CN106148218A - Aquatic product photosynthetic bacteria Tiny ecosystem and application thereof - Google Patents

Aquatic product photosynthetic bacteria Tiny ecosystem and application thereof Download PDF

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Publication number
CN106148218A
CN106148218A CN201510177916.9A CN201510177916A CN106148218A CN 106148218 A CN106148218 A CN 106148218A CN 201510177916 A CN201510177916 A CN 201510177916A CN 106148218 A CN106148218 A CN 106148218A
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China
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cgmcc
photosynthetic bacteria
rhodopseudomonas palustris
nitrite
present
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CN201510177916.9A
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CN106148218B (en
Inventor
汪攀
王蕊
李亚
刘婷
何增国
朱传忠
郑慧
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Beijing Da Bei Agricultural Science And Technology Group Ltd By Share Ltd
Jiujiang Dabei agricultural and Fishery Technology Co.,Ltd.
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Fujian Dabeinong Fisheries Science & Technology Co Ltd
TIANJIN CHANGNONG TECHNOLOGY Co Ltd
Beijing Dabeinong Technology Group Co Ltd
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Abstract

The present invention relates to microbial ecological agent technical field, be specifically related to a strain and improve the Rhodopseudomonas palustris of culture environment of aquatic products.The preserving number of Rhodopseudomonas palustris involved in the present invention is CGMCC No.9136.Having good result in terms of using the microbial ecological agent of CGMCC No.9136 of the present invention, the nitrite in degrading cultivation environment, within 24 hours, content of nitrite can be degraded 98%.

Description

Aquatic product photosynthetic bacteria Tiny ecosystem and application thereof
Technical field
The invention belongs to Tiny ecosystem applied technical field, be specifically related to a strain Rhodopseudomonas palustris Microbial ecological agent and application in aquaculture.
Background technology
In aquaculture, clear, rich, U.S.A of water body is to weigh the important indicator that breeding environment is good and bad. Clearly, water body to be made keeps nature limpid;Richness, refers to contain abundant algae in water body and swim dynamic Thing, provides sufficient bait as aquatic product animal;U.S.A i.e. refers to water body environment and aquatic animal Form ecological balance.And at present due to the high-density breeding in pond, cultivated animals feces and residual bait Constantly pile up, harmful microorganism and algal bloom, water body severe depletion of oxygen, cause fish and shrimp to float Head, the most dead phenomenon are extremely serious, and the use of a large amount of antibiotic can be made again in this case Become cultivated animals antibiotic content to exceed standard, and then affect the healthy of the mankind.Therefore, utilize It is extremely urgent that the method for biological restoration makes breeding water body reach nature, harmonious state.
Photosynthetic bacteria (Photosynthetic bacteria is called for short PSB) is to occur the earliest on the earth There is the prokaryote of original luminous energy synthetic system, be that a big class is not put The general name of the photosynthetic antibacterial of oxygen, for gram negative bacteria, is widely present in global biosphere Everywhere.Photosynthetic bacterial thallus is nontoxic, and protein content is up to 60%, containing vitamin B group, Ubiquinone and carotenoid.Because of its distinctive fixed nitrogen, suction hydrogen, carbon sequestration, utilize sulfide Deng metabolism mode at a lot of aspects, there is important practicality and economic worth.Support at Aquatic product Grow photosynthetic bacteria in industry to cannot be only used for purifying water, be also of high nutritive value because of it, can be as bait Feed additives, in preventing and treating fish and shrimp disease, promotes that the aspect such as growth, raising incubation rate all shows Good performance.Be flooded with diversiform photosynthetic bacteria preparation in the market, with liquid and It is in the majority that powdery exists, and according to laboratory to the analysis of multiple product on market, its quality is not to the utmost People's will, mainly shows as liquid bacterial agent color the most bright-coloured, thalline poor stability, and viable count is less, Even with the presence of find no photosynthetic bacteria, the urgent needs in market is with at present the most photosynthetic The uneven troubled waters of Production by Bacteria quality is to we provide the best opportunity, therefore, Develop excellent photosynthetic bacteria preparation significant.
Summary of the invention
The present invention provides a kind of photosynthetic bacteria microbial inoculum and application thereof.
It is an object of the invention to by the realization of techniques below mode scheme:
The present invention provides a strain Rhodopseudomonas palustris (Rhodopseudomonaspalustris) New strains, its preserving number is CGMCC No.9136.
The separation of the Rhodopseudomonas palustris (Rhodopseudomonaspalustris) of the present invention Qualification process is as follows:
1. preparation photosynthetic bacteria culture medium
Enrichment medium (/L): NaHCO32g, yeast powder 1g, K2HPO4·3H2O 0.2g, NH4Cl 1g, NaAc 3H2O 0.3g, MgSO4·7H2O 0.2g, NaCl 0.2g, Na2S2O3·5H2O 1g, pH7.0,121 DEG C of autoclaving 20min.
Isolation medium adds agar on the basis of enrichment medium, and final mass is divided into 1.5%, 121 DEG C of autoclaving 20min.
2. enrichment:
The muddy water sample come will be gathered it will be put in the anaerobism pipe of band screw socket added with enrichment medium In, wherein anaerobism pipe fills 15mL culture medium and 10mL pond water+bed mud, each covering 2-3cm paraffin oil.Intensity of illumination 1000lx~2000lx, 30 DEG C of Anaerobic culturel.
The most isolated and purified
After being enriched with one week, from the anaerobism pipe that culture medium reddens, take out 1mL liquid, take dilution Degree is 10-3、10-4, 10-5Liquid 100 μ L coated plate, within about 10 days, observed red bacterium Fall to growing;Choose red colonies flat board setting-out to pure.
4. identify
Picking list bacterium colony carries out 16S rRNA bacterium colony PCR, send Invitrogen to check order, sequence As shown in SEQ ID No.1.Record sequence on NCBI, carry out Blast comparison, tentatively reflect It is set to Rhodopseudomonas palustris Rhodopseudomonas palustris, with Rhodopseudomonas palustris The 16S rRNA gene similarity distance of R.palustrisATCC 17003 is 100%.This bacterium Poor growth, may occur in which single bacterium colony, size 0.5mm~1mm in 5 days;It is dark under anaerobic state Red colonies, Gram’s staining is negative rod-short, and solid plate is cultivated as rod-short (Fig. 1).
To be separated to during Rhodopseudomonas palustris Rhodopseudomonas palustris is preserved in State's Microbiological Culture Collection administration committee common micro-organisms center, is called for short CGMCC, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 Institute of Microorganism, Academia Sinica, preservation day Phase is on May 8th, 2014, and deposit number is respectively as follows: CGMCC No.9136.
Present invention also offers the microbial ecological agent comprising this Rhodopseudomonas palustris.Said preparation by Rhodopseudomonas palustris Rhodopseudomonas palustris CGMCC No.9136 fermentation and Become.
The present invention also provides for the preparation method of described microbial ecological agent, and it comprises the steps: natural pond Pool rhodopseudomonas CGMCC No.9136 seed liquor is inoculated with the inoculum concentration of volume ratio 1-10% In fermentation liquid, illumination cultivation 2-5 days.
Photosynthetic bacteria microbial inoculum in the present invention has degraded clearly to water nitrite content Effect, even if under dark condition, also has good degradation effect.Use CGMCC of the present invention The microbial ecological agent of No.9136, within 24 hours, content of nitrite can be degraded 98%, This effect can be reached without colleague's series products.
Accompanying drawing explanation
The bacterium colony figure of Fig. 1 Rhodopseudomonas palustris (Rhodopseudomonaspalustris).
Fig. 2 Asia acid sodium content canonical plotting.
The photosynthetic bacteria degradation effect to nitrite under the conditions of Fig. 3 two kinds.The left side is without Rhizoma Dioscoreae (peeled) The CGMCC No.9136 degraded to nitrite under part, the right is under illumination condition The CGMCC No.9136 degraded to nitrite.
Fig. 4 show shrimp culture pond nitrous degradation effect contrast under natural lighting.
Detailed description of the invention
Following example are used for illustrating the present invention, but are not limited to the scope of the present invention.
Prepared by embodiment 1 Rhodopseudomonas palustris bacterium solution
1. the formula (/L) of Rhodopseudomonas palustris amplification culture base:
CaCl20.1g, KH2PO40.5g, NH4Cl 1g, MgCl2·6H2O 0.5g, NaCl 1g, NaAc·3H2O 1g, two hydration sodium succinate 1g, yeast powder 0.3g, peptone 0.3g
2. amplification culture base incubation step
(1) each component is weighed according to culture medium prescription, with water dissolution, by culture medium subpackage To the PET bucket that 5L has disinfected in alcohol, every barrel of 4.5L, the red false unit cell in inoculation marsh Bacterium CGMCC No.9136500mL, capping.It is placed in electric filament lamp illumination cultivation 4 days in carton, Can be as the seed liquor of next step large-scale culture.
(3) each composition required for 1 ton of culture fluid is weighed, first complete with a small amount of tap water Dissolve, then be sufficiently mixed in drum with remaining tap water, inoculate 10 barrels of photosynthetic bacteria kinds Sub-liquid, is sub-packed in 5L transparent PET Plastic Drum capping after mixing.Illumination cultivation 5 days, treats Cultivate to rufous, i.e. obtain highly purified photosynthetic bacteria liquid.
The embodiment 2 Rhodopseudomonas palustris CGMCC No.9136 degradation capability to nitrite Test
1. experimental technique
(1) nitrite standard curve is done: prepare the nitrite solution of variable concentrations, put OD is measured under microplate reader550, make salinity and OD550Correspondence standard curve.
(2) culture medium of fall nitrous experiment: preparing 3 ° of saline, to be sub-packed in 150mL little by three In the bottle of angle, 50mL/ bottle, every bottle adds 0.05g sediment of pond.121 DEG C, high pressure steam sterilization 20min。
(3) dark condition and illumination condition be set, each under the conditions of experimental group and comparison are set Group, every strain bacterium 3 repetition, experimental group adds photosynthetic bacteria seed liquor 50 μ L, places respectively In dark with under illumination condition, place for 30 DEG C and cultivate.
(4) within every 24 hours, a content of nitrite is measured, method particularly includes: in ultra-clean work Taking 1mL liquid in station in the centrifuge tube of 1.5mL, 12,000rpm are centrifuged 2 minutes, inhale Take supernatant 200 μ L in 96 orifice plates, successively add each 20 μ L of Griess reagent I, II, quiet Putting 5min makes it react completely, is subsequently placed under microplate reader measurement OD550
(5) according to standard curve experiment with computing group and the nitrite concentration of matched group, measure 2 days, record experimental data was also analyzed.
2. experimental result
(1) nitrous content and OD550Standard curve making
Being illustrated in figure 2 nitrous content standard curve chart, the linearly dependent coefficient obtained is 0.998, The sodium nitrite concentration of the standard curve reaction showing this time to manufacture and OD550There is the strongest being correlated with Property.
(2) as it is shown on figure 3, described pool rhodopseudomonas CGMCC under dark and illumination condition No.9136 microbial inoculum all has obvious nitrite degradation effect.Under non-illuminated conditions, nitrous Content is degraded to minimum 0.05mg/L from 2.5mg/L;Under illumination condition, sodium nitrite from 2.3mg/L is degraded to the level of 0.005mg/L.
Comparative example 1 Rhodopseudomonas palustris CGMCC No.9136 and commercial like product application effect Fruit contrast
The three mouthfuls of cultivating pools choosing same cultivation factory do Contrast on effect, area 1 mu, cultivate close Spend 80,000 big every mu, water temperature 26 degrees Celsius, shrimp body length 4 centimetres.The full pond of 16:00 in afternoon is sprinkled Spill photosynthetic bacteria, every mu splash 1 barrel (5L), measure a water nitrite acid every 24 hours Salinity, surveys three days continuously.Result shows (Fig. 4), uses CGMCC No.9136, Within 24 hours, content of nitrite can be degraded 98%, and series products of going together can not reach this Effect.

Claims (3)

1. a strain Rhodopseudomonas palustris (Rhodopseudomonas palustris) new strains, Its preserving number is CGMCC No.9136.
2. containing the photosynthetic bacteria microbial ecological agent of bacterial strain described in claim 1, it is marsh The fermentation liquid of rhodopseudomonas CGMCC No.9136.
3. the preparation method of the photosynthetic bacteria microbial ecological agent described in claim 2, it includes Following steps: Rhodopseudomonas palustris CGMCC No.9136 seed liquor is with volume ratio 1-10% Inoculum concentration be inoculated in fermentation liquid, illumination cultivation 2-5 days.
CN201510177916.9A 2015-04-15 2015-04-15 Photosynthetic bacteria microecology for aquatic products and application thereof Active CN106148218B (en)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107805112A (en) * 2017-11-14 2018-03-16 汪逸凡 A kind of preparation method of STEVIA REBAUDIANA special fertilizer
CN113461179A (en) * 2021-08-06 2021-10-01 南京农业大学 Application of rhodopseudomonas palustris ZT-MG2 in eutrophic water body treatment

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101050446A (en) * 2007-03-21 2007-10-10 珠海市农业科学研究中心 Swamp Rhodopseudomonas of using nitrite nitrogen in high effect, and application
CN104498399A (en) * 2014-12-10 2015-04-08 湖南省植物保护研究所 Rhodopseudomonas palustris strain, biological agent and preparation method and application of biological agent

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101050446A (en) * 2007-03-21 2007-10-10 珠海市农业科学研究中心 Swamp Rhodopseudomonas of using nitrite nitrogen in high effect, and application
CN104498399A (en) * 2014-12-10 2015-04-08 湖南省植物保护研究所 Rhodopseudomonas palustris strain, biological agent and preparation method and application of biological agent

Non-Patent Citations (2)

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陈燕红等: "光照、氧气、pH和盐度对沼泽红假单胞菌2-8菌株生长和亚硝酸消除的影响", 《南方水产》 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107805112A (en) * 2017-11-14 2018-03-16 汪逸凡 A kind of preparation method of STEVIA REBAUDIANA special fertilizer
CN113461179A (en) * 2021-08-06 2021-10-01 南京农业大学 Application of rhodopseudomonas palustris ZT-MG2 in eutrophic water body treatment

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Address after: 100080, Zhongguancun building, No. 27 Zhongguancun street, Beijing, Haidian District, China, 14 floor

Patentee after: BEIJING DABEINONG TECHNOLOGY GROUP Co.,Ltd.

Patentee after: Fujian Dabei nonghuayou Aquatic Technology Group Co.,Ltd.

Patentee after: Tianjin DaBeiNong Biotechnology Co.,Ltd.

Address before: 100080, Zhongguancun building, No. 27 Zhongguancun street, Beijing, Haidian District, China, 14 floor

Patentee before: BEIJING DABEINONG TECHNOLOGY GROUP Co.,Ltd.

Patentee before: FUJIAN DABEINONG FISHERIES SCIENCE & TECHNOLOGY Co.,Ltd.

Patentee before: TIANJIN CHANGNONG SCIENCE & TECHNOLOGY CO.,LTD.

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Effective date of registration: 20220628

Address after: 332020 east of science and technology 1st Avenue and Gaoxin 8th Road, Gongqing urban industrial park, Jiujiang City, Jiangxi Province

Patentee after: Jiujiang Dabei agricultural and Fishery Technology Co.,Ltd.

Patentee after: Beijing Da Bei agricultural science and technology group Limited by Share Ltd

Address before: 100080, Zhongguancun building, No. 27 Zhongguancun street, Beijing, Haidian District, China, 14 floor

Patentee before: BEIJING DABEINONG TECHNOLOGY GROUP Co.,Ltd.

Patentee before: Fujian Dabei nonghuayou Aquatic Technology Group Co.,Ltd.

Patentee before: Tianjin DaBeiNong Biotechnology Co.,Ltd.