CN103773714A - Application of photosynthetic bacteria to improving sea farming water quality - Google Patents
Application of photosynthetic bacteria to improving sea farming water quality Download PDFInfo
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- CN103773714A CN103773714A CN201310730681.2A CN201310730681A CN103773714A CN 103773714 A CN103773714 A CN 103773714A CN 201310730681 A CN201310730681 A CN 201310730681A CN 103773714 A CN103773714 A CN 103773714A
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- photosynthetic bacterium
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- bacterium
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Abstract
The invention relates to the field of an application of microorganisms to improving the sea farming water quality and particularly relates to a photosynthetic bacterium SC01 and an application thereof. The photosynthetic bacterium is Rhodobactersphaeroides and is preserved in China General Microbiological Culture Collection Center on September 25, 2013; the preservation number is CGMCC No.8248. The photosynthetic bacterium can be applied to treating pollutants generated in a sea farming process. The strain provided by the invention has the characteristics of simple and feasible screening method, convenience in production, storage and transportation, obvious utilization effect and the like, and has a good application prospect.
Description
Technical field
The present invention relates to the Application Areas of microorganism in sea farming water correction, specifically a kind of photosynthetic bacterium SC01 and application thereof.
Background technology
The mariculture industry of China is through the fast development of many decades, in coastal economy and even the national economic development, play a part very important, follow the fast development of mariculture industry, the processing problem of the disease in breeding process and breeding wastewater becomes increasingly conspicuous, and becomes one of bottleneck of restriction industry sustainable health development.Particularly breeding wastewater can not get for a long time effectively processing and directly arranges extra large.In the seawater after cultivation, the nutritive ingredients such as Ammonia In Sea Water that bait is residual, the ight soil of aquaculture organism etc. causes, nitrite nitrogen, phosphoric acid salt are abundant, and arbitrarily row sea easily causes growing of the harmful organisms such as algae.In the time that envrionment conditions is suitable, some breed the outburst of red tide algae rapidly, usually cause the sharply decline of extra large water oxygen level, cause the aquaculture organism mortality of mariculture area, simultaneously, owing to containing the objectionable impuritiess such as microbiotic, sanitising agent, sterilant in breeding wastewater, arbitrarily arrange sea and likely cause the even minimizing of neritic organism diversity or disappearance of offshore, cause a series of serious ecological problems such as aqueous desert.
Therefore, develop some based on microbial technique and can be used in sea farming, be both conducive to aquaculture organism health, have the probiotics that can carry out cultivation water improvement, can greatly alleviate generation and the development of the problems referred to above.Consume thereby be conducive to reduce cultivation, reduce row's sea wind danger, increase farmers' income, promote long-term stability and the Sustainable development of industry.
The organism such as sulfide, ammonia that photosynthetic bacterium one class can be using light as the energy, utilize occurring in nature under anaerobism illumination or aerobic dark condition carries out photosynthetic microorganism as the hydrogen donor carbon source of holding concurrently.Photosynthetic bacterium is the prokaryotic organism that occur the earliest having original luminous energy synthetic system on the earth.In long-term evolutionary process, photosynthetic bacterium has formed the metabolic system of own uniqueness, has potential scientific research and using value aspect newtype drug, function enzyme and environmental improvement.
Aspect aquaculture, people have started to utilize photosynthetic bacterium to carry out the improvement of cultivation water and the practice of disease control aspect, but lack the theoretical direction of system.For bacterial classification select, the usage quantity of photosynthetic bacterium, the aspects such as use opportunity still do not have systematic theoretical direction at present, produce and use procedure in still there is larger blindness.
Summary of the invention
The invention provides photosynthetic bacterium SC01 and the application in sea farming water correction thereof.
For achieving the above object, the technical solution used in the present invention is:
A kind of photosynthetic bacterium SC01, the red bacterium of photosynthetic bacterium taxonomy called after class ball (Rhodobacter sphaeroides), on September 25th, 2013 in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, deposit number is CGMCC No.8248.Depositary institution address is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City.
Described strain culturing mode is:
1) sampling and process: get respectively plant area of turbot fry breeding factory waterways bottom seawater, pool wall dirt settling, pool bottom sludge, grow seedlings and culturing pool bottom seawater, dirt settling, pool wall and pond at the bottom of residual mud 5-10g, add the sterilizing seawater of 50-100ml, be placed in the aseptic triangular flask of 150-300ml, fully concussion 10-15min, remove the large particulate matters such as weeds, rock, be prepared into sample suspension liquid.
2) enrichment culture: sample thief suspension liquid 5-8mL, adds the aseptic enrichment culture liquid (KH that contains 35-50mL
2pO
40.025-0.5g/L, yeast extract paste 0.5-1.5g/L, Tryptones 0.02-0.05g, CaCl0.03-0.08g, NaAc0.5-1.5g/L, NH
4cl0.1-0.3g/L, MnSO
40.025-0.05g, MgSO
40.025-0.05g, FeSO
40.002-0.005g, pH value 6.8-7.4, is settled to 1000mL with distilled water) 100-150ml triangular flask in, add 10-15ml paraffin oil, sealed membrane sealing.28-30 ℃, 1500-3000lux illumination cultivation.Every 48h observes nutrient solution colour-change.This enrichment culture process continues to 30-60 days.
3) separation and Culture: when enrichment culture liquid obviously presents redness, get 2-5ml, add the liquid separation culture medium (K of 30-50ml
2hPO
40.025-0.5g/L, KH
2pO
40.025-0.5g/L, yeast extract paste 0.5-1.5g/L, NaAc0.5-1.5g/L, NH
4cl0.1-0.3g/L, Glu L-glutamic acid 0.002-0.005g, pH value 6.8-7.4, is settled to 1000mL with distilled water), 28-30 ℃, 1500-3000lux illumination, 3-5d is cultivated in 150-300rpm concussion.Now nutrient solution becomes scarlet by the naked eye, gets this nutrient solution of 50-200ul and is coated on solid separation culture medium flat board.Select red bacterium colony, repeat line and separate, to obtaining the consistent pure growth of colonial morphology, called after SC01 also preserves.Analyze through Physiology and biochemistry and 16s rDNA, determine that SC01 is a strain class ball photosynthetic bacterium.
This photosynthetic bacterium is the red bacterium of a kind ball (Rhodobacter sphaeroides).Gramstaining is negative, without gemma, without pod membrane.On LB flat board, it is red that bacterium colony becomes, circle, and intermediate projections, smooth moistening.
Photosynthetic bacterium of the present invention separates and obtains from turbot fry breeding factory.This photosynthetic bacterium gramstaining is negative, without gemma, without pod membrane.On LB flat board, it is red that bacterium colony becomes, circle, and intermediate projections, smooth moistening.The bacterial strain obtaining is carried out to 16SrDNA sequencing, adopt bacterium universal primer 27F and 1492R to carry out pcr amplification, to get PCR product 2～5 μ l and carry out 1wt% agarose gel electrophoresis, ultraviolet detection after EB dyeing. the PCR product of cutting glue recovery 1.5kb size with sepharose purification kit (purchased from the precious biotech firm in Dalian) carries out sequencing analysis.The 16SrDNA sequence sequencing result login GenBank of bacterial strain, and carry out homology comparison by the 16SrDNA sequence in BLAST and GenBank, by the 16SrDNA of the similar bacterial strain obtaining, utilize MEGA4.0 software, phylogenetic tree construction, analyzes the evolutionary degree of each isolated strains.Through identifying: be that one is the red bacterium of class ball (Rhodobacter sphaeroides).On September 25th, 2013, in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, deposit number was CGMCC No.8248.The GenBank number of logging in of this bacterial strain 16SrDNA is: KF791043.
The application of photosynthetic bacterium SC01, described photosynthetic bacterium can be applicable to process the pollutent producing in sea farming process.
Described photosynthetic bacterium put in breeding seawater and then improve water inlet and the discharge water of sea farming water, improving the survival rate of sea farming biology simultaneously.
The present invention has advantages of: photosynthetic bacterium of the present invention separates and obtains from the turbot fry breeding factory of Yantai City.The processing of this photosynthetic bacterium after to the water quality deterioration causing because of bait is residual, ight soil etc. in the Main Economic animal cultivation processes such as turbot has good effect.To low aeration rate or significant without the existence of main cultivated animals under aeration.Can be used for the improvement of water quality and the processing of breeding wastewater in aquaculture process, have a good application prospect.
Embodiment
Be described in detail around bacterial strain of the present invention and embodiment, but embodiments of the present invention are not limited to this.
The screening of photosynthetic bacterium
Screening and the cultivation of embodiment 1 photosynthetic bacterium
1) sampling and process: get respectively plant area of turbot fry breeding factory waterways bottom seawater, pool wall dirt settling, pool bottom sludge, grow seedlings and culturing pool bottom seawater, dirt settling, pool wall and pond at the bottom of residual mud 8g, add the sterilizing seawater of 100ml, be placed in the aseptic triangular flask of 300ml, fully concussion 15min, remove the large particulate matters such as weeds, rock, be prepared into sample suspension liquid.
2) enrichment culture: sample thief suspension liquid 5mL, adds the aseptic enrichment medium (KH that contains 50mL
2pO
40.5g/L, yeast extract paste 1.5g/L, Tryptones 0.05g, CaCl0.05g, NaAc1.0g/L, NH
4cl0.2g/L, MnSO
40.05g, MgSO
40.025g, FeSO
40.002g, pH value 7.2, is settled to 1000mL with distilled water) 150ml triangular flask in, add 10ml paraffin oil, sealed membrane sealing.28 ℃, 2000lux illumination cultivation.Every 48h observes nutrient solution colour-change.This enrichment culture process continues to 30 days.
3) isolation identification: when enrichment culture liquid obviously presents redness, get 2ml, add the liquid separation culture medium (K of 50ml
2hPO
40.5g/L, KH
2pO
40.5g/L, yeast extract paste 1.5g/L, NaAc1.0g/L, NH
4cl0.2g/L, L-glutamic acid 0.005g, pH value 7.4, be settled to 1000mL with distilled water), 28 ℃, 2000lux illumination, 300rpm shaking culture 3d, now nutrient solution becomes scarlet by the naked eye, gets this nutrient solution of 100ul and is coated on solid separation culture medium flat board (K
2hPO
40.5g/L, KH
2pO
40.5g/L, yeast extract paste 1.5g/L, NaAc1.0g/L, NH
4cl0.2g/L, L-glutamic acid 0.005g, agar powder 10g, pH value 7.4, is settled to 1000mL with distilled water.) upper, 28 ℃, 2000lux illumination, is inverted and cultivates 7d, selects red bacterium colony, repeats line on above-mentioned solid medium and separates, and to obtaining the consistent pure growth of colonial morphology, called after SC01 also preserves.Analyze through Physiology and biochemistry and 16s rDNA, determine that SC01 is the red bacterium of class ball (Rhodobacter sphaeroides).On September 25th, 2013, in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, deposit number was CGMCC No.8248.
4) fast culture: get 50ul-80 ℃ of frozen SC01 and be seeded to 10ml fast culture media (K
2hPO
40.25g/L, KH
2pO
40.25g/L, yeast extract paste 3.0g/L, NaAc2.0g/L, NH
4cl1.0g/L, pH value 6.8, is settled to 1000mL with Chen Haishui (removing throw out after staticly settling 14 days).) in, 28 ℃, 300rpm, cultivates 120h to OD under natural lighting
600to 1.0, using this as seed liquor, according to 5%(volume ratio) inoculum size is inoculated in seed liquor in fast culture media, and 28 ℃, 150rpm, cultivates 120h under natural lighting.
The impact of embodiment 2:SC01 on objectionable impuritiess such as ammonia nitrogens in breeding seawater
Get respectively 0,0.1,0.5,2,5,10ml OD
600to 1.0 SC01 nutrient solution, be added into respectively in the storage pond, holding pond of 50L seawater, wherein, think and add the non-control group of SC01 nutrient solution group, in each culturing pool, put into the healthy turbot that 10 health total lengths are 6-8cm.Indoor normal illumination, every day, conventional granulates feed addition was pressed every fish 0.5-0.8g, divided and added for 3 times.Respectively at adding the the 4th, 18,28 after SC01,40h water sampling, pH in test water, the variation (referring to table 1) of ammonia nitrogen, nitrite nitrogen, dissolved oxygen, activared carbon sulfur salts contg, by visible at each tested time point in table 1, the pH of each culturing pool and dissolved oxygen change not obvious.But along with the carrying out of breeding process, ammonia nitrogen in aquaculture water, nitrite nitrogen, the content of reactive phosphate raises gradually.
Simultaneously various objectionable impuritiess of the passing in time content in seawater raises gradually as shown in Table 1, especially obvious with ammonia nitrogen.Be 0.1,0.5 at dosage, in the culturing pool of 2.0ml SC01 nutrient solution, every detection index of seawater is all less than the control group that does not add SC01 nutrient solution, illustrates that having of appropriate SC01 is beneficial to the control of ammonia nitrogen in aquaculture water, nitrite nitrogen and reactive phosphate.But in the time that the dosage of SC01 increases to 5-10ml, the each monitoring index in seawater goes up not down, even higher than the control group that does not add SC01 nutrient solution.Visible because the bacterial number adding is too many, and the metabolite of bacterium and thalline itself makes water body be secondary polluted on the contrary.
The impact of table 1SC01 on objectionable impurities in breeding seawater
Embodiment 3: the impact on turbot mortality ratio under without aeration condition SC01
Get 0,0.1,0.5,2,5,10ml OD
600to 1.0 SC01 nutrient solution, be added into respectively in the storage pond, holding pond that contains 50L seawater, not add SC01 nutrient solution group as control group, in each culturing pool, putting into respectively 10 bodies long is 6-9cm, the healthy turbot seedling that mean body weight is 6-8g.Indoor normal illumination, each culturing pool per minute air flow is 6-8L, and the dosage of conventional granulates feed is according to every fish 0.8-1.0g every day, and every day, 7:00,13:00,19:00 divide and add for three times.Add after SC00148h, all experimental group and control group are all stopped to aeration 5-8h, observe the death condition (referring to table 2) of cultured fishes.Under stuffiness condition, the dissolved oxygen content in culturing pool declines rapidly.Stopping after aeration in 5h, do not add turbot seedling in the culturing pool of SC01 very fast all dead, and in the culturing pool that adds SC01 nutrient solution the death time of turbot seedling obviously postpone; The survival rate of turbot seedling exists directly related with SC01 add-on: in the 8h stopping after aeration, and none death of turbot seedling in the storage pond, holding pond that add-on is 1%; Add-on is that in 4% storage pond, holding pond, the mortality ratio of turbot seedling is 20%.And be less than 0.2% or while being greater than 10% when the addition of SC01, could not play good protection effect to turbot seedling.Therefore under suitable concentration, the existence of SC01 can consume the organism such as swill, ight soil in culturing pool, avoids the too high death that causes aquaculture organism of the concentration of narmful substances such as ammonia nitrogen.But too much the existence of SC01 may cause gathering of opsonigenous substances or by product again, unfavorable to aquaculture organism.Therefore to pay special attention to working concentration in actual use procedure, avoid having too much of a good thing.
Table 2 without turbot seedling under aeration condition in the mortality ratio of 8h
Claims (3)
1. a photosynthetic bacterium SC01, is characterized in that: photosynthetic bacterium is the red bacterium of class ball (Rhodobacter sphaeroides), preserves center preservation on September 25th, 2013 in Chinese microorganism, and deposit number is CGMCC No.8248.
2. an application of photosynthetic bacterium SC01 claimed in claim 1, is characterized in that: described photosynthetic bacterium can be applicable to process the pollutent producing in sea farming process.
3. by the application of photosynthetic bacterium SC01 claimed in claim 2, it is characterized in that: described photosynthetic bacterium put in breeding seawater and then improve water inlet and the discharge water of sea farming water, improving the survival rate of sea farming biology simultaneously.
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Cited By (3)
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| CN104388339A (en) * | 2014-10-27 | 2015-03-04 | 徐州工程学院 | Microecological preparation suitable for marine culture and application thereof |
| CN106676050A (en) * | 2017-03-17 | 2017-05-17 | 烟台寒武纪生物科技有限公司 | Method and application for preparing photosynthetic bacterium through photonic fermentation |
| CN115386521A (en) * | 2022-09-20 | 2022-11-25 | 广东海洋大学 | Photosynthetic bacterium and application thereof in prawn culture |
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| CN108384732B (en) * | 2018-02-09 | 2021-03-02 | 上海海洋大学 | Rhodobacter sphaeroides for reducing trichlorphon toxicity and application thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106676050A (en) * | 2017-03-17 | 2017-05-17 | 烟台寒武纪生物科技有限公司 | Method and application for preparing photosynthetic bacterium through photonic fermentation |
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| CN115386521B (en) * | 2022-09-20 | 2024-01-05 | 广东海洋大学 | Photosynthetic bacteria and application thereof in prawn culture |
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