705±59 p<0.0001 Species richness Shannon index 10.2±0.3 9.5±0.2 p<0.01 Diversity Simpson index 0.985±0.004 0.972±0.003 p<0.1 Species evenness

[0160] 2.2.2 Beta diversity

[0161] Beta diversity represents the abundance diversity of microorganisms between different samples. The results of the beta diversity of the rhizosphere soil of amaranth under different treatments are shown in the figure. As can be seen from the PCA analysis figure ( Figure 5 ), the beta diversity of the rhizosphere soil under the four treatments is quite different, with no overlapping part; as can be seen from the characteristic Venn diagram, the number of OTUs unique to Group 1, Group 2, Group 3 and Group 4 is 2572, 2513, 1722 and 1818 respectively, and the number of OTUs common to the four groups is 318, accounting for a low proportion, indicating that the similarity of the community structure between different samples is weak.

[0162] Sample clustering heat map ( Figure 6 ): GSF group and liquid fertilizer group (GLF) are clustered into one class, while Con group and CF group are clustered into another class, indicating that the flora structure of the organic fertilizer treatment is more similar, and is significantly different from the compound fertilizer / blank group.

[0163] 2.2.3 Changes in dominant flora at the phylum / class level

[0164] The dominant bacterial groups at the phylum / class level in the rhizosphere soil of amaranth in the blank group and the solid organic fertilizer group were detected. The results showed that (Table 11 and Figure 7 At the phylum level, the top 10 dominant bacterial groups in relative abundance reached over 90% in all 12 samples. The abundance of the top 10 dominant bacterial groups in the rhizosphere soil was higher in all three fertilizer treatments than in the blank treatment. The main dominant phyla in the rhizosphere soil were Proteobacteria, Acidobacteriota, Gemmatimonadota, and Firmicutes. At the phylum level, compared with the control (Con), compound fertilizer (CF) increased the abundance of Proteobacteria and Gemmatimonadota, while decreasing the abundance of Acidobacteria and Firmicutes. Liquid organic fertilizer (GLF) increased the abundance of Proteobacteria and decreased the abundance of Acidobacteria. Solid organic fertilizer (GSF) increased the abundance of Proteobacteria and decreased the abundance of Acidobacteria, Gemmatimonadota, and Firmicutes. Furthermore, all fertilizer treatments significantly decreased the abundance of Methylomirabilota. At the class level, the top 10 dominant bacterial groups in relative abundance reached 72.0%, 75.0%, 77.0% and 79% in the four treatments, respectively. Alphaproteobacteria, Gammaproteobacteria, Vicinamibacteria and Gemmatimonadetes were the dominant classes in the rhizosphere soil. After the soil treatment, the relative abundance of Alphaproteobacteria and Gemmatimonadota decreased, while the relative abundance of Bacilli increased significantly.

[0165] Table 11 Dominant bacterial groups at the phylum / class level

[0166]

[0167]

[0168] 2.2.4 Genus-level characteristic species and network analysis

[0169] Species abundance cluster heat map ( Figure 8 ): The genera of Halanaerobiaeota were the most abundant in the GSF group, which may be related to the anaerobic metabolism of the refractory carbon source in organic fertilizer; while the genera of Deferrisomatota were more abundant in the blank group, indicating that solid fertilizer changed the carbon cycle-related bacterial community.

[0170] Correlation network analysis ( Figure 9):GSF group, Sphingomonas, Bacillus, the abundance of the genus is higher, and is positively correlated with other genera (red line for the main), suggesting that these bacteria may form a synergistic metabolic network under the treatment of organic fertilizer, promote nutrient transformation.

[0171] In summary, solid organic fertilizer has a dual effect on microbial diversity: reduced diversity but enriched functional flora: solid fertilizer inhibits Acidobacteria, Sphingomonas, etc. Tolerant flora, but promotes Proteobacteria, Firmicutes, etc. Related to nutrient decomposition (such as nitrogen, phosphorus transformation) and plant growth (such as Bacillus), which may optimize flora function through "screening effect".

[0172] Potential impact of soil acidity: solid fertilizer application leads to soil pH of 3.54-4.34, which may inhibit neutral / alkaline bacteria, but enrich acidophilic bacteria (such as some genera in Proteobacteria), and long-term use should pay attention to acid-base balance.

[0173] Difference from compound fertilizer: the decline in microbial diversity under compound fertilizer treatment is smaller than that under solid fertilizer, but solid fertilizer can provide carbon sources for specific functional bacteria by increasing organic matter (such as GSF in the table above, which makes black soil organic matter reach 2.57%), which is beneficial to long-term soil fertility maintenance.

[0174] Example 4 Field planting amaranth experiment

[0175] 1. Field test design

[0176] Use the full red amaranth seeds produced by Nanjing Jiahua Agricultural Development Co., Ltd.

[0177] Treatment group: T1 (solid pomelo peel fermented organic fertilizer 25g / kg), T2 (fertilizer 2.5g / kg, market purchase of vegetable special fertilizer (Kunning king, fhf)), T3 (blank), 3 times of each group, planting area 4m x 1.5m.

[0178] Field planting method: one-time root application, other field management measures are carried out in accordance with the conventional way, and water is poured at regular intervals in the morning and evening.

[0179] 2. Detection index

[0180] The planting period is 65 days, and after 65 days, the fresh weight, dry weight, plant height of amaranth and soil microbial diversity (16S rRNA sequencing) are measured.

[0181] 3. Results

[0182] 3.1 Effect on growth

[0183] The fresh weight, dry weight, and plant height of each group are shown in Table 12.

[0184] Table 12 Growth indicators of each group

[0185] Treatment group Fresh weight (g / plant) Dry weight (g / plant) Plant height (cm) Yield (kg / mu) T1 141±8*** 42±3*** 62±4*** 2840±120** T2 122±6* 35±2* 53±3* 2300±90* T3 26±2 12±1 11±1 520±30

[0186] Note: compared with T3 group, *p<0.05, ***p<0.01.

[0187] 3.2 Effect on soil microorganisms

[0188] Alpha diversity: the ACE index of soil bacteria in the T1 group increased by 15.7% compared with the T2 group, and the Shannon index increased by 12.3%;

[0189] Dominant flora: the abundance of Bacillus in the rhizosphere soil of the T1 group increased by 32.6% compared with the T2 group, and the abundance of Acidobacteriota decreased by 18.4%, indicating that the soil fertility was improved.

[0190] 3.3 Effect on KEGG pathways

[0191] The KEGG pathway abundance changes are shown in Table 13.

[0192] Table 13 KEGG pathway abundance changes

[0193]

[0194]

[0195] Based on the above experimental results, the application provides a method for preparing organic microbial fertilizer by degrading substandard pomelo fruits using three strains (Microbacterium A5, HL-37, and TH-35). Through a segmented aerobic fermentation process, the prepared liquid microbial fertilizer is mixed with soybean meal / fecal fertilizer at a ratio of 1:1 and applied to the pomelo orchard, which can increase the weight of pomelo fruits by 10.2% and the sugar content by 5.3%, while eliminating the odor. When the solid organic fertilizer is used for vegetable planting, the yield is increased by 23.8% compared with chemical fertilizers, and the soil microbial structure is significantly improved. The application realizes the resource utilization of agricultural waste, and has the benefits of environmental protection and yield increase.

[0196] The above-described embodiments are only preferred modes of the application and do not limit the scope of the application. Without departing from the design spirit of the application, various modifications and improvements to the technical solutions of the application made by those skilled in the art shall fall within the protection scope of the claims of the application.

Claims

1. A method for preparing organic fertilizer using defective pomelo fruit, characterized in that: Include any of the following steps: Bacillus flexus HL-37 was inoculated into No. 1 pomelo fruit culture medium for cultivation to obtain HL-37 fermentation liquid; Bacillus cereus TH-35 was inoculated into No. 1 pomelo fruit culture medium for cultivation to obtain TH-35 fermentation liquid; Inoculating Microbacterium A5 into a No. 1 pomelo fruit culture medium for culture; then inoculating into a No. 2 pomelo fruit culture medium for culture, and then inoculating into a No. 3 pomelo fruit culture medium for culture, to obtain a Microbacterium A5 complete pomelo fruit fermentation liquid; Add the HL-37 fermentation liquid and the TH-35 fermentation liquid to the complete pomelo fruit fermentation liquid of Microbacterium A5, and continue fermenting to obtain liquid bacterial fertilizer; The liquid bacterial fertilizer is dried to obtain solid organic fertilizer; The No. 1 pomelo fruit culture medium: add 250g of defective pomelo fruit to 500mL of LB culture medium and add water to make the volume to 1L; The No. 2 pomelo fruit culture medium: add 400 g of defective pomelo fruit to 200 mL of LB culture medium, and add water to make the volume to 1 L; The No. 3 pomelo fruit culture medium: 500 g of defective pomelo fruit, add water to make the volume to 1 L.

2. The method according to claim 1, wherein The deposit number of the Bacillus flexus HL-37 is GDMCC No. 60045, the deposit number of the Bacillus cereus TH-35 is GDMCC No. 60044, and the deposit number of the Microbacterium A5 is CCTCC NO: M209174.

3. The method according to claim 1, wherein The inoculation amount of Microbacterium A5 is 5-15%.

4. The method according to claim 1, wherein The inlet amount of the HL-37 fermentation broth and the TH-35 fermentation broth is both 10%.

5. A liquid bacterial fertilizer prepared according to the method according to any one of claims 1 to 4.

6. a solid organic fertilizer prepared according to the method described in any one of claims 1 to 4.

7. Use of the liquid bacterial fertilizer as claimed in claim 5 in promoting the growth of pomelo.

8. The use according to claim 7, characterized in that The promoting of pomelo growth includes increasing the weight and / or sugar content of a single pomelo fruit.

9. Use of the solid organic fertilizer as claimed in claim 6 in promoting the growth of vegetables.

10. The use according to claim 9, characterized in that The promoting of vegetable growth includes increasing the fresh weight and / or plant height of the vegetables; The vegetables include amaranth.

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