CN104164392A - Bacillus licheniformis and application thereof in aquaculture - Google Patents
Bacillus licheniformis and application thereof in aquaculture Download PDFInfo
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- CN104164392A CN104164392A CN201410335919.6A CN201410335919A CN104164392A CN 104164392 A CN104164392 A CN 104164392A CN 201410335919 A CN201410335919 A CN 201410335919A CN 104164392 A CN104164392 A CN 104164392A
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- bacillus licheniformis
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Abstract
The invention relates to the technical field of functional microbial screening, and particularly provides a strain of bacillus licheniformis and application thereof in aquaculture. The strain is preserved in the China Center for Type Culture Collection (CCTCC) in June 24, 2014, the serial number is CCTCC NO:M2014277; the strain has the autotrophic and nitrosation performance, is simple in nutritional requirements, and can significantly reduce the content of ammonia nitrogen in water, and the removal rate of the ammonia nitrogen in the water 48h after the treatment is as high as 93.39%; in addition, the bacillus licheniformis can be used as feed additives in aquatic feed, and can effectively improve the feed utilization rate of penaeus vannamei boone, and the feed conversion ratio of the penaeus vannamei boone of an experimental group is decreased by 14.3% compared that of a control group, so that the bacillus licheniformis can be developed into aquaculture probiotics and sewage treatment agents, not only helps to improve the economic benefits of farmers, but also can significantly reduce the pollution to the water by high density cultivation, and is extensive in application prospect.
Description
Technical field
The present invention relates to functional microorganism triage techniques field, be specifically related to a bacillus licheniformis and the application in aquaculture thereof.
Background technology
In recent years, China's culture fishery fast development, intensive, high-density breeding scale expanding day, concentrated bait throwing in causes and in aquaculture water, accumulates a large amount of residual baits, ight soil and animals and plants corpse, cause water body nitrogen element content severe overweight, eutrophication aggravation, is destroyed breeding ecological environment, fish and shrimp diseases takes place frequently, and finally causes huge financial loss.Nitrosification is that ammonia-state nitrogen is converted into nitrite nitrogen, is the important ring in occurring in nature nitrogen cycle.Along with the highdensity development of aquaculture, daily ration, feeding quantity continues to rise, and occurring in nature denitrifying load continues to increase, and causes the accumulation of nitrogen.Research shows, the pathogenic main contributor of aquatic animal is the severe overweight of ammonia nitrogen content in aquaculture water, and especially, when pH is high, the toxicity of ammonia-state nitrogen is stronger.So the ammonia-state nitrogen in control aquaculture water becomes the emphasis of aquaculture water quality processing, is also the difficult point of current water conditioning.
Compared with physics denitrogenation, biological denitrificaion has its unique advantage: non-secondary pollution, denitrogenation is thorough, economic security etc.Because bio-denitrification technology has that cost is low, easy to operate, qualified discharge reliability is strong and the advantage such as non-secondary pollution, become gradually the major way of wastewater treatment.By selective medium screening and separating autotrophy nitrite bacteria, and various growth characteristics and the nitrogen removal characteristics of autotrophy nitrite bacteria are furtherd investigate, the denitrogenation processing of breeding wastewater, sanitary wastewater and industrial sewage is had to important theory value and practical significance.
Summary of the invention
The object of this invention is to provide the novel Bacillus licheniformis of a strain and the application in aquaculture thereof.Described Bacillus licheniformis is screened the water sample in salmon circulating water cultivation waste-water treatment tower, has autotrophy and nitrosification performance, can significantly reduce the ammonia-state nitrogen in water body, can also improve the utilization ratio of aquatic animal to feed, and application prospect is extensive.
One aspect of the present invention provides a bacillus licheniformis (Bacillus licheniformis) ZY-4, the Chinese Typical Representative culture collection center that has been preserved in Wuhan, China Wuhan University on June 24th, 2014, its deposit number is CCTCC NO:M2014277.
The application of above-mentioned Bacillus licheniformis ZY-4 in water body purification, for the treatment of breeding wastewater or sanitary sewage.
Above-mentioned Bacillus licheniformis ZY-4 can bacterium powder form make microbe additive.
The application of above-mentioned Bacillus licheniformis ZY-4 in aquatic feeds.
Beneficial effect of the present invention:
Bacillus licheniformis ZY-4 provided by the invention, has autotrophy and nitrosification performance, and nutritional needs is simple, can significantly reduce the content of ammonia-state nitrogen in water body, processes after 48h in water body the clearance of ammonia-state nitrogen up to 93.39%; In addition, described Bacillus licheniformis ZY-4 can be used as fodder additives and is applied in aquatic feeds, can effectively improve the utilization ratio of Penaeus vannamei to feed, and the feedstuff-meat ratio of experimental group prawn has reduced by 14.3% than control group.Described Bacillus licheniformis ZY-4, can be developed as aquatic products probiotics and sewage-treating agent, is not only conducive to improve raiser's economic benefit, can also significantly alleviate the pollution of high-density breeding to water body, and application prospect is extensive.
Embodiment
Separation, screening and the qualification of embodiment 1 bacterial strain
1, sample
Take from the water sample in east, Yantai ocean group salmon circulating water cultivation waste-water treatment tower;
2, autotrophy nitrosification substratum:
| Material title | Consumption |
| NaCl | 2.0g |
| MgSO 4·7H 2O | 0.5g |
| CaCO 3 | 5.0g |
| FeSO 4·7H 2O | 0.4g |
| K 2HPO 4 | 1.0g |
| Agar | 20.0g |
| H 2O | Be settled to 1000.0mL |
3, screening method
Adopt gradient dilution method that water sample is diluted to proper concn; The water sample of getting after 0.1mL dilution is evenly coated autotrophy nitrosification media surface, inserts constant incubator, cultivates after 2～3d at 30 DEG C, with the single bacterium colony on transfering loop picking substratum, carries out separation and purification, is primary dcreening operation bacterial strain.
This test altogether picking 10 strain list bacterium colonies, and called after ZY-1, ZY-2, ZY-3 according to this ..., ZY-10.Inoculate single bacterium colony to nutrient broth medium, under 30 DEG C, 160r/min condition, carry out after liquid culture 24h, aseptic centrifugal, bacterium mud is resuspended with physiological saline, make bacterium liquid, wherein bacteria concentration is 10
8～10
9cFU/mL.
In the ratio of 1% (v/v), above-mentioned bacterium liquid is inoculated into respectively to sterilized breeding wastewater (NH
4 +-N concentration is 2.12mg/L) in, under 30 DEG C, 160r/min condition, carry out shaking flask processing, the concentration of ammonia-state nitrogen in sampling and measuring experimental system after 48h, calculates the clearance of ammonia-state nitrogen.Concrete outcome is as shown in table 1:
The concentration of ammonia-state nitrogen adopts Na Shi method to measure;
Clearance=(starting point concentration-termination concentration)/starting point concentration × 100%.
Table 1 is processed after 48h NH in water body
4 +the removal situation of-N
| Strain number | Initial NH 4 +-N(mg/L) | Stop NH 4 +-N(mg/L) | Clearance |
| ZY-1 | 2.12 | 1.04 | 50.94％ |
| ZY-2 | 2.12 | 1.20 | 43.40％ |
| ZY-3 | 2.12 | 0.76 | 64.15％ |
| ZY-4 | 2.12 | 0.14 | 93.39％ |
| ZY-5 | 2.12 | 1.24 | 41.51％ |
| ZY-6 | 2.12 | 1.22 | 42.45％ |
| ZY-7 | 2.12 | 1.36 | 35.85％ |
| ZY-8 | 2.12 | 0.90 | 57.55％ |
| ZY-9 | 2.12 | 0.71 | 66.51％ |
| ZY-10 | 2.12 | 0.45 | 78.77％ |
Can find out from the data of table 1, the bacterial strain ZY-4 that the present invention screens is the strongest to the degradation capability of ammonia-state nitrogen, NH after processing 48h
4 +the clearance of-N is up to 93.39%.
4, identification of strains
1) colony morphology characteristic: above-mentioned bacterial strains ZY-4 is cultivated 18 hours on nutrient agar, be rendered as white, opaque, surface wettability, have the bacterium colony of gauffer, diameter 1mm, microscopy shows that its cell is rod-short.
2) utilize test kit to extract the genomic dna of above-mentioned bacterial strains ZY-4, utilize round pcr amplification 16SrRNA sequence, after order-checking, obtain the fragment that length is 1438bp.By BLAST compare of analysis, it, is identified and confirms that bacterial strain ZY-4 is Bacillus licheniformis (Bacillus licheniformis) up to 99% with the 16SrRNA sequence similarity degree of many bacillus licheniformis of announcing, consistent with biochemical identification result.
The 16S rRNA sequence of bacterial strain ZY-4 is as follows:
CGTGCTATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTCAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGCTTGATTGAACCGCATGGTTCAATTATAAAAGGTGGCTTTTACCTACCACTTACAAATGGACCCGCGGCGCATTACCTATTTGGGGAGGTAACGGCTCACCAAGGCAACAATGCGTACCCAACCTGAAAGGGTGATCGGCCACACGGGGACTGAAACACGGCCCAAACTCCTACGGAAGGCACCATTAGGGAATCTTCCGCATGGGACAAAAGTCTGACGGACCACCGCCCCGGGAGTGATGAAGGTTTTCGAATCGTAAAACTCTGTTGTTAGGGAAAAACAATTACCGTTCAAATAGGGCGGTACCTTGACGGTACCTAACCAAAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGCGCGCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTGAGGCGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGAGGGTTTCCGCCCTTTAGTGCTGCAGCAAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAACCCTAGAGATAGGGCTTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGGCAGAACAAAGGGCAGCGAAGCCGCGAGGCTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTGGAGCCAGCCGCCGAAGGTGAC
3) by above-mentioned bacterial strains ZY-4 called after Bacillus licheniformis (Bacillus licheniformis) ZY-4, the Chinese Typical Representative culture collection center that has been preserved in Wuhan, China Wuhan University on June 24th, 2014, its deposit number is CCTCC NO:M 2014277
Embodiment 2: Bacillus licheniformis ZY-4 processes aquaculture system experiment
Bacillus licheniformis ZY-4 is activated, and enlarged culturing, through liquid state fermentation, the dry approximately bacterium powder of 10,000,000,000/g of making of spraying.
Experiment is located at the hut shrimp aquaculture pond of Jiangsu Rudong to be carried out, and chooses three culturing pools that water-quality guideline is close parallel as three, with plastic cloth, the water body of culturing pool is separated respectively, and half water body is as experimental group, and second half water body as a control group.First NH in test experience group and control group water body respectively
4 +the starting point concentration of-N then adds the above-mentioned Bacillus licheniformis ZY-4 of 10ppm bacterium powder in experimental group water body, and control group does not add.The uninterrupted aeration of experimental session, and it is consistent respectively to organize aeration rate.Experimental temperature is 28-30 DEG C, measures respectively NH in each experimental group and control group water body after 48h
4 +the concentration of-N, averages the detected result of three parallel group, calculates clearance.Concrete outcome is as shown in table 2:
Table 2 is processed after 48h NH in culturing pool water body
4 +the removal situation of-N
| Group | Initial NH 4 +-N(mg/L) | Stop NH 4 +-N(mg/L) | Clearance |
| Control group | 2.17±0.03 | 2.14±0.02 | 1.38％ |
| Experimental group | 2.28±0.03 | 0.26±0.02 | 88.59％ |
From the result of table 2, Bacillus licheniformis ZY-4 bacterium powder provided by the invention is to the NH in the water body of hut shrimp aquaculture pond
4 +-N has very strong Degradation (p < 0.05), NH after processing 48h
4 +the clearance of-N is up to 88.59%, and the clearance of control group is only 1.38%.Thereby illustrate that the Bacillus licheniformis ZY-4 that the present invention screens can be widely used in aquaculture, for solving the too high problem of aquaculture water ammonia nitrogen content.
Ammonia-state nitrogen in energy degrading cultivation water, Bacillus licheniformis ZY-4 of the present invention is also very remarkable to the treatment effect of ammonia-state nitrogen in sanitary sewage, NH after processing 48h
4 +the clearance of-N is higher than 85%.
Embodiment 3 impacts of Bacillus licheniformis ZY-4 on aquatic animal feedstuff-meat ratio
Experiment is located at the hut shrimp aquaculture pond of Jiangsu Rudong, choose that water-quality guideline is normal, prawn healthy aquaculture pond, in cultivating pool, build the enclosure of 6 4m × 4m × 2m with plastic cloth, in enclosure, inject the water in this pond of equivalent, in enclosure, with the aeration stone of equivalent, aeration rate is regulated to uniformity.Randomly draw 3 enclosures as experimental group, other 3 as a control group.Throw in respectively healthy Penaeus vannamei 1000 tails for every group, before input, select at random 500 tail prawns and weigh from every group, three experimental group and control group are respectively averaged.
The commercially available prawn mixed feed of Feed selection of Penaeus vannamei, wherein adds Bacillus licheniformis ZY-4 bacterium powder of the present invention by 1 ‰ mass ratio in the feed of experimental group, and does not add in the feed of control group.The feed of throwing something and feeding every day 2 times, throw something and feed weight and the time homogeneous of feed of experimental group and control group causes, the feed weight that record is thrown something and fed at every turn.The uninterrupted aeration of experimental session, other management are all consistent.Feed after 20 days, select at random 500 tail prawns and weigh from every group, three experimental group and control group are respectively averaged, and then calculate feedstuff-meat ratio, and concrete outcome is as shown in table 3:
The impact of table 3 Bacillus licheniformis ZY-4 on aquatic animal feedstuff-meat ratio
Can find out from above-mentioned experimental result, Bacillus licheniformis ZY-4 of the present invention is applied in aquatic feeds as fodder additives, can effectively improve the utilization ratio of Penaeus vannamei to feed, the feedstuff-meat ratio of experimental group prawn has reduced by 14.3% (p < 0.05) than control group, and effect is remarkable.
To sum up, Bacillus licheniformis ZY-4 provided by the invention can effectively reduce the content of ammonia-state nitrogen in aquaculture water, reduce cultivated animals disease odds, can also serve as fodder additives simultaneously, significantly improve the utilization ratio of cultivated animals to feed, reduce feedstuff-meat ratio, not only contribute to improve raiser's economic benefit, can also alleviate the pollution of high-density breeding to water body, have a extensive future.
Claims (5)
1. a bacillus licheniformis, its deposit number is CCTCC NO:M2014277.
2. the application of Bacillus licheniformis claimed in claim 1 in water body purification.
3. Bacillus licheniformis claimed in claim 1 is in the application of preparing in microbial preparation.
4. a microbial preparation, described microbial preparation includes Bacillus licheniformis claimed in claim 1.
5. the application of microbial preparation claimed in claim 4 in aquatic feeds.
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| CN106745821A (en) * | 2017-02-22 | 2017-05-31 | 大连理工大学 | Sea-farming special microorganism water purification agent and preparation method thereof and application method |
| CN110157636A (en) * | 2019-04-01 | 2019-08-23 | 成都市农林科学院 | A kind of bacillus licheniformis, screening technique, the feed using and containing the bacillus licheniformis |
| CN110791454A (en) * | 2019-11-26 | 2020-02-14 | 中国科学院烟台海岸带研究所 | Efficient ammonia nitrogen degrading strain and application thereof |
| CN111100824A (en) * | 2020-01-21 | 2020-05-05 | 暨南大学 | Bacillus and application thereof in denitrification and desulfurization in aquaculture water |
| CN111676155A (en) * | 2020-05-31 | 2020-09-18 | 青岛玛斯特生物技术有限公司 | Bacillus licheniformis and application thereof in aquaculture |
| CN113088466A (en) * | 2021-04-06 | 2021-07-09 | 江苏祥豪实业股份有限公司 | Bacillus licheniformis with antagonistic odor-producing bacteria and deodorization effects and application thereof |
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105441358A (en) * | 2015-12-19 | 2016-03-30 | 佛山市艳晖生物科技有限公司 | On-site fermented bacillus licheniformis preparation for aquaculture farm and applications thereof |
| CN105441358B (en) * | 2015-12-19 | 2018-12-21 | 佛山市艳晖生物科技有限公司 | Bacillus licheniformis preparation and its application for the fermentation of aquatic farm scene |
| CN106745821A (en) * | 2017-02-22 | 2017-05-31 | 大连理工大学 | Sea-farming special microorganism water purification agent and preparation method thereof and application method |
| CN110157636A (en) * | 2019-04-01 | 2019-08-23 | 成都市农林科学院 | A kind of bacillus licheniformis, screening technique, the feed using and containing the bacillus licheniformis |
| CN110791454A (en) * | 2019-11-26 | 2020-02-14 | 中国科学院烟台海岸带研究所 | Efficient ammonia nitrogen degrading strain and application thereof |
| CN111100824A (en) * | 2020-01-21 | 2020-05-05 | 暨南大学 | Bacillus and application thereof in denitrification and desulfurization in aquaculture water |
| CN111100824B (en) * | 2020-01-21 | 2021-11-05 | 暨南大学 | Bacillus and application thereof in denitrification and desulfurization in aquaculture water |
| CN111676155A (en) * | 2020-05-31 | 2020-09-18 | 青岛玛斯特生物技术有限公司 | Bacillus licheniformis and application thereof in aquaculture |
| CN111676155B (en) * | 2020-05-31 | 2022-04-08 | 青岛玛斯特生物技术有限公司 | Bacillus licheniformis and application thereof in aquaculture |
| CN113088466A (en) * | 2021-04-06 | 2021-07-09 | 江苏祥豪实业股份有限公司 | Bacillus licheniformis with antagonistic odor-producing bacteria and deodorization effects and application thereof |
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