CN112458026A

CN112458026A - Bacillus subtilis capable of efficiently degrading vomitoxin and application thereof

Info

Publication number: CN112458026A
Application number: CN202011470276.8A
Authority: CN
Inventors: 孙育荣
Original assignee: Jiangsu Aomai Bio Technology Co ltd
Current assignee: Jiangsu Aomai Bio Technology Co ltd
Priority date: 2020-12-15
Filing date: 2020-12-15
Publication date: 2021-03-09

Abstract

The invention discloses a bacillus subtilis AM-05 strain for efficiently degrading vomitoxin, which is obtained by separating, screening and fermenting the bacillus subtilis AM-05 strain in wheat field soil infected with scab, and has good stability and high degradation rate when being applied to DON degradation.

Description

Bacillus subtilis capable of efficiently degrading vomitoxin and application thereof

Technical Field

The invention relates to bacillus subtilis AM-05 for efficiently degrading vomitoxin and application thereof, belonging to the field of biological feed additives in biological agriculture.

Background

The main component of vomitoxin is DON (deoxynivalenol), belongs to trichothecene compounds, and is mainly produced by fusarium graminearum, fusarium oxysporum, fusarium moniliforme, fusarium sporotrichioides, fusarium roseum, fusarium niveum and other fusarium. In addition, strains of the genus cephalosporium, the genus painterl, the genus trichoderma, etc. can produce the toxin. The trichothecene toxins are over 150 kinds, are strong immunosuppressants, and cause the typical symptom that the feed intake is reduced, so the toxins are also called as feed antifeedant toxins. Vomitoxin (DON) is one of the most important toxins, mainly from Fusarium, especially Fusarium graminearum and Fusarium yellow, also known as vomitoxin (vomitoxin, VT) because it can cause vomiting in pigs.

The toxin was first discovered in 1970 in a virus poisoned by primary gibberellic barley in Xiangchuan county of Japan, and in 1972, it was first isolated from gibberellic barley by Morooka et al, Japan, and Yoshizawa et al elucidated the structure of this novel mycotoxin and named 4-Deoxynivalenol (DON). The same chemical was isolated in 1973 by vesoder et al in the united states from corn contaminated with fusarium, which was named vomitoxin because it caused vomiting symptoms in pigs, and was subsequently found to be isolated from fusarium contaminated barley, gibberellin. Vomitoxin is a common mycotoxin in food, widely existing in nature, and 3 toxins commonly found in europe and north america are deoxynivalenol, 3-acetyldeoxynivalenol, and 15-deoxynivalenol. Because of their highly cytotoxic and immunosuppressive properties, they pose a threat to the health of humans and animals, especially with a significant impact on immune function. Depending on the dose of DON and the exposure time, immunosuppression or immunostimulation may be induced. When people ingest food contaminated by vomitoxin, acute poisoning symptoms such as anorexia, vomiting, diarrhea, fever, unstable standing, and slow response can be caused, and in severe cases, hematopoietic system is damaged to cause death. Because the grain proportion in the traditional Chinese diet habit is greatly higher than that in the west, the harm of vomitoxin is more prominent. In 1998, vomitoxin was listed as a class 3 carcinogen in an evaluation report published by the international cancer research institute. The European Union requires vomitoxin to be less than 1.0 mg/kg; chinese feed requirements are less than 1 ppm.

The vomitoxin has quite common pollution to Chinese grain raw materials and feed products, mycotoxin analysis reports from 2013 to 2019 of Jiangsu Austria feed mycotoxin engineering research center are summarized, 3632 corn samples are analyzed, the detection rate of the vomitoxin is 100%, the average value is 784.6ppm, and the highest value is 3652.3 ppm. And exceeding 53.97 percent. 2864 corn by-product samples were analyzed, and the detection rate of vomitoxin was 100%, the mean value was 1164.8ppm, and the highest value was 4402.7 ppm. And exceeding 76.52 percent. 4481 compound feed samples were analyzed, and the detection rate of vomitoxin was 100%, the mean value was 876.2ppm, and the maximum value was 3346.01 ppm. The standard exceeding rate is 22.35 percent.

The vomitoxin has stable chemical properties, can not be damaged in the processes of processing, storage and cooking generally, can be stored for a long time under the condition of a laboratory, keeps the toxicity unchanged, has stronger heat resistance, and has only a small amount of damage after being heated at the temperature of 121 ℃ for 25min under high pressure. The toxicity of the grain processed product is not influenced by the acidic environment, but the grain processed product containing vomitoxin can be damaged by adding alkali or high-pressure treatment, and the vomitoxin can not be completely removed due to technical limitation in the processing process, is mainly absorbed by gastrointestinal tracts, enters the liver through blood circulation for metabolism, is discharged with urine through the kidney, and causes wide damage to various organs. The half-life period of the vomitoxin in a human body is 2-4 h, the metabolism is rapid, and the vomitoxin and the metabolite thereof cannot be detected after 8 h.

At present, the detoxification method of the mildewed grains and feeds at home and abroad mainly comprises a physical adsorption method, a chemical detoxification method, a biological detoxification method and the like. The physical adsorption method has reversible DON adsorption and poor efficiency, and is easy to adsorb trace nutrient elements, thereby causing the loss of nutrient substances in cereals and feeds; chemical detoxification easily leaves some potential toxic substances, and causes secondary pollution to grains and feeds, so that the DON traditional detoxification method has certain limitation in practical application; the biological detoxification method is characterized in that DON is acted by enzyme released by microorganisms and is converted into a metabolite with lower toxicity, and the microbial degradation has the advantages of low cost, high efficiency, less nutrient loss, no secondary pollution, good ecological restoration and the like, so that great attention is paid.

Disclosure of Invention

The invention aims to solve the technical problem that aiming at the defects in the prior art, the bacillus subtilis AM-05 for efficiently degrading vomitoxin and the application thereof are provided.

The technical scheme for solving the technical problems is as follows:

a bacillus subtilis AM-05 for efficiently degrading vomitoxin is obtained by separating, screening and fermenting bacillus subtilis AM-05 strain in wheat field soil infected with gibberellic disease.

The technical scheme of the invention is further defined as follows:

further, in bacillus subtilis AM-05 capable of efficiently degrading vomitoxin, the preparation method of the strain is as follows:

(I) screening of bacterial species

(1) Taking field soil of wheat infected with gibberellic disease, shoveling off 5cm of the surface layer, taking 5-15 cm of the field soil, sampling by using a sterile instrument, putting the field soil into a sterile plastic bag or a sterile paper bag, binding, separating after sampling, shaking for 20-25min to fully mix a soil sample with water, dispersing cells, putting the sample into a triangular flask filled with 99ml of sterile water, treating the sample in a water bath at normal pressure of 80 ℃ for 1 hour, and taking out;

(2) sucking 1ml of soil suspension from a 1ml sterile pipette, adding into a large test tube containing 9ml of sterile water, mixing, sucking 1ml from the test tube with the sterile pipette, adding into another test tube containing 9ml of sterile water, mixing, and repeating the steps to obtain 10^-2、10^-3、10^-4、10^-5Soil solutions of different dilutions;

then using sterile straws respectively 10^-3、10^-4、10^-5Sucking 0.2ml of each of the three tubes of soil diluent, dripping the three tubes of soil diluent at the central position of the surface of the corresponding plate culture medium, coating the three tubes of soil diluent by using a sterile coater, standing the three tubes of soil diluent at room temperature for 5-10min, and immersing the bacteria liquid into the culture medium;

(3) inverting the plate culture medium in the step (2) into an incubator at 30 ℃ for culturing for 1-3 d;

(4) observing the surface of the bacterial colony to be rough and opaque, wherein the bacterial colony is in a dirty white or yellowish color, respectively selecting a few cells from the single bacterial colony of the strain to be inoculated on a seed culture medium, and culturing in an incubator at 30 ℃ to obtain a strain to be selected;

(5) screening the strains to be selected, detecting the degradation capability of vomitoxin, and screening to obtain a strain AM-05 with the highest degradation capability;

(6) supporting the strain culture with 30% glycerol tube, and storing in-80 deg.C refrigerator;

(II) fermentation of the bacterial species

And taking the separated bacillus subtilis AM-05, fermenting step by step, adding into a 15-ton fermentation tank, fermenting for 48 hours, and spray drying to obtain a bacillus subtilis AM-05 product of about 500 hundred million/g.

In Bacillus subtilis AM-05 for efficiently degrading vomitoxin, the preparation method of the culture medium in the step (2) comprises the following steps: peptone: 10-13g, yeast extract: 4-5g, NaCl: 9-12g, soluble starch: 1-2g, agar: 18-20g, using distilled water to fix the volume to 1000mL of 900-.

In the bacillus subtilis AM-05 for efficiently degrading vomitoxin, the preparation method of the culture medium in the step (4) comprises the following steps: bean cake powder: 10-13g, corn flour 1-3g and Na₂HPO₄：2-3g，(NH₄)₂SO₄:0.8-1.2g，NH₄Cl: 0.5-1g, using distilled water to fix the volume to 1000mL of 900-.

In the bacillus subtilis AM-05 for efficiently degrading vomitoxin, a fermentation culture medium required by the step-by-step fermentation in the step (II) comprises the following components in percentage by mass: bean cake powder: 70-80%, corn flour 5-10%, Na₂HPO₄：1-2%，(NH₄)₂SO₄：0.5-1%，NH₄Cl:0.5-1%1g，CaCl₂1-2%, alpha-amylase: 1-2%, and the sum of the above components is 100%.

The invention also designs an application of the bacillus subtilis AM-05 in degrading the vomitoxin, the bacillus subtilis AM-05 is added into the modified bentonite, then the yeast cell wall is added, and the special detoxicant vomitoxin can be obtained, and then the special detoxicant vomitoxin is mixed with the feed polluted by the vomitoxin;

the weight percentage of the material is as follows: bacillus subtilis AM-05: 10-30%, modified bentonite: 40-60%, yeast cell wall: 5-15%, and the sum of the above components is 100%.

The invention has the beneficial effects that:

the bacillus subtilis is widely distributed in soil and rotten organic matters, is easy to reproduce in the bacillus subtilis extract, has high growth speed and low requirement on nutrition, can efficiently secrete a plurality of proteins and metabolites, does not produce toxins, is a safe microorganism without pathogenicity, is obtained by firstly separating and screening bacillus subtilis AM-05 strain in the soil of wheat fields infected with gibberellic disease, and can efficiently degrade vomitoxin.

The microbial strain AM-05 with good DON degradation performance is obtained by separation and purification, and the screening, separation and purification technology is simple.

The strain has good stability for degrading DON, can be passaged in an optimized culture medium for 8 generations, has DON degradation rate of over 90 percent, and can effectively relieve the harm of vomitoxin proved by animal experiments.

The detoxifying agent vomit-ling applied by the bacillus subtilis AM-0 provided by the embodiment of the invention improves the growth performance of piglets, the detoxifying agent utilizes the bacillus subtilis to realize biological adsorption, and simultaneously, the yeast cell wall can be combined with mycotoxin to realize biological adsorption, so that the mycotoxin in the feed can be effectively removed.

Detailed Description

Example 1

In the embodiment, through separation and screening, bacillus subtilis AM-05 capable of efficiently degrading vomitoxin is obtained through separation, screening and fermentation from wheat field soil infected with gibberellic disease, and the specific preparation method is as follows:

(I) screening of bacterial species

(1) Taking wheat field soil infected with gibberellic disease, shoveling off 5cm of the surface layer, taking 5-15 cm of the wheat field soil, sampling according to the standard by using a sterile appliance, putting the wheat field soil into a sterile plastic bag or a sterile paper bag, binding, recording sampling time, sampling place, sampling environment and the like for examination, separating as soon as possible after sampling, shaking for twenty minutes to fully mix a soil sample and water, dispersing cells, putting the mixture into a triangular flask containing 99ml of sterile water, treating the sample in a water bath at normal pressure of 80 ℃ for 1 hour, and taking out;

(2) sucking 1ml of soil suspension from a 1ml sterile pipette, adding into a large test tube containing 9ml of sterile water, mixing, sucking 1ml from the test tube with the sterile pipette, adding into another test tube containing 9ml of sterile water, mixing, repeating the steps, and making into 10^-2、10^-3、10^-4、10^-5Soil solutions of different dilutions;

then using sterile straws respectively 10^-3、10^-4、10^-50.2ml of each soil diluent is absorbed by each of the three pipes, the three pipes are dripped at the central position of the surface of the corresponding plate culture medium, the three pipes are coated by an aseptic coater, the aseptic coater is held by the right hand and is horizontally placed on the surface of the plate culture medium, the bacterial suspension is slightly expanded outwards along the direction of a concentric circle to be uniformly distributed, and the three pipes are kept stand for 5-10min at room temperature to ensure that the bacterial liquid is immersed in the culture medium;

(4) observing that the surfaces of bacterial colonies are rough and opaque and are in dirty white or yellowish colors, respectively picking a small amount of cells from single bacterial colonies of the strains to inoculate the single bacterial colonies on a seed culture medium, culturing the single bacterial colonies in an incubator at 30 ℃ to obtain strains to be selected, and if the mixed bacteria are found, separating and purifying again; if the strain can not be better observed by naked eyes, gram staining is carried out on the strain, and the strain is observed under a microscope and compared with a standard strain;

identification of isolated strains

(a) Colony morphology identification

Selecting a fresh single colony of AM-05 cultured for 18-24 h for gram staining, observing the physiological morphology of a strain under a microscope, inoculating the single colony into an inorganic salt culture medium for overnight culture, centrifuging at 5000r/min for 5min, then discarding the supernatant, suspending the strain with distilled water, centrifuging at 5000r/min for 5min, repeating the operation once, discarding the supernatant, then suspending the strain with 2mL of sterile distilled water, freeze-drying to obtain strain powder, and observing by using a scanning electron microscope;

the AM-05 provided by the invention has the following morphological characteristics: after the strain is cultured on an LB flat plate at 37 ℃ for 24 hours, the colony is circular, the diameter is 1-2mm, the edge is neat, the surface is moist and glossy, and the colony is gray; observing under a microscope through gram staining, wherein the strain is a gram-negative bacterium, the shape of the strain is short rod-shaped, freeze-drying the centrifuged thallus, and observing the dried thallus powder by using SEM, wherein under the multiple of 20000 multiplied, the thallus is in the shape of short rod with irregular arrangement, two ends are blunt circles, and the size is 0.5 multiplied by 1-3 mu m.

(2) Analysis of biochemical Properties

The white bacterial strain is subjected to physiological and biochemical identification by referring to a common bacterial strain identification method in 'handbook of bacteria system identification'.

Oxidative fermentation of glucose and other sugar alcohols: and (3) inoculating a fresh single colony cultured for 18-24 hours AM-05 into Hufson and Lifson's culture medium by puncturing, sealing the culture medium with sterilized vaseline paraffin oil, sealing the culture medium with the thickness of 0.5-1 cm to isolate air, simultaneously designing an experimental group without oil sealing to verify the oxidative fermentation type of the strain, culturing the strain at room temperature for 1, 2, 3, 7 and 14 days, observing color change, and respectively replacing glucose with lactose, maltose, mannose, arabinose, fructose, xylose and sucrose to detect the sugar alcohol utilization condition of the strain.

(5) Screening the strains to be selected, detecting the degrading capability of vomitoxin, and screening to obtain the strains with the highest degrading capability;

(II) fermentation of the bacterial species

In this example, the method of disposing the medium in step (2): peptone: 10-13g, yeast extract: 4-5g, NaCl: 9-12g, soluble starch: 1-2g, agar: 18-20g, using distilled water to fix the volume to 1000mL of 900-.

In this example, the method for preparing the medium in step (4): bean cake powder: 10-13g, corn flour 1-3g and Na₂HPO₄：2-3g，(NH₄)₂SO₄:0.8-1.2g，NH₄Cl: 0.5-1g, using distilled water to fix the volume to 1000mL of 900-.

In the embodiment, the fermentation medium required for the stepwise fermentation in the step (II) comprises the following components in percentage by mass: bean cake powder: 70-80%, corn flour 5-10%, Na₂HPO₄：1-2%，(NH₄)₂SO₄：0.5-1%，NH₄Cl:0.5-1%1g，CaCl₂1-2%, alpha-amylase: 1-2%, and the sum of the above components is 100%.

The embodiment also relates to an application of the bacillus subtilis AM-05 in degrading vomitoxin, namely, the bacillus subtilis AM-05 is added into the modified bentonite, and then the yeast cell wall is added, and the mass percentages are as follows: bacillus subtilis AM-05: 10-30%, modified bentonite: 40-60%, yeast cell wall: 5-15%, the sum of the above components is 100%, obtaining the special detoxicant emetic for the vomitoxin, then mixing the detoxicant emetic with the feed polluted by the vomitoxin to effectively degrade the vomitoxin, and no other harmful chemical reagent is added into the feed, so that the change of the flavor of the feed and the loss of the nutritive value can not be caused.

The antidote emetic mixed in the application of the bacillus subtilis AM-05 in the embodiment is subjected to an in vitro and simulated in vivo evaluation method (HPLC method) Q/3201 AOMAI-BIO 10-2013:

1. vomitoxin standard stock solution

Weighing 1.0mg (accurate to 0.01 mg) of standard substance vomitoxin, placing in a 50ml volumetric flask, dissolving with methanol, fixing the volume to 50ml, and storing in a refrigerator at 4 ℃ for later use, wherein the concentration of the standard solution is 20 mug/ml;

2. vomitoxin standard working solution

Accurately transferring a certain amount of vomitoxin standard stock solution, diluting with methanol, and respectively preparing standard working solutions of 0 μ g/ml, 2 μ g/ml, 10 μ g/ml, 15 μ g/ml and 20 μ g/ml;

3. preparation of the samples

Sampling according to GB/T14699.1, selecting 500g of the detoxicant vomit medicine sample, reducing the sample to 100g by a quartile method, uniformly mixing the sample and the sample through a 0.28mm hole sieve, putting the mixture into a closed container, and keeping the mixture away from light and at room temperature for later use;

collecting pig small intestine intestinal chyme, centrifuging at 20000 rpm with a high speed centrifuge, extracting supernatant totally 10ml, and storing in a refrigerator at 4 deg.C for use;

4. preparation of sample solution

When the in vitro method is used for determination: weighing 0.2g (accurate to 0.002 g) of sample, placing the sample in a 100ml volumetric flask, performing double distilled water constant volume (obtaining 2mg/ml adsorbent solution), stirring for 24 hours in the dark under magnetic force, respectively and precisely absorbing 0.25ml adsorbent solution and 0.25ml vomitoxin standard substance with 10 microgram/ml in a 1.5ml centrifuge tube, placing the centrifuge tube on a constant temperature oscillation metal bath for 4 hours in the dark under 37 ℃, performing high-speed centrifugation for 10 minutes at 15000r/min, and taking supernatant fluid to filter for HPLC detection;

when the in vivo method is simulated: weighing 0.2g (accurate to 0.002 g) of sample, placing the sample in a 100ml volumetric flask, sucking 10ml of minced small intestine chyme supernatant, adding the mixture, then using double distilled water to fix the volume (obtaining 2mg/ml of adsorbent solution), stirring the mixture in a dark place and by magnetic force for 24 hours, respectively sucking 0.25ml of adsorbent solution and 0.25ml of 10 microgram/ml vomitoxin standard substance in a 1.5ml centrifugal tube, and placing the centrifugal tube on a constant temperature oscillation metal bath for 4 hours in a dark place at 37 ℃. Centrifuging at high speed of 15000r/min for 10min, collecting supernatant, filtering, and detecting by HPLC.

5. Measurement of

High performance liquid chromatography conditions: a chromatographic column: a C18 column with the length of 150mm, the inner diameter of 4.6mm and the diameter of the filler of 5 μm;

when vomitoxin is measured, the mobile phase is methanol: water =20:80, ultrasonic de-gassing, filtration through 0.45 μm filter membrane;

flow rate: 1ml/min, injection volume: 20 μ L, detection wavelength: vomitoxin 218nm, retention time: 10-15 min;

6. quantitative determination

And (3) standard curve preparation: injecting standard working solution into the chromatographic column, and drawing a standard curve by taking the concentration of the standard working solution as an abscissa and the area value of a chromatographic peak as an abscissa;

and injecting a sample solution into the chromatographic column to obtain a response value of a chromatographic peak area, and calculating the residual quantity of the mycotoxin in the sample solution according to a standard curve graph of the standard solution peak area.

The adsorption rate of the sample solution to the mycotoxin is calculated by the following formula:

X = m 1/ m 0* 100% = (m 0 -m 2)/ m 0* 100%

x: adsorption ratio (%) of the test sample to mycotoxin

m 0, addition amount of mycotoxin

m 1 adsorption amount of mycotoxin

m 2 residual quantity of mycotoxin

Taking the arithmetic mean value of the results of two parallel measurements as the measurement result, wherein the absolute difference value of the results of the parallel measurements does not exceed 0.2 percent of the arithmetic mean value, and the degradation rate of the obtained vomitoxin by the vomitoxin can reach more than 90 percent.

Influence of emetic on growth performance of piglets

In this experiment, 28 weaned piglets (3 weeks old) were selected and randomly allocated to 4 treatment groups, 7 piglets per group, and each piglet was raised in 1 metabolic cage. The group 1 is fed with a Base (BD), the daily ration of the group 1 meets the nutritional requirement of NRC (2012), and the other 3 groups are fed with BD respectively added with 1.0mg/kg DON, 3.0mg/kg DON and 3.0mg/kg DON +0.05% emetolin (the emetolin prepared by the invention).

The experimental groups are shown in table 1:

TABLE 1 test grouping

Group of	Treatment of
		Treatment group 1	Basic ration (BD, Control)
Treatment group 2	Basal Diet (BD) +1mg/kg DON (1.0 DON)
		Treatment group 3	Basal Diet (BD) +3mg/kg DON (3.0 DON)
Treatment group 4	Basal Diet (BD) +3mg/kg DON +0.05% emetic (3.0 DON + A)

The weekly weight and feed intake of piglets during the test period are recorded, and the daily gain and the feed conversion rate are calculated.

The content of each toxin in the daily ration is shown in table 2 specifically:

TABLE 2 actual content of several toxins in each group of diets¹

	Control	1.0 DON	3.0 DON	3.0 DON+A
					DON,mg/kg	<0.1	1.022	3.124	3.201
AFB₁, mg/kg	0.002	0.005	0.071	0.050
					ZEA, mg/kg	<0.01	0.071	0.598	0.531

Note: the detection lines of 1DON, AFB1 and ZEA are respectively 100, 0.5 and 10 mug/kg; DON is the HPLC measurement value; AFB1 and ZEA are ELISA measured values; -is not detected.

The growth performance index is shown in table 3:

TABLE 3 influence of emetic toxins and emetics on growth performance of piglets

	Control	1.0 DON	3.0 DON	3.0 DON+A
					Initial body weight, kg	6.29 ± 0.20	6.24 ± 0.17	6.56 ± 0.20	6.20 ± 0.15
1-2 weeks
					Body weight gain, kg	2.73 ± 0.65^a	1.07 ± 0.39^b	1.08 ± 0.59^b	2.09 ± 0.40^ab
Feed intake, kg	3.90 ± 0.54^a	2.39 ± 0.30^b	2.41 ± 0.42^b	3.15 ± 0.42^ab
					Feed/Gain, g/g	1.77 ± 0.29	2.16 ± 0.43	1.77 ± 0.53	1.70 ± 0.20
3-4 weeks
					Body weight gain, g	7.08 ± 0.27^a	5.95 ± 0.24^b	4.66 ± 0.59^c	5.50 ± 0.16^bc
Feed intake, g	10.39 ± 0.42^a	9.31 ± 0.51^ab	7.29 ± 0.93^c	8.66 ± 0.61^bc
					Feed/Gain,g/g	1.47± 0.03	1.57 ± 0.07	1.56 ± 0.11	1.57 ± 0.09
1-4 weeks
					Body weight gain, g	9.81 ± 0.88^a	7.02 ± 0.44^b	5.73 ± 1.17^b	7.59 ± 0.54^b
Feed intake, g	14.29 ± 0.93^a	11.70 ± 0.75^b	9.70 ± 1.34^b	11.81 ± 0.96^ab
					Feed/Gain,g/g	1.48 ± 0.05^a	1.67 ± 0.05^b	1.98 ± 0.26^b	1.56 ± 0.08^ab

Note: results are expressed as mean ± sem (n = 7), with the difference indicated by the lower case letters on the shoulder marks of the same row being significant (P < 0.05).

As shown in Table 3, the DON added to the diet at 1.0mg/kg was significantly reduced as compared with the control group (P <0.05) piglet body weight gain (16.0-60.8%) and/or feed intake (18.1-38.7%) at 1-2 weeks, 3-4 weeks and 1-4 weeks and significant increase(s) ((16.0-60.8)P <0.05) feed-meat ratio of 1-4 weeks (12.8%). Adding 1 with daily ration.Compared with the group with 0mg/kg DON, the addition of 3.0mg/kg DON in the daily ration can further reduce (P <0.05) piglet 3-4 weeks body weight gain (21.2%) and feed intake (21.7%). It is worth noting that 0.05% of emetic detoxicant is added into the daily ration, so that the harm of 3.0mg/kg DON to the weight gain, feed intake and/or feed conversion ratio of piglets in 1-2 weeks, 3-4 weeks and 1-4 weeks can be effectively relieved, and the emetic effectively improves the growth performance of the piglets, thereby improving the economic benefit.

In addition to the above embodiments, the present invention may have other embodiments. All technical solutions formed by adopting equivalent substitutions or equivalent transformations fall within the protection scope of the claims of the present invention.

Claims

1. The bacillus subtilis AM-05 for efficiently degrading emetic toxin according to claim 1, wherein: the bacillus subtilis AM-05 strain is obtained by separating, screening and fermenting wheat field soil infected with gibberellic disease.

2. The Bacillus subtilis AM-05 for degrading vomitoxin with high efficiency according to claim 1, which is prepared by the following specific method:

(I) screening of bacterial species

(II) fermentation of the bacterial species

3. The bacillus subtilis AM-05 for efficiently degrading emetic toxin according to claim 2, wherein: the preparation method of the culture medium in the step (2) comprises the following steps: peptone: 10-13g, yeast extract: 4-5g, NaCl: 9-12g, soluble starch: 1-2g, agar: 18-20g, using distilled water to fix the volume to 1000mL of 900-.

4. The bacillus subtilis AM-05 for efficiently degrading emetic toxin according to claim 2, wherein: the preparation method of the culture medium in the step (4) comprises the following steps: bean cake powder: 10-13g, corn flour 1-3g and Na₂HPO₄：2-3g，(NH₄)₂SO₄:0.8-1.2g，NH₄Cl: 0.5-1g, using distilled water to fix the volume to 1000mL of 900-.

5. The bacillus subtilis AM-05 for efficiently degrading emetic toxin according to claim 2, wherein: the fermentation medium required by the step-by-step fermentation in the step (II) comprises the following components in percentage by mass: bean cake powder: 70-80%, corn flour 5-10%, Na₂HPO₄：1-2%，(NH₄)₂SO₄：0.5-1%，NH₄Cl:0.5-1%1g，CaCl₂1-2%, alpha-amylase: 1-2%, and the sum of the above components is 100%.

6. Use of bacillus subtilis AM-05 according to any one of claims 1 to 5 for the degradation of emetic toxin, wherein: firstly adding bacillus subtilis AM-05 into the modified bentonite, then adding yeast cell walls to obtain the special detoxicant emetic for the vomitoxin, and then mixing the detoxicant emetic with the feed polluted by the vomitoxin;