CN116333912A - Bacillus coagulans with broad-spectrum antibacterial effect and application thereof - Google Patents
Bacillus coagulans with broad-spectrum antibacterial effect and application thereof Download PDFInfo
- Publication number
- CN116333912A CN116333912A CN202211395783.9A CN202211395783A CN116333912A CN 116333912 A CN116333912 A CN 116333912A CN 202211395783 A CN202211395783 A CN 202211395783A CN 116333912 A CN116333912 A CN 116333912A
- Authority
- CN
- China
- Prior art keywords
- bacillus coagulans
- strain
- bacillus
- nitrogen
- broad
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 241000193749 Bacillus coagulans Species 0.000 title claims abstract description 92
- 229940054340 bacillus coagulans Drugs 0.000 title claims abstract description 92
- 230000000844 anti-bacterial effect Effects 0.000 title abstract description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 9
- 239000003674 animal food additive Substances 0.000 claims abstract description 8
- 238000004519 manufacturing process Methods 0.000 claims description 14
- 241000894006 Bacteria Species 0.000 claims description 9
- 108090000623 proteins and genes Proteins 0.000 claims description 9
- 238000002360 preparation method Methods 0.000 claims description 7
- 108020004465 16S ribosomal RNA Proteins 0.000 claims description 6
- 102000004169 proteins and genes Human genes 0.000 claims description 5
- 108700022487 rRNA Genes Proteins 0.000 claims description 5
- 238000001840 matrix-assisted laser desorption--ionisation time-of-flight mass spectrometry Methods 0.000 claims description 4
- 239000000022 bacteriostatic agent Substances 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims description 2
- 230000000694 effects Effects 0.000 abstract description 20
- 238000012216 screening Methods 0.000 abstract description 15
- JVMRPSJZNHXORP-UHFFFAOYSA-N ON=O.ON=O.ON=O.N Chemical compound ON=O.ON=O.ON=O.N JVMRPSJZNHXORP-UHFFFAOYSA-N 0.000 abstract description 11
- 244000052616 bacterial pathogen Species 0.000 abstract description 11
- MBMQEIFVQACCCH-UHFFFAOYSA-N trans-Zearalenon Natural products O=C1OC(C)CCCC(=O)CCCC=CC2=CC(O)=CC(O)=C21 MBMQEIFVQACCCH-UHFFFAOYSA-N 0.000 abstract description 9
- MBMQEIFVQACCCH-QBODLPLBSA-N zearalenone Chemical compound O=C1O[C@@H](C)CCCC(=O)CCC\C=C\C2=CC(O)=CC(O)=C21 MBMQEIFVQACCCH-QBODLPLBSA-N 0.000 abstract description 9
- 241000607598 Vibrio Species 0.000 abstract description 7
- XKMRRTOUMJRJIA-UHFFFAOYSA-N ammonia nh3 Chemical compound N.N XKMRRTOUMJRJIA-UHFFFAOYSA-N 0.000 abstract description 7
- 241000193468 Clostridium perfringens Species 0.000 abstract description 6
- 241000588724 Escherichia coli Species 0.000 abstract description 6
- 241000607142 Salmonella Species 0.000 abstract description 6
- 230000015556 catabolic process Effects 0.000 abstract description 6
- 238000006731 degradation reaction Methods 0.000 abstract description 6
- 244000005700 microbiome Species 0.000 abstract description 6
- 241000191967 Staphylococcus aureus Species 0.000 abstract description 5
- 238000009360 aquaculture Methods 0.000 abstract description 5
- 244000144974 aquaculture Species 0.000 abstract description 5
- 241000272525 Anas platyrhynchos Species 0.000 abstract description 4
- 241000588771 Morganella <proteobacterium> Species 0.000 abstract description 3
- 230000005764 inhibitory process Effects 0.000 abstract description 3
- 244000144972 livestock Species 0.000 abstract description 3
- 244000144977 poultry Species 0.000 abstract description 3
- -1 bacteriostats Substances 0.000 abstract 1
- 230000001112 coagulating effect Effects 0.000 abstract 1
- 238000004321 preservation Methods 0.000 abstract 1
- 239000001963 growth medium Substances 0.000 description 19
- 238000012360 testing method Methods 0.000 description 18
- 102000004190 Enzymes Human genes 0.000 description 15
- 108090000790 Enzymes Proteins 0.000 description 15
- 229940088598 enzyme Drugs 0.000 description 15
- 206010012735 Diarrhoea Diseases 0.000 description 14
- 239000003242 anti bacterial agent Substances 0.000 description 14
- 229940088710 antibiotic agent Drugs 0.000 description 14
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 12
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 12
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 12
- 230000001580 bacterial effect Effects 0.000 description 11
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 10
- 241000193830 Bacillus <bacterium> Species 0.000 description 9
- 241000238557 Decapoda Species 0.000 description 9
- 230000000968 intestinal effect Effects 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- 241001465754 Metazoa Species 0.000 description 8
- 241000282887 Suidae Species 0.000 description 8
- 230000034994 death Effects 0.000 description 8
- 238000000855 fermentation Methods 0.000 description 8
- 230000004151 fermentation Effects 0.000 description 8
- 239000007788 liquid Substances 0.000 description 8
- 238000000034 method Methods 0.000 description 8
- 239000006041 probiotic Substances 0.000 description 8
- 235000018291 probiotics Nutrition 0.000 description 8
- 239000003833 bile salt Substances 0.000 description 7
- 230000002354 daily effect Effects 0.000 description 7
- 230000012010 growth Effects 0.000 description 7
- 239000002609 medium Substances 0.000 description 7
- 150000007524 organic acids Chemical class 0.000 description 7
- 230000004083 survival effect Effects 0.000 description 7
- 241000607272 Vibrio parahaemolyticus Species 0.000 description 6
- 239000002253 acid Substances 0.000 description 6
- 230000008901 benefit Effects 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 229910052757 nitrogen Inorganic materials 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 239000011780 sodium chloride Substances 0.000 description 6
- 206010059866 Drug resistance Diseases 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 5
- 230000002401 inhibitory effect Effects 0.000 description 5
- 239000004310 lactic acid Substances 0.000 description 5
- 235000014655 lactic acid Nutrition 0.000 description 5
- 238000001819 mass spectrum Methods 0.000 description 5
- 239000004382 Amylase Substances 0.000 description 4
- 102000013142 Amylases Human genes 0.000 description 4
- 108010065511 Amylases Proteins 0.000 description 4
- 108091005804 Peptidases Proteins 0.000 description 4
- 241000700605 Viruses Species 0.000 description 4
- 235000019418 amylase Nutrition 0.000 description 4
- 229940093761 bile salts Drugs 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 235000021050 feed intake Nutrition 0.000 description 4
- 230000036039 immunity Effects 0.000 description 4
- 239000007924 injection Substances 0.000 description 4
- 238000002347 injection Methods 0.000 description 4
- 230000001105 regulatory effect Effects 0.000 description 4
- 231100000419 toxicity Toxicity 0.000 description 4
- 230000001988 toxicity Effects 0.000 description 4
- 108010059892 Cellulase Proteins 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- 239000004365 Protease Substances 0.000 description 3
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 3
- 238000010521 absorption reaction Methods 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 229940106157 cellulase Drugs 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 238000011156 evaluation Methods 0.000 description 3
- 239000012458 free base Substances 0.000 description 3
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 3
- 210000004211 gastric acid Anatomy 0.000 description 3
- 230000002496 gastric effect Effects 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- 230000006799 invasive growth in response to glucose limitation Effects 0.000 description 3
- 239000006916 nutrient agar Substances 0.000 description 3
- 235000005985 organic acids Nutrition 0.000 description 3
- 230000001737 promoting effect Effects 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 208000031295 Animal disease Diseases 0.000 description 2
- 241000194108 Bacillus licheniformis Species 0.000 description 2
- 244000063299 Bacillus subtilis Species 0.000 description 2
- 235000014469 Bacillus subtilis Nutrition 0.000 description 2
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 2
- 206010010774 Constipation Diseases 0.000 description 2
- 206010012742 Diarrhoea infectious Diseases 0.000 description 2
- ULGZDMOVFRHVEP-RWJQBGPGSA-N Erythromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 ULGZDMOVFRHVEP-RWJQBGPGSA-N 0.000 description 2
- 238000003794 Gram staining Methods 0.000 description 2
- 206010018910 Haemolysis Diseases 0.000 description 2
- 241000238553 Litopenaeus vannamei Species 0.000 description 2
- 241000607683 Salmonella enterica subsp. enterica serovar Pullorum Species 0.000 description 2
- 241000607594 Vibrio alginolyticus Species 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000003115 biocidal effect Effects 0.000 description 2
- 229910052791 calcium Inorganic materials 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 208000001848 dysentery Diseases 0.000 description 2
- 230000004064 dysfunction Effects 0.000 description 2
- 230000037406 food intake Effects 0.000 description 2
- 210000004051 gastric juice Anatomy 0.000 description 2
- 230000008588 hemolysis Effects 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 230000000529 probiotic effect Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 210000002784 stomach Anatomy 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- KDYFGRWQOYBRFD-UHFFFAOYSA-N succinic acid Chemical compound OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 2
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 description 1
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 description 1
- 241000607528 Aeromonas hydrophila Species 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 208000035143 Bacterial infection Diseases 0.000 description 1
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical class [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 1
- 208000035240 Disease Resistance Diseases 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 241000607473 Edwardsiella <enterobacteria> Species 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 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
- 241000238631 Hexapoda Species 0.000 description 1
- 241000186660 Lactobacillus Species 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
- 102000004882 Lipase Human genes 0.000 description 1
- 108090001060 Lipase Proteins 0.000 description 1
- 239000004367 Lipase Substances 0.000 description 1
- 239000004100 Oxytetracycline Substances 0.000 description 1
- 241000589540 Pseudomonas fluorescens Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- 108010059993 Vancomycin Proteins 0.000 description 1
- CKUAXEQHGKSLHN-UHFFFAOYSA-N [C].[N] Chemical compound [C].[N] CKUAXEQHGKSLHN-UHFFFAOYSA-N 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 238000005273 aeration Methods 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000009604 anaerobic growth Effects 0.000 description 1
- 238000009635 antibiotic susceptibility testing Methods 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 208000022362 bacterial infectious disease Diseases 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- CYDMQBQPVICBEU-UHFFFAOYSA-N chlorotetracycline Natural products C1=CC(Cl)=C2C(O)(C)C3CC4C(N(C)C)C(O)=C(C(N)=O)C(=O)C4(O)C(O)=C3C(=O)C2=C1O CYDMQBQPVICBEU-UHFFFAOYSA-N 0.000 description 1
- 229960004475 chlortetracycline Drugs 0.000 description 1
- CYDMQBQPVICBEU-XRNKAMNCSA-N chlortetracycline Chemical compound C1=CC(Cl)=C2[C@](O)(C)[C@H]3C[C@H]4[C@H](N(C)C)C(O)=C(C(N)=O)C(=O)[C@@]4(O)C(O)=C3C(=O)C2=C1O CYDMQBQPVICBEU-XRNKAMNCSA-N 0.000 description 1
- 235000019365 chlortetracycline Nutrition 0.000 description 1
- 229960002227 clindamycin Drugs 0.000 description 1
- KDLRVYVGXIQJDK-AWPVFWJPSA-N clindamycin Chemical compound CN1C[C@H](CCC)C[C@H]1C(=O)N[C@H]([C@H](C)Cl)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 KDLRVYVGXIQJDK-AWPVFWJPSA-N 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 239000002361 compost Substances 0.000 description 1
- IQFVPQOLBLOTPF-HKXUKFGYSA-L congo red Chemical compound [Na+].[Na+].C1=CC=CC2=C(N)C(/N=N/C3=CC=C(C=C3)C3=CC=C(C=C3)/N=N/C3=C(C4=CC=CC=C4C(=C3)S([O-])(=O)=O)N)=CC(S([O-])(=O)=O)=C21 IQFVPQOLBLOTPF-HKXUKFGYSA-L 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 102000038379 digestive enzymes Human genes 0.000 description 1
- 108091007734 digestive enzymes Proteins 0.000 description 1
- 230000035622 drinking Effects 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 201000006549 dyspepsia Diseases 0.000 description 1
- 210000002969 egg yolk Anatomy 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 229960003276 erythromycin Drugs 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 238000001125 extrusion Methods 0.000 description 1
- 238000009313 farming Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000004727 humoral immunity Effects 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000009654 indole test Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 239000007927 intramuscular injection Substances 0.000 description 1
- 238000010255 intramuscular injection Methods 0.000 description 1
- 229910052740 iodine Inorganic materials 0.000 description 1
- 239000011630 iodine Substances 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 229950007634 kitasamycin Drugs 0.000 description 1
- 229940039696 lactobacillus Drugs 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- XYJOGTQLTFNMQG-KJHBSLKPSA-N leucomycin V Chemical compound CO[C@H]1[C@H](O)CC(=O)O[C@H](C)C\C=C\C=C\[C@H](O)[C@H](C)C[C@H](CC=O)[C@@H]1O[C@H]1[C@H](O)[C@@H](N(C)C)[C@H](O[C@@H]2O[C@@H](C)[C@H](O)[C@](C)(O)C2)[C@@H](C)O1 XYJOGTQLTFNMQG-KJHBSLKPSA-N 0.000 description 1
- 235000019421 lipase Nutrition 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 239000001630 malic acid Substances 0.000 description 1
- 235000011090 malic acid Nutrition 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 230000007269 microbial metabolism Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000006919 modified lb Substances 0.000 description 1
- 230000001937 non-anti-biotic effect Effects 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 229960000625 oxytetracycline Drugs 0.000 description 1
- IWVCMVBTMGNXQD-PXOLEDIWSA-N oxytetracycline Chemical compound C1=CC=C2[C@](O)(C)[C@H]3[C@H](O)[C@H]4[C@H](N(C)C)C(O)=C(C(N)=O)C(=O)[C@@]4(O)C(O)=C3C(=O)C2=C1O IWVCMVBTMGNXQD-PXOLEDIWSA-N 0.000 description 1
- 235000019366 oxytetracycline Nutrition 0.000 description 1
- 238000009304 pastoral farming Methods 0.000 description 1
- 230000008855 peristalsis Effects 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
- 239000002574 poison Substances 0.000 description 1
- 238000012257 pre-denaturation Methods 0.000 description 1
- 235000013406 prebiotics Nutrition 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 235000019260 propionic acid Nutrition 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- IUVKMZGDUIUOCP-BTNSXGMBSA-N quinbolone Chemical compound O([C@H]1CC[C@H]2[C@H]3[C@@H]([C@]4(C=CC(=O)C=C4CC3)C)CC[C@@]21C)C1=CCCC1 IUVKMZGDUIUOCP-BTNSXGMBSA-N 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 230000007480 spreading Effects 0.000 description 1
- 238000003892 spreading Methods 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000001384 succinic acid Substances 0.000 description 1
- 235000019722 synbiotics Nutrition 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- IWVCMVBTMGNXQD-UHFFFAOYSA-N terramycin dehydrate Natural products C1=CC=C2C(O)(C)C3C(O)C4C(N(C)C)C(O)=C(C(N)=O)C(=O)C4(O)C(O)=C3C(=O)C2=C1O IWVCMVBTMGNXQD-UHFFFAOYSA-N 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- MYPYJXKWCTUITO-LYRMYLQWSA-N vancomycin Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C2C=C3C=C1OC1=CC=C(C=C1Cl)[C@@H](O)[C@H](C(N[C@@H](CC(N)=O)C(=O)N[C@H]3C(=O)N[C@H]1C(=O)N[C@H](C(N[C@@H](C3=CC(O)=CC(O)=C3C=3C(O)=CC=C1C=3)C(O)=O)=O)[C@H](O)C1=CC=C(C(=C1)Cl)O2)=O)NC(=O)[C@@H](CC(C)C)NC)[C@H]1C[C@](C)(N)[C@H](O)[C@H](C)O1 MYPYJXKWCTUITO-LYRMYLQWSA-N 0.000 description 1
- 229960003165 vancomycin Drugs 0.000 description 1
- MYPYJXKWCTUITO-UHFFFAOYSA-N vancomycin Natural products O1C(C(=C2)Cl)=CC=C2C(O)C(C(NC(C2=CC(O)=CC(O)=C2C=2C(O)=CC=C3C=2)C(O)=O)=O)NC(=O)C3NC(=O)C2NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(CC(C)C)NC)C(O)C(C=C3Cl)=CC=C3OC3=CC2=CC1=C3OC1OC(CO)C(O)C(O)C1OC1CC(C)(N)C(O)C(C)O1 MYPYJXKWCTUITO-UHFFFAOYSA-N 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 150000003722 vitamin derivatives Chemical class 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- 230000004584 weight gain Effects 0.000 description 1
- 235000019786 weight gain Nutrition 0.000 description 1
Images
Classifications
-
- 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
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/22—Bacillus
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P1/00—Disinfectants; Antimicrobial compounds or mixtures thereof
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
- A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- 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
-
- 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
- C12R2001/07—Bacillus
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- Environmental Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Polymers & Plastics (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Tropical Medicine & Parasitology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Dentistry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Hydrology & Water Resources (AREA)
- Environmental & Geological Engineering (AREA)
- Water Supply & Treatment (AREA)
- Molecular Biology (AREA)
- Physiology (AREA)
- Animal Husbandry (AREA)
- Agronomy & Crop Science (AREA)
- Food Science & Technology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to the technical field of functional microorganism screening and application, in particular to a novel bacillus coagulans strainBacillus coagulans) And applications thereof. The preservation number of the coagulating spore rod is CCTCC NO: m20221235 the strain has broad-spectrum antibacterial capability and good denitrification performance, has remarkable inhibition effect on pathogenic bacteria such as escherichia coli, salmonella, clostridium perfringens, staphylococcus aureus, duck in-service Morganella, vibrio and the like, and has good denitrification effect on pathogenic bacteria such as escherichia coli, salmonella, clostridium perfringens and the likeThe removal rate of ammonia nitrogen and nitrite nitrogen reaches 100%, the zearalenone can be efficiently degraded, the degradation rate reaches 80.5%, and the ammonia nitrogen and nitrite nitrogen can be widely applied to the field of livestock and poultry or aquaculture as feed additives, bacteriostats, water regulators and the like, and the application prospect is wide.
Description
Technical Field
The invention relates to the technical field of functional microorganism screening, in particular to bacillus coagulans with a broad-spectrum antibacterial effect and application thereof.
Background
The microecological preparation is prepared by applying the microecological principle under the guidance of microecological theory and utilizing living microorganisms beneficial to a host or bioactive substances generated by microbial metabolism through a special process so as to achieve the aim of regulating the microecological balance of the organism. Microecologics are currently divided into 3 types, namely probiotics, prebiotics and synbiotics. In the 60 s of the 20 th century, with the gradual appearance of potential hazards to farming due to long-term or irregular use of antibiotics, microecologics have only received sufficient attention in livestock farming. After 90 s, aquaculture rapidly progresses, and aquaculture animal diseases are endangered, and aquatic microecologics gradually become research hotspots.
The microecological preparation is added into the feed, so that the immunity and disease resistance of animals can be improved, the production performance and the cultivation benefit of the animals are improved, the pollution to the environment caused by misuse and abuse of antibiotics and various chemical veterinary medicines can be reduced, and good ecological benefit is brought. In the aspect of aquatic products, besides the microecological preparation for feeding, the microecological preparation for regulating water and improving the bottom has the functions of purifying water quality, improving intestinal microbial communities of aquatic products, improving digestion and absorption capacity, promoting animal growth, improving the expression level of specific immune related genes, enhancing the capability of organisms to resist virus and bacterial infection and the like. Therefore, in recent years, the use of microecologics in animal production has been expanding.
The nutrition value of the feeding microecology is mainly expressed in that: 1) Regulating gastrointestinal microecological balance, and correcting intestinal flora dysfunction, thereby reducing infectious diarrhea and non-infectious diarrhea caused by bacteria, viruses and protozoa, because these diarrhea are all related to intestinal flora dysfunction; 2) Inhibiting the proliferation of pathogenic bacteria including amine-producing spoilage bacteria in the intestine, thereby maintaining optimal combination and stability of intestinal flora; 3) Synthesizing amylase, cellulase, protease and other enzymes and promoting the secretion of host digestive enzymes, thereby promoting the digestion and absorption of feed and reducing diarrhea caused by dyspepsia; 4) The vitamin is synthesized, the absorption of calcium and lactose is promoted, and the constipation and diarrhea of piglets are prevented, so that the health care effect on the piglets is realized; 5) The organic acid produced by metabolism can promote gastrointestinal peristalsis, and the microecology can moisten the stool, so that constipation of sows can be prevented; 6) The microecology can induce effective mucosal immunity and humoral immunity reaction, enhance the immunity of a host, reduce the endotoxin content in blood and reduce the occurrence of diseases.
At present, 2 main feeding micro-ecologies widely applied in animal production in China are bacillus and lactobacillus. In practical application, the 2-size feeding strain is found to have obvious advantages and has the defect of being yet to be improved when being added as an exogenous beneficial bacterium in animal intestinal tracts. The bacillus has the greatest advantages of being resistant to bile salts, gastric acids and alkali, particularly high temperature and extrusion, and can survive in granulated feed. In addition, the microbial enzyme has good enzyme production capacity and broad-spectrum antibacterial capacity. However, bacillus does not produce lactic acid, and can not grow well in anaerobic environment such as intestinal canal, so that the bacillus can not play a direct role in intestinal canal micro-ecological system, and the effect of adding probiotics as feed in feed is limited. Bacillus subtilis and Bacillus licheniformis are the main strains for producing feed bacillus preparations at present. For example, zhang Bingsheng and the like screen a bacillus subtilis SD-KC-001, which has stronger acid resistance, salt resistance and heat resistance, strong growth capacity, can produce enzymes such as amylase, protease and the like with good antibacterial capacity. Through feeding animals, the strain can improve the weight gain, reduce the drug cost, reduce the death rate and the like after being added into the feed. Zhou Yi and the like screen a bacillus licheniformis DN01 strain, have strong inhibition effects on aquatic pathogenic bacteria aeromonas hydrophila, pseudomonas fluorescens and Edwardsiella, can obviously promote the growth of crucian, and greatly improve the survival rate after toxicity attack.
At present, development of a feeding microecological strain with multiple functions, strong stability and excellent effect is still an important research and development point in the field.
Disclosure of Invention
The invention aims to provide a novel bacillus coagulansBacillus coagulans) And applications thereof. The bacillus coagulans has broad-spectrum antibacterial capability, has remarkable inhibition effect on pathogenic bacteria such as escherichia coli, salmonella, clostridium perfringens, staphylococcus aureus, duck in-service bacillus, vibrio and the like, and can be widely applied to the field of cultivation.
The bacillus coagulans provided by the invention is bacillus coagulans NR 372%Bacillus coagulans NR 372) strain which has been preserved in China center for type culture collection (CCTCC NO) of university of Wuhan in China, 8 th month of 2022: m20221235.
The bacillus coagulans NR372 strain provided by the invention has a MALDI-TOF-MS protein fingerprint shown in figure 2 and a rRNA gene fingerprint shown in figure 3.
The 16s rRNA sequence of the bacillus coagulans NR372 strain provided by the invention is SEQ ID NO:1.
the invention provides application of bacillus coagulans NR372 strain in preparation of a bacteriostat.
The bacillus coagulans NR372 strain provided by the invention is applied to water quality purification.
The bacillus coagulans NR372 strain provided by the invention is applied to feed production.
The invention also provides a feed additive, which comprises the bacillus coagulans NR372 strain.
The live bacterial amount of the bacillus coagulans NR372 strain in the feed additive is not less than 10 8 CFU/g。
The bacillus coagulans NR372 obtained by screening has strong comprehensive antibacterial effect, and has remarkable inhibitory effect on pathogenic bacteria such as escherichia coli, salmonella, clostridium perfringens, staphylococcus aureus, duck in-service Morganella, vibrio and the like, wherein the bacillus coagulans has the strongest antibacterial effect on salmonella pullorum, vibrio eurovirens and vibrio alginolyticus, and the diameter of a bacteriostasis zone reaches 2.8-3.5cm.
The bacillus coagulans NR372 has strong gastric acid tolerance, and the survival rate can reach 50% after 4 hours; the strain is completely tolerant to artificial intestinal juice, and the survival rate is up to 100% after 12 hours; the highest tolerance concentration of the strain to bile salts is 1%, and the survival rate after 4 hours is up to more than 90%.
The bacillus coagulans NR372 has strong comprehensive acid production and enzyme production capability, at least can produce 8 organic acids and 4 enzymes, and the content of lactic acid and acetic acid in fermentation liquor is highest and reaches 7309.23 ug/mL and 5501.28 ug/mL respectively; the strain has good denitrification performance, the removal rate of ammonia nitrogen and nitrite nitrogen reaches 100%, and the degradation rate of zearalenone can be efficiently degraded to 80.5%.
The bacillus coagulans NR372 can effectively improve the production performance of weaned pigs, obviously reduce the diarrhea rate of the weaned pigs and has obviously better effect than antibiotics. Compared with the treatment group 1 fed with antibiotics, the diarrhea index of piglets in the treatment group fed with bacillus coagulans is reduced by 20-40%, and the effect is very remarkable.
Bacillus coagulans NR372 can obviously reduce the death rate of Litopenaeus vannamei infected with vibrio parahaemolyticus. Compared with the toxicity attacking group, the death rate of the prawns in the probiotics treatment group fed with the bacillus coagulans NR372 is reduced by 80.1 percent, and unexpected technical effects are achieved.
The bacillus coagulans NR372 provided by the invention is widely applied to the field of livestock and poultry or aquaculture as a feed additive, a bacteriostatic agent, a water regulating agent and the like, and has a wide application prospect.
Drawings
FIG. 1 is a colony morphology of NR372 strain;
FIG. 2 is a protein mass spectrum peak diagram of NR372 strain;
FIG. 3 is a rRNA gene fingerprint.
Detailed Description
The bacillus coagulans NR372 provided by the invention meets the regulation requirement, and is identified by multiphase taxonomy, and the bacillus coagulans NR372 is a newly discovered strain. The bacillus coagulans NR372 provided by the invention has remarkable antibacterial effect on various common pathogenic bacteria and has important application value.
The applicant uses bacillus coagulans NR372 @ 4 days of 2022Bacillus coagulans NR 372) strain is preserved in China center for type culture collection (CCTCC NO) of university of Wuhan, china: m20221235.
The screening method of the present invention is not limited to the examples, but known screening methods can be used to achieve the screening purpose, and the screening description of the examples is only illustrative of the present invention and is not intended to limit the scope of the present invention. Modifications and substitutions to methods, procedures, or conditions of the present invention without departing from the spirit and nature of the invention are intended to be within the scope of the present invention.
The present invention will be described in detail with reference to specific examples.
EXAMPLE 1 isolation and screening of strains
1. Sample source:
and 20 parts of compost fermented feed.
2. Bacterial strain primary screening
1g of the sample is weighed and placed in 9 mL sterile normal saline, and evenly mixed for 10min at 200rpm to obtain bacterial suspension; placing the bacterial suspension in a water bath kettle at 80 ℃ for 10min to remove non-spore bacteria; inoculating 1% of the strain into a modified LB culture medium, culturing at 37 ℃ for 20 hours, and performing dilution coating on a modified calcium carbonate identification plate; selecting single colonies which are provided with a calcium dissolving ring and well grown, and respectively carrying out gram staining, spore staining and contact enzyme tests; and selecting 4 strains which are positive in gram staining, spore-producing and positive in contact enzyme, wherein the strains are YB95, YB115, YB202 and YB266 respectively.
3. Bacterial strain re-screening
Selecting more than 20 pathogenic bacteria which often cause animal diseases in the industries of livestock, poultry, aquatic products and pets: and performing bacteriostasis tests on escherichia coli, salmonella, clostridium perfringens, staphylococcus aureus, duck in-service bacillus, vibrio and the like, and evaluating the bacteriostasis capacity of 4 strains of YB95, YB115, YB202 and YB266 obtained through primary screening according to the diameter of a bacteriostasis ring.
The operation flow is as follows: and taking a sterile plate, pouring 18-20 mL of sterile nutrient agar culture medium, uniformly spreading the culture medium in the plate, placing the culture medium on a horizontal table top to solidify, taking the plate as a bottom layer, inverting the plate to evenly divide the region, and marking the to-be-added substance. And (3) taking a proper amount of sterile semi-solid nutrient agar culture medium (the agar content is 1%), cooling to 48-50 ℃, adding 1-2 mL of pathogenic bacteria suspension into each 50-100 mL of culture medium, and respectively adding 5mL of semi-solid nutrient agar culture medium into each plate to uniformly spread on a bottom layer to serve as a bacterial layer. And after being placed on a horizontal table for cooling, 4-6 oxford cups are uniformly arranged in each 1 plate at equal distance. The diameters of the bacteriostasis circles are measured after 0.2mL of the strain bacterial liquid which is cultured by expanding is respectively dripped into oxford cups in each double-layer plate and is cultured for 24 hours at 37 ℃, and the specific results are shown in Table 1.
TABLE 1 antibacterial Effect of strains
As shown in the results of Table 1, the YB202 strain obtained by screening has the best comprehensive antibacterial effect, and has remarkable inhibitory effect on pathogenic bacteria such as escherichia coli, salmonella, clostridium perfringens, staphylococcus aureus, morganella anatipestifer, vibrio and the like, wherein the antibacterial effect on salmonella pullorum, vibrio eurovirens and vibrio alginolyticus is strongest, and the diameter of a bacteriostasis ring reaches 2.8-3.5cm. The applicant designated this strain as NR372 and further identified and functionally evaluated.
EXAMPLE 2 identification of NR372 Strain
1. Colony morphology identification
As shown in FIG. 1, NR372 strain forms a round colony with the diameter of 2-3mm after being cultured on MRS culture medium for 24-48 hours, and the colony is white and glossy, and has moist and flat surface and neat edge; the NR372 bacteria form in rod shape, single, paired or chain arrangement, spore end growth.
2. Physiological and biochemical identification
The NR372 strain was subjected to physiological and biochemical identification, and the results are shown in Table 2.
Physiological and biochemical characteristics of NR372 strain and standard bacillus coagulansBacillus coagulans) Substantially identical, initially identified as bacillus coagulans.
TABLE 2 physiological and biochemical tables of strains
| Biochemical characterization | NR372 | Standard Bacillus coagulans |
| Indole test | - | - |
| Starch hydrolysis | + | + |
| Gelatin liquefaction | - | - |
| Yolk reaction | - | - |
| Hydrolysis of amino acids | - | - |
| Anaerobic growth | + | + |
| NaCl 2% | + | + |
| |
+ | + |
| NaCl 7% | - | - |
| |
- | - |
Note that: positive +; + a 90-100% of strains are positive; negative.
3. 16S rRNA identification
The genome of the NR372 strain was extracted using the kit. The genome was then used as a template to amplify its 16S rRNA using specific primers 27F and 1492R.
27F：5’-AGAGTTTGATCATGGCTCAG-3’；
1492R：5’-TAGGGTTACCTTACGACTT-3’。
The PCR system comprises: 0.7. Mu.l 27F, 0.7. Mu.l 14992R, 4. Mu.l template DNA, 17.5. Mu.l SuperMiX and 12.1. Mu.l water. The PCR reaction conditions were set as follows: (1) 94 ℃ for 5min; (2) pre-denaturation at 94℃for 30s; (3) 55 ℃ for 30s; (4) 72 ℃ for 1min; performing the loop of steps (2) to (4) 35; (5) at 72℃for 10min. The amplified PCR product is subjected to 1% agarose gel electrophoresis detection, and the result shows that the size of the PCR product is about 1500bp, thereby meeting the requirements.
The PCR amplified product was sent to sequencing company for sequencing, and the result shows that the 16s rRNA sequence of NR372 strain is SEQ ID NO:1. the sequence was subjected to BLAST alignment in NCBI database, which showed the highest similarity to Bacillus coagulans. Thus, the NR372 strain was preliminarily determined to be Bacillus coagulans.
4. MALDI-TOF-MS profile identification
A small amount of NR372 single colony is coated on a target plate in a film form; adding 1 mu L of lysate in the mass spectrum sample pretreatment kit, and naturally airing at room temperature; adding 1 mu L of matrix solution in the mass spectrum sample pretreatment kit to cover the sample, and naturally airing at room temperature; and (5) placing the sample target into a mass spectrometer for identification. The identification result shows that the NR372 strain is bacillus coagulans, and the protein mass spectrum peak diagram is shown in figure 2.
5. Ribo-Printer full-automatic microorganism gene fingerprint identification
And (3) performing on-machine identification on the NR372 strain according to the operation instruction of a full-automatic microorganism gene fingerprint identification system to obtain an rRNA gene fingerprint diagram of the NR372 strain, wherein the rRNA gene fingerprint diagram is shown in figure 3. By comparison with the fingerprint patterns of known standard strain libraries, the NR372 strain has a similarity with bacillus coagulans of more than 90%, so that the strain is identified as bacillus coagulans.
In summary, the applicant uses three molecular biology methods of 16S rRNA identification, MALDI-TOF-MS protein mass spectrum identification, ribo-Printer full-automatic microorganism gene fingerprint identification to identify NR372 strain, and the identification results are consistent. And then combining colony morphology and physiological and biochemical characteristics of the NR372 strain, the applicant determines that the NR372 strain is bacillus coagulans and names the bacillus coagulans NR 372%Bacillus coagulans NR372）。
The applicant has found that the above Bacillus coagulans NR372 @ 4 of 2022Bacillus coagulans NR 372) is preserved in China center for type culture collection (CCTCC NO) of university of Wuhan in China: m20221235.
EXAMPLE 3 safety evaluation of Bacillus coagulans NR372
1. Hemolysis:
the activated bacillus coagulans NR372 strain is inoculated on a blood plate, and after the blood plate is cultivated for 24 hours at the constant temperature of 37 ℃, whether a transparent hydrolysis ring is generated around a colony is observed.
The results show that: no hydrolytic circle was produced around the colonies, indicating that the Bacillus coagulans NR372 strain did not possess hemolysis.
2. Drug resistance:
the drug resistance of the strain may cause potential safety hazards in the production and application process. To prevent the presence of drug resistance of bacillus coagulans NR372, its antibiotic susceptibility was studied. The minimum inhibitory concentration (MIC value) of NR372 in 7 common antibiotics was determined according to CLSI antibiotic susceptibility test gradient dilution, the antibiotic used was Sigma standard, and the susceptibility to antibiotics was evaluated with reference to EFSA (2012) standard. See table 3 for specific results.
TABLE 3 minimum inhibitory concentration (MIC value) in antibiotics
| Chloramphenicol | Clindamycin | Gentamicin | Streptomycin | Erythromycin | Vancomycin | Tetracycline |
| 4（S） | 0.08（S） | 1（S） | 2（S） | 1（S） | 2（S） | 1（S） |
Note that: unit ug/ml, S represents sensitivity according to the strain sensitivity to antibiotics evaluation standard EFSA (2012); r represents drug resistance.
From the results in Table 3, it can be seen that the Bacillus coagulans NR372 provided by the invention is sensitive to 7 common antibiotics, and has no drug resistance.
EXAMPLE 4 Bacillus coagulans NR372 test for resistance to Artificial gastrointestinal fluids and bile salts
(1) Test for artificial gastric juice resistance
The bacillus coagulans NR372 bacterial liquid in logarithmic growth phase is prepared according to the following ratio of 1:9 volume ratio is inoculated into artificial gastric juice with pH of 2.0 and artificial intestinal juice with pH of 7.6 respectively, after standing culture is carried out for 4h and 12h respectively at 37 ℃, MRS culture medium is poured for culture, and control group is physiological saline. The viable count was compared before and after plate count, and each sample was repeated 3 times. (2) Test for bile salt resistance
The bacillus coagulans NR372 bacterial liquid in logarithmic growth phase is prepared according to the following ratio of 1:9, respectively inoculating the culture media into MRS liquid culture media with the content of bile salt of 0.1%, 0.2%, 0.3%, 0.5% and 1% by volume, standing at 37 ℃ for 4h, pouring the MRS culture media for culture, taking the culture media which are not subjected to bile salt treatment as a control group, comparing the numbers of viable bacteria before and after the culture media are counted by a flat plate, and repeating each sample for 3 times.
The above results show that: the bacillus coagulans NR372 has strong gastric acid tolerance, and the survival rate can reach 50% after 4 hours; the strain is completely tolerant to artificial intestinal juice, and the survival rate is up to 100% after 12 hours; the highest tolerance concentration of the strain to bile salts is 1%, and the survival rate after 4 hours is up to more than 90%.
EXAMPLE 5 enzymatic Properties of Bacillus coagulans NR372
Inoculating the activated bacillus coagulans NR372 into MRS liquid culture medium, and culturing for 24 hours at 37 ℃ and 220r/min to prepare a bacteria testing solution for later use.
The bacterial solutions are respectively inoculated on a screening culture medium containing amylase, protease, cellulase and lipase, and are cultured for 24 hours at 37 ℃. After the culture of the cellulase screening culture medium is finished, 0.2% Congo red solution is added for dyeing for 30min, and then 1mol/L NaCl is used for washing for 10min, and then the size of an enzyme producing ring is measured. The diameter of the enzyme-producing ring was measured after the amylase screening medium was changed in color by adding a dilute iodine solution, and the results are shown in Table 4.
TABLE 4 evaluation of Bacillus coagulans NR372 enzyme production Effect
| Enzyme species | Protease enzyme | Amylase enzyme | Cellulase enzymes | Lipase enzyme |
| Diameter of enzyme producing ring (mm) | 28 | 25 | 25 | 26 |
As can be seen from the results in Table 4, the Bacillus coagulans NR372 enzyme producing effect provided by the present invention was excellent.
EXAMPLE 6 acid Forming Properties of Bacillus coagulans NR372
6.1 pH change
The activated Bacillus coagulans NR372 was inoculated into 100mL of MRS liquid medium and cultured at 37℃and 200rpm for 24 hours. Samples were taken at 0h, 4h, 8h, 12h, 16h, 18h, 24h, respectively, to determine the pH of the fermentation broth.
The results show that: during 0-12h, the pH of the Bacillus coagulans NR372 broth dropped rapidly, and at 24h the pH dropped to 4.3.
6.2 Determination of organic acid content
And (3) centrifuging the bacillus coagulans NR372 fermentation liquor after 24h fermentation for 10min at 5000r/min, filtering the obtained supernatant with a 0.22um filter membrane, and then performing liquid chromatography to analyze the content of organic acids such as lactic acid, acetic acid and the like in the fermentation liquor. The acid standard curve was determined with the standard concentration (. Mu.g/mL) on the abscissa and the peak area on the ordinate. Substituting the peak area of the sample into a standard curve, and calculating the content of acid in the sample. The results are shown in Table 5.
TABLE 5 analysis of organic acid content in Bacillus coagulans NR372 fermentation broth
| Organic acid | Content (ug/mL) |
| Lactic acid | 7309.23 |
| Acetic acid | 5501.28 |
| Citric acid | 1600.98 |
| Succinic acid | 1534.01 |
| Formic acid | 1001.05 |
| Propionic acid | 902.98 |
| Malic acid | 3093.90 |
| Oxalic acid | 1234.20 |
As can be seen from the data in Table 5, the Bacillus coagulans NR372 provided by the invention has strong comprehensive acid production capability, at least 8 organic acids can be produced, and the contents of lactic acid and acetic acid in fermentation liquor are highest and reach 7309.23 ug/mL and 5501.28 ug/mL respectively, so that unexpected technical effects are achieved.
EXAMPLE 7 Bacillus coagulans NR372 Effect on removal of ammoniacal nitrogen and nitrite nitrogen
Bacillus coagulans NR372 was inoculated into MRS liquid medium and activated at 37℃and 200rpm for 24 hours. 1ml of the bacterial liquid was centrifuged at 4000rpm for 5min. The precipitate was washed three times with 0.9% physiological saline and inoculated into 50ml of a complex nitrogen source medium (glucose 0.1g, (NH) 4 ) 2 SO 4 0.01g，NaNO 2 0.01g，K 2 HPO 4 ·3H 2 O 1g，KH 2 PO 4 0.3g，MgSO 4 ·7H 2 O 0.25g，FeSO 4 ·7H 2 O 0.05g，MnSO 4 ·4H 2 O0.01g,NaCl 30g,pH 8.0), the initial concentration of ammoniacal nitrogen and nitrite nitrogen in the composite nitrogen source culture medium is 2mg/L, and the carbon-nitrogen ratio is 10; blank control was not doneAfter 48h of treatment and shaking culture at 37 ℃ and 150rpm, centrifuging at 4500rpm for 5min to obtain a supernatant, and measuring the content of ammonia nitrogen and nitrite nitrogen in the supernatant by referring to the method described in national standard GB 17378.4-2007. The removal rates of Bacillus coagulans NR372 for ammoniacal nitrogen and nitrite nitrogen were calculated according to the following formulas, respectively.
Ammonia nitrogen removal (%) = (X1-X2)/X1X 100%.
Nitrite nitrogen removal (%) = (Y1-Y2)/y1×100%.
X1 is the ammonia nitrogen content in the blank control after 48 hours of culture, and X2 is the ammonia nitrogen content in the test group culture medium after 48 hours of culture; y1 is the nitrite nitrogen content in the blank control after 48h of culture, and Y2 is the nitrite nitrogen content in the test group medium after 48h of culture.
The results show that the bacillus coagulans NR372 provided by the invention has good denitrification performance, and has unexpected technical effects on the removal rate of ammoniacal nitrogen and nitrite nitrogen reaching 100%.
Example 8 degradation Effect of Bacillus coagulans NR372 on zearalenone
40. Mu.L of Bacillus coagulans NR372 fermentation broth was inoculated into 4 mL of MRS liquid medium containing Zearalenone (ZEN) (10. Mu.g/mL), and incubated at 37℃and 200rpm for 48 hours with no inoculation of ZEN (10. Mu.g/mL) of MRS liquid medium as a control. According to the specification of the zearalenone ELISA detection kit, the culture solution is treated, the residual amount of ZEN in the culture solution is detected by an enzyme-labeled instrument, and the degradation rate is calculated.
Degradation rate (%) =
×100%。
The result shows that the bacillus coagulans NR372 provided by the invention can efficiently degrade zearalenone, and the degradation rate reaches 80.5%.
Example 9 application Effect of Bacillus coagulans NR372 on the prevention of diarrhea in weaned piglets
1. Experiment site:
animal houses of Qingdao university.
2. And (3) test design:
healthy commercial castrated boars 40 with average weight of 7.5 and kg are selected (26+/-1) for weaning at the day, and the average is divided into 4 treatment groups, and 10 piglets in each treatment group are fed into one colony house for 4 colony houses.
Grouping of test treatment groups:
control group: the basic diet group is free from adding any antibiotics and bacillus coagulans;
treatment group 1: basic ration + antibiotics (kitasamycin 50 g/t, oxytetracycline 50 g/t, aureomycin 75 g/t);
treatment group 2: basic ration + Bacillus coagulans NR372 (1X 10) 7 CFU/g）；
Treatment group 3: basic ration + Bacillus coagulans NR372 (5X 10) 7 CFU/g）。
The test pigs are fed according to the weaned pig management scheme, basic ration is fed for the first 3 days, free feeding and drinking are carried out, only insect expelling and immunization are carried out on the test pigs, the 4 th morning is subjected to empty stomach weighing, and the test period is 10 days. 3 times daily feeding and free drinking water are carried out in the test period, and the temperature of the pig house is reasonably controlled. Test 10d, all test pigs were tested for body weight, feed intake, and for analysis of growth performance indicators such as average daily gain, average daily feed intake, feed conversion ratio, diarrhea index, etc. The results are shown in Table 6.
The diarrhea index is calculated by taking a cage as a statistical unit as follows:
diarrhea index (%) =number of diarrhea piglets/(number of piglets×days of test) ×100%.
TABLE 6 influence of Bacillus coagulans NR372 on growth performance of weaned piglets
| Index (I) | Control | Treatment group | 1 | Treatment group 2 | Treatment group 3 |
| Initial weight (kg) | 7.52 | 7.53 | 7.51 | 7.50 | |
| Last weight (kg) | 9.21 b | 9.54 a | 9.45 a | 9.56 a | |
| Average daily gain (g/d) | 168 b | 200 a | 193 a | 206 a | |
| Average daily feed intake (g/d) | 216 ab | 242 ab | 240 ab | 258 a | |
| Feed to meat ratio | 1.29 a | 1.21 b | 1.24 ab | 1.25 ab | |
| Diarrhea index (%) | 18 a | 10 b | 8 b | 6 b |
As can be seen from the data in table 6, the average daily gain, daily feed intake, feed conversion ratio and diarrhea index were significantly improved for each treatment group fed with antibiotics and bacillus coagulans, as compared to the control group. Compared with the treatment group 1 fed with antibiotics, the diarrhea index of piglets in the treatment group fed with bacillus coagulans is reduced by 20-40%, and the effect is very remarkable. Therefore, the bacillus coagulans NR372 provided by the invention can effectively improve the production performance of weaned pigs, obviously reduce the diarrhea rate of the weaned pigs, and has obviously better effect than antibiotics.
EXAMPLE 10 protective Effect of Bacillus coagulans NR372 on vannamei
1. Source of vannamei:
some breeding factory in Laiyi city
2. Experiment site:
a blue biological aquaculture room.
3. Experiment design:
healthy, disease-free and close-specification litopenaeus vannamei was selected and randomly divided into 4 groups of 3 replicates, 10 replicates each, and 21d cultivation experiments were performed in a 60L glass aquarium. During the experiment, each day was 8:00 and 16: feeding for two times at 00, wherein the average water temperature is about 25 ℃, the salinity is about 8.0, the pH is about 8.0, the continuous trace aeration is carried out, the dissolved oxygen is kept above 5mg/L, and the test is carried out according to the following groups after 3 days of feeding without any abnormal condition.
(1) Control group: feeding a non-antibiotic basic material, and normally feeding without toxicity attack;
(2) Benefit (benefit)Group of bacteria: feeding antibiotic-free base material and bacillus coagulans NR372 (3×10) 7 CFU/g), normal feeding is carried out without toxicity attack;
(3) Attack the poison group: feeding antibiotic-free base material, intramuscular injection of Vibrio parahaemolyticus culture solution with injection concentration of 1.0X10 6 CFU/mL, injection volume 25. Mu.L;
(4) Probiotic treatment group: feeding antibiotic-free base material and bacillus coagulans NR372 (3×10) 7 CFU/g), after 3 days of continuous feeding, the Vibrio parahaemolyticus culture solution was intramuscular injected at a concentration of 1.0X10 6 CFU/mL, injection volume was 25. Mu.L. The basic material containing bacillus coagulans NR372 is fed once every 3d, and the basic material is normally fed in the rest time.
The vitality, ingestion and death of the prawns in each group are recorded every day, and the death rate is counted after 21 d. The specific results are shown in Table 7.
TABLE 7 influence of Bacillus coagulans NR372 on mortality of Vernaeus vannamei
| Grouping | Mortality rate of |
| Control group | 4.95% |
| Probiotics group | 2.35% |
| Toxin eliminating group | 66.7% |
| Probiotic treatment group | 13.3% |
From the results in table 7, the mortality of the probiotics group prawns fed with bacillus coagulans NR372 was reduced by 52.5% compared to the control group. After the prawn of the virus attack group is injected with vibrio parahaemolyticus, the prawn shows reduced ingestion, redness of appendages, empty stomach, hepatopancreatic enlargement, immobility or roaming along the wall of a cultivation barrel, and weak vitality until the prawn dies finally, and the 21d death rate is up to 66.7%; the death rate of the prawns in the probiotics treatment group fed with the bacillus coagulans NR372 before and after the injection of the vibrio parahaemolyticus is only 13.3 percent, which is reduced by 80.1 percent compared with that of the virus attack group.
The result shows that the bacillus coagulans NR372 provided by the invention can effectively improve the immunity of the prawns, inhibit the growth and proliferation of pathogenic bacteria such as vibrio parahaemolyticus and the like, greatly reduce the death rate of the prawns and achieve unexpected technical effects.
Claims (9)
1. The bacillus coagulans is characterized in that the collection number of the bacillus coagulans is CCTCC NO: m20221235.
2. The bacillus coagulans of claim 1, wherein the bacillus coagulans has a MALDI-TOF-MS protein fingerprint as shown in fig. 2 and a rRNA gene fingerprint as shown in fig. 3.
3. The bacillus coagulans of claim 1, wherein the bacillus coagulans has a 16s rRNA sequence of SEQ ID NO:1.
4. the use of bacillus coagulans according to claim 1 for the preparation of a bacteriostatic agent.
5. The use of the bacillus coagulans according to claim 1 for water purification.
6. The use of bacillus coagulans according to claim 1 in feed production.
7. A feed additive comprising the bacillus coagulans of claim 1.
8. The feed additive according to claim 7, wherein the viable bacteria amount of Bacillus coagulans in the feed additive is not less than 10 8 CFU/g。
9. Use of the feed additive according to claim 7 or 8 in feed production.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211395783.9A CN116333912A (en) | 2022-11-09 | 2022-11-09 | Bacillus coagulans with broad-spectrum antibacterial effect and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211395783.9A CN116333912A (en) | 2022-11-09 | 2022-11-09 | Bacillus coagulans with broad-spectrum antibacterial effect and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116333912A true CN116333912A (en) | 2023-06-27 |
Family
ID=86893522
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211395783.9A Pending CN116333912A (en) | 2022-11-09 | 2022-11-09 | Bacillus coagulans with broad-spectrum antibacterial effect and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116333912A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117143770A (en) * | 2023-08-30 | 2023-12-01 | 广州格拉姆生物科技有限公司 | Wettman coagulans GLM336 and application thereof |
| CN117286079A (en) * | 2023-11-22 | 2023-12-26 | 广东容大生物股份有限公司 | Enteric-derived condensation Wittman's bacterium and application thereof, feeding microecological preparation comprising same and preparation method thereof |
| CN117305160A (en) * | 2023-09-19 | 2023-12-29 | 无锡爱科派生物科技有限公司 | Compound bacillus coagulans microbial inoculum and preparation method and application thereof |
-
2022
- 2022-11-09 CN CN202211395783.9A patent/CN116333912A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117143770A (en) * | 2023-08-30 | 2023-12-01 | 广州格拉姆生物科技有限公司 | Wettman coagulans GLM336 and application thereof |
| CN117143770B (en) * | 2023-08-30 | 2024-03-08 | 广州格拉姆生物科技有限公司 | Wettman coagulans GLM336 and application thereof |
| CN117305160A (en) * | 2023-09-19 | 2023-12-29 | 无锡爱科派生物科技有限公司 | Compound bacillus coagulans microbial inoculum and preparation method and application thereof |
| CN117286079A (en) * | 2023-11-22 | 2023-12-26 | 广东容大生物股份有限公司 | Enteric-derived condensation Wittman's bacterium and application thereof, feeding microecological preparation comprising same and preparation method thereof |
| CN117286079B (en) * | 2023-11-22 | 2024-03-15 | 广东容大生物股份有限公司 | Enteric-derived condensation Wittman's bacterium and application thereof, feeding microecological preparation comprising same and preparation method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106282072B (en) | Compound lactobacillus microecological preparation and preparation method and application thereof | |
| CN112011481B (en) | Lactobacillus reuteri for preventing and treating bacterial diarrhea of livestock and poultry and application thereof | |
| CN116333912A (en) | Bacillus coagulans with broad-spectrum antibacterial effect and application thereof | |
| CN105062933A (en) | Lactobacillus reuteri and application thereof | |
| CN113170842B (en) | Composite microecological preparation for preventing and treating necrotic enteritis of poultry and application thereof | |
| CN114085789B (en) | Pediococcus pentosaceus MA.WTPQJ01 and application thereof | |
| CN113549574B (en) | Bacillus coagulans and application thereof | |
| CN114134083B (en) | Bacillus bailii and application thereof | |
| CN115094012B (en) | Preparation method and application of bacillus coagulans BC-HYC strain microbial inoculum | |
| CN114874949B (en) | Clostridium butyricum, fermentation product thereof, microbial inoculum containing clostridium butyricum and animal feed additive | |
| CN114806944B (en) | Lactobacillus plantarum LP11, fermentation broth thereof, and preparation method and application thereof | |
| CN114806975A (en) | Microecological preparation containing enterogenous probiotics as well as preparation method and application of microecological preparation | |
| CN113088468B (en) | Lactobacillus casei Ma. GLRGJ1 and application thereof | |
| CN117210370A (en) | Bacillus subtilis for producing bacteriocin and application thereof | |
| CN117448213A (en) | Lactobacillus plantarum for inhibiting clostridium perfringens and its progeny and application | |
| CN112741210A (en) | Biological preparation for improving animal organism immunity function and preparation method thereof | |
| CN116064324B (en) | Lactobacillus rhamnosus, culture method thereof and application thereof in preventing and treating diarrhea and enteritis | |
| CN117801981A (en) | Screening and application of probiotics lactic acid bacteria for promoting growth of broiler chickens | |
| CN114891705B (en) | Acid-producing Min pig source lactobacillus reuteri and culture method and application thereof | |
| CN113980838A (en) | Bacillus subtilis capable of efficiently and directionally expressing bacteriocin M6 and application thereof | |
| CN109161501B (en) | Feeding bacillus licheniformis and application thereof | |
| CN111548965A (en) | Donkey-derived bacillus pumilus and application thereof in preparation of medicine for treating diarrhea of donkey colt | |
| CN115992066B (en) | Lactobacillus paracasei NHB-LpD2 for liquid fermented feed and application | |
| RU2203947C1 (en) | Strain of bacterium bacillus licheniformis used for preparing probiotic preparation designated for prophylaxis and treatment of gastroenteric diseases in animals, poultry and fishes | |
| RU2184774C1 (en) | Strain of bacterium bacillus subtilis used for preparing probiotic preparation designated for prophylaxis and treatment of gastroenteric diseases in animals, poultry and piscine |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |