CN102994426A - Bacillus licheniformis and applications thereof - Google Patents
Bacillus licheniformis and applications thereof Download PDFInfo
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- CN102994426A CN102994426A CN2012104886811A CN201210488681A CN102994426A CN 102994426 A CN102994426 A CN 102994426A CN 2012104886811 A CN2012104886811 A CN 2012104886811A CN 201210488681 A CN201210488681 A CN 201210488681A CN 102994426 A CN102994426 A CN 102994426A
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- 241000894006 Bacteria Species 0.000 claims abstract description 29
- 239000000843 powder Substances 0.000 claims abstract description 18
- 241001465754 Metazoa Species 0.000 claims abstract description 12
- 238000002360 preparation method Methods 0.000 claims description 8
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 claims description 6
- 239000006041 probiotic Substances 0.000 claims description 6
- 235000018291 probiotics Nutrition 0.000 claims description 6
- 239000000654 additive Substances 0.000 claims description 4
- 229910000019 calcium carbonate Inorganic materials 0.000 claims description 3
- 238000000855 fermentation Methods 0.000 claims description 3
- 230000004151 fermentation Effects 0.000 claims description 3
- 230000004913 activation Effects 0.000 claims description 2
- 238000012258 culturing Methods 0.000 claims description 2
- 239000007788 liquid Substances 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 abstract description 10
- 230000008901 benefit Effects 0.000 abstract description 7
- 244000144972 livestock Species 0.000 abstract description 4
- 244000144977 poultry Species 0.000 abstract description 4
- 239000003674 animal food additive Substances 0.000 abstract description 3
- 238000004321 preservation Methods 0.000 abstract description 2
- 238000006243 chemical reaction Methods 0.000 abstract 1
- 230000036039 immunity Effects 0.000 abstract 1
- 241000287828 Gallus gallus Species 0.000 description 21
- 235000013330 chicken meat Nutrition 0.000 description 7
- 210000000056 organ Anatomy 0.000 description 7
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- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 4
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- 235000019786 weight gain Nutrition 0.000 description 4
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- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
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- 238000000926 separation method Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 2
- 241000726221 Gemma Species 0.000 description 2
- 208000010359 Newcastle Disease Diseases 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 230000003115 biocidal effect Effects 0.000 description 2
- 210000001669 bursa of fabricius Anatomy 0.000 description 2
- 238000011161 development Methods 0.000 description 2
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- 210000001198 duodenum Anatomy 0.000 description 2
- 238000004043 dyeing Methods 0.000 description 2
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 108020004465 16S ribosomal RNA Proteins 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 238000003794 Gram staining Methods 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 102000005686 Serum Globulins Human genes 0.000 description 1
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- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 description 1
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- 239000008273 gelatin Substances 0.000 description 1
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- 238000009654 indole test Methods 0.000 description 1
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- 239000002054 inoculum Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
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- 231100000957 no side effect Toxicity 0.000 description 1
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Abstract
The invention provides a bacillus licheniformis, especially a Bacillus licheniformis HB1 with a preservation number of CCTCC NO:M 2012480. The selected Bacillus licheniformis is used for preparing bacteria powder; and the prepared bacteria powder is used as a feed additive which is added to livestock feed with addition being 0.01-0.5wt%. after being mixed with the feed, the bacteria powder of the selected Bacillus licheniformis is fed to animals, the production performances of the animals, such as the daily gain, feed conversion ratio and immunity can be effectively improved, so that the bacteria powder of Bacillus licheniformis can be used as a feed additive and is applied to livestock and poultry feed, thereby favorably increasing the economic benefit and having wide application prospects.
Description
Technical field
The invention belongs to the microbe to screen applied technical field, be specifically related to a bacillus licheniformis and application thereof.
Background technology
On poultry industry, microbiotic also is used to promote growth of animal for a long time except being used to prevent and treat courses of infection, thereby improves animal products output.Antibiotic use has brought huge economic benefit, but has also occurred many drawbacks simultaneously; The resistance pathogenic bacteria is increased, and the residual problem such as exceed standard of animal products medicine.Along with the attention of society to food safety, people are also improving constantly the negative effect consciousness that antibiotic feed additive brings.In order to tackle these problems, the biological growth promotion of employing probiotics micro-ecological formulation, the immune measure of accent become current livestock and poultry cultivation focus and come into one's own.Probiotics preparation can by replenishing, adjust or keep the microecological balance in the animal intestinal, reach the purpose of disease preventing and treating, promotion health or raising production performance; And probiotics preparation has no side effect, has no drug resistance, noresidue, and therefore, probiotics micro-ecological formulation obtains the favor of aquaculture deeply.This also makes the novel probiotic bacterium that filters out excellent property become study hotspot.
Summary of the invention
The purpose of this invention is to provide a bacillus licheniformis and the application in livestock and poultry cultivation thereof.Described Bacillus licheniformis separates in the small intestine of the healthy broiler chicken of 21 ages in days.The lichen bacillus ferments of the present invention is prepared into the bacterium powder, is applied in the animal and fowl fodder as fodder additives, can significantly improve the production performance of cultivated animals.
One aspect of the present invention provides a bacillus licheniformis, this bacterial strain called after Bacillus licheniformis HB1(Bacillus licheniformis HB1), be preserved on November 26th, 2012 that " Chinese Typical Representative culture collection " center ", its deposit number are CCTCC NO:M 2012480.
The Bacillus licheniformis of the present invention's screening is used for preparing the bacterium powder; Preparation process is: with Bacillus licheniformis HB1 activation, enlarged culturing, through 35 ~ 37 ℃ of liquid state fermentations, the centrifugal collection of 5000 ~ 8000rpm bacterium mud adds the light calcium carbonate carrier, and the bacterium powder that the bacterium amount is 200 ~ 1,000 hundred million/g is made in oven dry.
Above-mentioned bacterium powder is as the application of fodder additives, is to be that 0.01% ~ 0.5% ratio is added in the animal and fowl fodder by mass percentage.
The bacterium powder of the Bacillus licheniformis of the present invention screening and the cultivated animals of feeding after feed mixes, the production performances such as its day weight gain, feedstuff-meat ratio, immunology have effectively been improved, therefore, it can be applied in the animal and fowl fodder as fodder additives, be beneficial to the increase economic benefit, have broad application prospects.
Embodiment
Separation, the purifying of embodiment 1 Bacillus licheniformis HB1
1, separation and purifying:
1) get the healthy broiler chicken of 21 ages in days, after slaughtering, take out rapidly little intestinal segment, shred into the long segment of 1cm, place the sterilized water that adds granulated glass sphere, concussion mixing 15min, and kill the thalline of non-gemma form with 80 ℃ of water bath processing;
2) with treatment solution with 10 times of gradient dilutions, get the diluent of suitable gradient and coat on the nutrient agar plate, 37 ℃ of incubated overnight;
3) provoke the bacterial strain that bacterium colony is large, growth is fast, the purifying of further ruling.
2, the genus kind of bacterial strain is identified:
With 1 bacillus that is separated to, called after HB1 carries out biochemical identification.This bacterial strain is positive through gramstaining, and is shaft-like, and gemma is ellipticity, is positioned at middle part or inclined to one side end; Physiological and biochemical property: V-P tests (+), Starch Hydrolysis (+), and casein hydrolysis (+), gelatin hydrolysis (+), nitrate reduction (+), indole test (-), 15 ~ 55 ℃ of growth temperature ranges, pH scope 5 ~ 9 is higher than 7%NaCl and does not grow.Biochemical identification result shows that this genus bacillus is Bacillus licheniformis (Bacillus licheniformis); Adopt simultaneously this bacterium 16S rRNA sequence of pcr amplification, the Bacillus licheniformis homology reaches 99% in compare of analysis and the ncbi database, and this bacterium HB1 is accredited as Bacillus licheniformis (Bacillus licheniformis), and the result is consistent with Physiology and biochemistry.
This Bacillus licheniformis (Bacillus licheniformis) HB1 delivered on November 26th, 2012 " preservation of Chinese Typical Representative culture collection " center ", deposit number is: CCTCC NO:M 2012480.
The preparation of embodiment 2 Bacillus licheniformis HB1 bacterium powder
1, seed liquor preparation: the single bacterium colony of picking Bacillus licheniformis HB1, be inoculated in the 50mL nutrient broth medium, pH is 7.0 ± 0.5,37 ℃, 200rpm shaking culture 24h is seed liquor;
2, fermentation culture: the inoculum size with 5% is seeded in the new nutrient broth, and 37 ℃, the 200rpm shaking culture when sporulation reaches 90%, finishes to cultivate;
3, collect thalline: with 5000rpm, the 10min condition is removed fermented supernatant fluid, and it is for subsequent use to collect bacterium mud;
4, oven dry: with light calcium carbonate and bacterium mud stirring and evenly mixing, 50 ℃ of oven dry in loft drier namely get Bacillus licheniformis HB1 bacterium powder.
Embodiment 3 Bacillus licheniformis HB1 are to the growth of meat chicken performance impact
Purchase 1 age in days Luo Shi, 308 white plumage meat cocks, be divided into control group and experimental group, 16 repetitions of each treatment group, each repeats 8 plumage chickens, the production test phase is 35 days, and wherein control group is the general goods daily ration of feeding, and experimental group is to feed to add the general goods daily ration of 0.01% Bacillus licheniformis HB1 bacterium powder, day weight gain, daily ingestion amount, feedstuff-meat ratio, surviving rate in the experiment with measuring process, fatten the index variation situation, experimental result is as shown in table 1:
Table 1 Bacillus licheniformis HB1 is on the impact of meat chicken production performance
By the broiler feeding test-results of table 1 as can be known, the full phase, experimental group broiler chicken day weight gain contrast group has improved 3.24%, and feedstuff-meat ratio reduces by 0.02 on the control group basis, with contrast difference significantly (P＜0.05).In the test index, experimental group day weight gain, daily ingestion amount, feedstuff-meat ratio, surviving rate, fatten the index aspect and all be better than control group.
The above results shows, adds Bacillus licheniformis HB1 of the present invention in the feed, can significantly improve the production performance of broiler chicken, has not only reduced the production cost of broiler chicken, has also improved the production efficiency of broiler chicken, has increased raiser's economic benefit.
Embodiment 4 Bacillus licheniformis HB1 are on the impact of broiler chicken immune level and immune organ thereof
When feeding the Luo Shi 308 white plumage meat cocks among the embodiment 3 to 35 age in days, carry out following operation: 1) experimental group and control group are got respectively 12 chicken blood samplings at random, and separation of serum detects serum globulin (IGg) content and newcastle disease (ND) antibody horizontal; 2) experimental group and control group are got at random respectively 12 chickens and are slaughtered, and take whole thymus gland, the fabricius bursa, spleen, weigh, and calculate Immune Organs Index; Then get the immune organ pathological material of disease, 10% formalin is fixed, paraffin section, and HE dyeing, microscopy is observed the development condition of each immune organ, and experimental result is as shown in table 2.
Table 2 Bacillus licheniformis HB1 is on the impact of broiler chicken immune level and immune organ thereof
As can be seen from Table 2, it is 2.26 times of control group that experimental group records IGg content, in improving blood aspect the IGg content apparently higher than control group (P＜0.01), have the significance advantage.
The result of Microscopic observation Development of Immune Organs situation shows that experimental group obviously is better than control group.Wherein index is 1.62 times of control group under the thymus gland mirror of experimental group, and index is 1.42 times of control group under the fabricius bursa mirror.
The above results shows, adds Bacillus licheniformis HB1 of the present invention in feed, can significantly promote the growth of broiler chicken immune organ and tissue structure, improves the immunological competence of broiler chicken.
The impact that embodiment 5 Bacillus licheniformis HB1 grow Intestine of Broiler
When feeding the Luo Shi 308 white plumage meat cocks of embodiment 3 to 35 age in days, experimental group and control group are got respectively duodenum, jejunum, the ileum of 5 chickens at random, 10% formalin is fixed, paraffin section, duodenum, jejunum, ileum are got the privileged site pathological material of disease, and every section intestines are got respectively 5 serial section, HE dyeing, microscopy is observed each section intestinal tube and is grown and healthy state, measures, records institute's villous and crypts length in each section, and experimental result is as shown in table 3:
Table 3 Intestine of Broiler developmental state statistics
By table 3 experimental result as can be known, the Duodenal villi length of experimental group is apparently higher than control group.Each of experimental group section intestinal tube crypts shortens, and the aspects such as epithelial cell surviving rate rising have obvious advantage.
From villus length and crypts lenth ratio result, can find out that the duodenal villus length of experimental group and crypts lenth ratio are apparently higher than control group, otherness is (P＜0.05) significantly; In the villus length of jejunum and the crypts lenth ratio, experimental group is higher than control group (P〉0.05); In the villus length of ileum and the crypts lenth ratio, experimental group is apparently higher than control group, and otherness is (P＜0.05) significantly.
Analysis-by-synthesis, the villus length of experimental group and crypts lenth ratio have advantage.And the villus length of control group and crypts lenth ratio are very low.
The above results shows, adds Bacillus licheniformis HB1 of the present invention in feed, can significantly promote the intestinal growth of broiler chicken, conforms to embodiment 3 meat chicken production performance indexs.
Claims (6)
1. a bacillus licheniformis is Bacillus licheniformis (Bacillus licheniformis) HB1, and its deposit number is CCTCC NO:M 2012480.
2. the application of Bacillus licheniformis claimed in claim 1 in the preparation probiotics preparation.
3. bacterium powder, described bacterium powder prepares with Bacillus licheniformis claimed in claim 1.
4. the preparation method of bacterium powder claimed in claim 3, its step is as follows: with Bacillus licheniformis activation claimed in claim 1, enlarged culturing, through 35 ~ 37 ℃ of liquid state fermentations, the centrifugal collection of 5000 ~ 8000rpm bacterium mud, add the light calcium carbonate carrier, the bacterium powder that the bacterium amount is 200 ~ 1,000 hundred million/g is made in oven dry.
5. bacterium powder claimed in claim 3 is as the application of fodder additives.
6. application as claimed in claim 5 is to be that 0.01% ~ 0.5% ratio is added in the animal and fowl fodder by mass percentage with the bacterium powder.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201210488681.1A CN102994426B (en) | 2012-11-26 | 2012-11-26 | Bacillus licheniformis and applications thereof |
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| CN201210488681.1A CN102994426B (en) | 2012-11-26 | 2012-11-26 | Bacillus licheniformis and applications thereof |
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| CN102994426B CN102994426B (en) | 2014-03-12 |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103667128A (en) * | 2013-11-27 | 2014-03-26 | 北京昕大洋科技发展有限公司 | Bacillus licheniformis and its use |
| CN109722397A (en) * | 2018-12-10 | 2019-05-07 | 中国农业科学院棉花研究所 | Bacillus cercus YUPP-10 microbial inoculum and its preparation method and application |
| CN110862944A (en) * | 2019-11-29 | 2020-03-06 | 青岛蔚蓝生物集团有限公司 | Bacillus licheniformis and application thereof in removal of zearalenone |
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| CN101210228A (en) * | 2006-12-29 | 2008-07-02 | 新疆农业科学院微生物应用研究所 | Bacillus licheniformis for degrading cellulose and application thereof in straw fermentation |
| CN102293340A (en) * | 2011-08-17 | 2011-12-28 | 北京大北农科技集团股份有限公司 | Compound premix able to improve immune function for pigs |
| CN102533587A (en) * | 2011-11-28 | 2012-07-04 | 北京大北农科技集团股份有限公司 | Bacillus licheniformis and fungicide, feed additive and premix thereof |
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2012
- 2012-11-26 CN CN201210488681.1A patent/CN102994426B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101210228A (en) * | 2006-12-29 | 2008-07-02 | 新疆农业科学院微生物应用研究所 | Bacillus licheniformis for degrading cellulose and application thereof in straw fermentation |
| CN102293340A (en) * | 2011-08-17 | 2011-12-28 | 北京大北农科技集团股份有限公司 | Compound premix able to improve immune function for pigs |
| CN102533587A (en) * | 2011-11-28 | 2012-07-04 | 北京大北农科技集团股份有限公司 | Bacillus licheniformis and fungicide, feed additive and premix thereof |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103667128A (en) * | 2013-11-27 | 2014-03-26 | 北京昕大洋科技发展有限公司 | Bacillus licheniformis and its use |
| CN103667128B (en) * | 2013-11-27 | 2015-09-16 | 北京昕大洋科技发展有限公司 | A kind of Bacillus licheniformis and uses thereof |
| CN109722397A (en) * | 2018-12-10 | 2019-05-07 | 中国农业科学院棉花研究所 | Bacillus cercus YUPP-10 microbial inoculum and its preparation method and application |
| CN110862944A (en) * | 2019-11-29 | 2020-03-06 | 青岛蔚蓝生物集团有限公司 | Bacillus licheniformis and application thereof in removal of zearalenone |
| CN110862944B (en) * | 2019-11-29 | 2022-08-30 | 青岛蔚蓝生物集团有限公司 | Bacillus licheniformis and application thereof in removal of zearalenone |
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