CN116925982B - Rhizobium JY1-1 capable of producing acid and application thereof - Google Patents
Rhizobium JY1-1 capable of producing acid and application thereof Download PDFInfo
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- CN116925982B CN116925982B CN202311180287.6A CN202311180287A CN116925982B CN 116925982 B CN116925982 B CN 116925982B CN 202311180287 A CN202311180287 A CN 202311180287A CN 116925982 B CN116925982 B CN 116925982B
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- 241000589180 Rhizobium Species 0.000 title claims abstract description 30
- 239000002253 acid Substances 0.000 title claims description 17
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 claims abstract description 46
- 238000000034 method Methods 0.000 claims abstract description 30
- 241000237502 Ostreidae Species 0.000 claims abstract description 23
- 235000020636 oyster Nutrition 0.000 claims abstract description 23
- 239000010815 organic waste Substances 0.000 claims abstract description 17
- 244000005700 microbiome Species 0.000 claims abstract description 6
- 239000002361 compost Substances 0.000 claims description 59
- 239000000843 powder Substances 0.000 claims description 53
- 238000000855 fermentation Methods 0.000 claims description 36
- 230000004151 fermentation Effects 0.000 claims description 36
- 230000001580 bacterial effect Effects 0.000 claims description 17
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 17
- 210000003608 fece Anatomy 0.000 claims description 12
- 239000010871 livestock manure Substances 0.000 claims description 12
- 239000010902 straw Substances 0.000 claims description 12
- 241000287828 Gallus gallus Species 0.000 claims description 11
- 235000019764 Soybean Meal Nutrition 0.000 claims description 10
- 239000004455 soybean meal Substances 0.000 claims description 10
- 239000000463 material Substances 0.000 claims description 9
- 238000004321 preservation Methods 0.000 claims description 8
- 239000008223 sterile water Substances 0.000 claims description 7
- 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 claims description 4
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 239000007788 liquid Substances 0.000 claims description 4
- 239000002893 slag Substances 0.000 claims description 4
- 238000009629 microbiological culture Methods 0.000 claims description 3
- 238000009264 composting Methods 0.000 abstract description 44
- 239000002689 soil Substances 0.000 abstract description 29
- 230000008569 process Effects 0.000 abstract description 15
- 238000012216 screening Methods 0.000 abstract description 13
- 239000005416 organic matter Substances 0.000 abstract description 9
- 230000015556 catabolic process Effects 0.000 abstract description 7
- 238000006731 degradation reaction Methods 0.000 abstract description 7
- 239000011575 calcium Substances 0.000 abstract description 6
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 abstract description 5
- 229910052791 calcium Inorganic materials 0.000 abstract description 5
- 238000011161 development Methods 0.000 abstract description 5
- 230000001737 promoting effect Effects 0.000 abstract description 5
- 230000000813 microbial effect Effects 0.000 abstract description 4
- 230000003213 activating effect Effects 0.000 abstract description 2
- 230000035558 fertility Effects 0.000 abstract description 2
- 239000003337 fertilizer Substances 0.000 abstract description 2
- 230000002053 acidogenic effect Effects 0.000 abstract 1
- QJZYHAIUNVAGQP-UHFFFAOYSA-N 3-nitrobicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid Chemical compound C1C2C=CC1C(C(=O)O)C2(C(O)=O)[N+]([O-])=O QJZYHAIUNVAGQP-UHFFFAOYSA-N 0.000 description 38
- 239000004021 humic acid Substances 0.000 description 36
- PUKLDDOGISCFCP-JSQCKWNTSA-N 21-Deoxycortisone Chemical compound C1CC2=CC(=O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@@](C(=O)C)(O)[C@@]1(C)CC2=O PUKLDDOGISCFCP-JSQCKWNTSA-N 0.000 description 25
- FCYKAQOGGFGCMD-UHFFFAOYSA-N Fulvic acid Natural products O1C2=CC(O)=C(O)C(C(O)=O)=C2C(=O)C2=C1CC(C)(O)OC2 FCYKAQOGGFGCMD-UHFFFAOYSA-N 0.000 description 25
- 239000002509 fulvic acid Substances 0.000 description 25
- 229940095100 fulvic acid Drugs 0.000 description 25
- 239000007787 solid Substances 0.000 description 18
- 229910052500 inorganic mineral Inorganic materials 0.000 description 15
- 239000011707 mineral Substances 0.000 description 15
- 238000011282 treatment Methods 0.000 description 14
- 238000005259 measurement Methods 0.000 description 13
- 239000000047 product Substances 0.000 description 9
- 230000008859 change Effects 0.000 description 8
- 239000003895 organic fertilizer Substances 0.000 description 8
- 241000196324 Embryophyta Species 0.000 description 7
- 239000000706 filtrate Substances 0.000 description 7
- 239000002994 raw material Substances 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 239000000203 mixture Substances 0.000 description 6
- 239000006228 supernatant Substances 0.000 description 6
- 108020004465 16S ribosomal RNA Proteins 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 238000002156 mixing Methods 0.000 description 5
- 239000002699 waste material Substances 0.000 description 5
- 241000589174 Bradyrhizobium japonicum Species 0.000 description 4
- 230000009286 beneficial effect Effects 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 239000003607 modifier Substances 0.000 description 4
- 238000001139 pH measurement Methods 0.000 description 4
- 238000005070 sampling Methods 0.000 description 4
- 238000012360 testing method Methods 0.000 description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 3
- 229910021529 ammonia Inorganic materials 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 239000008367 deionised water Substances 0.000 description 3
- 229910021641 deionized water Inorganic materials 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 238000003837 high-temperature calcination Methods 0.000 description 3
- 230000007774 longterm Effects 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 230000000877 morphologic effect Effects 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 238000011084 recovery Methods 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 239000003516 soil conditioner Substances 0.000 description 3
- 238000005303 weighing Methods 0.000 description 3
- 235000008733 Citrus aurantifolia Nutrition 0.000 description 2
- 238000003794 Gram staining Methods 0.000 description 2
- 241001522196 Ostrea edulis Species 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 235000011941 Tilia x europaea Nutrition 0.000 description 2
- 238000009825 accumulation Methods 0.000 description 2
- 230000001174 ascending effect Effects 0.000 description 2
- 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 2
- 238000001816 cooling Methods 0.000 description 2
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- 238000001514 detection method Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000004090 dissolution Methods 0.000 description 2
- 238000000227 grinding Methods 0.000 description 2
- 239000003864 humus Substances 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 239000004571 lime Substances 0.000 description 2
- 230000035800 maturation Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 150000007524 organic acids Chemical class 0.000 description 2
- 238000004064 recycling Methods 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 238000009423 ventilation Methods 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- RSPISYXLHRIGJD-UHFFFAOYSA-N OOOO Chemical compound OOOO RSPISYXLHRIGJD-UHFFFAOYSA-N 0.000 description 1
- 238000013494 PH determination Methods 0.000 description 1
- 244000046052 Phaseolus vulgaris Species 0.000 description 1
- 235000010627 Phaseolus vulgaris Nutrition 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 239000002154 agricultural waste Substances 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
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- 230000002708 enhancing effect Effects 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000002386 leaching Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000010298 pulverizing process Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 239000013535 sea water Substances 0.000 description 1
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- 239000011780 sodium chloride Substances 0.000 description 1
- 239000002910 solid waste Substances 0.000 description 1
- 238000010563 solid-state fermentation Methods 0.000 description 1
- 238000001694 spray drying Methods 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 239000003643 water by type Substances 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
- C05D—INORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C; FERTILISERS PRODUCING CARBON DIOXIDE
- C05D3/00—Calcareous fertilisers
- C05D3/02—Calcareous fertilisers from limestone, calcium carbonate, calcium hydrate, slaked lime, calcium oxide, waste calcium products
-
- 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
- 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
- C12R2001/41—Rhizobium
-
- 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
Abstract
The invention discloses an acidogenic rhizobium JY1-1 and application thereof, belonging to the technical field of agricultural microorganisms. The rhizobium JY1-1 obtained by screening is separated from plant rhizosphere soil in oyster shell deposit, can be well adapted to the environment of oyster shell deposit, can efficiently play the roles of activating calcium and promoting composting process, and does not pollute the environment; in addition, rhizobia JY1-1 obtained by screening can be added into oyster shell-organic waste co-composting, so that the composting process is quickened, the organic matter degradation is promoted, the soil fertility is increased, the rhizobia JY1-1 has the potential of being used as a functional microbial fertilizer, and has important significance for promoting green agricultural development.
Description
Technical Field
The invention relates to rhizobia and application thereof, in particular to rhizobia JY1-1 with higher acid production capacity and application of the rhizobia as a beneficial bacterial strain for co-composting and synergism of oyster shell powder, belonging to the technical field of agricultural microorganisms.
Background
In recent years, with the increase of market demands and the improvement of aquaculture technologies, oyster yields have increased dramatically, and the number of waste oyster shells generated therewith has also increased dramatically. At present, the use of oysters is limited to edible parts, only a small amount of non-edible oyster shells are used as feeds, ornaments, precious medicinal materials or soil improvers, and most of the non-edible oyster shells are piled up as solid wastes in landfill sites or sent back to the sea. Because oyster shells are difficult to biodegrade, they cause serious pollution to soil, natural waters and marine ecosystems. The stacked oyster shells occupy a large amount of land resources, and residues on the discarded oyster shells are easy to be degraded and generate malodor, so that the surrounding environment is greatly polluted, and the health of residents is threatened. The pollution range is also increased if the waste oyster shells are caught in the seawater. Therefore, how to treat the waste oyster shells is a problem which is urgently needed to be solved at present.
The oyster shell contains more than 90% of CaCO 3 ，CaCO 3 Is a good natural calcium source, has lower leaching loss rate after rain, smaller migration depth and better calcium effectiveness in the soil of the plough layer. At present, oyster shells are mainly treated by crushing and high-temperature calcination.
The most convenient treatment process is crushing, the crushed oyster shell powder has only different structure from oyster shell, and the main component is CaCO 3 . After oyster shell powder is applied to soil, the oyster shell powder is stable in structure and is insoluble in water, so that the oyster shell powder can be absorbed and utilized by plants after long-term transformation. In addition, long-term application of oyster shell powder can also cause soil hardening and even risk soil acid recovery.
After high temperature calcination, caCO in oyster shell 3 Is decomposed into CaO, since CaO is liable to react with H 2 O reacts to produce Ca (OH) 2 The alkalinity is enhanced, so that the oyster shell powder calcined at high temperature can only improve the acidity of soil for a short period of time. In addition, the high-temperature calcination can inactivate microorganisms on oyster shells, and the obtained product is similar to lime, and can consume soil organic matters when directly applied to soil.
The agricultural waste is also produced in the development process of agriculture, and the environment is polluted. The most common method for recycling agricultural organic waste is composting. The cost of composting is low, the operation is simple, and the waste can be recycled. Research shows that the oyster shell powder with small proportion is addedThe organic acid can produce CaCO in oyster shell 3 And (5) corroding to activate the calcium. The functions of the soil organic acid are mainly to improve the properties of plant rhizosphere soil, such as: (1) Regulating the pH value of soil, influencing the conversion of soil phosphorus, enhancing the microbial activity of the soil and the like; (2) Promoting mineral dissolution, and having stronger weathering effect on partial primary minerals and secondary minerals; (3) Takes part in the soil forming process, influences the mineral dissolution speed and the microbial activity and further acts on the soil forming process; (4) Promote the absorption of plant nutrients, increase crop yield, etc.
The soil conditioner mainly comprises a mineral conditioner, a single-application organic fertilizer, a mineral and organic fertilizer combined application and organic-inorganic co-composting. Wherein:
(1) Mineral improver: mixing a plurality of minerals in proportion, and then roasting at high temperature to prepare the soil conditioner which is a weak alkaline conditioner. The mineral modifier is used independently, wherein minerals only act on surface soil, and soil hardening and acid recovery can be caused after long-term use;
(2) Single-application organic fertilizer: common organic fertilizer soil modifiers include straw, plant ash, manure and the like, and the organic fertilizer must be thoroughly decomposed before being applied to the soil;
(3) Mineral and organic fertilizer are applied: because the mineral modifier is singly used for a long time to lead to soil hardening and acid recovery, researches show that the effect of lime and the organic fertilizer are better than that of singly applying the organic fertilizer and the mineral modifier;
(4) Organic-inorganic co-composting: if oyster shell powder (inorganic) is added into organic waste for co-composting, research shows that the co-composted oyster shell powder is applied into soil, and the improvement effect is better than that of the former soil conditioner. However, when co-composting oyster shell powder with organic waste, the co-composting process is slow and the fermentation process is long, usually taking 1-2 years to be effective, because no microorganism coordinates with each other.
Disclosure of Invention
The first object of the present invention is to: provides a rhizobium strain with higher acid-producing capability, and is a beneficial strain for co-composting and synergism of oyster shell powder because a large amount of acid is produced in the fermentation process of the rhizobium strain.
A second object of the present invention is to: by utilizing the beneficial strain, two oyster shell-organic wastes which can accelerate the composting process and shorten the fermentation process (which is effective usually about 1 month) are prepared for co-composting.
In order to achieve the first object, the present invention adopts the following technical scheme:
acid-producing rhizobium JY1-1, wherein the rhizobium JY1-1 is preserved in the China general microbiological culture Collection center of the China Committee for culture Collection of microorganisms, and is classified and named as rhizobiumRhizobium sp.The preservation number is CGMCC NO.27052, the preservation date is 2023, 4 and 10 days, and the preservation address is Beijing in China.
In order to achieve the second object, the present invention adopts the following technical scheme:
the first oyster shell-organic waste co-compost consists of oyster shell powder, sugar slag, soybean meal and rhizobia JY1-1, wherein the mass percentages of the oyster shell powder, the sugar slag and the soybean meal are respectively 40%, 56% and 4%, the rhizobia JY1-1 accounts for 0.5-1.0% of the dry weight of the whole system, and the initial water content of the whole pile material is 60%.
The second oyster shell-organic waste co-compost consists of oyster shell powder, chicken manure, straw and the rhizobia JY1-1, wherein the mass percentages of the oyster shell powder, the chicken manure and the straw are respectively 40%, 16.28% and 43.72%, the rhizobia JY1-1 accounts for 0.5-1.0% of the dry weight of the whole system, and the initial water content of the whole pile material is 60%.
Preferably, rhizobia JY1-1 exists in the form of bacterial powder, and is specifically prepared by the following method: rhizobium JY1-1 is selected, dissolved in sterile water, inoculated in a fermentation medium, cultured for 48 hours at 30 ℃ and 200rpm, and the fermentation liquid is spray-dried to prepare rhizobium JY1-1 bacterial powder.
Preferably, the fermentation medium is formulated as follows: glucose 50g/L, yeast powder 5g/L, KH 2 PO 4 ·H 2 O 1.4g/L，MgSO 4 ·7H 2 O 0.5g/L，（NH 4 ） 2 SO 4 10g/L，FeSO 4 ·7H 2 O 0.03g/L，ZnSO 4 0.03g/L。
The invention has the advantages that:
(1) The rhizobium JY1-1 obtained by screening is separated from plant rhizosphere soil in oyster shell deposit, can be well adapted to the environment of oyster shell deposit, can efficiently play the roles of activating calcium and promoting composting process, and does not pollute the environment;
(2) The rhizobium JY1-1 obtained by screening can be added into oyster shell-organic waste co-composting, so that the composting process is quickened, the organic matter degradation is promoted, the soil fertility is increased, the rhizobium JY1-1 has the potential of being used as a functional microbial fertilizer, and has important significance for promoting green agricultural development;
(3) The oyster shell-organic waste co-compost provided by the invention is safe and environment-friendly in raw materials, realizes recycling of waste resources while improving agricultural environment, and is simple in preparation method operation and easy to popularize and apply.
Drawings
FIG. 1 is a graph showing the result of gram staining of strain JY1-1 obtained by screening in accordance with the present invention;
FIG. 2 is a diagram showing the analysis of the strain development tree of strain JY1-1 obtained by screening in the present invention;
FIG. 3 is a graph showing the pH change of fermentation broth of strain JY1-1 obtained by screening in the present invention;
FIG. 4 is a graph showing the pH value of the co-compost of rhizobia JY1-1, oyster shell powder, sugar residue and soybean meal;
FIG. 5 is a graph showing the variation of the loss rate of volatile solids in the co-compost of rhizobia JY1-1, oyster shell powder, sugar dreg and soybean meal;
FIG. 6 is a graph showing the variation of total humic acid content in rhizobia JY1-1, oyster shell powder, sugar residue and soybean meal co-compost;
FIG. 7 is a graph showing the change of the content of humic acid in the co-compost of rhizobia JY1-1, oyster shell powder, sugar residue and soybean meal;
FIG. 8 is a graph showing the variation of the fulvic acid content of rhizobia JY1-1, oyster shell powder, sugar dreg and soybean meal co-compost;
FIG. 9 is a graph showing the pH value of the co-compost of rhizobia JY1-1, oyster shell powder, chicken manure and straw;
FIG. 10 is a graph showing the variation of the loss rate of volatile solids in rhizobia JY1-1, oyster shell powder, chicken manure and straw co-compost;
FIG. 11 is a graph showing the variation of total humic acid content in rhizobia JY1-1, oyster shell powder, chicken manure and straw co-compost;
FIG. 12 is a graph showing the variation of the content of humic acid in rhizobia JY1-1, oyster shell powder, chicken manure and straw co-compost;
FIG. 13 is a graph showing the variation of the fulvic acid content of rhizobia JY1-1, oyster shell powder, chicken manure and straw co-compost.
Detailed Description
The invention is described in detail below with reference to the drawings and the specific embodiments.
1. Isolation and screening of strains
1. Isolation of strains
Air drying and grinding plant rhizosphere soil in oyster shell deposit, which is a soil sample collected from Bohai sea, 1g of the plant rhizosphere soil is dissolved in 9mL of sterile water, and gradient diluting to 10 -6 Each of the suction concentrations was 10 -3 、10 -4 、10 -5 、10 -6 Respectively coating 0.2mL of diluted bacterial liquid on a screening culture medium, inversely culturing for 48h at 30 ℃, picking out bacterial strains which generate larger blue or purple transparent rings, repeatedly streaking and separating to obtain pure bacterial strains, and preserving the pure bacterial strains in a test tube for standby.
The formula of the screening culture medium is as follows: KNO (KNO) 3 0.1g/L，NaCl 0.5g/L，K 2 HPO 4 ·3H 2 O 0.5g/L，MgSO 4 ·7H 2 O 0.5g/L，FeSO 4 ·7H 2 O0.01 g/L, soluble starch 20g/L, agar 15g/L, congo red 0.2g/L. The deionized water is used for fixing the volume to 1000mL, the pH is adjusted to 7.0-7.4, and the sterilization is carried out for 15min at the temperature of 121 ℃.
2. Screening of strains
Picking a ring of the pure strain obtained in the previous step, dissolving in 10mL of sterile water, respectively taking 0.2mL of the pure strain to inoculate in 18 fermentation media with the volume of 50mL, culturing at 30 ℃ and 200rpm, taking out a bottle every 8h, centrifuging at 12000rpm for 10min, and measuring the pH value of the filtrate. The strain with the lowest pH value (most acid production) of the final filtrate is the beneficial strain of oyster shell co-composting synergy, and is marked as JY1-1.
The formula of the fermentation medium is as follows: glucose 50g/L, yeast powder 5g/L, KH 2 PO 4 ·H 2 O 1.4g/L，MgSO 4 ·7H 2 O 0.5g/L，（NH 4 ） 2 SO 4 10g/L，FeSO 4 ·7H 2 O 0.03g/L，ZnSO 4 0.03g/L. The deionized water is used for fixing the volume to 1000mL, the pH is adjusted to 7.0-7.4, and the sterilization is carried out for 15min at the temperature of 121 ℃.
2. Identification of strains
1. Morphological identification
Morphological identification was performed on strain JY1-1. The identification results are as follows: the colony forms are convex, sticky, milky white and semitransparent, congo red is not absorbed by the colony on a screening culture medium, and the bacterial cells are gram-negative, rod-shaped and arranged into single or double spores without spores when observed by an optical microscope. The gram staining results are shown in FIG. 1.
2. 16S rDNA sequence homology analysis
A colony PCR method was used to amplify the 16S rDNA fragment of strain JY1-1. Through detection, the 16S rDNA of the strain JY1-1 has the nucleotide sequence shown as SEQ ID NO. 1. And (3) performing sequence alignment on the 16S rDNA of the strain JY1-1 to obtain a strain development tree analysis chart shown in figure 2.
By combining morphological characteristics of the strain and 16S rDNA sequence analysis, the strain JY1-1 was identified as rhizobiumRhizobium sp.OOOO. The strain is preserved in China general microbiological culture collection center (CGMCC) with the preservation number of CGMCC No.27052 and the preservation address of Beijing, china, for 4 months and 10 days.
3. Acid production test of strains
Taking rhizobium japonicum JY1-1, dissolving in 10mL of sterile water, respectively taking 0.2mL of the solution, inoculating the solution into 10 fermentation media with the volume of 50mL, culturing the solution at 30 ℃ and 200rpm, taking out one bottle every 8h, centrifuging the solution at 12000rpm for 10min, and measuring the pH value of the filtrate.
The measurement results of the pH value of the filtrate are shown in Table 1.
TABLE 1 determination of pH of filtrate
The pH value detection result of the filtrate is plotted to obtain a pH value change condition diagram of the fermentation broth of the strain JY1-1 shown in FIG. 3. As can be seen from fig. 3: rhizobia JY1-1 has higher acid-producing capability.
4. Preparation of fungus powder
Selecting rhizobium japonicum JY1-1, dissolving in 10mL of sterile water, inoculating 0.2mL of the rhizobium japonicum into a fermentation medium with the volume of 50mL, culturing at 30 ℃ and 200rpm for 48h, and performing spray drying on the fermentation broth to obtain rhizobium japonicum JY1-1 bacterial powder.
5. Preparation of oyster shell powder
Pulverizing Concha Ostreae, and grinding into powder to obtain Concha Ostreae powder.
6. Co-composting solid state fermentation
1. Adding the rhizobium JY1-1 bacterial powder prepared in the previous step into oyster shell powder and sugar residue and soybean meal co-compost to perform solid fermentation
The physicochemical properties of oyster shell powder, sugar residue and soybean meal are shown in Table 2.
TABLE 2 physicochemical Properties of the raw materials
Note that: a is based on wet basis mass; b is based on dry mass.
(1) Test design
The proportions of the raw materials of the co-compost are shown in Table 3.
TABLE 3 ratio of Co-compost raw materials (mass percent)
Mixing sugar slag, bean pulp and oyster shell powder according to the table 3, adding rhizobia JY1-1 bacterial powder prepared in the previous step according to the proportion of 0.5% and 1.0% of the dry weight of the whole system, and setting the initial water content of the whole pile material to 60% (w/w) by taking the rhizobia JY1-1 as a Control (CK) and carrying out 3 treatments. No repetition is provided.
After the materials are mixed, the mixture is filled into a 5L aerobic composting reactor, the fermentation time lasts for 35 days, and the ventilation conditions of all treatment groups are the same.
The method comprises the steps of carrying out turning operation on the 0 th day, the 3 rd day, the 6 th day, the 9 th day, the 14 th day, the 21 st day, the 28 th day and the 35 th day respectively, pouring the mixture in the bottle into another container during each turning operation, fully mixing, collecting compost samples by adopting a multipoint sampling method, sampling for 8 times, preserving in two parts, and preserving one fresh sample in a refrigerator at 4 ℃ for measuring fermentation indexes such as pH value, organic matters and the like; and the other part is naturally air-dried (dry sample) for standby, and is used for measuring fermentation indexes such as total humic acid (HS), humic Acid (HA), fulvic Acid (FA) and the like.
(2) Method for measuring fermentation index
(1) Method for measuring pH value
2g of fresh sample is weighed, 20mL of deionized water is added, shaking is carried out for 30min at 25 ℃ and 200rpm, a filter paper is rapidly filtered to obtain filtrate, and a pH meter (pH-100, force, shanghai) is used for measuring the pH value.
(2) Method for measuring organic matter
Expressed as Volatile Solid (VS) content, 10mL of crucible was baked at 105℃for 0.5h in an oven, cooled to room temperature, weighed, repeatedly operated until the crucible weight was constant, recorded as m 1 Weighing a proper amount of fresh sample, placing into a crucible, firstly placing into a baking oven, baking at 105 ℃ for 12 hours, cooling to room temperature, weighing, and marking as m 2 Burning in muffle furnace at 550 deg.C for 6 hr, cooling to room temperature, weighing and marking as m 3 The calculation formula of the VS content and the loss rate thereof is as follows:
wherein:VS 0 volatile solid for fermentation at initial timeThe content of the body is set to be equal to the total content of the body,VS t is the content of volatile solids after fermentation over time t.
(3) Method for measuring total humic acid (HS), humic Acid (HA) and Fulvic Acid (FA)
Weigh 1g of dry sample and use 20mL of Na 4 P 2 O 7 ·10H 2 Mixed solution of O solution and NaOH solution (Na 4 P 2 O 7 ·10H 2 The final concentrations of O and NaOH are 0.1 mol/L), oscillating for 12 hours at room temperature of 200rpm, centrifuging for 10 minutes at 4000rpm, taking the supernatant, repeating the above operation three times, storing the supernatant in a refrigerator at 4 ℃ each time, finally mixing the three supernatants, adding concentrated hydrochloric acid to adjust the pH value to 7, filtering the supernatant with a 0.45 mu m filter membrane, and detecting to obtain the total humic acid (HS) content. 10mL of the supernatant with the pH value adjusted is taken in a 50mL centrifuge tube, the pH value is adjusted to 1 by 6mol/L HCl solution, the mixture is placed in a refrigerator at the temperature of 4 ℃ for 12 hours and is centrifuged, the precipitate is Humic Acid (HA), and the supernatant is Fulvic Acid (FA). The contents of HS, HA and FA were determined using a TOC analyzer (vario TOC select, elementar, germany).
(3) Measurement results of fermentation index
(1) Measurement of pH value
TABLE 4 determination of the pH of compost samples
The results of the pH measurement of the compost samples were plotted to obtain a graph of the pH change of the co-compost shown in FIG. 4.
As can be seen from FIG. 4, the initial pH of the treated groups (0.5% JY1-1 group and 1.0% JY1-1 group) to which rhizobia JY1-1 was added was lower than that of the control group (CK group) to which rhizobia JY1-1 was not added, and the initial pH of the treated groups was decreased with the increase in the ratio of rhizobia JY1-1, because rhizobia JY1-1 produced acid during fermentation. As the fermentation proceeds, the pH of the compost rises to a maximum value on day 3 due to degradation of the acid and accumulation of ammonia, followed by a gradual decrease in pH (mainly due to evaporation of ammonia). In the later stage of composting, the pH value change of the compost tends to be stable. When composting is finished, the pH values of the treatment group and the control group are in the range of 7.4-8.5, which indicates that rhizobia JY1-1 has no negative effect on the overall pH value of the compost, and the final compost product also meets the maturation requirement.
(2) Results of measurement of organic matter
TABLE 5 organic matter measurement results of compost samples (%)
The organic matter measurement results of the compost samples were plotted to obtain a graph showing the change in the co-compost volatile solid loss rate shown in fig. 5.
As can be seen from fig. 5: the increase of the volatile solid loss rate in the early stage of composting is faster, which indicates that the organic matters are rapidly degraded, the increase of the volatile solid degradation efficiency of a control group (CK group) in the later stage of composting is slow, and the volatile solid loss rate of a treatment group (0.5% JY1-1 group and 1.0% JY1-1 group) added with rhizobium JY1-1 is still kept at a higher level, which indicates that the addition of rhizobium JY1-1 improves the degradation efficiency of the organic matters, accelerates the composting process, and increases the volatile solid loss rate along with the increase of the ratio of the rhizobium JY1-1, wherein the volatile solid loss rate of the 1.0% JY1-1 group is up to 46.54%.
(3) Measurement results of HS, HA and FA
TABLE 6 determination of HS, HA and FA for compost samples (g/kg)
The results of the HS, HA and FA measurements of the compost samples were plotted to obtain a graph of the total humic acid content in the co-compost shown in FIG. 6, a graph of the humic acid content in the co-compost shown in FIG. 7 and a graph of the fulvic acid content in the co-compost shown in FIG. 8, respectively.
As can be seen from fig. 6: after aerobic composting treatment, the content of total humic acid (HS) in all groups is in an ascending trend, and the content of HS in the final composting product is respectively as follows: CK group 97.65g/kg, 0.5% JY1-1 group 112.06g/kg, 1.0 JY1-1 group 129.74g/kg.
As can be seen from fig. 7: after aerobic composting treatment, the content of all groups of Humic Acid (HA) generally rises, and the content of HA in the final compost products is respectively as follows: CK group 97.65g/kg, 0.5% JY1-1 group 112.06g/kg, 1.0% JY1-1 group 129.74g/kg.
As can be seen from fig. 8: after aerobic composting, the content of the Fu-rienoic acid (FA) in all groups generally decreases, and the content of the FA in the final compost products is respectively as follows: CK group 52.66g/kg, 0.5% JY1-1 group 52.90g/kg, 1.0% JY1-1 group 51.21g/kg.
Obviously, the humus of the group with the rhizobium JY1-1 is higher than that of the group CK, which indicates that the addition of the rhizobium JY1-1 can promote the decomposition of minerals and the release of nutrients in the compost.
2. Adding the rhizobium JY1-1 bacterial powder prepared in the previous step into oyster shell powder and chicken manure and straw co-compost to perform solid fermentation
The physicochemical properties of oyster shell powder, chicken manure and straw are shown in Table 7.
TABLE 7 physicochemical Properties of the raw materials
Note that: a is based on wet basis mass; b is based on dry mass.
(1) Test design
The proportions of the raw materials of the co-compost are shown in Table 8.
Table 8 the proportions (mass percent) of the raw materials of the co-compost
Chicken manure, straw and oyster shell powder are firstly mixed according to the table 8, then rhizobium JY1-1 bacterial powder prepared in the prior art is added according to the proportion of 0.5 percent and 1.0 percent of the dry weight of the whole system, and rhizobium JY1-1 is not added as a Control (CK), wherein the initial water content of the whole pile material is set to be 60 percent (w/w), and the total number of treatments is 3. No repetition is provided.
After the materials are mixed, the mixture is filled into a 50L aerobic composting reactor, the fermentation time lasts for 35 days, and the ventilation conditions of all treatment groups are the same.
Turning over the mixture in the bottle on the 0 th day, the 3 rd day, the 7 th day, the 10 th day, the 17 th day, the 26 th day and the 35 th day respectively, pouring the mixture in the bottle into another container during each turning over operation, fully mixing, collecting compost samples by adopting a multipoint sampling method, sampling for 7 times, storing in two parts, storing one fresh sample in a refrigerator at 4 ℃, and measuring fermentation indexes such as pH value, organic matters and the like; and the other part is naturally air-dried (dry sample) for standby, and is used for measuring fermentation indexes such as total humic acid (HS), humic Acid (HA), fulvic Acid (FA) and the like.
(2) Method for measuring fermentation index
The method for measuring the pH value and organic matters of the fresh sample and the method for measuring the HS, HA and FA of the dry sample are the same, and are not repeated.
(3) Measurement results of fermentation index
(1) Measurement of pH value
TABLE 9 determination of the pH of compost samples
The results of the pH measurement of the compost samples were plotted to obtain a graph of the pH change of the co-compost shown in FIG. 9.
As can be seen from fig. 9: the initial pH of both the treatment and control groups was in the range of 7.07-8.89, with the treatment group being slightly lower than the control group. As the fermentation proceeds, the pH of each group of compost tends to rise first and then drop slightly as a whole, and the pH of the treated group is slightly lower than that of the control group, due to degradation of the acid and accumulation of ammonia. When composting is finished, the pH values of the treatment group and the control group are both in the range of 8.00-9.00, which indicates that rhizobia JY1-1 has no negative effect on the overall pH value of the compost, and the final compost product also meets the maturation requirement.
(2) Results of measurement of organic matter
Table 10 organic matter measurement results (%)
The organic matter measurement results of the compost samples were plotted to obtain a graph showing the change in the co-compost volatile solid loss rate shown in fig. 10.
As can be seen from fig. 10: the volatile solid loss rate of the pre-composting treatment group is faster than that of the control group, and the volatile solid loss rate of all groups in the later composting period is slower, but the volatile solid loss rate of the treatment group is higher than that of the control group, wherein the volatile solid loss rate of 1.0% JY1-1 group is as high as 43.19%.
(3) Measurement results of HS, HA and FA
TABLE 11 determination of HS, HA and FA for compost samples (g/kg)
The results of the HS, HA and FA measurements of the compost samples were plotted to obtain a graph of the total humic acid content in the co-compost shown in FIG. 11, a graph of the humic acid content in the co-compost shown in FIG. 12 and a graph of the fulvic acid content in the co-compost shown in FIG. 13, respectively.
As can be seen from fig. 11: in the aerobic composting process, the total humic acid (HS) content after composting of all groups is in an ascending trend, and the HS content in the final composting products is respectively as follows: CK group 130.95g/kg, 0.5% JY1-1 group 137.64g/kg, 1.0% JY1-1 group 144.83g/kg.
As can be seen from fig. 12: in the aerobic composting process, the content of Humic Acid (HA) after composting of all groups generally rises, and the content of HA in the final composting products is respectively as follows: CK 86.21g/kg, 0.5% JY1-1 93.09g/kg, 1.0% JY1-1 108.01g/kg.
As can be seen from fig. 13: in the aerobic composting process, the content of Fulvic Acid (FA) after composting of all groups generally shows a descending trend, and the content of FA in the final composting products is respectively as follows: CK group 44.74g/kg, 0.5% JY1-1 group 44.55g/kg, 1.0% JY1-1 group 36.82g/kg.
Obviously, the humus of the group with the rhizobium JY1-1 is higher than that of the group CK, which indicates that the addition of the rhizobium JY1-1 can promote the decomposition of minerals and the release of nutrients in the compost.
To sum up: the rhizobia JY1-1 obtained by screening is added into oyster shell-organic waste co-composting, so that the co-composting process of oyster shell and organic waste can be accelerated, and the degradation of organic matters can be promoted.
It should be noted that the above examples are only examples for clearly illustrating the present invention, and are not limiting to the embodiments of the present invention. Other variations or modifications of the above teachings will be apparent to those of ordinary skill in the art. Not all embodiments are exhaustive. All obvious changes or modifications which are obvious from the technical proposal of the invention are still within the protection scope of the invention.
Claims (7)
1. The acid-producing rhizobium JY1-1 is characterized in that the rhizobium JY1-1 is preserved in the China general microbiological culture Collection center of the China Committee for culture Collection of microorganisms, and is classified and named as rhizobiumRhizobium sp.The preservation number is CGMCC NO.27052, the preservation date is 2023, 4 and 10 days, and the preservation address is Beijing in China.
2. The oyster shell-organic waste co-compost is characterized by comprising 40% by mass, 56% by mass and 4% by mass of oyster shell powder, 56% by mass of sugar slag and 4% by mass of soybean meal and rhizobia JY1-1 which account for 0.5-1.0% by dry weight of the whole system, and the initial water content of the whole pile material is 60%.
3. The oyster shell-organic waste co-compost of claim 2, wherein the rhizobia JY1-1 exists in the form of bacterial powder, and is prepared by the following method:
rhizobium JY1-1 is selected, dissolved in sterile water, inoculated in a fermentation medium, cultured for 48 hours at 30 ℃ and 200rpm, and the fermentation liquid is spray-dried to prepare rhizobium JY1-1 bacterial powder.
4. The oyster shell-organic waste co-compost of claim 3, wherein the fermentation medium has a formula of:
glucose 50g/L, yeast powder 5g/L, KH 2 PO 4 ·H 2 O 1.4g/L，MgSO 4 ·7H 2 O 0.5g/L，（NH 4 ） 2 SO 4 10g/L，FeSO 4 ·7H 2 O 0.03g/L，ZnSO 4 0.03g/L。
5. The oyster shell-organic waste co-compost is characterized by comprising 40% of oyster shell powder, 16.28% of chicken manure, 43.72% of straw and 1-1% of rhizobia JY1-1 in percentage by mass, wherein the rhizobia JY1-1 accounts for 0.5-1.0% of the dry weight of the whole system, and the initial water content of the whole pile material is 60%.
6. The oyster shell-organic waste co-compost of claim 5, wherein the rhizobia JY1-1 exists in the form of bacterial powder, and is prepared by the following method:
rhizobium JY1-1 is selected, dissolved in sterile water, inoculated in a fermentation medium, cultured for 48 hours at 30 ℃ and 200rpm, and the fermentation liquid is spray-dried to prepare rhizobium JY1-1 bacterial powder.
7. The oyster shell-organic waste co-compost of claim 6, wherein the fermentation medium has a formula of:
glucose 50g/L, yeast powder 5g/L, KH 2 PO 4 ·H 2 O 1.4g/L，MgSO 4 ·7H 2 O 0.5g/L，（NH 4 ） 2 SO 4 10g/L，FeSO 4 ·7H 2 O 0.03g/L，ZnSO 4 0.03g/L。
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| US5281530A (en) * | 1987-08-12 | 1994-01-25 | Mycogen Corporation | Genes encoding nematode-active toxins cloned from bacillus thuringiensis isolate PS17 |
| CA2907438A1 (en) * | 2013-03-15 | 2014-09-18 | Spogen Biotech Inc. | Plant growth-promoting bacteria and methods of use |
| CN109640649A (en) * | 2016-03-16 | 2019-04-16 | 斯波根生物技术公司 | Promote the method for plant health using resolvase and the microorganism for being overexpressed enzyme |
| CN116348589A (en) * | 2020-08-03 | 2023-06-27 | 巴斯夫欧洲公司 | Materials and methods for improving plant health |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5281530A (en) * | 1987-08-12 | 1994-01-25 | Mycogen Corporation | Genes encoding nematode-active toxins cloned from bacillus thuringiensis isolate PS17 |
| CA2907438A1 (en) * | 2013-03-15 | 2014-09-18 | Spogen Biotech Inc. | Plant growth-promoting bacteria and methods of use |
| CN109640649A (en) * | 2016-03-16 | 2019-04-16 | 斯波根生物技术公司 | Promote the method for plant health using resolvase and the microorganism for being overexpressed enzyme |
| CN116348589A (en) * | 2020-08-03 | 2023-06-27 | 巴斯夫欧洲公司 | Materials and methods for improving plant health |
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