CN105941975A - Broiler feed on basis of lactobacillus johnsonii BS15 and method for preparing broiler feed - Google Patents
Broiler feed on basis of lactobacillus johnsonii BS15 and method for preparing broiler feed Download PDFInfo
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Abstract
The invention discloses broiler feed on the basis of lactobacillus johnsonii BS15 and a method for preparing the broiler feed. The broiler feed on the basis of the lactobacillus johnsonii BS15 contains 1*10<8>-1*10<9> cfu/kg of the lactobacillus johnsonii BS15. The service time of the boiler feed starts from one day after chicks are hatched to the broiler slaughtering time; the broiler feed and feed are mixed with each other and then are fed to broilers. The broiler feed and the method have the advantages that the problem of drug residues due to antibiotics can be solved, and gaps in the field after the antibiotics are forbidden can be filled; the intestinal tracts of the broilers can be regulated, the purpose of allowing the broilers to healthfully grow can be achieved, and growth promotion and disease prevention functions are integrated by a novel prevention and control process; the broiler feed is used for assisting in establishing healthful digestive systems of the broilers, and accordingly deterioration of the meat quality of the broilers can be prevented when sub-clinical necrotic enteritis occurs.
Description
Technical field
The invention belongs to broiler breeding technical field, particularly relate to a kind of broiler fodder based on Yue Shi lactobacillus BS15 and preparation method thereof.
Background technology
Necrotic enteritis (Necrotic enteritis, NE) is one of problem of meriting attention most of broiler breeding industry, bacillus perfringens (Clostridium perfringens is called for short CP) be that main cause causes.CP is to antibiotic sensitive, and at Systems in Certain Developed Countries because antibiotic is as the disabling of growth promoter, the sickness rate of NE increases day by day, and as a example by the U.S., the economic loss that NE has caused reaches 3,000,000,000 dollars every year, and averagely every broiler loses 0.05 dollar.In addition the alimentary toxicosis case load caused by A type CP has occupied the 2nd in countries such as American and Britain.Broiler NE is divided into clinical type and Subclinical, all can damage intestinal mucosa, damages intestinal health.Clinical type NE is mainly in the morbidity of chicken 2 to 6 week, and day, mortality rate was up to 1%, and general mortality rate is between 10% to 40%.But in recent years, the Subclinical Types that NE presents is by higher concern, and slowly and the price of deed reduces mainly to show as broiler growing way, is not result in the death of chicken.Subclinical NE, owing to not having obvious clinical manifestation, is therefore more easy to cause economic loss in process of production and strengthen the epidemic prevention difficulty of disease.The statistics of all states of European Union shows, the economic loss that subclinical NE brings exceeds nearly 33% than the NE of classical symptom.The generation of subclinical NE will cause being decreased obviously of chicken meat quality, is mainly reflected in that moisture easily lost by Carnis Gallus domesticus, the holding time shortens and the level of various nutrient substance including fatty acid declines.And owing to subclinical NE is bought by consumer without obvious pathological changes, the Carnis Gallus domesticus causing a large amount of meat impaired and eats.Carnis Gallus domesticus as being eaten one of most meat, be the mankind important polyunsaturated fatty acid source, polyunsaturated fatty acid (especially EPA and DHA) then be prevention human cardiovascular system disease important nutrient.Along with China progresses into the rear antibiotic epoch, the Prevention Research for subclinical NE also becomes the most urgent.From the point of view of world wide, the antibiotic substitute searched out at present mainly includes the interpolation of some organic acid such as the enzyme preparation such as probiotic bacteria, lysozyme and such as butanoic acid.But, improve, about finding a kind of effective antibiotic substitute, the problem that subclinical NE meat quality of table poultry declines, but have no report.Probiotic bacteria is the means preventing subclinical NE to occur, and meanwhile, has a large amount of report to prove that some probiotic bacterias can improve chicken meat quality during normally raising table hens.But, the most never there is relevant report to prove probiotic bacteria and there is the report improving subclinical NE morbidity chicken meat, meanwhile, in current existing report, not deeply, Research foundation lacks the limitation also leading in subsequent applications in the research of used probiotic bacteria self.Yue Shi lactobacillus (Lactobacillus johnsonii) is a kind of amphimicrobian Gram positive bacillus, most Yue Shi lactobacilluss are considered probiotic bacteria, being used for the production of fermented dairy product etc., meanwhile, part Yue Shi lactobacillus has the effect reducing cholesterol.Yue Shi lactobacillus BS15 is isolatable from herdsman's soil yoghourt on Hongyuan grassland, Aba Tibetan Autonomous Prefecture, Sichuan Province, and early-stage Study confirms that it has the effect (patent of invention number: 201410156594.5) of good prevention mice non-alcoholic fatty liver disease.Carry out improving in the research process of subclinical NE meat quality of table poultry at us, it has been found that in feedstuff, add BS15, the situation that meat quality of table poultry declines can be efficiently controlled.In view of BS15 as a kind of probiotic bacteria, important antibiotic substitute can be considered, simultaneously, in probiotic bacteria research field, BS15 has good Research foundation, therefore, in the rear antibiotic epoch, utilize BS15 that the chicken meat quality of subclinical NE morbidity broiler is improved, the blank of this field of food safety can be made up.
Summary of the invention
It is an object of the invention to provide a kind of broiler fodder based on Yue Shi lactobacillus BS15 and preparation method thereof, aim to solve the problem that and utilize BS15 that the chicken meat quality of subclinical NE morbidity broiler is improved, make up the problem of drug residue caused by the technological gap of field of food safety and antibiotic.
The present invention is achieved in that a kind of broiler fodder based on Yue Shi lactobacillus BS15, described broiler fodder 1 × 10 based on Yue Shi lactobacillus BS158-1×109The Yue Shi lactobacillus BS15 of cfu/kg.
Further, described broiler fodder based on Yue Shi lactobacillus BS15 uses the time to be to hatch 1 age in days from chickling to deliver for sale to broiler and butcher.
Further, described broiler fodder based on Yue Shi lactobacillus BS15 mix with feedstuff realization broiler is fed.
Another object of the present invention is to provide the preparation method of a kind of described broiler fodder based on Yue Shi lactobacillus BS15, described preparation method comprises the following steps:
Step one, takes out the Yue Shi lactobacillus BS15 strain that preserves from Culture Collection, and strain melts defrosting at 25 DEG C, connects 1-2 ring in modified MRS solid medium with inoculating loop, is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h；
Step 2, picking gained list bacterium colony, sterile working is seeded to MRS liquid culture medium, is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h, obtains initial bacterium solution；
Step 3, gained bacterium solution is carried out 10 times of dilutions with physiological saline solution, take bacterium solution 200 μ L and be inoculated in aseptic modified MRS solid medium with rubbing method, after static 1min, after being positioned over 36-37 DEG C of biochemical cultivation case constant temperature culture 12h again, clump count on statistics flat board, and calculate antibacterial viable count contained by every milliliter in gained bacterium solution；
Step 4, according in per kilogram feedstuff 1 × 108-109Cfu viable bacteria calculates, by the feedstuff blender mixing and stirring of bacterium solution Yu corresponding weight, then by mixed fodder subpackage, in lucifuge, be dried, storage under low temperature environment.
Further, described MRS solid medium is: peptone 10.00g/L, Carnis Bovis seu Bubali cream 5.00g/L, yeast extract 4.00g/L, glucose 20.00g/L, tween 80 1.00mL/L, sodium acetate 5.00g/L, dibasic ammonium citrate 0.20g, dipotassium hydrogen phosphate 2.00g/L, magnesium sulfate 0.20g/L, manganese sulfate 0.05g/L, solid medium need to add agar 17.00g, adds aquesterilisa to 1000mL, regulation pH to 6.2,121 DEG C of sterilizing 20min.
Further, in described calculating gained bacterium solution, antibacterial viable count formula contained by every milliliter is as follows:
Every milliliter of original bacteria liquid viable count=A × B × 5
In formula, A: same dilution factor more than three repeats flat-plate bacterial colony number；
B: extension rate.
Broiler fodder based on Yue Shi lactobacillus BS15 that the present invention provides and preparation method thereof, purpose is for improving the chicken meat quality of subclinical NE broiler a kind of effective means of offer, i.e. Yue Shi lactobacillus BS15 interpolation in broiler fodder, and then solve the problem in terms of this field food safety.The present invention has adapted to progressively forbid adding in feedstuff at present the inexorable trend of antibiotic, the problem solving drug residue caused by antibiotic, and compensate for the blank in antibiotic disabling this field later；It addition, the present invention is not in the case of broiler occurs subclinical NE, still can regulate Intestine of Broiler, make the target of broiler more healthy growth, be a kind of brand-new prevention and controls incorporating growth promotion and prevention function of diseases.
The present invention relates to the brand-new purposes of Yue Shi lactobacillus BS15, it is as a kind of feed additive using BS15, for assisting broiler to set up more healthy digestive system, thus prevent the broiler meat when subclinical NE occurs to decline, using method is to realize feeding broiler by the mode that BS15 mixes with feedstuff, the use time is to hatch shell (1 age in days) from chickling to deliver for sale to broiler and butcher, and consumption controls 1 × 108-1×109Cfu/kg feedstuff.A kind of broiler fodder based on Yue Shi lactobacillus BS15.
Present invention firstly provides and prevent the meat of subclinical NE morbidity broiler to decline problem, and searched out currently the only a kind of antibiotic substitute that can apply to this target；(2) theoretical according to intestinal microecology, BS15 can improve Bacterial community in Intestine of Broiler, improve intestinal environment, it is thus possible to have the effect simultaneously preventing multiple intestinal tract disease；(3) interpolation of BS15 does not constitute any drug residue problem to broiler itself, and itself possesses effect of nursing intestinal, it is ensured that food health.
Accompanying drawing explanation
Fig. 1 is the preparation method flow chart of the broiler fodder based on Yue Shi lactobacillus BS15 that the embodiment of the present invention provides.
Detailed description of the invention
In order to make the purpose of the present invention, technical scheme and advantage clearer, below in conjunction with embodiment, the present invention is further elaborated.Should be appreciated that specific embodiment described herein, only in order to explain the present invention, is not intended to limit the present invention.
Below in conjunction with the accompanying drawings the application principle of the present invention is explained in detail.
The deposit number of the Yue Shi lactobacillus BS15 Lactobacillus johnsonii BS15 of the embodiment of the present invention is CCTCC No.M2013663；Depositary institution: China typical culture collection center, is preserved in China, Wuhan, Wuhan University；The culture of Yue Shi lactobacillus BS15 was by December in 2013 13 days, and storage life is 30 years.
As it is shown in figure 1, the preparation method of the broiler fodder based on Yue Shi lactobacillus BS15 of the embodiment of the present invention comprises the following steps:
S101: the Yue Shi lactobacillus BS15 strain preserved from Culture Collection is taken out, strain melts defrosting at 25 DEG C, 1-2 ring is connect in modified MRS solid medium (modified MRS culture medium formula: peptone 10.00g/L with inoculating loop, Carnis Bovis seu Bubali cream 5.00g/L, yeast extract 4.00g/L, glucose 20.00g/L, tween 80 1.00mL/L, sodium acetate 5.00g/L, dibasic ammonium citrate 0.20g, dipotassium hydrogen phosphate (K2HPO4) 2.00g/L, magnesium sulfate (MgSO4·7H20) 0.20g/L, manganese sulfate (MnSO4·10H20) 0.05g/L, solid medium need to add agar 17.00g, adds aquesterilisa to 1000mL, regulates pH to 6.2,121 DEG C of sterilizing 20min), it is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h；
S102: picking gained list bacterium colony, sterile working is seeded to MRS liquid culture medium, is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h, obtains initial bacterium solution；
S103: gained bacterium solution is carried out a series of 10 times of dilutions with physiological saline solution, takes each dilution gradient and (take 10-6、10-7With 10-8) bacterium solution 200 μ L is inoculated in aseptic modified MRS solid medium with rubbing method, after static 1min, then after being positioned over 36-37 DEG C of biochemical cultivation case constant temperature culture 12h, clump count on statistics flat board, and calculate antibacterial viable count contained by every milliliter in 2. gained bacterium solution.Computing formula is as follows:
Every milliliter of original bacteria liquid viable count=A × B × 5
In formula, A: same dilution factor more than three repeats flat-plate bacterial colony number,
B: extension rate.
According to count results, the bacterium solution obtained is diluted to 2 × 10 with physiological saline solution8CFU/ml, is placed in 4 DEG C of Refrigerator stores after packing standby.
S104: according in per kilogram feedstuff 1 × 108-109Cfu viable bacteria calculates, by feedstuff blender (blender need to clean totally in advance, it is ensured that remains without mycete, the antibiotic etc.) mixing and stirring of bacterium solution and corresponding weight, then by mixed fodder subpackage, and storage under lucifuge, dry, low temperature environment.
Below in conjunction with specific embodiment, the application effect of the present invention is explained in detail.
Result obtained by the embodiment of the present invention, is based on following condition and completes: be randomly divided into four groups by male for 1 age in days AA broiler chicken strong young 480: the BS15 prevention group (basal diet+1 × 10 of matched group (basal diet), subclinical NE group (basal diet) and two various dose8Cfu/kg and 1 × 109The BS15 bacterium solution spice of cfu/kg daily ration).Subclinical NE group and BS15 are prevented establishment make subclinical NE model (fill in 15 ages in days and feed 10 multiple dose eimeria tenella live vaccine；Fill every day in 18-21 age in days and feed A type bacillus perfringens once (1.0 × 108Cfu CP/mL, 1mL/ plumage).
The embodiment 1 detection (Carnis Gallus domesticus proportion, yellowish pink, pH value, moisture holding capacity, tenderness) to Carnis Gallus domesticus basis pork quality traits
After chicken is butchered, peel off chest muscle and lower limb flesh, calculate muscle proportion；Take left pectoral and lower limb flesh, using the CR-300 portable colour difference meter detection L* value (brightness value) at the thickest and regular place of muscle, a* value (red scale value) and b* value (yellow value degree), every chicken takes three secondary data and is as the criterion with arithmetical average；Using PH-STAR portable pH tester to carry out the same area of homonymy chest muscle and lower limb flesh puncturing detection, every chicken takes three secondary data and is as the criterion with arithmetical average, the pH value after record chest muscle 40min and 24h；The method surveying drip loss is used to measure muscle moisture holding capacity, right side monoblock pectoralis major is peeled off, it is cut into a block length 5cm along muscle fiber vertical direction with scalpel, wide 3cm, the strip meat sample of thick 2cm, meat sample one end is hooked with iron wire after correct amount (W1) (being accurate to 0.0001g), make muscle fiber (the muscle fiber direction of muscle should keep consistent with attraction of gravitation action direction) vertically downward, it is suspended in the plastic bag of inflation, tighten bag mouth, meat sample is avoided to contact with bag wall, 24h is preserved under the conditions of 4 DEG C, take out meat sample and weigh (W2), calculate drip loss, formula is: drip loss (%)=(W1-W2)/W1 × 100%；Utilizing TA XT plus experimental analysis instrument to detect tenderness, carry out shearing manipulation, the muscle that every pretreatment is good is cut 2 times, and every piece of pectoralis major cuts 3 strips, calculates the arithmetic mean of instantaneous value of 6 shearing forces as final reference value.
Result shows, BS15 significantly improves the detection of Carnis Gallus domesticus every basis pork quality traits that subclinical NE causes.
Table 1. Yue Shi lactobacillus BS15 significantly improves the basic meat change that subclinical NE causes
a,bThe gap that each row data subscript numeral all differs is notable. and (P < 0.05) result presents with mean+/-standard error form.
Embodiment 2: the detection to Carnis Gallus domesticus shelf cycle (i.e. oxidation resistance)
Test 42d often group and randomly draw 12, take left side pectoralis major and-20 DEG C of freezen protective of lower limb flesh, when drawing materials, ensure positions are conformed to each other, after animal experiment completes, detect corresponding index.Muscle is made 10% tissue homogenate, uses vigor and the content of MDA of SOD, GSH-Px, CAT in test kit (building up biotechnology company purchased from Nanjing) detection muscle.
Table 2. Yue Shi lactobacillus BS15 can extend the shelf cycle (raising oxidation resistance) of subclinical NE Muscle of chicken
SOD=superoxide dismutase；CAT=catalase；GSH-Px=glutathion peroxidase；MDA=malonaldehyde
a,bThe gap that each row data subscript numeral all differs is notable. and (P < 0.05) result presents with mean+/-standard error form.
Result shows, Yue Shi lactobacillus BS15 can extend the shelf cycle (raising oxidation resistance) of subclinical NE Muscle of chicken
Embodiment 3: to the detection of intramuscular fat, inosinic acid, cholesterol and content of triglyceride in Carnis Gallus domesticus
Test 42d often group and randomly draw 12, take same area pectoralis major and each one piece of lower limb flesh, liquid nitrogen cryopreservation after tinfoil parcel, after go to-20 DEG C of refrigerators preserve, and use soxhlet extraction methods to extract intramuscular fat after animal experiment terminates；Take same area pectoralis major and each one piece of lower limb flesh; liquid nitrogen cryopreservation after tinfoil parcel; after go in-20 DEG C of refrigerators preserve; and as early as possible all samples is made after 42d animal experiment terminates 10% tissue homogenate, freezing is sent and is used ELISA kit to carry out the mensuration of inosinic acid level by Shanghai Hu Feng Chemical Co., Ltd.；Institute's blood sampling is separated out serum, uses T-CHOL (TC) and triglyceride (TG) in automatic clinical chemistry analyzer detection chest muscle, lower limb flesh and liver.
Result shows, Yue Shi lactobacillus BS15 can significantly improve intramuscular fat, inosinic acid, cholesterol and the change of content of triglyceride in subclinical NE Carnis Gallus domesticus.
Table 3.BS15 significantly improves intramuscular fat, inosinic acid, cholesterol and the change of content of triglyceride in subclinical NE Carnis Gallus domesticus
a,bThe gap that each row data subscript numeral all differs is notable. and (P < 0.05) result presents with mean+/-standard error form.
Embodiment 4: to the detection of fatty acid composition in Carnis Gallus domesticus
Test 42d often group and randomly draw 12, take same area pectoralis major one piece, using HP6890 gas chromatograph to detect FA composition in chest muscle, detected index includes C14:0, C16:0, C16:1n-7, C18:0, C18:2n-6, C18:3n-3, C20:2n-6, C20:4n-6, C20:5n-3, C22:4n-6, C22:5n-3, C22:6n-3, total satisfied fatty acid, total monounsaturated fatty acid, total polyunsaturated fatty acid, n-6 polyunsaturated fatty acid and n-3 polyunsaturated fatty acid.
Result shows, Yue Shi lactobacillus BS15 can significantly improve the change of subclinical NE Fatty Acid in Muscle Tissues of Chicken content, it is particularly possible to efficiently controls the content of polyunsaturated fatty acid, it is ensured that value of chicken.
Table 4. Yue Shi lactobacillus BS15 significantly improves the change (unit: mg/100g dired chicken meat material) of subclinical NE Fatty Acid in Muscle Tissues of Chicken content
SFA: satisfied fatty acid；MUFA: monounsaturated fatty acid；PUFA: polyunsaturated fatty acid
The gap that a, b each row data subscript numeral all differs is notable. and (P < 0.05)
Result presents with mean+/-standard error form.
The foregoing is only presently preferred embodiments of the present invention, not in order to limit the present invention, all any amendment, equivalent and improvement etc. made within the spirit and principles in the present invention, should be included within the scope of the present invention.
Claims (6)
1. a broiler fodder based on Yue Shi lactobacillus BS15, it is characterised in that institute
State per kilogram broiler fodder based on Yue Shi lactobacillus BS15 and comprise 1 × 108-1×109
The Yue Shi lactobacillus BS15 of cfu/kg.
2. broiler fodder based on Yue Shi lactobacillus BS15 as claimed in claim 1,
It is characterized in that, described broiler fodder based on Yue Shi lactobacillus BS15 uses the time to be certainly
Chickling hatches 1 age in days to be delivered for sale to broiler and butchers.
3. broiler fodder based on Yue Shi lactobacillus BS15 as claimed in claim 1,
It is characterized in that, described broiler fodder based on Yue Shi lactobacillus BS15 mixes real with feedstuff
Now broiler is fed.
4. a broiler based on Yue Shi lactobacillus BS15 as claimed in claim 1 is raised
The preparation method of material, it is characterised in that described preparation method comprises the following steps:
Step one, the Yue Shi lactobacillus BS15 bacterium that will preserve from Culture Collection
Planting and take out, strain melts defrosting at 25 DEG C, connects 1-2 ring with inoculating loop solid in modified MRS
Body culture medium, is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h；
Step 2, picking gained list bacterium colony, sterile working is seeded to modified MRS liquid culture
Base, is placed in 36-37 DEG C of biochemical cultivation case constant temperature culture 12h, obtains initial bacterium solution；
Step 3, carries out 10 times of dilutions by gained bacterium solution with physiological saline solution, takes bacterium solution
200 μ L are inoculated in aseptic modified MRS solid medium with rubbing method, after static 1min, then
After being positioned over 36-37 DEG C of biochemical cultivation case constant temperature culture 12h, clump count on statistics flat board, and
Calculate antibacterial viable count contained by every milliliter in gained bacterium solution；
Step 4, according in per kilogram feedstuff 1 × 108-109Cfu viable bacteria calculates, by bacterium solution and phase
Answer the feedstuff blender mixing and stirring of weight, then by mixed fodder subpackage, in lucifuge,
It is dried, stores under low temperature environment.
5. preparation method as claimed in claim 4, it is characterised in that described MRS solid
Culture medium is: peptone 10.00g/L, Carnis Bovis seu Bubali cream 5.00g/L, yeast extract 4.00g/L, Fructus Vitis viniferae
Sugar 20.00g/L, tween 80 1.00mL/L, sodium acetate 5.00g/L, dibasic ammonium citrate 0.20g,
Dipotassium hydrogen phosphate 2.00g/L, magnesium sulfate 0.20g/L, manganese sulfate 0.05g/L, solid medium
Agar 17.00g need to be added, add aquesterilisa to 1000mL, regulation pH to 6.2,121 DEG C of sterilizings
20min。
6. preparation method as claimed in claim 4, it is characterised in that described calculating gained
In bacterium solution, contained by every milliliter, antibacterial viable count formula is as follows:
Every milliliter of original bacteria liquid viable count=A × B × 5
In formula, A: same dilution factor more than three repeats flat-plate bacterial colony number；
B: extension rate.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113018320A (en) * | 2021-02-04 | 2021-06-25 | 四川农业大学 | Application of lactobacillus johnsonii BS15 in preparation of medicine for preventing and/or treating intestinal injury caused by chronic fluorosis |
| CN115316570A (en) * | 2022-08-11 | 2022-11-11 | 四川农业大学 | Biological feed for breeding hermetia illucens and preparation method thereof |
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| CN1265564A (en) * | 1997-06-03 | 2000-09-06 | 卡尔皮斯株式会社 | Method for administering viable microorganism composition for poultry |
| CN102578416A (en) * | 2012-02-23 | 2012-07-18 | 北京大北农科技集团股份有限公司 | Application of lactobacillus johnsonii in preparation of feed used for improving livestock immunity level |
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| CN1265564A (en) * | 1997-06-03 | 2000-09-06 | 卡尔皮斯株式会社 | Method for administering viable microorganism composition for poultry |
| CN102578416A (en) * | 2012-02-23 | 2012-07-18 | 北京大北农科技集团股份有限公司 | Application of lactobacillus johnsonii in preparation of feed used for improving livestock immunity level |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113018320A (en) * | 2021-02-04 | 2021-06-25 | 四川农业大学 | Application of lactobacillus johnsonii BS15 in preparation of medicine for preventing and/or treating intestinal injury caused by chronic fluorosis |
| CN115316570A (en) * | 2022-08-11 | 2022-11-11 | 四川农业大学 | Biological feed for breeding hermetia illucens and preparation method thereof |
| CN115316570B (en) * | 2022-08-11 | 2023-09-01 | 四川农业大学 | Biological feed for culturing hermetia illucens and preparation method thereof |
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