CN116515679A - 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 - Google Patents
一株嗜热嗜气解硫胺素芽孢杆菌及其应用 Download PDFInfo
- Publication number
- CN116515679A CN116515679A CN202310246130.2A CN202310246130A CN116515679A CN 116515679 A CN116515679 A CN 116515679A CN 202310246130 A CN202310246130 A CN 202310246130A CN 116515679 A CN116515679 A CN 116515679A
- Authority
- CN
- China
- Prior art keywords
- thermophilic
- bacillus
- thiobacillus
- composting
- microbial inoculum
- 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
- 241000193830 Bacillus <bacterium> Species 0.000 title claims abstract description 23
- 241000605118 Thiobacillus Species 0.000 claims abstract description 22
- 230000001737 promoting effect Effects 0.000 claims abstract description 16
- 241001281729 Aneurinibacillus thermoaerophilus Species 0.000 claims abstract description 10
- 238000004321 preservation Methods 0.000 claims abstract description 4
- 239000002068 microbial inoculum Substances 0.000 claims description 20
- 239000002361 compost Substances 0.000 claims description 16
- 241000193385 Geobacillus stearothermophilus Species 0.000 claims description 14
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 12
- 230000001580 bacterial effect Effects 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 239000003795 chemical substances by application Substances 0.000 claims description 8
- 239000011780 sodium chloride Substances 0.000 claims description 6
- 238000012258 culturing Methods 0.000 claims description 4
- 239000008367 deionised water Substances 0.000 claims description 4
- 229910021641 deionized water Inorganic materials 0.000 claims description 4
- 239000001888 Peptone Substances 0.000 claims description 3
- 108010080698 Peptones Proteins 0.000 claims description 3
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 3
- 235000019319 peptone Nutrition 0.000 claims description 3
- 239000000843 powder Substances 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 238000011081 inoculation Methods 0.000 claims 1
- 238000009264 composting Methods 0.000 abstract description 25
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 abstract description 16
- 230000015556 catabolic process Effects 0.000 abstract description 10
- 238000006731 degradation reaction Methods 0.000 abstract description 10
- 229910052757 nitrogen Inorganic materials 0.000 abstract description 8
- 238000012216 screening Methods 0.000 abstract description 5
- 230000012010 growth Effects 0.000 abstract description 4
- JZRWCGZRTZMZEH-UHFFFAOYSA-N Thiamine Natural products CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 abstract description 2
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 abstract description 2
- 229960003495 thiamine Drugs 0.000 abstract description 2
- 235000019157 thiamine Nutrition 0.000 abstract description 2
- 239000011721 thiamine Substances 0.000 abstract description 2
- 239000003337 fertilizer Substances 0.000 description 21
- 230000000813 microbial effect Effects 0.000 description 16
- 239000000243 solution Substances 0.000 description 13
- 241000196324 Embryophyta Species 0.000 description 7
- 244000005700 microbiome Species 0.000 description 6
- 239000005416 organic matter Substances 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- 241000894006 Bacteria Species 0.000 description 5
- 238000000034 method Methods 0.000 description 5
- 235000015097 nutrients Nutrition 0.000 description 5
- 239000002689 soil Substances 0.000 description 5
- 230000000694 effects Effects 0.000 description 4
- 238000001962 electrophoresis Methods 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 230000008635 plant growth Effects 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 238000012271 agricultural production Methods 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 239000002054 inoculum Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 238000009629 microbiological culture Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 239000000575 pesticide Substances 0.000 description 2
- 235000010958 polyglycerol polyricinoleate Nutrition 0.000 description 2
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 238000005070 sampling Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 241000605272 Acidithiobacillus thiooxidans Species 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000609240 Ambelania acida Species 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 102000018997 Growth Hormone Human genes 0.000 description 1
- 108010051696 Growth Hormone Proteins 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 241000589157 Rhizobiales Species 0.000 description 1
- 241000589180 Rhizobium Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 241000607479 Yersinia pestis Species 0.000 description 1
- 241000589651 Zoogloea Species 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 244000000005 bacterial plant pathogen Species 0.000 description 1
- 239000010905 bagasse Substances 0.000 description 1
- 230000002457 bidirectional effect Effects 0.000 description 1
- 230000000853 biopesticidal effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 230000001351 cycling effect Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 210000003608 fece Anatomy 0.000 description 1
- 235000003891 ferrous sulphate Nutrition 0.000 description 1
- 239000011790 ferrous sulphate Substances 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 239000000122 growth hormone Substances 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000020169 heat generation Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 239000010871 livestock manure Substances 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 244000000010 microbial pathogen Species 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- 235000010333 potassium nitrate Nutrition 0.000 description 1
- 239000004323 potassium nitrate Substances 0.000 description 1
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 1
- -1 pplication Species 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F11/00—Other organic fertilisers
- C05F11/08—Organic fertilisers containing added bacterial cultures, mycelia or the like
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F17/00—Preparation of fertilisers characterised by biological or biochemical treatment steps, e.g. composting or fermentation
- C05F17/20—Preparation of fertilisers characterised by biological or biochemical treatment steps, e.g. composting or fermentation using specific microorganisms or substances, e.g. enzymes, for activating or stimulating the treatment
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F3/00—Fertilisers from human or animal excrements, e.g. manure
-
- 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
-
- 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
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W30/00—Technologies for solid waste management
- Y02W30/40—Bio-organic fraction processing; Production of fertilisers from the organic fraction of waste or refuse
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Wood Science & Technology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Microbiology (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Virology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明公开了一株嗜热嗜气解硫胺素芽孢杆菌，命名为嗜热嗜气解硫胺素芽孢杆菌F‑6(Aneurinibacillus thermoaerophilus F‑6)，保藏号为CGMCCNO.26032。本发明嗜热嗜气解硫胺素芽孢杆菌为首次从堆肥高温期样品中筛选分离得到的，该菌株耐高温，生长繁殖快，能促进堆肥腐熟；进一步的，本发明提供的含嗜热嗜气解硫胺素芽孢杆菌能够提高堆肥温度，加速有机质降解，提高凯氏氮含量，促进堆肥腐熟。
Description
技术领域
本发明涉及植物促生菌技术领域，具体涉及一株嗜热嗜气解硫胺素芽孢杆菌及其应用。
背景技术
化肥在现代农业生产中发挥着巨大作用。但是随着化肥使用量的增加，其利用率逐年降低。农药和化肥的残留，向环境释放了大量的有害物质，污染了土壤、水源和食品，对人类健康及生存环境构成了极大威胁，要求保护生态环境和生产安全食品的呼声日益强烈。因此，开发利用替代化学肥料的新肥源，以适应发展绿色农业和绿色食品的需要已是当务之急。而微生物肥料可使土壤中无效营养有效化，预防和控制农作物病害，减少农药和化肥的使用，是解决土壤、水源和食品污染的根本途经，被普遍认为是一种环境友好、经济有效的提高作物产量的方法。
微生物肥料是一类含有微生物活体的特定制品，应用于农业生产中，能够获得特定的肥料效应，其中制品中活微生物起关键作用。目前，一般将微生物肥料制品分为两大类：一类是狭义的微生物肥料，指通过微生物的生命活动，增加了植物营养元素的供应量，包括土壤和生产环境中植物营养元素的供应总量，导致植物营养状况的改善，进而增加产量。这一类微生物肥料的代表是根瘤菌肥；另一类是广义的微生物肥料，指通过其中微生物的生命活动，不但能提高植物营养元素的供应量，还能产生植物生长激素，促进植物对营养元素的吸收利用或有拮抗某些病原微生物的致病作用，减轻农作物病虫害间接提高作物产量。与化学肥料相比，微生物肥料具有以下优点：不破坏土壤结构；保护生态、不污染环境，对人畜无毒无害；肥效持久；提高作物产量，改善作物品质；成本低廉，经济有效。
植物根际促生菌(Plant growth promoting rhizobacteria，PGPR)是一类能高密度定殖在植物根际的微生物，兼有抑制植物病原菌、根际有害微生物，以及促进植物生长并增加作物产量的作用。作为生物肥料和生物农药的重要资源库，PGPR的研究和应用已经起到了举足轻重的作用。从资源再生利用的角度来研究和开发微生物肥料则对资源综合利用和环境保护更具有现实意义。
发明内容
为解决上述技术问题，本发明提供一株嗜热嗜气解硫胺素芽孢杆菌及其应用，旨在得到一种加速有机质降解，提高凯氏氮含量，促进堆肥腐熟，快捷方便的腐熟、堆肥方法。
一株嗜热嗜气解硫胺素芽孢杆菌，命名为嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)，于2022年11月02日保藏于中国微生物菌种保藏管理委员会普通微生物中心，保藏号为CGMCC NO.26032。
如上所述嗜热嗜气解硫胺素芽孢杆菌在促进堆肥腐熟中的应用。
如上所述嗜热嗜气解硫胺素芽孢杆菌的菌剂在促进堆肥腐熟中的应用。
其中，所述嗜热嗜气解硫胺素芽孢杆菌的菌剂的制备方法为将所述的嗜热嗜气解硫胺素芽孢杆菌F-6按1％(v/v)的接种量接种到500mL灭菌后的LB培养液中，在50℃，180rpm摇床中培养24h，收集培养液，即得菌剂。
其中，所述的LB培养液组成为蛋白胨10g，酵母粉5g，氯化钠10g，去离子水1L。
与现有技术相比，本发明的有益效果：
本发明嗜热嗜气解硫胺素芽孢杆菌为首次从堆肥高温期样品中筛选分离得到的，该菌株耐高温，生长繁殖快，能促进堆肥腐熟；进一步的，本发明提供的含嗜热嗜气解硫胺素芽孢杆菌能够提高堆肥温度，加速有机质降解，提高凯氏氮含量，促进堆肥腐熟。
保藏信息说明
嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)于2022年11月02日保藏于中国微生物菌种保藏管理委员会普通微生物中心(简称CGMCC)，保藏号为CGMCC NO：26032。
附图说明
图1为温度随堆肥时间变化曲线；室温(Room)，空白组(CK)，实验组(F-6)。
图2为pH随堆肥时间变化曲线；空白组(CK)，实验组(F-6)。
图3为含水率随堆肥时间变化曲线；空白组(CK)，实验组(F-6)。
图4为有机质含量随堆肥时间变化曲线；空白组(CK)，实验组(F-6)。
图5为凯氏氮含量随堆肥时间变化曲线；空白组(CK)，实验组(F-6)。
图6为嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)菌落图。
图7为筛选出菌株的凝胶电泳图谱。
具体实施方式
下面结合附图具体实施方式进行详细描述，但应当理解本发明的保护范围并不受具体实施方式的限制。实施例中采用的原料、试剂若无特殊说明，皆为市售所得。
实施例1
嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)的筛选
1.实验材料与设备
1.1主要仪器与设备
表1主要仪器与设备
1.2培养基
改良高氏一号培养基：硝酸钾1.0g，磷酸二氢钾0.5g，硫酸镁0.5g硫酸亚铁0.01g，氯化钠0.5g，可溶性淀粉20.0g，琼脂15.0g，去离子水1.0L；
LB培养液：蛋白胨10g，酵母粉5g，氯化钠10g，去离子水1L。
实施例2
嗜热嗜气解硫胺素芽孢杆菌F-6的分离筛选
2.1取样
筛菌的样品为4天的堆肥样品，在堆体温度较高处多点取样(在堆体表面下方10-20cm)，样品混匀后装入样品袋中于-4℃的冰箱里保存。
2.2耐高温菌的筛选及纯化
称取在冰箱保存的样品10g，加入到装有90mL灭菌处理后的NaCl溶液(0.9％w/v)的三角瓶中，置于摇床上振荡30min，打散菌胶团，使细菌呈单细胞状态分散于溶液中；将灭菌处理后的NaCl溶液分别加到7支试管中(同样进行灭菌处理)，每支加入9mL，取震荡后的菌液1mL分别加入上述试管，即分别稀释到10-1、10-2、10-3、10-4、10-5、10-6、10-7，再分别将各稀释度下的菌液取0.1mL涂布在改良高氏一号培养基平板上，50℃恒温培养48h；每种稀释梯度的平板均作5个平行；可以观察到细菌群落在10-5稀释度下长势较好，在5个平行平板中选取菌落较大的进行分离培养并多次划线纯化。如图6所示，多次划线纯化后所得编号为F-6的菌株菌落形状不规则，菌落较小，乳白色，表面无光泽，半透明，有粘稠感，菌落扁平，边缘为霜花放射状。
实施例3
菌株的鉴定
鉴定对象：实施例2中多次划线纯化后所得编号为F-6的菌株；
3.1提取DNA
DNA使用杭博《Biospin细菌基因组DNA提取试剂盒》提取，具体操作步骤详见说明书；
3.2PCR扩增
使用诺唯赞2×Max Master Mix(Dye Plus)进行PCR扩增，引物、反应体系、PCR循环条件如表2、表3和表4所示：
表2 PCR扩增引物
表3反应体系
表4 PCR循环条件
PCR产物电泳条件
1.0％琼脂糖凝胶，电压120V，电泳30min，每孔上样1u L，电泳图谱见图7。
表5电泳顺序
| 泳道 | 1 | 2 | 3 |
| 样品编号 | F-6(1) | F-6(2) | F-6(3) |
3.3.测序及比对结果
PCR扩增产物送至广州擎科生物技术有限公司进行双向测序，实施例2多次划线纯化后所得编号为F-6的菌株的序列如SEQ ID NO.1所示；
在NCBI进行BLASTN比对，得菌种鉴定结果，结果见表6；
表6比对结果
| 菌株编号 | 相似性最高的菌株 | 一致性 |
| F-6 | Aneurinibacillus thermoaerophilus | 100％ |
由上表所示，菌株F-6与Aneurinibacillus thermoaerophilus相似度达到100％，因此，将菌株F-6命名为嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillusthermoaerophilus F-6)。
实施例4
含嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)菌株的菌剂在促进堆肥发酵中的应用
4.1菌剂制备
把嗜热嗜气解硫胺素芽孢杆菌F-6种子液按1％(v/v)的接种量，接种到500mL灭菌后的LB培养液中，在摇床(50℃，180rmp)中培养24h，收集培养液即为菌剂；添加菌剂组为实验组(F-6)，空白组(CK)添加500mL灭菌后的LB培养液；
4.2堆肥实验
堆肥原料选用猪粪和蔗髓、干草、木糠，质量比12.3:1.7:1.2:1，共20kg；堆肥进行39d，分别在第0，4，10，17，26，36d取样，测定了温度，pH，含水率，有机质，凯氏氮；整个堆肥过程在0d实验组接种菌剂一次(空白组在0d接种等体积的LB培养液)，堆体翻堆4次，分别是4d、10d、17d、26d；
4.3实验结果
嗜热嗜气解硫胺素芽孢杆菌F-6耐高温，增长繁殖快；与不添加菌剂的堆肥相比(CK)，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的堆肥具有以下优势：
(1)从堆肥过程的温度看，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的高温期堆体温度比不添加菌剂的堆体温度高，有利于灭杀堆体中病原体及加速堆肥物料的高温降解，促进堆体发酵(图1)；
(2)由图2可知，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的堆肥的pH值在嗜热期下降明显，说明菌剂的添加促进了堆肥过程中大量有机物降解增加了有机酸的产生；
(3)由图3可知，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的堆肥水分降得更快，即添加菌剂的堆体微生物反应更为剧烈，产热更多带走大量的水分；
(4)从图4有机质(OM)的降解率看，不添加菌剂的OM从82.42％下降到69.73％，降解率为15.40％，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的OM从81.74％下降到65.92％，降解率为19.35％；表明嗜热嗜气解硫胺素芽孢杆菌F-6菌剂显著促进有机质的降解，并且在嗜热期下降速率更快，与温度在嗜热期更高一致，间接表明嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的添加使得微生物活动更为剧烈；
(5)从图5凯氏氮可以看出，嗜热嗜气解硫胺素芽孢杆菌F-6菌剂的添加显著的增加了凯氏氮的含量。
综上所述，添加嗜热嗜气解硫胺素芽孢杆菌F-6菌剂能够提高堆肥嗜热期温度，加速有机质降解和水分蒸发，提高凯氏氮的含量，促进堆肥发酵。
前述对本发明的具体示例性实施方案的描述是为了说明和例证的目的。这些描述并非想将本发明限定为所公开的精确形式，并且很显然，根据上述教导，可以进行很多改变和变化。对示例性实施例进行选择和描述的目的在于解释本发明的特定原理及其实际应用，从而使得本领域的技术人员能够实现并利用本发明的各种不同的示例性实施方案以及各种不同的选择和改变。本发明的范围意在由权利要求书及其等同形式所限定。
Claims (5)
1.一株嗜热嗜气解硫胺素芽孢杆菌，其特征在于：所述的嗜热嗜气解硫胺素芽孢杆菌命名为嗜热嗜气解硫胺素芽孢杆菌F-6(Aneurinibacillus thermoaerophilus F-6)，保藏号为CGMCC NO.26032。
2.如权利要求1所述的嗜热嗜气解硫胺素芽孢杆菌在促进堆肥腐熟中的应用。
3.如权利要求1所述的嗜热嗜气解硫胺素芽孢杆菌的菌剂在促进堆肥腐熟中的应用。
4.根据权利要求3所述嗜热嗜气解硫胺素芽孢杆菌的菌剂在促进堆肥腐熟中的应用，其特征在于：所述嗜热嗜气解硫胺素芽孢杆菌的菌剂的制备方法为将所述的嗜热嗜气解硫胺素芽孢杆菌F-6按1％的接种量接种到灭菌后的LB培养液中，摇床中培养，收集培养液，即得菌剂。
5.根据权利要求4所述嗜热嗜气解硫胺素芽孢杆菌的菌剂在促进堆肥腐熟中的应用，其特征在于：所述的LB培养液组成为蛋白胨10g，酵母粉5g，氯化钠10g，去离子水1L。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310246130.2A CN116515679A (zh) | 2023-03-15 | 2023-03-15 | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310246130.2A CN116515679A (zh) | 2023-03-15 | 2023-03-15 | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116515679A true CN116515679A (zh) | 2023-08-01 |
Family
ID=87389337
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310246130.2A Pending CN116515679A (zh) | 2023-03-15 | 2023-03-15 | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116515679A (zh) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101098520B1 (ko) * | 2011-06-21 | 2011-12-26 | 문홍순 | 아네우리니바실러스 서모아에로필러스 균주를 이용한 유기성 폐기물의 퇴비화를 위한 미생물 제제 및 이의 제조방법 그리고 이를 이용한 유기성 폐기물의 퇴비화 방법 |
| CN111117939A (zh) * | 2020-02-24 | 2020-05-08 | 中微泽美环保科技(杭州)有限公司 | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 |
| CN113969246A (zh) * | 2021-10-27 | 2022-01-25 | 江西省科学院微生物研究所 | 一种保氮菌株、复合菌剂及复合菌剂制备方法和应用 |
-
2023
- 2023-03-15 CN CN202310246130.2A patent/CN116515679A/zh active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101098520B1 (ko) * | 2011-06-21 | 2011-12-26 | 문홍순 | 아네우리니바실러스 서모아에로필러스 균주를 이용한 유기성 폐기물의 퇴비화를 위한 미생물 제제 및 이의 제조방법 그리고 이를 이용한 유기성 폐기물의 퇴비화 방법 |
| CN111117939A (zh) * | 2020-02-24 | 2020-05-08 | 中微泽美环保科技(杭州)有限公司 | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 |
| CN113969246A (zh) * | 2021-10-27 | 2022-01-25 | 江西省科学院微生物研究所 | 一种保氮菌株、复合菌剂及复合菌剂制备方法和应用 |
Non-Patent Citations (1)
| Title |
|---|
| 石娟;周攀;陈意超;刘晓风;李东: "一株耐盐嗜热菌Aneurinibacillus thermoaerophilus H7的分离及其油脂降解特性", 应用与环境生物学报, no. 001, 31 December 2021 (2021-12-31), pages 214 - 219 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Maji et al. | Humic acid rich vermicompost promotes plant growth by improving microbial community structure of soil as well as root nodulation and mycorrhizal colonization in the roots of Pisum sativum | |
| CN106978366B (zh) | 一种混合菌剂及其在促进堆肥腐熟中的应用 | |
| CN107586745A (zh) | 一种畜禽粪便堆肥除臭保氮菌株、菌剂及其制备方法和应用 | |
| CN101024591A (zh) | 利用微生物菌剂快速发酵猪粪生产商品有机肥的方法 | |
| CN112375720B (zh) | 一种枯草芽孢杆菌及其应用 | |
| CN108117453A (zh) | 利用猪养殖废弃物制作生物有机肥的方法 | |
| CN102703363A (zh) | 一株甲基营养性芽孢杆菌utm401及其应用 | |
| CN110437000A (zh) | 一种微生物肥料及其制备方法和应用 | |
| CN106699474A (zh) | 一种新型生物有机肥及其制备方法 | |
| Agarwal et al. | Importance of biofertilizers in agriculture biotechnology | |
| CN106883067A (zh) | 草莓专用抗枯萎病生物有机肥的制备方法 | |
| CN104789494B (zh) | 采用强化垃圾堆肥微生物菌剂提高草皮抗盐性的方法 | |
| CN104909846B (zh) | 一种含有解淀粉芽孢杆菌hfj‑7的复合鸡粪腐熟发酵剂及其应用 | |
| CN104560817B (zh) | 一株产植酸酶的嗜热地衣芽孢杆菌utm102及其应用 | |
| CN116286523A (zh) | 一株枯草芽孢杆菌及其应用 | |
| CN104774788B (zh) | 垃圾堆肥中草坪耐盐强化复合微生物菌群的制备方法及应用 | |
| CN106938949A (zh) | 一种负载高浓度枯草芽孢杆菌餐厨垃圾有机肥的制作方法 | |
| CN110257486B (zh) | 一种基于纤维素酶基因表征堆肥腐熟度的方法 | |
| CN116515679A (zh) | 一株嗜热嗜气解硫胺素芽孢杆菌及其应用 | |
| CN107500884A (zh) | 用于发酵畜禽粪便生产生物有机肥的复合制剂 | |
| CN108373982B (zh) | 一株indicus不动杆菌及其解磷方法 | |
| CN116240141A (zh) | 一株热红短芽孢杆菌及其应用 | |
| CN106905063A (zh) | 一种负载高浓度多粘类芽孢杆菌餐厨垃圾有机肥的制作方法 | |
| CN110184332B (zh) | 一种处理高纤维物料的堆肥菌剂复配方法及其应用 | |
| Sebayang et al. | Population dynamics of Azotobacter sp., Azospirillum sp., Bacillus subtilis, Talaromyces pinophilus, Trichoderma asperellum, and Syncephalastrum racemosum in the medium with various earthworm applications |
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 |