CN105483050A - Lactobacillus plantarum strain and application thereof in fermented feed - Google Patents
Lactobacillus plantarum strain and application thereof in fermented feed Download PDFInfo
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- CN105483050A CN105483050A CN201511029973.9A CN201511029973A CN105483050A CN 105483050 A CN105483050 A CN 105483050A CN 201511029973 A CN201511029973 A CN 201511029973A CN 105483050 A CN105483050 A CN 105483050A
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- 240000006024 Lactobacillus plantarum Species 0.000 title claims abstract description 17
- 235000013965 Lactobacillus plantarum Nutrition 0.000 title abstract description 7
- 229940072205 lactobacillus plantarum Drugs 0.000 title abstract description 7
- 238000004321 preservation Methods 0.000 claims abstract description 9
- 239000000843 powder Substances 0.000 claims description 34
- 241000196324 Embryophyta Species 0.000 claims description 26
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 22
- 235000013312 flour Nutrition 0.000 claims description 19
- 239000007788 liquid Substances 0.000 claims description 18
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 claims description 16
- 239000000463 material Substances 0.000 claims description 14
- 244000068988 Glycine max Species 0.000 claims description 13
- 235000010469 Glycine max Nutrition 0.000 claims description 13
- 235000007164 Oryza sativa Nutrition 0.000 claims description 13
- 238000004458 analytical method Methods 0.000 claims description 13
- 244000005700 microbiome Species 0.000 claims description 13
- 235000009566 rice Nutrition 0.000 claims description 13
- 210000000582 semen Anatomy 0.000 claims description 12
- 235000013339 cereals Nutrition 0.000 claims description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- 238000000855 fermentation Methods 0.000 claims description 10
- 230000004151 fermentation Effects 0.000 claims description 10
- 239000000203 mixture Substances 0.000 claims description 10
- 230000001580 bacterial effect Effects 0.000 claims description 8
- 235000012054 meals Nutrition 0.000 claims description 8
- 239000002054 inoculum Substances 0.000 claims description 6
- 238000002156 mixing Methods 0.000 claims description 6
- 239000011782 vitamin Substances 0.000 claims description 5
- 235000013343 vitamin Nutrition 0.000 claims description 5
- 229940088594 vitamin Drugs 0.000 claims description 5
- 229930003231 vitamin Natural products 0.000 claims description 5
- 235000019733 Fish meal Nutrition 0.000 claims description 4
- 244000017020 Ipomoea batatas Species 0.000 claims description 4
- 235000002678 Ipomoea batatas Nutrition 0.000 claims description 4
- 244000046052 Phaseolus vulgaris Species 0.000 claims description 4
- 235000010627 Phaseolus vulgaris Nutrition 0.000 claims description 4
- 235000019764 Soybean Meal Nutrition 0.000 claims description 4
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 4
- 229930006000 Sucrose Natural products 0.000 claims description 4
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 4
- 241000482268 Zea mays subsp. mays Species 0.000 claims description 4
- 150000001413 amino acids Chemical class 0.000 claims description 4
- YYRMJZQKEFZXMX-UHFFFAOYSA-L calcium bis(dihydrogenphosphate) Chemical compound [Ca+2].OP(O)([O-])=O.OP(O)([O-])=O YYRMJZQKEFZXMX-UHFFFAOYSA-L 0.000 claims description 4
- 239000001506 calcium phosphate Substances 0.000 claims description 4
- 239000004467 fishmeal Substances 0.000 claims description 4
- 229910000150 monocalcium phosphate Inorganic materials 0.000 claims description 4
- 235000019691 monocalcium phosphate Nutrition 0.000 claims description 4
- 150000003839 salts Chemical class 0.000 claims description 4
- 239000004455 soybean meal Substances 0.000 claims description 4
- 239000004575 stone Substances 0.000 claims description 4
- 235000013619 trace mineral Nutrition 0.000 claims description 4
- 239000011573 trace mineral Substances 0.000 claims description 4
- 150000003722 vitamin derivatives Chemical class 0.000 claims description 4
- 235000001014 amino acid Nutrition 0.000 claims description 2
- 235000002639 sodium chloride Nutrition 0.000 claims description 2
- 240000007594 Oryza sativa Species 0.000 claims 4
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 abstract description 18
- 239000004310 lactic acid Substances 0.000 abstract description 9
- 235000014655 lactic acid Nutrition 0.000 abstract description 9
- 239000002253 acid Substances 0.000 abstract description 7
- 230000003031 feeding effect Effects 0.000 abstract description 4
- 244000144972 livestock Species 0.000 abstract description 4
- 238000009629 microbiological culture Methods 0.000 abstract description 4
- 235000013336 milk Nutrition 0.000 abstract description 4
- 239000008267 milk Substances 0.000 abstract description 4
- 210000004080 milk Anatomy 0.000 abstract description 4
- 238000004519 manufacturing process Methods 0.000 abstract description 3
- 230000004060 metabolic process Effects 0.000 abstract description 3
- 244000052616 bacterial pathogen Species 0.000 abstract 1
- 230000003385 bacteriostatic effect Effects 0.000 abstract 1
- 230000029087 digestion Effects 0.000 abstract 1
- 230000036039 immunity Effects 0.000 abstract 1
- 230000000968 intestinal effect Effects 0.000 abstract 1
- 241000186660 Lactobacillus Species 0.000 description 25
- 229940039696 lactobacillus Drugs 0.000 description 25
- 241000894006 Bacteria Species 0.000 description 14
- 239000002609 medium Substances 0.000 description 14
- 241000209094 Oryza Species 0.000 description 9
- 230000001954 sterilising effect Effects 0.000 description 9
- 238000002474 experimental method Methods 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 230000003053 immunization Effects 0.000 description 6
- 235000015097 nutrients Nutrition 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 6
- 206010012735 Diarrhoea Diseases 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 4
- 239000005030 aluminium foil Substances 0.000 description 4
- 241001465754 Metazoa Species 0.000 description 3
- 230000000052 comparative effect Effects 0.000 description 3
- 210000000936 intestine Anatomy 0.000 description 3
- 244000144977 poultry Species 0.000 description 3
- 229940081969 saccharomyces cerevisiae Drugs 0.000 description 3
- 238000004659 sterilization and disinfection Methods 0.000 description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 2
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 230000000844 anti-bacterial effect Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 230000037396 body weight Effects 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 229910052791 calcium Inorganic materials 0.000 description 2
- 235000019621 digestibility Nutrition 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 230000000369 enteropathogenic effect Effects 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 206010025482 malaise Diseases 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 201000010538 Lactose Intolerance Diseases 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 241000344273 bacterium 36 Species 0.000 description 1
- 230000034303 cell budding Effects 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 235000020247 cow milk Nutrition 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000003640 drug residue Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 238000002386 leaching Methods 0.000 description 1
- 239000011344 liquid material Substances 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 239000006872 mrs medium Substances 0.000 description 1
- 235000019629 palatability Nutrition 0.000 description 1
- 238000009928 pasteurization Methods 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 101150080777 pheS gene Proteins 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000006041 probiotic Substances 0.000 description 1
- 230000000529 probiotic effect Effects 0.000 description 1
- 235000018291 probiotics Nutrition 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000012797 qualification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000011218 seed culture Methods 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- 230000007480 spreading Effects 0.000 description 1
- 238000003892 spreading Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- HRXKRNGNAMMEHJ-UHFFFAOYSA-K trisodium citrate Chemical compound [Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O HRXKRNGNAMMEHJ-UHFFFAOYSA-K 0.000 description 1
- 229940038773 trisodium citrate Drugs 0.000 description 1
- 239000007222 ypd medium Substances 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
- C12R2001/25—Lactobacillus plantarum
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/80—Food processing, e.g. use of renewable energies or variable speed drives in handling, conveying or stacking
- Y02P60/87—Re-use of by-products of food processing for fodder production
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicinal Chemistry (AREA)
- Fodder In General (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a lactobacillus plantarum strain. The lactobacillus plantarum strain is characterized by being preserved in the China General Microbiological Culture Collection Center (CGMCC) with the register number of CGMCCNo.11721 and the preservation date of Nov. 23rd, 2015, and the CGMCC is located in the Institute of Microbiology, Chinese Academy of Science, No.3, first yard, Beichenxi Road, Chaoyang District, Beijing. The strain has high growth performance, high acid production performance and high bacteriostatic performance, and products fermented by means of the strain have a good feeding effect. Growth of enteric pathogenic bacteria can be inhibited through lactic acid metabolism, so that the immunity of livestock is improved remarkably, the intestinal microecological environment is improved, morbidity is reduced, and the digestion rate and the milk yield are increased to a certain extent.
Description
Technical field
The present invention relates to microbe application and feed preparation manufacture field, particularly relating to a kind of for promoting the plant lactobacillus of animal immunizing power and the application in fermented feed thereof.
Background technology
Fermented feed has good palatability, safe, pollution-free, drug residue free, the feature such as easy to digest.By feeding, fermented feed can promote animal immunizing power, reduces the generation of disease, reduces diarrhea rate, mortality ratio etc., can reduce or the use of substitute antibiotics.
Plant lactobacillus fermenting process can utilize the lactose in raw material to produce lactic acid, and lactic acid is combined the absorption that can promote calcium with calcium, and therefore for the colony having lactose intolerance, be especially in and generate budding colony fast, fermented feed has great advantage.Feed creates various lactobacillus meta-bolites through lactobacillus-fermented, and comprise vitamins, enzyme and various biologically active substances etc., fermented feed is easier to digest and assimilate.But the fermentation capacity of current plant lactobacillus is limited, the production application poor effect in fermented feed.
Summary of the invention
In view of this, the object of the invention is to proposition one lactobacillus plantarum and the application in fermented feed thereof, to promote the immunizing power of animal, reduce the generation of disease, reduce diarrhea rate, mortality ratio etc.
Based on above-mentioned purpose, plant lactobacillus Lactobacillusplantarum provided by the invention is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation place is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, what preservation was registered on the books is numbered CGMCCNo.11721, and preservation date is on November 23rd, 2015.
The present invention also provides the application of a kind of above-mentioned thing Bacterium lacticum Lactobacillusplantarum in fermented feed.
In some embodiments of the invention, described application comprises:
Add water mixing in cereal materials, and obtained fermention medium, is inoculated in described plant lactobacillus Lactobacillusplantarum in described fermention medium and ferments, obtained fermented liquid;
Non-fermented raw mixture is mixed with fermented liquid, obtained fermented feed.
In some embodiments of the invention, described plant lactobacillus Lactobacillusplantarum and Saccharomyces Cerevisiae in S accharomycescerevisiae is inoculated in described fermention medium carries out cooperative fermentation.
In some embodiments of the invention, described yeast is that Saccharomyces Cerevisiae in S accharomycescerevisiaeSa-10 has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, CGMCCNo.6120.
In some embodiments of the invention, described cereal materials is popcorn, dehulled soybean meal or fermented bean dregs, sweet potato powder or analysis for soybean powder, rice flour or Semen setariae powder, whole meal flour, and the weight percentage of described popcorn, dehulled soybean meal or fermented bean dregs, sweet potato powder or analysis for soybean powder, rice flour or Semen setariae powder, whole meal flour and water is followed successively by 1 ~ 20%, 1 ~ 50%, 1 ~ 20%, 1 ~ 20%, 1 ~ 20% and 50 ~ 90%.
In some embodiments of the invention, described cereal materials is rice flour, analysis for soybean powder and Semen Maydis powder, and the weight percentage of described rice flour, analysis for soybean powder and Semen Maydis powder is followed successively by 1 ~ 30%, 1 ~ 30% and 40 ~ 80%.
In some embodiments of the invention, the inoculum size of described plant lactobacillus Lactobacillusplantarum is 1% ~ 3%.
In some embodiments of the invention, described non-fermented raw mixture comprises amino acid, VITAMIN, trace element, salt, stone flour, monocalcium phosphate, white sugar and super steam fish meal.
Bacterial classification provided by the invention growth, produce sour, antibacterial etc. in show good performance, its leavened prod has good feeding effect.Its metabolism lactic acid can suppress enteropathogenic bacteria growing, thus significantly can promote the immunizing power of livestock and poultry, improves intestine microenvironment, reduces sickness rate, and brings certain lifting to the aspect such as digestibility, milk yield.
Embodiment
For making the object, technical solutions and advantages of the present invention clearly understand, below in conjunction with specific embodiment, the present invention is described in more detail.
The separation andpreconcentration of embodiment 1 plant lactobacillus Lactobacillusplantarum
Described plant lactobacillus EM-12 bacterial strain of stating is deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center on November 23rd, 2015, deposit number is: CGMCCNo.11721, depositary institution address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode: 100101.
Plant lactobacillus EM-12 provided by the invention makes separation screening in fermentation cow's milk by oneself from the Inner Mongol four prince's flag herdsman to obtain, and concrete isolation identification step is as follows:
1) get rice meal, analysis for soybean powder and Semen Maydis powder mixs by weight the ratio of 1:1:1, get said mixture and to add water mixing according to the ratio of material water weight ratio 1:4, obtain substratum;
2) get the fermentation milk sample containing viable bacteria, the inoculum size according to 2% is inoculated in described substratum, fully shakes mixing, 36 DEG C of anaerobism, quiescent culture, incubation time 24 hours; Above-mentioned nutrient solution is repeatedly gone down to posterity 3 times in this substratum, obtains the lactobacillus strain being applicable to fermented cereal class;
3) get above-mentioned nutrient solution 1mL, inoculation gradient dilution also coats the dull and stereotyped upper 36 DEG C of cultivation 72h of MRS; Picking individual colonies meets bacterium 36 DEG C of cultivation 18h in MRS liquid nutrient medium, uses in order to preservation and qualification.
4) bacterial classification is delivered to Institute of Microorganism, Academia Sinica to identify, comprehensively analyze according to bacterium cell form, biochemical character, 16SrRNA gene order, pheS gene order etc., be accredited as plant lactobacillus (Lactobacillusplantarum).
Cellular form and the physical and chemical experiment of this bacterial strain the results are shown in following table:
16SrDNA sequencing results:
GCGTGCTAATACATGCAGTCGAACGAACTCTGGTATTGATTGGTGCTTGCATCATGATTTACATTTGAGTGAGTGGCGAACTGGTGAGTAACACGTGGGAAACCTGCCCAGAAGCGGGGGATAACACCTGGAAACAGATGCTAATACCGCATAACAACTTGGACCGCATGGTCCGAGTTTGAAAGATGGCTTCGGCTATCACTTTTGGATGGTCCCGCGGCGTATTAGCTAGATGGTGGGGTAACGGCTCACCATGGCAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGGTTTCGGCTCGTAAAACTCTGTTGTTAAAGAAGAACATATCTGAGAGTAACTGTTCAGGTATTGACGGTATTTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCTTCGGCTCAACCGAAGAAGTGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGTATGGGTAGCAAACAGGATTAGATACCCTGGTAGTCCATACCGTAAACGATGAATGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCTACGCGAAGAACCTTACCAGGTCTTGACATACTATGCAAATCTAAGAGATTAGACGTTCCCTTCGGGGACATGGATACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATTATCAGTTGCCAGCATTAAGTTGGGCACTCTGGTGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAGTTGCGAACTCGCGAGAGTAAGCTAATCTCTTAAAGCCATTCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAACACCCAAAGTCGGTGGGGTAACCTTTTAGGAACCAGCCGCCTAAGTGACAGA
PheS gene sequencing result:
GGCGACAATTACAAGACGTGCTACTACGCACGCAGACGTCTGCTGATCAGCCGCGGTCACTTGAAAATCACGATTTTTCTAAAGGACCGCTGAAGGTCTTGTCACCTGGCCGCGTTTATCGGCGTGATACGGATGATGCAACCCATTCCCATCAATTTCATCAAATTGAAGGGTTAGTCGTGGACAAGCATATTACGATGGCTGATTTGAAGGGCACCTTAATTCTGGTTGCCAAGACTTTGTTTGGCGATCAATTCGATGTTCGGCTACGGCCAAGCTTCTTTCCATTCACGGAACCATCCGTAGAAGCTGATGTAACTTGCTTTAATTGCAATGGCAAGGGCTGTGCAATCTGTAAGCAAACGGGTTGGATCGAAGTACTGGGTGCCGGCATGGTTCACCCCCACGTGTTAGAAATGTCTGGCATTGATCCAGAAGAATATGGTGGTTTTGCTTTCGGTCTAGGGA
The mensuration of embodiment 2 plant lactobacillus EM-12 thalli growth performance and product acid activity
(1) preparation of substratum:
Get a certain amount of cereal materials and be crushed to 1.0mm, to add water mixing in the ratio of material water weight ratio 1:3, then by mixed liquor high-temperature sterilization, then be cooled to about 36 DEG C, acquisition fermention medium.Wherein, described sterilising conditions is 121 DEG C of sterilizings, 20 minutes time.Described cereal materials comprises rice meal, analysis for soybean powder and Semen Maydis powder, and the weight ratio of described rice flour, analysis for soybean powder and Semen Maydis powder is 1:1:1.
(2) acquisition of fermented bacterium:
Get this lactobacillus strain of glycerine pipe preservation, be seeded to MRS substratum, 36 DEG C of quiescent culture 20 hours, obtain lactobacillus strain.Wherein, described MRS liquid nutrient medium is composed as follows: 10g peptone, 5g extractum carnis, 4g yeast leaching powder, 20g glucose, 2g dipotassium hydrogen phosphate, 5g sodium acetate, 2g trisodium citrate, 1mL tween 80,0.2g magnesium sulfate, 0.05g manganous sulfate add 1000mL distilled water, regulate pH to 6.5,121 DEG C of sterilizing 20min.
(3) by obtained lactobacillus strain by volume 2% inoculum size be inoculated in the fermention medium of step (1), 36 DEG C of standing Anaerobic culturel, incubation time 20 hours, obtain fermented liquid.Get after fermention medium dilutes 10 times, under 600nm, measure absorbance, absorbance is 0.76 ± 0.02.Get fermented liquid centrifugal mensuration pH and lactic acid content, wherein pH value is 3.80 ± 0.05, lactic acid content 7.5 ± 0.5g/L.
The mensuration of comparative example 2 plant lactobacillus type strain product acid activity
Get China Committee for Culture Collection of Microorganisms's General Microbiological Culture preservation center plant lactobacillus type strain, measure its product acid activity according to the step (1) of embodiment 2, (2) and (3).
(1) with the step (1) of embodiment 2.
(2) with the step (2) of embodiment 2.
(3) get after seed culture medium dilutes 10 times, under 600nm, measure absorbance, absorbance is 0.62 ± 0.04; Get fermented liquid centrifugal mensuration pH and lactic acid content, wherein pH value is 4.10 ± 0.05, lactic acid content 6.0 ± 0.5g/L.
Visible, plant lactobacillus EM-12 provided by the invention has higher product acid activity, can make an addition in feed as souring agent.
The guaranteeing the quality of embodiment 3 plant lactobacillus EM-12, antipollution experiment
(1) non-fermented raw mixture is prepared: by weight percentage, amino acid 2%, VITAMIN 0.5%, trace element 0.5%, salt 1.0%, stone flour 3.0%, monocalcium phosphate 3.0%, white sugar 30% and super steam fish meal 60% are mixed, and to be crushed to granularity be 1.0mm, obtained non-fermented raw mixture.
(2) fermented liquid is prepared according to the step of embodiment 2.
(3) be 70%, 30% to mix fermented liquid and non-fermented raw mixture according to weight percentage, obtain fermented soln.
(4) adopt plastics Soft Roll to pack, adopt the general pasteurization of food service industry to carry out sterilizing, sterilization time controlled at 60 minutes.
(5) guarantee the quality, antipollution experiment
1. the fermented soln of acquisition is poured in triangular flask, the uncovered incubator being placed in 30 DEG C of juxtaposition; 2. solution is poured in aluminium foil bag, and heat sealing, be placed in 30 DEG C of incubators.Wherein, the feed liquid 1. keeps 72 hours, and free from extraneous odour, pH are unchanged.2. without swollen bag, free from extraneous odour, pH were unchanged in 72 hours for the aluminium foil bag in.
What comparative example 3 plant lactobacillus type strain product was sour guarantees the quality, antipollution experiment
Get China Committee for Culture Collection of Microorganisms's General Microbiological Culture preservation center plant lactobacillus type strain, carry out guaranteeing the quality according to the step (1) of embodiment 3, (2), (3), (4) and (5), antipollution experiment.
(1)-(4) are with step (1)-(4) of embodiment 3.
(5) guarantee the quality, antipollution experiment
1. the fermented soln of acquisition is poured in triangular flask, the uncovered incubator being placed in 30 DEG C of juxtaposition; 2. solution is poured in aluminium foil bag, and heat sealing, be placed in 30 DEG C of incubators.Wherein, the feed liquid 1. keeps 72 hours, and smell acid is smelly, and pH significantly declines.2. the 72 hours obvious swollen bags of the aluminium foil bag in, smell acid is smelly, pH significantly declines.
Visible, plant lactobacillus EM-12 provided by the invention have guarantee the quality preferably, contamination resistance.
It should be noted that, in another embodiment, the preparation of described non-fermented raw mixture comprises the following steps: by weight percentage, amino acid 3%, VITAMIN 0.4%, trace element 0.6%, salt 1.5%, stone flour 2.3%, monocalcium phosphate 3.6%, white sugar 38.6% and super steam fish meal 50% are mixed, and to be crushed to granularity be 1.2mm, obtained non-fermented raw mixture.
The feeding effect of embodiment 4 fermented feed
The fermented feed prepared by aforesaid method is used for the feeding experiment of weanling pig, and the uniform 28 age in days weanling pigs of Stochastic choice 100 health, body weight, are divided into control group and test group, often organizes 5 repetitions, each repetition 10,14 days trial periods.Control group fed powder, test group within first 4 days, feed fermentation liquid sucking pig material, within 5-7 days, be transited into the powder identical with control group gradually, the powder identical with control group of feeding for 8-14 days.The body weight of statistics piglet and diarrhoea data are as following table.
As seen from the above table, the piglet of liquid material of feeding comparatively is fed the many weightening finishes 9.0% (1.29kg) of piglet of powder, and diarrhea rate reduces by 70.6%.
The present invention significantly can promote the immunizing power of livestock and poultry, improves intestine microenvironment, reduces diarrhea rate.
Embodiment 5 works in coordination with the fermentation situation of yeast
Spreading cultivation of fermented bacterium: select yeast and plant lactobacillus EM-12, cultivate spread cultivation through primary inclined plane cultivation, two-stage liquid respectively, step is as follows:
(1) primary inclined plane is cultivated
Bacterium lacticum is transferred in MRS medium slant, cultivate 24 hours in 36 DEG C; Yeast saccharomyces cerevisiae is transferred in YPD medium slant, cultivate 20 hours in 30 DEG C;
(2) secondary liquid is cultivated
Get the fresh Bacterium lacticum inclined plane inoculating that step (1) obtains to cultivate in liquid nutrient medium, seal with aseptic sealed membrane, be placed in constant incubator, 36 DEG C of quiescent culture 20h obtain one-level Bacterium lacticum seed liquor, get primary seed solution in 5% ratio be seeded in secondary seed medium, the same first order seed of culture condition, obtains the seed liquor of Bacterium lacticum;
Getting the fresh yeast saccharomyces cerevisiae inclined-plane that step (1) obtains is forwarded in liquid nutrient medium, 30 DEG C of shaking tables cultivate 18 hours, obtain one-level yeast saccharomyces cerevisiae seed liquor, get primary seed solution and be inoculated in secondary seed medium by 5%, the same first order seed of culture condition; Obtain the seed liquor of yeast saccharomyces cerevisiae.
(3) preparation of substratum:
Get a certain amount of cereal materials and be crushed to 1.0mm, to add water mixing in the ratio of material water weight ratio 1:4, by mixed liquor high-temperature sterilization, and be cooled to about 36 DEG C, acquisition fermention medium.Wherein, described sterilising conditions is 121 DEG C of sterilizings, 20 minutes time.Described cereal materials comprises rice meal, analysis for soybean powder and Semen Maydis powder, and the weight ratio of described rice flour, analysis for soybean powder and whole meal flour is 3:1:1.
(4) get the seed liquor of yeast saccharomyces cerevisiae, the seed liquor of lactobacillus strain respectively, access same fermention medium by the inoculum size of total inoculum size 4%.Culture temperature 34 DEG C, shaking speed 200rpm.Wherein, yeast and Bacterium lacticum inoculate volume ratio 1:1.Ferment 20 hours, yeast viable count is 1.1 × 10
8cfu/ml, living preparation of lactobacillus number 2.2 × 10
9cfu/ml.
In the present embodiment, described yeast is yeast saccharomyces cerevisiae (Saccharomycescerevisiae) Sa-10, has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, CGMCCNo.6120.
Comparative example 5
Plant lactobacillus EM-12 is replaced with China Committee for Culture Collection of Microorganisms's General Microbiological Culture preservation center plant lactobacillus type strain, other steps are with embodiment 5.Ferment 20 hours, yeast viable count is 0.8 × 10
8cfu/ml, living preparation of lactobacillus number 1.8 × 10
9cfu/ml.
Visible, this bacterial strain has superior growth performance more when the probiotic bacterium of collaborative yeast class carries out fermentation culture, is therefore more suitable for the cooperative fermentation with yeast class bacterial classification, having and more having research and development, using potentiality when preparing multiple bacteria compound fermentation feed.
Bacterial classification provided by the invention growth, produce sour, antibacterial etc. in show good performance, its leavened prod has good feeding effect.Its metabolism lactic acid can suppress enteropathogenic bacteria growing, thus significantly can promote the immunizing power of livestock and poultry, improves intestine microenvironment, reduces sickness rate, and brings certain lifting to the aspect such as digestibility, milk yield.
Those of ordinary skill in the field are to be understood that: the discussion of above any embodiment is only exemplary, and not intended to be implies that the scope of the present disclosure (comprising claim) is limited to these examples; Under thinking of the present invention, also can combine between technical characteristic in above embodiment or different embodiment, step can realize with random order, and there are other changes many of different aspect of the present invention as above, and they do not provide in details for the sake of simplicity.Therefore, within the spirit and principles in the present invention all, any omission made, amendment, equivalent replacement, improvement etc., all should be included within protection scope of the present invention.
Claims (9)
1. a lactobacillus plantarum Lactobacillusplantarum, it is characterized in that, described bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation place is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, what preservation was registered on the books is numbered CGMCCNo.11721, and preservation date is on November 23rd, 2015.
2. the application of a plant lactobacillus Lactobacillusplantarum as claimed in claim 1 in fermented feed.
3. application according to claim 2, is characterized in that, comprising:
Add water mixing in cereal materials, and obtained fermention medium, is inoculated in described plant lactobacillus Lactobacillusplantarum in described fermention medium and ferments, obtained fermented liquid;
Non-fermented raw mixture is mixed with fermented liquid, obtained fermented feed.
4. application according to claim 3, is characterized in that, is all inoculated in described fermention medium by described plant lactobacillus Lactobacillusplantarum and Saccharomyces Cerevisiae in S accharomycescerevisiae and carries out cooperative fermentation.
5. application according to claim 4, is characterized in that, described yeast is that Saccharomyces Cerevisiae in S accharomycescerevisiaeSa-10 has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, CGMCCNo.6120.
6. application according to claim 3, it is characterized in that, described cereal materials is popcorn, dehulled soybean meal or fermented bean dregs, sweet potato powder or analysis for soybean powder, rice flour or Semen setariae powder, whole meal flour, and the weight percentage of described popcorn, dehulled soybean meal or fermented bean dregs, sweet potato powder or analysis for soybean powder, rice flour or Semen setariae powder, whole meal flour and water is followed successively by 1 ~ 20%, 1 ~ 50%, 1 ~ 20%, 1 ~ 20%, 1 ~ 20% and 50 ~ 90%.
7. application according to claim 3, is characterized in that, described cereal materials is rice flour, analysis for soybean powder and Semen Maydis powder, and the weight percentage of described rice flour, analysis for soybean powder and Semen Maydis powder is followed successively by 1 ~ 30%, 1 ~ 30% and 40 ~ 80%.
8. the application according to claim 3 or 4, is characterized in that, the inoculum size of described plant lactobacillus Lactobacillusplantarum is 1% ~ 3%.
9. application according to claim 3, is characterized in that, described non-fermented raw mixture comprises amino acid, VITAMIN, trace element, salt, stone flour, monocalcium phosphate, white sugar and super steam fish meal.
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Denomination of invention: A Plant Lactobacillus Strain and Its Application in Fermented Feed Granted publication date: 20181218 Pledgee: Beijing technology intellectual property financing Company limited by guarantee Pledgor: BEIJING ENHALOR INTERNATIONAL TECH CO.,LTD. Registration number: Y2024990000166 |