Gluconobacter oxydans, application thereof and method for rapidly fermenting apple enzyme fertilizer

Abstract

The invention belongs to the technical field of microorganisms, and relates to gluconobacter oxydans and application thereof and a method for rapidly fermenting an apple enzyme fertilizer, the strain is preserved in China General Microbiological Culture Collection Center (CGMCC) on July 16, 2024, and the preservation number is CGMCC No.31321; the preservation address is No.3, Yard 1, Beichen West Road, Chaoyang District, Beijing. The gluconobacter oxydans C2 strain has good environmental adaptability and can be used for quickly fermenting apple pulp. The gluconobacter oxydans C2 strain provided by the invention can be used for preparing an apple solid enzyme fertilizer independently or together with a combined microbial agent, can promote the growth of crops, and has a good application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and in particular relates to a strain of Gluconobacter oxidans and an application thereof and a method for quickly fermenting apple enzyme fertilizer. Background Art

[0002] In recent years, with the strong support of the government, the apple industry has developed rapidly. However, due to the single variety structure and the overly concentrated maturity period, the storage and sales pressure of fruits after harvest is relatively high. In addition, the market sales of middle and lower fruits with poor appearance are poor. Improper storage or random discarding can easily breed various fungi, leading to environmental deterioration. This vicious cycle has seriously affected the development of the industry. To solve these problems, deep processing of middle and lower fruits is an effective method. By producing products such as juice, jam, dried apples, and apple cider vinegar, not only the added value of the products is increased, but also the waste of middle and lower fruits can be greatly reduced. At the same time, environmentally friendly treatment of apples that are improperly stored or have no sales market, such as making compost, feed, or preparing enzyme fertilizers through biotechnology treatment, is also a good solution.

[0003] The preparation process of apple enzyme fertilizer is to use apples as organic matter and prepare enzyme fertilizer through fermentation. Enzyme fertilizer is rich in various organic nutrients, which can improve soil fertility, improve soil structure and promote crop growth. However, the fermentation time of apple enzyme fertilizer usually ranges from several weeks to months, and regular monitoring and stirring are required during this period, which not only increases management and labor costs, but also may bring pollution risks and operational difficulties. These shortcomings limit its use and need further research and improvement to improve production efficiency and market competitiveness. Summary of the invention

[0004] In view of the above technical problems, the present invention provides a strain of Gluconobacter oxidans and its application and a method for rapidly fermenting apple enzyme fertilizer. The strain can rapidly reduce the pH value and sugar content of apple pulp and is used for fermenting and preparing apple enzyme fertilizer. The prepared apple enzyme fertilizer can be used as base fertilizer and topdressing for field crops, fruit trees and vegetable crops, and has the effect of promoting crop yield increase.

[0005] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions:

[0006] A strain of Gluconobacter oxydans is C2, and its classification name is Gluconobacter oxydans (Gluconobacter oxydans). It was deposited in the General Microbiology Center of China Culture Collection Administration on July 16, 2024, and its deposit number is CGMCC No. 31321; the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0007] The Gluconobacter oxydans C2 provided by the present invention has a large amount of dehydrogenases connected to the respiratory chain on its cell membrane, and the catalytic activity center is located in the periplasmic space of the cell membrane, and the substrate to be oxidized can be oxidized without being transported to the inside of the cell, and the incomplete oxidation product will be released into the culture medium. Therefore, the strain has the ability of efficient oxidation, can quickly decompose the organic matter in the substrate waiting for oxidation of apple pulp, produce abundant organic acids and other beneficial ingredients, and can significantly reduce the pH value and sugar content of the substrate to be oxidized within 48h. It is used for the fermentation of apple enzyme fertilizer, and the fermentation speed can be greatly accelerated, which is suitable for fermenting and preparing apple enzyme fertilizer.

[0008] Its 16S rDNA sequence (as shown in SEQ ID No.1) is as follows:

[0009]

[0010] In a second aspect, the present invention further provides a bacterial agent comprising the above-mentioned Gluconobacter oxydans C2 and a biologically acceptable microbial agent carrier and / or auxiliary material.

[0011] The Gluconobacter oxydans C2 provided by the present invention can be prepared into bacterial agents of different dosage forms by selecting suitable carriers and / or excipients to improve the stability, activity and use effect of the bacterial agent to meet the needs of different production. The dosage form of the bacterial agent includes but is not limited to liquid, powder, granule, tablet and capsule; the excipients include but are not limited to glycerol, polyethylene glycol, starch, glucose, maltodextrin, sucrose, lactose, skim milk powder, gum arabic and chitosan; the carrier includes but is not limited to culture medium, water, starch, cellulose and gelatin.

[0012] Exemplarily, a liquid bacterial agent of Gluconobacter oxidans C2 is provided, which is composed of Gluconobacter oxidans C2 and MRS liquid culture medium, and the preparation method is as follows:

[0013] The activated Gluconobacter oxidans C2 was inoculated into MRS liquid culture medium for expansion culture until the early stage of the stable phase, thereby obtaining the Gluconobacter oxidans C2 liquid bacterial agent.

[0014] Exemplarily, a Gluconobacter oxidans C2 bacterial powder is provided, which is composed of Gluconobacter oxidans C2, soluble starch, oligoxylose, and skimmed milk powder, and the preparation method is as follows:

[0015] Preparation of freeze-dried protective agent: Take soluble starch, oligoxylose and skimmed milk powder, dissolve the three in water, mix well, and make the mass percentage concentration of soluble starch in the obtained mixed solution be 5%-15%, the mass percentage concentration of oligoxylose is 5%-20%, and the mass percentage concentration of skimmed milk powder is 6%-20%.

[0016] The activated Gluconobacter oxidans C2 is inoculated into an MRS liquid culture medium and cultured until the beginning of the stable period, and the bacteria are obtained by centrifugation; the bacteria are evenly mixed with a freeze-drying protective agent to prepare a bacterial suspension, and the volume of the mixed solution is adjusted so that the bacterial concentration reaches an appropriate concentration, and the Gluconobacter oxidans C2 bacterial powder is prepared through a freeze-drying process.

[0017] Furthermore, the bacterial agent also contains one or more other microorganisms.

[0018] The Gluconobacter oxydans C2 provided by the present invention can be used in combination with other microorganisms to optimize the fermentation process of apple enzyme fertilizer by utilizing the metabolic characteristics and advantages of different strains. For example, lactic acid bacteria, Bacillus subtilis, yeast, etc. can be selected to form a composite microbial agent with Gluconobacter oxydans C2.

[0019] Furthermore, the microorganism is a soil microorganism.

[0020] Soil microorganisms are bacteria and fungi in the soil that can improve soil properties, repair soil, and have growth-promoting properties for plants. Common ones include symbiotic nitrogen-fixing bacteria, mycorrhizal fungi, and plant rhizosphere growth-promoting bacteria. The oxidizing gluconobacter C2 provided by the present invention and soil microorganisms can significantly improve the fermentation efficiency and fertilizer quality of apple enzyme fertilizers through scientific and reasonable combinations, and optimize the growth environment and production benefits of crops. For example, one or more soil microorganisms such as actinomycetes, Bacillus subtilis, Bacillus licheniformis, and Bacillus amyloliquefaciens can be selected to form a composite microbial agent with oxidizing gluconobacter C2.

[0021] In a third aspect, the present invention also provides the use of the above-mentioned Gluconobacter oxydans C2 and the above-mentioned bacterial agent in the preparation of apple fermentation products.

[0022] Specifically, the Gluconobacter oxydans C2 and its bacterial agent provided by the present invention can be used to prepare apple cider vinegar. Apple juice is fermented to produce apple cider through primary fermentation, and then secondary fermentation is carried out by Gluconobacter oxydans C2 to convert ethanol into acetic acid, and finally produce apple cider vinegar.

[0023] The Gluconobacter oxidans C2 and its bacterial agent provided by the present invention can also be used to prepare apple enzyme. In the preparation of apple enzyme, the Gluconobacter oxidans C2 can convert sugars and other organic substances in apples into a variety of organic acids, enzymes and other metabolites, thereby increasing the biological activity and nutritional value of the apple enzyme and improving its flavor and taste.

[0024] Furthermore, the application is application in the preparation of apple enzyme fertilizer.

[0025] The Gluconobacter oxidans C2 and its bacterial agent provided by the present invention can be used to prepare apple enzyme fertilizer. Gluconobacter oxidans C2 can be used for fermentation to quickly oxidize sugars in apples, generate rich organic acids, and convert apples and their byproducts into high-quality organic fertilizers.

[0026] In a fourth aspect, the present invention further provides an apple enzyme fertilizer, which is obtained by fermenting apples or apple by-products as fermentation substrates with the above-mentioned Gluconobacter oxidans C2 or a microbial combination comprising the above-mentioned Gluconobacter oxidans C2.

[0027] The apple by-products are secondary products produced during the apple processing process or the unused parts during the processing process. Apple by-products include but are not limited to the following categories: apple pomace, apple peel, apple juice pomace and apple waste liquid.

[0028] The apples or apple by-products may be processed, chopped or crushed to maximize the surface area for utilization by Gluconobacter oxidans C2.

[0029] There are many ways to inoculate Gluconobacter oxidans C2 into the fermentation matrix using apples or apple byproducts. The specific inoculation method can be determined according to the production equipment, process requirements and target product characteristics. Optionally, direct inoculation: the cultured Gluconobacter oxidans C2 bacterial solution is directly added to the fermentation matrix; adding fermentation agents: the prepared Gluconobacter oxidans C2 bacterial powder, microcapsules or other forms of bacterial agents are added to the fermentation matrix.

[0030] In the process of preparing apple enzyme fertilizer, in addition to the main raw material apple or apple by-products, some auxiliary materials can also be selected to optimize the fermentation process and improve the quality of apple enzyme fertilizer. For example, sugars such as brown sugar and honey promote the growth of microorganisms; minerals such as magnesium sulfate and zinc sulfate promote the growth and metabolism of microorganisms and improve enzyme activity; pH ​​regulators such as citric acid and calcium carbonate maintain the pH value of the fermentation environment; stabilizers such as gelatin and guar gum increase the viscosity of the fermentation liquid and prevent stratification; nutritional supplements such as amino acids and vitamins are helpful for the reproduction of microorganisms.

[0031] Furthermore, the apple enzyme fertilizer is an apple liquid enzyme fertilizer, comprising the following raw materials:

[0032] Apple pulp, apples or apple by-products and water in a mass ratio of 2-5:5-8;

[0033] Sugars, the mass of which is 1% to 5% of the mass of the apple pulp;

[0034] The fermentation agent is the above-mentioned Gluconobacter oxidans C2 or a microbial combination comprising the above-mentioned Gluconobacter oxidans C2; based on the volume of apple pulp, the accession amount of the Gluconobacter oxidans C2 is 1×10 8 -1×10 9 CFU / L.

[0035] The preparation of apple liquid enzyme fertilizer can be optimized by controlling the amount of water added and adjusting the concentration of apple pulp; preferably, the mass ratio of apples or apple by-products to water is 3:7.

[0036] As the main carbon source of microorganisms, the sugar can promote the growth and metabolic activity of fermentation bacteria and help the fermentation process. In the process of preparing apple enzyme fertilizer, brown sugar is one of the common added sugars, which can not only reduce the preparation cost, but also improve the nutritional value and fertilizer efficiency of apple enzyme fertilizer. Preferably, the mass of brown sugar is 1%-5% of the mass of the apple pulp.

[0037] Furthermore, the apple enzyme fertilizer is an apple solid enzyme fertilizer, characterized in that it comprises the following raw materials:

[0038] A fermentation substrate comprising apples or apple byproducts and agricultural byproducts in a mass ratio of 2-4:1;

[0039] Sugars, the mass of which is 1%-5% of the mass of the fermentation substrate;

[0040] The fermentation agent is the above-mentioned Gluconobacter oxidans C2 or a microbial combination comprising the above-mentioned Gluconobacter oxidans C2; based on the volume of the fermentation substrate, the access amount of the Gluconobacter oxidans C2 is 1×10 9 -1×10 10 CFU / L.

[0041] In the present invention, agricultural and sideline products refer to secondary products or ancillary products produced in the agricultural production process in addition to the main agricultural products (such as grains, vegetables, fruits, etc.), including but not limited to rice bran, soybean meal, straw, bean dregs, bran, corn cobs, vegetable waste and sawdust. Preferably, the fermentation substrate includes apples or apple byproducts and bran in a mass ratio of 2-4:1.

[0042] The preferred mass of brown sugar for preparing the above-mentioned apple solid enzyme fertilizer is 1%-5% of the mass of the fermentation substrate.

[0043] In a fifth aspect, the present invention also provides a method for preparing liquid enzyme fertilizer by rapid fermentation using apple, comprising the following steps:

[0044] Step 1: crushing apples or apple byproducts into apple puree and mixing with tap water, with the mass ratio of apple puree to tap water being 2-5:5-8, to obtain apple pulp;

[0045] Step 2: adding 1% to 5% brown sugar to the apple pulp obtained in step 1, and mixing well to obtain an apple fermentation substrate;

[0046] Step 3: Inoculate the above-mentioned live bacteria of Gluconobacter oxidans C2 into the apple fermentation substrate obtained in step 2. The inoculation amount of the live bacteria of Gluconobacter oxidans C2 is 1×10 8 -1×10 9 CFU / L, 30-40℃, 100-300r / min, culture until the concentration of Gluconobacter C2 in the fermentation substrate reaches 1×10 8 -1×10 9 CFU / g, and apple fermentation liquid was obtained;

[0047] Step 4: Take the apple fermentation liquid obtained in step 3 and incubate it at 30-40° C. until the pH is less than or equal to 3 and the sugar content is less than or equal to 3, then terminate the fermentation to obtain apple liquid enzyme fertilizer.

[0048] The apple liquid enzyme fertilizer in step 4 can be used directly, or can be used as liquid fertilizer after filtering to remove solid residue, or can be further processed into solid fertilizer.

[0049] In step 3, the oxidized gluconic acid rod C2 was inoculated into the apple fermentation substrate and cultured to a bacterial cell concentration of 1×10 8 -1×10 9 CFU / g is used as a standard to control the culture time. At this time, the bacteria are in a rapid growth stage and can consume a lot of sugar. Generally, at 35℃, 150-200r / min, and culture for 24h, the concentration of Gluconobacter C2 in the fermentation substrate can reach 1×10 8 -5×10 8 CFU / g.

[0050] In step 4, the pH of the apple fermentation liquid is less than or equal to 3, and the sugar content is less than or equal to 3 as the fermentation endpoint judgment standard. Generally, after culturing at 35° C. for 24 hours, the pH of the apple fermentation liquid is less than 3, and the sugar content is less than 3.

[0051] In a sixth aspect, the present invention also provides a rapid fermentation method using apple solid enzyme fertilizer, comprising the following operations:

[0052] Step 1: crushing apples or apple by-products into apple puree and mixing with bran, wherein the mass ratio of apple puree to bran is 2-4:1, adding 1%-5% brown sugar, and mixing well to obtain a solid fermentation base;

[0053] Step 2: Inoculate the above-mentioned Gluconobacter oxidans C2 into the solid-state fermentation base. Based on the volume of the solid-state fermentation base, the inoculation amount of the Gluconobacter oxidans C2 is 1×10 9 -1×10 10 CFU / L, after mixing, closed fermentation was carried out at 30-40°C, and cultured until the pH was less than or equal to 4 to obtain apple fermentation solid;

[0054] Step 3: The apple fermentation solid obtained in step 2 is openly cultured at 30-40° C. and statically cultured until the pH is greater than or equal to 6.9 and the sugar content is less than or equal to 1.5, and the fermentation is terminated to obtain apple solid enzyme fertilizer.

[0055] In step 2, oxidized gluconate rod C2 is inoculated into the solid fermentation medium and cultured until the pH of the solid fermentation medium is less than or equal to 4, and this is used as a standard to control the culture time. Generally, at 35°C, the pH of the solid fermentation medium can reach less than 4 when the closed fermentation is cultured for 30 days.

[0056] In step 3, the pH of the apple fermentation solid is greater than or equal to 6.9, and the sugar content is less than or equal to 1.5, as the fermentation endpoint judgment standard. Generally, after static culture at 35°C for 30 days, the pH of the apple fermentation solid can reach greater than 6.9 and the sugar content can reach less than 1.5.

[0057] In the process of preparing apple enzyme fertilizer, finished microbial preparations or enzyme preparations can be added as fermentation promoters to promote the decomposition of organic matter, improve fermentation efficiency, and improve the nutritional content and fertilizer efficiency of fertilizers. Preferably, in step 2, Gluconobacter oxidans C2 and composite probiotics can be introduced into the solid fermentation base; the composite probiotics can be 0.1%-1% of Gendun No. 1.

[0058] The Root Shield No. 1 is a product of Hebei Mumeituli Technology Co., Ltd. and contains ≥ 200 million Bacillus licheniformis / mL.

[0059] In a seventh aspect, the present invention also provides a method for preparing apple solid enzyme fertilizer, comprising the following operations:

[0060] Step 1: crushing apples or apple by-products into apple puree and mixing with bran, wherein the mass ratio of apple puree to bran is 2-4:1, adding 1%-5% brown sugar, and mixing well to obtain a solid fermentation base;

[0061] Step 2: Inoculate the above-mentioned Gluconobacter oxidans C2 into the solid-state fermentation base. Based on the volume of the solid-state fermentation base, the inoculation amount of the Gluconobacter oxidans C2 is 1×10 9 -1×10 10 Pile evenly, cover and seal, turn the pile once when the temperature is higher than 40°C, perform anaerobically fermentation until the pH is greater than or equal to 6.9 and the sugar content is less than or equal to 1.0, remove the film, spread out and dry in the shade until the moisture content is less than 20%, and obtain apple solid enzyme fertilizer.

[0062] In step 2, the solid fermentation base pH is greater than or equal to 6.9 and the sugar content is less than or equal to 1.0 as the fermentation endpoint judgment standard. Generally, after anaerobic fermentation at 35°C for 90 days, the solid fermentation base can reach a pH greater than 6.9 and a sugar content less than 1.0.

[0063] Preferably, in step 2, Gluconobacter oxidans C2 and compound probiotics can be introduced into the solid-state fermentation base, and the compound probiotics can be prepared by using 0.1%-1% of the root shield combined microbial agent.

[0064] The Root Shield combination microbial agent is a product of Hebei Mumei Tuli Technology Co., Ltd. It is composed of two finished products, Root Shield No. 1 and Root Shield No. 2, mixed in a 1:1 ratio, and contains a variety of soil microorganisms, including Bacillus licheniformis, Bacillus amyloliquefaciens, etc.; the Root Shield No. 2 contains Bacillus amyloliquefaciens ≥ 200 million / mL.

[0065] In an eighth aspect, the present invention also provides the application of the apple enzyme fertilizer and the apple enzyme fertilizer produced by the above fermentation method in promoting crop yield increase, wherein the apple enzyme fertilizer is used as basal fertilizer and / or topdressing for field crops, fruit trees or vegetable crops. BRIEF DESCRIPTION OF THE DRAWINGS

[0066] Figure 1 The morphological characteristics of Gluconobacter oxydans C2; A is the plate image of colony morphology; B is the microscopic morphology image;

[0067] Figure 2 is the phylogenetic tree of Gluconobacter oxydans C2;

[0068] Figure 3 The growth conditions of Gluconobacter oxidans C2 in Example 1 at different pH values. DETAILED DESCRIPTION

[0069] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0070] The strains used in the following examples of the present invention are derived from the following:

[0071] The Gluconobacter oxydans C2 strain was deposited in the General Microbiology Center of the China Culture Collection Administration on July 16, 2024, and its deposit number is CGMCC No. 31321; the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0072] The culture medium used in the following examples of the present invention is as follows:

[0073] MRS medium: 5g beef extract, 20g glucose, 5g soy peptone, 5g yeast extract, 20g lactose, 10g calcium carbonate, 20g agar, 1L water. Add 15g agar powder to the solid medium, add the above ingredients to distilled water, heat to dissolve, adjust the pH to 6.0, and sterilize at 121℃ for 20 minutes after packaging.

[0074] The sources of the bacterial agents used in the following embodiments of the present invention are as follows:

[0075] Gendun No. 1 and Gendun No. 2 were purchased from Hebei Mumei Tuli Technology Co., Ltd.

[0076] Unless otherwise specified, other experimental materials and instruments can be purchased through commercial channels.

[0077] Example 1

[0078] This embodiment provides a strain of Gluconobacter oxydans C2, which is isolated from crushed apple puree.

[0079] 1. Isolation and screening of Gluconobacter oxydans C2

[0080] Take 1-2g of crushed applesauce and inoculate it into 0.9% sterile saline after sterilization and mix it thoroughly. Dilute the sample 10 times with sterile saline to form different concentration gradients from 10-1 to 10-8. Take 200μL of each concentration and spread it evenly on the MRS agar plate. Repeat three times for each concentration. The coated culture medium is inverted and cultured in a constant temperature incubator at 35°C for 48h. The acetic acid bacteria colonies are identified according to their morphology and color characteristics. Single colonies that can produce transparent circle morphology characteristics on the plate are selected, and the plate streaking method is used for further separation and purification. After streaking and purification 2-3 times, a single strain is obtained, and a strain with acid-producing ability is isolated and obtained, named C2, which is stored in a refrigerator at 4°C.

[0081] 2. Identification of Gluconobacter oxydans C2 strain

[0082] 2.1 Colony and bacterial morphology of strain C2

[0083] Colony morphology of the strain: milky white, round, with neat edges, smooth surface, and slightly raised. Figure 1 -A; Microscopic morphology of bacteria: obligate aerobic Gram-negative bacteria, short rod-shaped, spore-forming, see Figure 1 -B.

[0084] 2.2 Sequencing analysis of 16S rDNA of strain C2

[0085] The strains obtained by screening were streaked onto MRS medium and cultured at 35°C for 2 days. The cells were collected and sent to Beijing Liuhe BGI Gene Technology Co., Ltd. for 16S rDNA sequencing. The 16S rDNA sequence of strain C2 was compared on the EzBioCloud website, and the result was Gluconobacter oxydans, with a homology of 99.72%. The phylogenetic tree was constructed using MEGA software, based on the 16S rDNA sequence, using the Neighbour-Joining method, and the Btoostrap value was 1000. The phylogenetic tree was constructed as shown in the figure. Figure 2 It can be seen that strain C2 has the highest affinity with Gluconobacter oxydans.

[0086] In summary, combined with 16S rDNA sequence analysis and physiological and biochemical identification results, C2 was identified as a Gluconobacter oxydans strain and named Gluconobacter oxydans C2.

[0087] The Gluconobacter oxydans C2 strain was deposited in the General Microbiology Center of China Culture Collection Administration on July 16, 2024, and its deposit number is CGMCC No. 3121; the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0088] 3. Determination of salt resistance, acid and alkali resistance and temperature resistance of Gluconobacter oxydans C2 strain

[0089] 3.1 Salt tolerance test

[0090] Gluconobacter oxydans C2 was inoculated into 100 mL of MRS liquid culture medium and cultured in a shaker at 35°C and 200 rpm for 2 days. 100 μL was spread on MRS plates containing different salt concentrations (0-14%), with 3 parallel plates in each group. The plates were placed in a 35°C incubator and cultured for 7 days. The growth was observed and recorded every day. The results are shown in Table 1.

[0091] Table 1 Salt tolerance of Gluconobacter oxidans C2

[0092] Salt concentration 0 0.5％ 1％ 2％ 3％ 4％ 5％ 6％ Growth +++ +++ +++ ++++ +++ +++ +++ +++ Salt concentration 7％ 8％ 9％ 10％ 11％ 12％ 13％ 14％ Growth +++ +++ +++ ++ + + - -

[0093] Gluconobacter oxydans C2 grew vigorously on MRS plates with a salt concentration of 2% and could tolerate a salt concentration of 12%.

[0094] 3.2 Acid and alkali resistance test

[0095] Gluconobacter oxydans C2 was inoculated into 100 mL MRS liquid culture medium and cultured in a shaker at 35°C and 200 rpm for 12 h. 1% of the culture medium was inoculated into MRS liquid culture medium with different pH values ​​(1-11) and 3 replicates were placed in a shaker at 35°C and 200 rpm for 1 day. The OD values ​​were measured. 600nm The absorbance value of Figure 3 .

[0096] like Figure 3 As shown, Gluconobacter oxydans C2 can grow in the range of pH = 2-10, with the optimum pH being 6.0, and grows well under weak alkaline conditions (OD 600 nm =0.88).

[0097] 3.3 Temperature resistance test

[0098] Gluconobacter oxydans C2 was inoculated into 100 mL of MRS liquid culture medium, and cultured in a shaker at 35°C and 200 rpm for 12 h. Then, 1% of the culture medium was inoculated into MRS liquid culture medium at 15°C, 20°C, 25°C, 30°C, 35°C, 40°C, 45°C, 50°C, 55°C, and 60°C, and cultured at 200 rpm for 1 day. The OD 600 nm The results are shown in Table 2.

[0099] Table 2 Growth temperature of Gluconobacter oxidans C2

[0100] temperature 15℃ 20℃ 25℃ 30℃ 35℃ 40℃ 45℃ 50℃ 55℃ 60℃ <![CDATA[OD 600 ]]> 0 0.347 0.678 0.989 1.146 1.052 0.665 0.332 0.087 0

[0101] The growth temperature range of Gluconobacter oxydans C2 is 20-55℃, and the optimal growth temperature is 35℃.

[0102] Example 2

[0103] This embodiment provides a Gluconobacter oxydans C2 liquid bacterial agent, which is composed of Gluconobacter oxydans C2 and MRS liquid culture medium, and is specifically prepared by the following steps:

[0104] (1) Streak Gluconobacter oxydans C2 onto MRS plates and culture at 35°C for 2 days.

[0105] (2) A single colony of Gluconobacter oxydans C2 on the MRS plate was selected and inoculated into MRS liquid culture medium and cultured at 35°C and 200 rpm for 12 h.

[0106] (3) After the seed culture was completed, the inoculum size was 1% and transferred into MRS liquid culture medium and cultured at 35° C. and 200 rpm for 12 h to obtain the liquid bacterial agent of Gluconobacter oxidans C2.

[0107] The test showed that the number of viable bacteria in the inoculant was 2.1-2.6×10 8 CFU / mL.

[0108] Example 3

[0109] The present embodiment provides a Gluconobacter oxydans C2 bacterial powder, which is composed of Gluconobacter oxydans C2, soluble starch, oligoxylose and skimmed milk powder, and is specifically prepared by the following steps:

[0110] (1) Soluble starch, xylooligosaccharides and skimmed milk powder are dissolved in water and mixed to make the mass percentage concentration of soluble starch, xylooligosaccharides and skimmed milk powder in the obtained mixed solution 10%, 14% and 12% respectively.

[0111] (2) Streak Gluconobacter oxydans C2 onto an MRS plate and culture at 35°C for 2 days.

[0112] (3) A single colony of Gluconobacter oxydans C2 on the MRS plate was selected and inoculated into MRS liquid culture medium and cultured at 35°C and 200 rpm for 12 h.

[0113] (4) After the seed culture was completed, the inoculum was transferred into MRS liquid culture medium at 1% and cultured at 35° C. and 200 rpm for 12 h to obtain the culture medium of Gluconobacter oxidans C2.

[0114] (5) The culture solution of Gluconobacter oxydans C2 was centrifuged at 6000 rpm / min for 10 min at 4°C to collect bacterial sludge, which was then washed twice with 0.85% saline. The obtained bacterial sludge was added to a protective agent to prepare a bacterial suspension, and the volume was adjusted so that the bacterial concentration reached 1×10 9 -1×10 10 CFU / mL, and prepared into Gluconobacter oxydans C2 powder by freeze-drying process. The freeze-drying parameters were -80℃, vacuum degree 1.0pa, and freezing time 24h.

[0115] The test showed that the number of viable bacteria in the inoculant was 3.5-6.5×10 7 CFU / g.

[0116] Example 4

[0117] This embodiment provides an apple liquid enzyme fertilizer and a method for preparing the apple liquid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0118] Step 1: Crush apples into applesauce and mix with tap water in a mass ratio of applesauce to tap water of 2:8 to obtain apple pulp.

[0119] Step 2: Add 5% brown sugar to the apple pulp and mix well to obtain an apple fermentation substrate.

[0120] Step 3: 200 mL of apple fermentation substrate was inoculated with live Gluconobacter oxidans C2 bacteria. The inoculation amount of the Gluconobacter oxidans C2 bacteria was 1×10 8 CFU / L, 35℃, 150r / min, culture for 24h to obtain apple fermentation liquid.

[0121] Step 4: Incubate the apple fermentation liquid at 35°C for 24 hours to obtain apple liquid enzyme fertilizer.

[0122] Example 5

[0123] This embodiment provides an apple liquid enzyme fertilizer and a method for preparing the apple liquid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0124] Step 1: Crush apples into applesauce and mix with tap water in a mass ratio of 3:7 to obtain apple pulp.

[0125] Step 2: Add 2% brown sugar to the apple pulp and mix well to obtain an apple fermentation substrate.

[0126] Step 3: 1‰ of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2 was inoculated into 200 mL of apple fermentation substrate. Based on the volume of apple pulp, the inoculation amount of Gluconobacter oxidans C2 was about 2.1-2.6×10 8 CFU / L, 35℃, 150r / min, culture for 24h to obtain apple fermentation liquid.

[0127] Step 4: Incubate the apple fermentation liquid at 35°C for 24 hours to obtain apple liquid enzyme fertilizer.

[0128] Example 6

[0129] This embodiment provides an apple liquid enzyme fertilizer and a method for preparing the apple liquid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0130] Step 1: Crush the apples into applesauce and mix with tap water. The mass ratio of applesauce to tap water is 5:5.

[0131] Step 2: Add 1% brown sugar to the apple pulp and mix well to obtain an apple fermentation substrate.

[0132] Step 3: 200 mL of apple fermentation substrate was inoculated with the bacterial powder of Gluconobacter oxidans C2 prepared in Example 3, and the amount of bacterial powder added was adjusted so that the inoculated amount of Gluconobacter oxidans C2, based on the volume of apple pulp, was 1×10 9 CFU / L, 35℃, 150r / min, culture for 24h to obtain apple fermentation liquid.

[0133] Step 4: Take the apple fermentation liquid, incubate it at 35°C for 24 hours to obtain apple liquid enzyme fertilizer.

[0134] Comparative Example 1

[0135] Comparative Example 1 provides an apple liquid enzyme fertilizer. The mass ratio of applesauce to tap water and the amount of brown sugar added are the same as those in Experimental Example 5, except that the Gluconobacter oxidans C2 strain is not added and the fertilizer is naturally fermented for 48 hours.

[0136] Verification Example 1

[0137] The changes in sugar content and pH value of apple pulp liquid before and after fermentation were measured in Comparative Example 1 and Example 5, with three replicates in each group. The results are shown in the following table.

[0138] Table 3 Gluconobacter oxidans C2 liquid fermentation effect

[0139]

[0140] It can be seen from Table 3 that the pH and sugar content of apple pulp liquid decreased slightly under natural fermentation; but the fermentation results after adding Staphylococcus oxidans were very obvious, with a significant reduction in pH and sugar content, and the cycle was extremely short, meeting the conditions for preparing apple liquid enzyme fertilizer.

[0141] Example 7

[0142] This embodiment provides an apple solid enzyme fertilizer and a method for preparing the apple solid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0143] Step 1: Crush apples into apple puree and mix with bran, with the mass ratio of apple puree to bran being 2:1, add 5% brown sugar, mix well, and obtain a solid fermentation base.

[0144] Step 2: 400 mL of solid-state fermentation medium was inoculated with Gluconobacter oxidans C2. Based on the volume of the solid-state fermentation medium, the inoculation amount of live bacteria of Gluconobacter oxidans C2 was 1×10 9 CFU / L, mixed and then closed fermentation was carried out at 35°C. After static culture for 30 days, apple fermentation solid was obtained.

[0145] Step 3: Take the apple fermentation solid obtained in step 3, culture it in an open environment, and culture it at 35° C. for 30 days to obtain apple solid enzyme fertilizer.

[0146] Example 8

[0147] This embodiment provides an apple solid enzyme fertilizer and a method for preparing the apple solid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0148] Step 1: Crush apples into apple puree and mix with bran, with the mass ratio of apple puree to bran being 4:1. Add 1% brown sugar and mix well to obtain a solid fermentation base.

[0149] Step 2: 400 mL of solid-state fermentation medium was inoculated with Gluconobacter oxidans C2. Based on the volume of the solid-state fermentation medium, the inoculation amount of live bacteria of Gluconobacter oxidans C2 was 1×10 10 CFU / L, mixed and then closed fermentation was carried out at 35°C. After static culture for 30 days, apple fermentation solid was obtained.

[0150] Step 3: Take the apple fermentation solid obtained in step 3, culture it in an open environment, and culture it at 35° C. for 30 days to obtain apple solid enzyme fertilizer.

[0151] Example 9

[0152] This embodiment provides an apple solid enzyme fertilizer and a method for preparing the apple solid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0153] Step 1: Crush apples into apple puree and mix with bran, with the mass ratio of apple puree to bran being 3:1. Add 1% brown sugar and mix well to obtain a solid fermentation base.

[0154] Step 2: 1% of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2 was inoculated into 400 mL of solid fermentation medium. Based on the volume of the solid fermentation medium, the inoculation amount of live bacteria of Gluconobacter oxidans C2 was 2.1-2.6×10 9 CFU / L; after mixing, closed fermentation was carried out at 35°C, and after static culture for 30 days, apple fermentation solid was obtained.

[0155] Step 3: Take the apple fermentation solid obtained in step 3, culture it in an open environment, and culture it at 35° C. for 30 days to obtain apple solid enzyme fertilizer.

[0156] Example 10

[0157] This embodiment provides an apple solid enzyme fertilizer and a method for preparing the apple solid enzyme fertilizer by rapid fermentation. The specific steps are as follows:

[0158] Step 1: Crush apples into apple puree and mix with bran, with the mass ratio of apple puree to bran being 3:1. Add 1% brown sugar and mix well to obtain a solid fermentation base.

[0159] Step 2: 1% of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2 and 1% of Gendun No. 1 were inoculated into 400 mL of solid fermentation medium. Based on the volume of the solid fermentation medium, the inoculation amount of live Gluconobacter oxidans C2 was 2.1-2.6×10 9 CFU / L; after mixing, closed fermentation was carried out at 35°C, and after static culture for 30 days, apple fermentation solid was obtained.

[0160] Step 3: Take the apple fermentation solid obtained in step 3, culture it in an open environment, and culture it at 35° C. for 30 days to obtain apple solid enzyme fertilizer.

[0161] Comparative Example 2

[0162] Comparative Example 2 provides an apple solid enzyme fertilizer. The mass ratio of applesauce to tap water and the amount of brown sugar added are the same as those in Experimental Example 9, except that the Gluconobacter oxidans C2 strain is not added and the fertilizer is naturally fermented for 60 days.

[0163] Verification Example 2

[0164] The changes in sugar content and pH value of apple pulp before and after solid fermentation were measured in Comparative Example 2 and Examples 9 and 10, with three replicates in each group. The results are shown in the following table.

[0165] Moisture determination: Take 20g of fermented fruit puree and place it on a flat plate. Dry it at 105℃ for 4h, then weigh it and calculate the moisture content.

[0166] Sugar and pH determination: Take 20g of fermented fruit puree and put it in a disposable plastic cup. Add 40mL of distilled water and stir evenly to determine the pH. After standing, remove the supernatant and determine the sugar content.

[0167] The experimental results are as follows: As can be seen from Table 4, the pH and sugar content of apple pulp under natural fermentation decreased at a low level; but the fermentation results after adding Staphylococcus oxidans were obvious, and the pH and sugar content were greatly reduced, meeting the conditions for preparing apple solid enzyme fertilizer.

[0168] When used as an auxiliary bacterial agent for simultaneous fermentation, the pH reduction was the most significant, and the sugar reduction was also better than that of the natural fermentation group. Therefore, it can also be used as another method for rapid fermentation of apple solid enzyme fertilizer. Both have good prospects.

[0169] Table 4 Solid fermentation effect of Gluconobacter oxidans C2

[0170]

[0171]

[0172] Embodiment 11

[0173] This embodiment provides an apple solid enzyme fertilizer, which is prepared by the following steps.

[0174] Step 1: Apples are crushed into apple puree and mixed with bran. The mass ratio of apple puree to bran is 2:1. The moisture content of the material is about 65%. 1% brown sugar is added and mixed to obtain a solid fermentation base.

[0175] Step 2: 10L of solid-state fermentation base was inoculated with 1% of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2. Based on the volume of the solid-state fermentation base, the inoculation amount of live bacteria of Gluconobacter oxidans C2 was 2.1-2.6×10 9 CFU / L, turn it over evenly and pile it into a 40 cm high and 100 cm wide pile, cover it with plastic film and cover it tightly to prevent air leakage, turn it over once when the temperature is higher than 40℃, and ferment it anaerobicly for 90 days. Open the plastic film, spread it out and dry it in the shade until the moisture content is less than 20%, and then get solid apple enzyme fertilizer.

[0176] After fermentation, the pH value rises from the initial 5.55 to 7.24, the sugar content drops from 4.9 to 0.9, and the moisture content drops from 65.5% to 16.8%. It has no unpleasant odor and can be used as solid apple enzyme fertilizer.

[0177] Example 12

[0178] This embodiment provides a solid enzyme fertilizer for apples. The specific steps are as follows:

[0179] Step 1: Apples are crushed into apple puree and mixed with bran. The mass ratio of apple puree to bran is 4:1. The moisture content of the material is about 65%. 1% brown sugar is added and mixed to obtain a solid fermentation base.

[0180] Step 2: 10L solid fermentation base was inoculated with 1% of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2 and 1% of the root shield combined microbial agent. Based on the volume of the solid fermentation base, the inoculation amount of the live bacteria of Gluconobacter oxidans C2 was 2.1-2.6×10 9 CFU / L, turn it over evenly and pile it into a 40 cm high and 100 cm wide pile, cover it with plastic film and cover it tightly to prevent air leakage, turn it over once when the temperature is higher than 40℃, and ferment it anaerobicly for 90 days. Open the plastic film, spread it out and dry it in the shade until the moisture content is less than 20%, and then get solid apple enzyme fertilizer.

[0181] After fermentation, the pH rises from the initial 5.47 to 7.38, the sugar content drops from 4.8 to 0.7, and the water content drops from 63.24% to 15.56%. It has no unpleasant odor and can be used as solid apple enzyme fertilizer.

[0182] Example 13

[0183] This embodiment provides a solid enzyme fertilizer for apples. The specific steps are as follows:

[0184] Step 1: Apples are crushed into apple puree and mixed with bran. The mass ratio of apple puree to bran is 4:1. The moisture content of the material is about 65%. 1% brown sugar is added and mixed to obtain a solid fermentation base.

[0185] Step 2: 10L of solid-state fermentation base was inoculated with 1% of the liquid bacterial agent of Gluconobacter oxidans C2 prepared in Example 2 and 0.1% of the root shield combined microbial agent. Based on the volume of the solid-state fermentation base, the inoculation amount of live bacteria of Gluconobacter oxidans C2 was 2.1-2.6×10 9 CFU / L, turn it over evenly and pile it into a 40 cm high and 100 cm wide pile, cover it with plastic film and cover it tightly to prevent air leakage, turn it over once when the temperature is higher than 40℃, and ferment it anaerobicly for 90 days. Open the plastic film, spread it out and dry it in the shade until the moisture content is less than 20%, and then get solid apple enzyme fertilizer.

[0186] After fermentation, the pH value rises from the initial 5.57 to 7.34, the sugar content drops from 4.9 to 0.8, and the moisture content drops from 64.21% to 17.36%. It has no unpleasant odor and can be used as solid apple enzyme fertilizer.

[0187] Embodiment 14

[0188] This embodiment provides the use of apple liquid enzyme fertilizer prepared by rapid fermentation of Gluconobacter oxydans C2 in increasing crop yield.

[0189] The apple liquid enzyme fertilizer prepared in Example 5 was used as fertilizer to fertilize tomatoes. Field tests were conducted to verify its yield-increasing effect.

[0190] The experiment was conducted in a greenhouse using a completely randomized block design. Five treatment groups were set up for tomato (Yuansheng), with three replicates in each group. The area of ​​each block was 4.20m×7.00m=29.4m 2 , large row 80cm, small row 60cm, plant spacing 40cm for tomato planting. The fields of the five treatment groups used the same base bed, and the same person carried out field management and pest control according to conventional methods; treatment groups 1-4 were all fertilized conventionally, among which treatment groups 2-4 were fertilized with apple liquid enzyme fertilizer 2.5kg / mu, 5kg / mu, and 10kg / mu respectively at the time of planting on the basis of conventional fertilization; treatment group 5 was used as a blank control and only field management and pest control were carried out.

[0191] Conventional fertilization method: Before transplanting, apply diammonium phosphate at 45kg / mu in the furrows; after the seedlings have grown, apply diammonium phosphate at 7kg / mu in the furrows; when the ears and fruits begin to swell, apply top dressing in combination with watering, 12kg / mu of compound fertilizer (16-6-22), and stop top dressing 30 days before pulling out the seedlings.

[0192] The yield of each plot was counted, and the data were analyzed using Duncan's new multiple range test method of DPS software. The results are shown in the following table.

[0193] Table 5 Tomato plot yield statistics (yield unit: kg)

[0194]

[0195]

[0196] Table 6 Statistics of soluble solids in tomatoes

[0197]

[0198] Table 7 Statistics of ascorbic acid in tomatoes

[0199]

[0200] Table 8 Statistics of tomato lycopene

[0201]

[0202] Note: In the columns corresponding to the P values ​​of the significant differences in Tables 5-8, there is no significant difference between the groups with the same letter, and there is a significant difference between the groups with different letters.

[0203] As can be seen from the table above, treatment 3 (conventional fertilization + the above enzyme fertilizer 2.5L / mu) is significantly better than other treatments, with an increase of 11.92% compared with conventional fertilization (treatment 1) and 20.29% compared with the blank control (treatment 5). Soluble solids increased by 28.20% compared with the blank control, ascorbic acid increased by 35.45% compared with the blank control, and lycopene increased by 29.72% compared with the blank control. In addition, the tomato fruits of treatment 3 had more uniform color change during the same period compared with other treatments, and the fruits were larger and more solid.

[0204] Embodiment 15

[0205] This embodiment provides the use of apple solid enzyme fertilizer prepared by fermentation with Gluconobacter oxydans C2 in increasing crop yield.

[0206] The apple solid enzyme fertilizer prepared in Example 13 was used as fertilizer to fertilize tomatoes. Field tests were conducted to verify its yield-increasing effect. The verification method was the same as that in Example 14.

[0207] Table 9 Tomato plot yield statistics (yield unit: kg)

[0208]

[0209] Table 10 Statistics of soluble solids in tomatoes

[0210]

[0211] Table 11 Statistics of ascorbic acid in tomatoes

[0212]

[0213] Table 12 Statistics of tomato lycopene

[0214]

[0215] Note: In the columns corresponding to the P values ​​of the significant differences in Tables 9-12, there is no significant difference between the groups with the same letter, but there is a significant difference between the groups with different letters.

[0216] As can be seen from the table above, treatment 3 (conventional fertilization + the above enzyme fertilizer 5kg / mu) is significantly better than other treatments, with a yield increase of 9.30% compared with conventional fertilization (treatment 1) and a yield increase of 28.58% compared with the blank control (treatment 5). Soluble solids increased by 38.02% compared with the blank control, ascorbic acid increased by 28.58% compared with the blank control, and lycopene increased by 40.20% compared with the blank control. In addition, the tomato fruits of treatment 3 had more uniform color change during the same period compared with other treatments, and the fruits were larger and more solid.

[0217] In summary, the Gluconobacter oxydans C2 strain provided by the present invention has good adaptability to the environment, has strong salt resistance, acid and alkali resistance, and temperature resistance, can quickly ferment apple pulp, significantly reduce the pH value and sugar content of apple pulp in a short time, and is suitable for fermenting and preparing apple enzyme fertilizer. At the same time, the apple enzyme fertilizer prepared by the Gluconobacter oxydans C2 strain provided by the present invention has the effect of promoting the increase of crop yield, can be used as base fertilizer and topdressing for field crops, fruit trees and vegetable crops, and has good application prospects.

[0218] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modification, equivalent substitution or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A strain of Gluconobacter oxydans C2, characterized in that: Its classification name is Gluconobacter oxydans. It was deposited in the General Microbiology Center of China Culture Collection on July 16, 2024, and its deposit number is CGMCC No.31321. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

2. A bacterial agent, characterized in that The invention comprises the Gluconobacter oxidans C2 as claimed in claim 1 and a biologically acceptable microbial agent carrier and / or auxiliary material.

3. The bacterial agent according to claim 2, characterized in that The bacterial agent further comprises one or more other microorganisms; preferably, the microorganisms are soil microorganisms.

4. Use of the Gluconobacter oxydans C2 described in claim 1 and the bacterial agent described in claims 2-3 in the preparation of apple fermentation products; preferably, use in the preparation of apple enzyme fertilizer.

5. An apple enzyme fertilizer, characterized in that: The apple enzyme fertilizer is prepared by fermenting apples or apple byproducts as fermentation substrates with the Gluconobacter oxidans C2 described in claim 1 or a microbial combination comprising the Gluconobacter oxidans C2 described in claim 1.

6. The apple enzyme fertilizer according to claim 5, characterized in that: The apple enzyme fertilizer is an apple liquid enzyme fertilizer, comprising the following raw materials: Apple pulp, comprising apples or apple byproducts and water in a mass ratio of 2-5:5-8; Sugars, the mass of which is 1% to 5% of the mass of the apple pulp; The fermentation agent is the Gluconobacter oxidans C2 described in claim 1 or a microbial combination comprising the Gluconobacter oxidans C2 described in claim 1; based on the volume of apple pulp, the accession amount of the Gluconobacter oxidans C2 is 1×10 8 -1×10 9 CFU / L.

7. The apple enzyme fertilizer according to claim 5, characterized in that: The apple enzyme fertilizer is an apple solid enzyme fertilizer, characterized in that it comprises the following raw materials: A fermentation substrate comprising apples or apple byproducts and agricultural byproducts in a mass ratio of 2-4:1; Sugars, the mass of which is 1%-5% of the mass of the fermentation substrate; The fermentation agent is the Gluconobacter oxidans C2 described in claim 1 or a microbial combination comprising the Gluconobacter oxidans C2 described in claim 1; based on the volume of the fermentation substrate, the access amount of the Gluconobacter oxidans C2 is 1×10 9 -1×10 10 CFU / L.

8. A rapid fermentation method for apple liquid enzyme fertilizer, characterized in that: The following steps are involved: Step 1: crushing apples or apple byproducts into apple puree and mixing with water, with the mass ratio of apple puree to water being 2-5:5-8 to obtain apple pulp; Step 2: adding 1%-5% brown sugar to the apple pulp obtained in step 1, mixing well, and obtaining an apple fermentation substrate; Step 3: Inoculate the live bacteria of Gluconobacter oxidans C2 described in claim 1 into the apple fermentation substrate obtained in step 2, and the inoculation amount of the live bacteria of Gluconobacter oxidans C2 is 1×10 8 -1×10 9 CFU / L, 30-40℃, 100-300r / min, culture until the concentration of Gluconobacter C2 in the fermentation substrate reaches 1×10 8 -1×10 9 CFU / g, and apple fermentation liquid was obtained; Step 4: Take the apple fermentation liquid obtained in step 3 and incubate it at 30-40° C. until the pH is less than or equal to 3 and the sugar content is less than or equal to 3, then terminate the fermentation to obtain apple liquid enzyme fertilizer.

9. A rapid fermentation method for apple solid enzyme fertilizer, characterized in that: The following steps are involved: Step 1: crushing apples or apple by-products into apple puree and mixing with bran, wherein the mass ratio of apple puree to bran is 2-4:1, adding 1%-5% brown sugar, and mixing well to obtain a solid fermentation base; Step 2: Introduce the Gluconobacter oxidans C2 described in claim 1 into the solid-state fermentation base, and the amount of the Gluconobacter oxidans C2 introduced is 1×10 9 -1×10 10 CFU / L, after mixing, closed fermentation was carried out at 30-40°C, and cultured until the pH was less than or equal to 4 to obtain apple fermentation solid; Step 3: The apple fermentation solid obtained in step 2 is openly cultured at 30-40° C. and statically cultured until the pH is greater than or equal to 6.9 and the sugar content is less than or equal to 1.5, and the fermentation is terminated to obtain apple solid enzyme fertilizer.

10. A method for preparing apple solid enzyme fertilizer, characterized in that: The following steps are involved: Step 1: crushing apples or apple by-products into apple puree and mixing with bran, wherein the mass ratio of apple puree to bran is 2-4:1, adding 1%-5% brown sugar, and mixing well to obtain a solid fermentation base; Step 2: Inoculate the Gluconobacter oxidans C2 described in claim 1 into the solid-state fermentation substrate. The inoculation amount of the Gluconobacter oxidans C2 is 1×10 9 -1×10 10 Pile evenly, cover and seal, turn the pile once when the temperature is higher than 40°C, perform anaerobically fermentation until the pH is greater than or equal to 6.9 and the sugar content is less than or equal to 1.0, remove the film, spread out and dry in the shade until the moisture content is less than 20%, and obtain apple solid enzyme fertilizer.

11. The apple enzyme fertilizer according to any one of claims 5 to 7, the apple enzyme fertilizer prepared by the rapid fermentation method according to claim 8, the apple enzyme fertilizer prepared by the rapid fermentation method according to claim 9, or the apple enzyme fertilizer prepared by the preparation method according to claim 10 is used in promoting crop yield increase, characterized in that: The apple enzyme fertilizer is used as base fertilizer and / or topdressing fertilizer for field crops, fruit trees or vegetable crops.

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