CN108517307A - A kind of method that modified clay couples use inhibition algal grown with algal control bacterium - Google Patents
A kind of method that modified clay couples use inhibition algal grown with algal control bacterium Download PDFInfo
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Abstract
The present invention provides a kind of method that modified clay couples use inhibition algal grown with algal control bacterium.The modified clay is that cetyl trimethylammonium bromide is modified sodium montmorillonite.The algal control bacterium can be bacillus cereus Bacillus cereus G4, abbreviation G4 bacterial strains.It is preserved in Guangdong Province's Culture Collection on December 13rd, 2017, deposit number is respectively GDMCC NO.60299.Experimental data shows：Modified clay and being used in combination for algal control bacterium (G4) have obvious inhibiting effect to frustule, continue the algal control time up to 168 hours, have no the rebound of frustule quantity.Modification clay source used in the present invention is wide, it is simple to prepare, of low cost and easily operated；The combination of modified clay and algal control bacterium enables red tide more thoroughly to inhibit；Modified clay and algal control bacterium combine the method for inhibiting red tide, not will produce secondary pollution, are a kind of environmentally friendly methods.
Description
Technical field
The invention belongs to marine environmental protection fields, and in particular to a kind of modified clay is coupled with algal control bacterium using inhibiting algae
The method of class growth.
Background technology
Red tide is one of serious global Marine Natural Disaster of one kind, has caused highest attention all over the world.Closely
China coastal seas maritime environment pollution aggravates over year, and local Environment Management of Eutrophication outstanding problem, breakout of red tide is increasingly frequent, gives nearshore waters
Serious ecology, resource and environmental problem are brought, and causes huge economic loss, is strand ecological safety and coastal economy
The significant threat of sustainable development.Therefore, the red-tide control method for studying novel high-efficiency environment friendly is in current red tide prevention and control field
One of significant problem urgently to be resolved hurrily.
Although having developed the control technology of some red tides in previous research, such as physical method, chemical method and life
Object administering method, but all there is certain defect in these methods, and regulation effect is not satisfactory.Such as the yellow mud in physical method
Flocculence needs larger dosage just can play the effect of inhibition, this increases the difficulty and cost of operation to a certain extent
Input；Some chemical methodes such as algicide also brings direct or indirect danger while killing algae to other biologies
Evil, cannot embody environment friendly feature；Macro-organism (fish) of some biological methods as launched the algae that ingests, there is also
Break food web frame, destroys the ecological risk that population is stablized.
In the method for existing control red tide, including Physical, chemical method and biological method, these methods are often single to be made
With having exposed some shortcomings in practical application.It is in particular in：Physical administer red tide can only achieve in the early stage have it is stronger
Inhibiting effect does not remove algae thoroughly, and the outburst of algae has the possibility of rebound；And utilization chemical method such as copper agent,
Although the chemistry algicide such as herbicide can directly kill algae, the specificity of these chemical substances is poor, and is easy enrichment
Secondary pollution is caused in food chain；The use of some biological methods such as takes the photograph the intervention of algae biology, and can cause disturbing for population
It moves, bring ecological risk；And biological method takes effect relatively slowly, is not easy to form the effect got instant result.
In recent years, the discovery of algae-lysing bacterium (algicidal bacteria) brings new dawn to the improvement of red tide.
Its important component as aquatic ecosystem biotic population structure and function, it is non-to maintaining algae bio amount balance to have
Normal important role.Studies have shown that withering away for red tide may there are related with algae-lysing bacterium.Algae-lysing bacterium is as red-tide control
Effective microbe, caused more and more to pay close attention to.But the population advantage of algae in red tide phenomenon is faced, algae-lysing bacterium
Dosage demand is very big, it is difficult to meet the needs of actual environment improvement.
Therefore how by existing technology progress efficient coupling, the intrinsic advantage of various technologies is given full play to, exploitation is a kind of
Integrated, efficient, environmentally friendly algal control method is always the effort target of researcher.
Invention content
The object of the present invention is to provide a kind of methods inhibiting algal grown.
It is provided by the present invention inhibit algal grown method, be：Algae sedimentation agent and algal control bacterium are added to pending
In algae solution, coupling processing inhibits algal grown.
In the above method, the algae sedimentation agent can be modified clay.
The modified clay can be prepared by the method included the following steps：It is molten that dilute hydrochloric acid is added into modifying agent
Liquid adds water, obtains modifying agent hydrochloric acid solution after to be modified dose of dissolving, is added to the modifying agent hydrochloric acid solution under stirring viscous
In soil, modified clay is obtained.
The modifying agent can be the high alkyl compound of high efficiency and hypotoxicity.
The modifying agent concretely cetyl trimethylammonium bromide and/or tributyl cetyl phosphonium bromide, preferably
For cetyl trimethylammonium bromide.
The dilute hydrochloric acid solution can be the dilute hydrochloric acid of mass concentration 1%.
The proportioning of the modifying agent and dilute hydrochloric acid solution can be 10mg：1mL.
The water being added is seawater, the seawater preferably filtered.
The proportioning of the modifying agent and water can be 10mg：10mL.
The clay can be sodium montmorillonite.
The mass ratio of the clay and the modifying agent in modifying agent hydrochloric acid solution can be 1g：100mg.
Concretely cetyl trimethylammonium bromide is modified sodium montmorillonite to the modified clay.
In the above method, the algal control bacterium can be bacillus cereus Bacillus cereus G4, abbreviation G4 bacterial strains.
The bacillus cereus Bacillus cereus G4 are detached from the fungal component of algae border and are obtained, and in 2017
On December 13, in is preserved in Guangdong Province's Culture Collection, and (abbreviation GDMCC, address are：Xianlie Middle Road, Guangzhou City 100
Number, Guangdong Microbes Inst, 5 building, No. 59 building, postcode：510070), deposit number is GDMCC NO.60299.
In the above method, concentration of the modified clay in pending algae solution can be 5-15mg/mL, concretely 10-15
mg/ml、10mg/ml。
Final concentration of the algal control bacterium in pending algae solution can be 105-107Cells/mL, concretely 106 cells/
mL。
Algae in the pending algae solution concretely breakout of red tide algae --- this Shi Shi algaes (Scrippsiella of taper
Trochoidea, cell pyriform is 16~36 μm long, 20~23 μm wide).
Experimental data shows：Modified clay and being used in combination for algal control bacterium (G4) have obvious inhibiting effect to frustule, hold
The continuous algal control time up to 168 hours, has no the rebound of frustule quantity.
Modification clay source used in the present invention is wide, it is simple to prepare, of low cost and easily operated；Modified clay and
The combination of algal control bacterium enables red tide more thoroughly to inhibit；Modified clay and algal control bacterium combine the method for inhibiting red tide,
Secondary pollution is not will produce, is a kind of environmentally friendly method.
Biomaterial preservation explanation
The Classification And Nomenclature of biomaterial：Bacillus (Bacillus cereus G4)
The strain number of biomaterial：G4
Depositary institution's title of biomaterial：Guangdong Province's Culture Collection
The depositary institution of biomaterial is referred to as：GDMCC
The depositary institution address of biomaterial：Xianlie Middle Road, Guangzhou City 100, Guangdong Microbes Inst, No. 59 building 5
Building
The preservation date of biomaterial：On December 13rd, 2017
The collection of biomaterial is registered on the books number：GDMCC NO.60299
Description of the drawings
Fig. 1 is the sedimentation shape that cetyl ammonium bromide is modified 120h frustules after clay concentration is handled at different dosages
State.
Fig. 2 is the sedimentation of 120h frustules after tributyl cetyl bromination P Modification clay is handled at different dosages
State.
Fig. 3 is influence of the modified clay to frustule photosynthetic efficiency.
Fig. 4 is influence of the modified clay to frustule chlorophyll.
Fig. 5 is influence of the coupled method to frustule photosynthetic efficiency Fm/Fv.
Fig. 6 is influence of the coupled method to Chlorophylls.
Fig. 7 is the morphological feature comparison diagram of blank group and coupling systems frustule.
Specific implementation mode
Below by specific embodiment, the present invention will be described, but the present invention is not limited thereto.
Experimental method used in following embodiments is conventional method unless otherwise specified；Institute in following embodiments
Reagent, biomaterial etc., are commercially available unless otherwise specified.
Experiment algae used is Scrippsiella trochoidea (Scrippsiella trochoidea) (by this experiment from Shenzhen Yantian
It detaches and obtains in the marine site of port)
The nutritive salt formula of algae culturing liquid is (containing inorganic salt quality in every liter of seawater)：NaNO337.5mg NaH2PO4
2.5mg, Fe-EDTA 2.5mg (FeCl31.6g+EDTA 0.9g), Tyiamine Hd element 5 μ g, Biotin VH, 0.025 μ g,
VB12 0.025μg,CuSO4.5H2O 0.0098μg,ZnSO4.7H2O 0.022μg,CaCl2.6H2O 0.01μg,
MgCl2.4H2O 0.180μg,Na2MoO4.2H2O,0.0063μg.21 DEG C ± 1 DEG C of cultivation temperature, intensity of illumination 3000Lx, brightness
Compare 12h:12h.
The chlorophyll and photosynthetic efficiency of frustule are carried out using phytoplankton classification luminoscope (PHYTO-PAM).Specific behaviour
Work is as follows, takes 2mL algae solutions to be packed into measuring cup and is placed in magazine, 5min dark adaptations are carried out to frond, opens Phyto-PAM modulation
Pulse fluoro instrument wavelength is that 520nm intensity is 0.1 μm of ol/ (m2S) brown detection light.Measurement process is by Phytowin softwares
Control opens and measures light (ML), opens saturation pulse key after optical signal stabilization, writes down Fv/Fm values, as photosynthetic efficiency
Yield values.(ChII a) measurement of chlorophyll levels is also carried out using the instrument.
The selection of modifying agent and clay.Modifying agent uses cetyl trimethylammonium bromide and tributyl cetyl bromination
Two kinds of phosphorus.Clay has selected sodium montmorillonite.Montmorillonite is a kind of three-decker clay mineral, due to its lattice defect, table
Reason on the hydroxyl structure of face, this particle surface is negatively charged in water, but can prepare and have to frustule with sodium ion chelating
The cationic clay of stronger adsorption capacity.
The preparation method of modified clay：The above two modifying agent for taking 100mg, is added the dilute hydrochloric acid solution of 10mL1%, stirs
Mixing or shaking makes modifying agent dissolve, rear that filtration seawater 100mL is added, and obtains the modifying agent hydrochloric acid solution of 1mg/mL, then weigh 1g
Sodium montmorillonite is added the modifying agent hydrochloric acid solution of 100mL, that is, configures modified clay while stirring.
Strain isolation comes from algae border symbiotic microorganism with screening sample, by Shenzhen Graduate School of Tsinghua University culture, bacterial strain
Cultural method uses LB culture mediums.Algae border microorganism gushes the red tide sample of field outburst from Shen Zhendong.
LB solid mediums：Tryptone 10g, yeast extract 5g, sodium chloride 10g, deionized water 1000ml, agar powder
15g, pH7.0.
LB liquid medium：Tryptone 10g, yeast extract 5g, sodium chloride 10g, deionized water 1000ml,
pH7.0。
Solid LB condition of culture：After coating, by tablet as in 30 DEG C of incubators, cultivate 24-72 hours.
Liquid LB condition of culture：30 DEG C, for 24 hours in constant-temperature table, 180 revs/min.
The selection of example 1, tested algae class
This Shi Shi algae of the breakout of red tide algae that we have selected Guangdong marine site common in the present invention --- taper
(Scrippsiella trochoidea, cell pyriform is 16~36 μm long, 20~23 μm wide), the Trentepohlia is in dinoflagellate class, extensively
It is distributed in offshore and river mouth, burst can cause water body ischaemia when being proliferated, danger is brought to marine organisms and aquaculture
Evil.2010 so far continuous 7 years of Guangdong seacoast the taper of different scales this formula algae red tide occurs, it has also become the high frequency that red tide occurs
Species.By taking the 12 days~August of August in 2013 30 days as an example, Shenzhen dam light and east gushes-Yantian Harbor Free Trade one with occur large-scale taper this
Family name's algae red tide, 19 days duration, 15 square kilometres of maximum area.Therefore, the tested species selected in this invention are taper
Si Shi algaes.
The preparation of example 2, modified clay
Several clay types of document report are compared, we have selected sodium montmorillonite.Montmorillonite is a kind of
Three-decker clay mineral, due to its lattice defect, the reason in surface hydroxyl structure, this particle surface is in negative electricity in water
Property, but can prepare the cationic clay for having stronger adsorption capacity to red tide cell with sodium ion chelating.
Modifying agent has selected the high alkyl compound of high efficiency and hypotoxicity, after the types of different modifying agent is to clay alteration
Effect has a significant impact, we select two kinds of lower modifying agent of toxicity：Cetyl trimethylammonium bromide and tributyl 16
Two kinds of modifying agent of alkyl bromination phosphorus.
The preparation of modified clay：The modifying agent for taking 100mg, is added the dilute hydrochloric acid solution of 10mL1%, and stirring or concussion make to change
Property agent dissolving, the rear seawater 100mL that filtration is added obtains the modifying agent hydrochloric acid solution of 1mg/mL, then weigh 1g sodium cover it is de-
Soil is added the modifying agent hydrochloric acid solution of 100mL, that is, configures modified clay while stirring.Compared in the past using ultra-pure water as solvent
Way, We conducted some improvement, are configured by solvent of seawater.Purpose is to expand the use scope of modified clay；
The modification clay followed by prepared can preferably match briny environment and Phytoplankton ＆ Suspension.From seawater from the point of view of the result of preliminary experiment to changing
The character of property clay has no significant effect.
Example 3 is modified settlement action of the clay to frustule
By Fresh and the modification clay of concentration in gradient, it is added in the frustule culture bottle of 50ml, two kinds of modifications
The concentration of clay is respectively：Cetyl ammonium bromide be modified clay concentration be 15mg/ml (height), 10mg/ml (in), 5mg/ml
(low)；Tributyl cetyl bromination P Modification clay concentration is：30mg/ml (height), 20mg/ml (in), 10mg/ml (low),
Concussion shakes up.To be added the algae solution of modified clay in 21 DEG C ± 1 DEG C, intensity of illumination 3000Lx, Light To Dark Ratio 12h:Under 12h environment
Culture observation, per observation algae solution clarity for 24 hours and sedimentation degree.
Fig. 1 is the sedimentation shape that cetyl ammonium bromide is modified 120h frustules after clay concentration is handled at different dosages
State.
Fig. 2 is the sedimentation of 120h frustules after tributyl cetyl bromination P Modification clay is handled at different dosages
State.
By Fig. 1,2 it is found that modified clay has apparent flocculation to frustule, with its flocculating effect of the raising of concentration
Significantly.
From figure 1 it appears that the 1. group cetyl ammonium bromide be modified clay at 15mg/ml (high concentration), algae is thin
Born of the same parents sink to Tissue Culture Flask bottom substantially, and algae cultivates upper liquid clarification；At 10mg/ml (middle concentration), frustule part is sunk to the bottom, algae
Culture solution is in light yellow and relatively clarification；At 5mg/ml (low concentration), frustule small part is sunk to the bottom, and algae culturing liquid is in yellow；And
Blank group is obviously than experimental group color depth.The color contrast of above-mentioned various concentration algae culturing liquid can be seen that the addition of modified clay
There is apparent settlement action to frustule, and dose dependent is presented.
From Fig. 2 kinds as can be seen that the 2. group tributyl cetyl bromination P Modification clay in 30mg/ml (high concentration)
When, frustule part is deposited to Tissue Culture Flask bottom, and algae culturing liquid is in yellow and relatively clarifies；At 20mg/ml (middle concentration),
Frustule is further sunk to the bottom, and algae culturing liquid is in yellow；At 10mg/ml (low concentration), frustule is sunk to the bottom on a small quantity, and algae culturing liquid is in
It yellow and less clarifies, above three groups of color distortions are little, and compared with 1. group, less apparent to the effect of settling of algae.
The influence of example 4, modified clay to frustule photosynthetic efficiency and chlorophyll
Go out more efficient modification clay-cetyl ammonium bromide by the experimental selection of example 3 and is modified clay.In concentration
Middle concentration 10mg/ml is selected, because the modification clay of high concentration and the modification clay of middle concentration are to the inhibition of frustule
It is close, in order to save usage amount and cost, therefore select the dose concentration of 10mg/ml.
Take laboratory passage culture algae (initial density be 5.4 × 105A/mL), it is sub-packed in the cell of 9 100mL
(per bottled liquid 50mL) in culture bottle.Experiment sets three test groups (every group 3 parallel).Three test groups are respectively blank group
(without modified clay and algal control bacterium), experimental group 1 (the cetyl bromination P Modification of tributyl containing 10mg/ml clay), experiment
Group 2 (cetyl ammonium bromide containing 10mg/ml is modified clay).Frustule concentration and chlorophyll in liquid is continuously monitored after packing to contain
Amount.
It adds in two kinds of modified clays to the algae solution of culture, to algal inhibition, the results are shown in Figure 3.
Early stage cetyl ammonium bromide has obvious inhibiting effect, photosynthetic efficiency to frustule as seen in Figure 3
Fv/Fm values fall below 0.07 after 24 hours, but have apparent rebound with the extension frustule Fv/Fm values of time, until 120
After hour (Fv/Fm values are respectively 0.52 and 0.51) substantially similar to control group.And tributyl cetyl phosphonium bromide is thin to algae
The not apparent inhibiting effect of the photosynthetic efficiency of born of the same parents, Fv/Fm values remain basically stable with control group.
It adds in two kinds of modified clays to the algae solution of culture, the influence to frustule chlorophyll is as shown in Figure 4.
Experimental group (- 168h for 24 hours, interval is for 24 hours), content point of chlorophyll in entire experimental period as can be seen from Figure 4
It is not：208.31 μ g/L, 225.45 μ g/L, 250.37 μ g/L, 286.82 μ g/L, 253.55 μ g/L, 234.36 μ g/L and
227.87μg/L.After adding tributyl cetyl bromination P Modification clay, chlorophyll concentration is substantially reduced after 24h,
Its value is 107.23 μ g/L；And after 48h, the reducing effect of chlorophyll has a rebound, and value is respectively： 211.94μg/L、
234.93 μ g/L, 222.87 μ g/L, 185.71 μ g/L, 180.33 μ g/L and 176.41 μ g/L.Cetyl bromination in contrast
Ammonium is modified clay has apparent inhibiting effect to frustule chlorophyll, reduces 20 times in entire experimental period compared with the control
More than, the content of chlorophyll is respectively：6.47μg/L、9.71μg/L、10.69μg/L、 14.39μg/L、12.46μg/L、
11.84 μ g/L and 11.16 μ g/L.
Complex chart 3 and Fig. 4's as a result, it may be seen that cetyl ammonium bromide be modified clay effect it is more preferable, thus
In subsequent coupling experiment, we have selected this kind of clay to carry out subsequent experiment.
The screening of example 5, algal control bacterial strain
I) it is separately cultured with LB culture mediums from the microorganism of algae border and obtains monoclonal bacterial strain, co-cultured later with algae, detection
The quantity and chlorophyll content of frustule, select the bacterial strain of rejection ability.
Ii) 16s r-RNA sequence homology analysis
The isolated bacterial strain G4 of conventional method culture above-mentioned steps one extracts the total DNA of bacterial strain as gene magnification mould
Plate is expanded using universal primer, and wherein forward primer is：27f:
5'-AGAGTTTGATCMTGGCTCAG-3'；Reverse primer is：1492R:
5'-TACGGYTACCTTTGTTACGACTT-3' is carried out in 30 μ L reaction systems, and reaction system is as follows：
Reaction condition：95 DEG C of pre-degeneration 5min；95 DEG C of denaturation 30sec, 55 DEG C of annealing 30sec, 72 DEG C of extension 1min, totally 35
A cycle；72 DEG C of extension 10min.PCR product 1% detected through gel electrophoresis, deposition condition：+ 1% Ago-Gel of 3 μ L samples,
Marker bands form：100bp；250bp；500bp；750bp；1000bp；2000bp； 3000bp；5000bp.750bp bands
A concentration of 20ng/ μ L, are shown as highlighting band, remaining band concentration is 10ng/ μ L.
Examining order is completed by Beijing six directions Hua Da Gene Tech. Company Limited.
Sequence analysis is by comparing National Center for Biotechnology Information ncbi database (http://
Www.ncbi.nlm.nih.gov it) completes.
In view of 16s rDNA sequence homology analysis as a result, the bacterial strain that step 1 separation screening obtains is accredited as following knot
Fruit：
Bacillus cereus Bacillus cereus G4；Abbreviation G4 bacterial strains (its sequence signature is shown in sequence 1 in sequence table)
Above-mentioned bacterial strains be preserved in on December 13rd, 2017 Guangdong Province's Culture Collection (abbreviation GDMCC,
Address is：Xianlie Middle Road, Guangzhou City 100, Guangdong Microbes Inst, 5 building, No. 59 building, postcode：510070), preservation is registered
Number be respectively GDMCC NO.60299.Specific preservation is described as follows：
I) preservation explanation
Strain name：Bacillus
Latin name：(Bacillus cereus G4)
Strain number：G4
Preservation mechanism：Guangdong Province's Culture Collection
Preservation mechanism is referred to as：GDMCC
Address：Xianlie Middle Road, Guangzhou City 100, Guangdong Microbes Inst, 5 building, No. 59 building
Preservation date：On December 13rd, 2017
Collection is registered on the books number：GDMCC NO.60299
Sequence signature is as follows：
>1-GGGTGCTATACATGCAGTCGAGCGAATGGATTAAGAGCTTGCTCTTATGAAGTTAGCGGCGGACGG
GT GAGTAACACGTGGGTAACCTGCCCATAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATAA
CATTTTGAACCGCATGGTTCGAAATTGAAAGGCGGCTTCGGCTGTCACTTATGGATGGACCCGCGTCGCA
TTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCAACGATGCGTAGCCGACCTGAGAGGGTGATCGGCC
ACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGA
AAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGCTTTCGGGTCGTAAAACTCTGTTGTTAGGGAAGA
ACAAGTGCTAGTTGAATAAGCTGGCACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCA
GCAGCCGCGGTAATACGTAGGTGGCAAGCGTTATCCGGAATTATTGGGCGTAAAGCGCGCGCAGGTGGTT
TCTTAAGTCTGATGTGAAAGCCCACGGCTCAACCGTGGAGGGTCATTGGAAACTGGGAGACTTGAGTGCA
GAAGAGGAAAGTGGAATTCCATGTGTAGCGGTGAAATGCGTAGAGATATGGAGGAACACCAGTGGCGAA
GGCGACTTTCTGGTCTGTAACTGACACTGAGGCGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTG
GTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGAGGGTTTCCGCCCTTTAGTGCTGAAGTTAACGC
ATTAAGCACTCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTCAAAGGAATTGACGGGGGCCCGCACA
AGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAA
CCCTAGAGATAGGGCTTCTCCTTCGGGAGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCG
TGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCATCATTTAGTTGGGCACTC
TAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCT
GGGCTACACACGTGCTACAATGGACGGTACAAAGAGCTGCAAGACCGCGAGGTGGAGCTAATCTCATAA
AACCGTTCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGCTGGAATCGCTAGTAATCGCGGAT
CAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACA
CCCGAAGTCGGTGGGGTAACCTTTTTGGAGCCAGCCGCCTAAGGT-1438
The coupling effect of algae restraint of example 6, modified clay and algal control bacterium
The rebound of modified clay soils is mainly avoided in the use of coupled method.The cetyl ammonium bromide newly matched is modified
Algae solution is added in clay (10mg/ml), adds appropriate bacterium solution and ensures final concentration 106The bacterial strain of cells/mL or so, addition is
G4 bacteriums；Title is：Bacillus cereus Bacillus cereus G4 are detached by this laboratory early period from the fungal component of algae border
It obtains.
To algal inhibition, the results are shown in Figure 5 after coupled method use.Control group photosynthetic efficiency Fv/Fm values are entirely being tested
Amplitude of variation in period is between 0.38-0.56, and experimental group " cetyl ammonium bromide is modified clay+G4 " is to frustule
Photosynthetic efficiency has apparent inhibiting effect, and Fv/Fm values are in the range of 0.01-0.02, part-time point (such as 72h and 96h) light
It closes efficiency and is reduced to 0.
The results are shown in Figure 6 for influence of the coupled method to Chlorophylls.It is modified clay and bacterium as seen in Figure 6
Being used in combination for (G4) of kind has obvious inhibiting effect to frustule, continues the algal control time up to 168 hours, has no frustule quantity
Rebound.From the point of view of entire experimental period (- 168h for 24 hours, interval is for 24 hours), the content of control group chlorophyll is respectively：205.2μg/
L, 220.6 μ g/L, 240.11 μ g/L, 267.34 μ g/L, 289.24 μ g/L, 310.08 μ g/L and 340.63 μ g/L.When coupling makes
With rear, chlorophyll concentration is substantially reduced after 24h, ranging from 68.59-95.2 μ g/L of the value in two experimental groups it
Between；And after 48h, the reducing effect of chlorophyll is kept, until its chlorophyll test value is horizontal less than 20 μ g/L when experimental endpoints, and
Whole process has no rebound.
In addition, we by microexamination compare the morphological feature of experimental group and control group frustule, (Fig. 7, left figure are
Control group frustule, right figure are coupling systems frustule).
As can be seen from Figure 7, control group frustule form is complete, and cell outline is clear, uniform coloring；And coupling systems frustule goes out
Now cracking, ghostization are serious, and form fragmentation, most cells are killed by algae-lysing bacterium G4.
<110>Shenzhen Graduate School of Tsinghua University
<120>A kind of method that modified clay couples use inhibition algal grown with algal control bacterium
<130> GNCAQ181120
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1438
<212> DNA
<213>Artificial sequence
<220>
<223>
<400> 1
gggtgctata catgcagtcg agcgaatgga ttaagagctt gctcttatga agttagcggc 60
ggacgggtga gtaacacgtg ggtaacctgc ccataagact gggataactc cgggaaaccg 120
gggctaatac cggataacat tttgaaccgc atggttcgaa attgaaaggc ggcttcggct 180
gtcacttatg gatggacccg cgtcgcatta gctagttggt gaggtaacgg ctcaccaagg 240
caacgatgcg tagccgacct gagagggtga tcggccacac tgggactgag acacggccca 300
gactcctacg ggaggcagca gtagggaatc ttccgcaatg gacgaaagtc tgacggagca 360
acgccgcgtg agtgatgaag gctttcgggt cgtaaaactc tgttgttagg gaagaacaag 420
tgctagttga ataagctggc accttgacgg tacctaacca gaaagccacg gctaactacg 480
tgccagcagc cgcggtaata cgtaggtggc aagcgttatc cggaattatt gggcgtaaag 540
cgcgcgcagg tggtttctta agtctgatgt gaaagcccac ggctcaaccg tggagggtca 600
ttggaaactg ggagacttga gtgcagaaga ggaaagtgga attccatgtg tagcggtgaa 660
atgcgtagag atatggagga acaccagtgg cgaaggcgac tttctggtct gtaactgaca 720
ctgaggcgcg aaagcgtggg gagcaaacag gattagatac cctggtagtc cacgccgtaa 780
acgatgagtg ctaagtgtta gagggtttcc gccctttagt gctgaagtta acgcattaag 840
cactccgcct ggggagtacg gccgcaaggc tgaaactcaa aggaattgac gggggcccgc 900
acaagcggtg gagcatgtgg tttaattcga agcaacgcga agaaccttac caggtcttga 960
catcctctga caaccctaga gatagggctt ctccttcggg agcagagtga caggtggtgc 1020
atggttgtcg tcagctcgtg tcgtgagatg ttgggttaag tcccgcaacg agcgcaaccc 1080
ttgatcttag ttgccatcat ttagttgggc actctaaggt gactgccggt gacaaaccgg 1140
aggaaggtgg ggatgacgtc aaatcatcat gccccttatg acctgggcta cacacgtgct 1200
acaatggacg gtacaaagag ctgcaagacc gcgaggtgga gctaatctca taaaaccgtt 1260
ctcagttcgg attgtaggct gcaactcgcc tacatgaagc tggaatcgct agtaatcgcg 1320
gatcagcatg ccgcggtgaa tacgttcccg ggccttgtac acaccgcccg tcacaccacg 1380
agagtttgta acacccgaag tcggtggggt aacctttttg gagccagccg cctaaggt 1438
Claims (9)
1. bacillus cereus Bacillus cereus G4, in Guangdong Province, the deposit number of Culture Collection is
GDMCC NO.60299。
2. applications of the bacillus cereus Bacillus cereus G4 described in claim 1 in inhibiting algal grown.
3. a kind of product inhibiting algal grown, including algae sedimentation agent and algal control bacterium.
4. the product according to claim 3 for inhibiting algal grown, it is characterised in that：The algae sedimentation agent is modified viscous
Soil；And/or
The algal control bacterium is bacillus cereus Bacillus cereus G4 described in claim 1.
5. the product according to claim 4 for inhibiting algal grown, it is characterised in that：Under the modified clay is by including
The method for stating step is prepared：Dilute hydrochloric acid solution is added into modifying agent to add water after to be modified dose of dissolving, obtain modifying agent
The modifying agent hydrochloric acid solution is added in clay by hydrochloric acid solution under stirring, obtains modified clay.
6. the product according to claim 5 for inhibiting algal grown, it is characterised in that：The modifying agent is cetyl three
Methyl bromide ammonium and/or tributyl cetyl phosphonium bromide；
The dilute hydrochloric acid solution is the dilute hydrochloric acid of mass concentration 1%；
The proportioning of the modifying agent and dilute hydrochloric acid solution is 10mg：1mL；
The water being added is seawater；
The proportioning of the modifying agent and water can be 10mg：10mL；
The clay is sodium montmorillonite；
The mass ratio of the clay and the modifying agent in modifying agent hydrochloric acid solution is 1g：100mg.
7. a kind of method inhibiting algal grown, is：Algae sedimentation agent and algal control bacterium are added in pending algae solution, at coupling
It manages to inhibit algal grown.
8. according to the method described in claim 7, it is characterized in that：In the method, the algae sedimentation agent is modified clay；
A concentration of 5-15mg/mL of the modified clay in pending algae solution；
The algal control bacterium in pending algae solution final concentration of 105-107cells/mL。
9. method according to claim 7 or 8, it is characterised in that：Algae in the pending algae solution is breakout of red tide
This Shi Shi algae Scrippsiella trochoidea of algae --- taper.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111296490A (en) * | 2020-02-20 | 2020-06-19 | 福建师范大学 | Method for inhibiting chain-like gymnodinium by coupling coagulant and algae inhibiting bacteria fermentation liquor |
| CN113321318A (en) * | 2021-06-02 | 2021-08-31 | 中国科学院海洋研究所 | Method for improving efficiency of clay in treating harmful algal blooms based on microbial composite modification |
| CN114958706A (en) * | 2022-04-11 | 2022-08-30 | 中国科学院海洋研究所 | Method for improving capacity of bacillus for removing red tide algae cells |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105236539A (en) * | 2015-10-28 | 2016-01-13 | 中国科学院海洋研究所 | Modified clay capable of eliminating red tide with high efficiency |
| CN105779349A (en) * | 2016-04-06 | 2016-07-20 | 中国水产科学研究院南海水产研究所 | Bacillus cereus strain JZBC1 for dissolving pond dominant dinoflagellate-scrippsiella trochoidea and application thereof |
| CN106190898A (en) * | 2016-07-12 | 2016-12-07 | 中国水产科学研究院南海水产研究所 | A kind of industrialization liquid fermentation method of the bacillus cereus JZBC1 dissolving pond dinoflagellate |
| CN107032465A (en) * | 2017-05-05 | 2017-08-11 | 中国科学院海洋研究所 | A kind of efficient modification clay composite material for removing microalgae red tide and preparation method thereof |
| CN107986412A (en) * | 2017-12-28 | 2018-05-04 | 中国科学院海洋研究所 | A kind of modification clay systems administered for breeding water body harmful algal bloom |
-
2018
- 2018-05-22 CN CN201810493097.2A patent/CN108517307B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105236539A (en) * | 2015-10-28 | 2016-01-13 | 中国科学院海洋研究所 | Modified clay capable of eliminating red tide with high efficiency |
| CN105779349A (en) * | 2016-04-06 | 2016-07-20 | 中国水产科学研究院南海水产研究所 | Bacillus cereus strain JZBC1 for dissolving pond dominant dinoflagellate-scrippsiella trochoidea and application thereof |
| CN106190898A (en) * | 2016-07-12 | 2016-12-07 | 中国水产科学研究院南海水产研究所 | A kind of industrialization liquid fermentation method of the bacillus cereus JZBC1 dissolving pond dinoflagellate |
| CN107032465A (en) * | 2017-05-05 | 2017-08-11 | 中国科学院海洋研究所 | A kind of efficient modification clay composite material for removing microalgae red tide and preparation method thereof |
| CN107986412A (en) * | 2017-12-28 | 2018-05-04 | 中国科学院海洋研究所 | A kind of modification clay systems administered for breeding water body harmful algal bloom |
Non-Patent Citations (3)
| Title |
|---|
| JING LI ET AL.: ""An investigation of the space distribution of Ulva microscopic propagules and ship-based experiment of mitigation using modified clay"", 《MARINE POLLUTION BULLETIN》 * |
| 宁华等: ""溶藻细菌应用于生物杀藻剂的研究进展"", 《净水技术》 * |
| 闫新亚等: ""不同有机改性黏土对海洋卡盾藻去除作用的研究"", 《安全与环境学报》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111296490A (en) * | 2020-02-20 | 2020-06-19 | 福建师范大学 | Method for inhibiting chain-like gymnodinium by coupling coagulant and algae inhibiting bacteria fermentation liquor |
| CN113321318A (en) * | 2021-06-02 | 2021-08-31 | 中国科学院海洋研究所 | Method for improving efficiency of clay in treating harmful algal blooms based on microbial composite modification |
| CN113321318B (en) * | 2021-06-02 | 2022-11-08 | 中国科学院海洋研究所 | Method for improving efficiency of clay in treating harmful algal blooms based on microbial composite modification |
| CN114958706A (en) * | 2022-04-11 | 2022-08-30 | 中国科学院海洋研究所 | Method for improving capacity of bacillus for removing red tide algae cells |
| WO2023197743A1 (en) * | 2022-04-11 | 2023-10-19 | 中国科学院海洋研究所 | Method for improving ability of bacillus species to remove red tide algae cells |
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