Screening Of Alginolytic Bacteria,preparation Of Compound Bacteria And Its Application In Aquaculture Tail Water

With the development of China's industry,environmental problems have come one after another,among which the eutrophication of lake water seriously affects the ecological environment and life health and safety.The microbial algae control method has good application prospects due to its advantages of good ecological safety and strong specificity.In this study,Microcystis aeruginosa,which is common in eutrophic water,was used as the research object of biological algae control,and R₁bacteria screened from the indigenous snails in Taihu Lake was used as the test strain.The growth kinetics and algae lysis kinetics of R₁ bacteria were constructed,and then the dual effect functional bacteria XR₁ for algae lysis,nitrogen and phosphorus removal was prepared by protoplast fusion.Finally,the subsurface flow wetland system was successfully applied to yangjiaxu crab pond demonstration base in Qianhuang town of Changzhou City,achieving the goal of zero tailwater discharge.In terms of the algae dissolving characteristics of algae dissolving R₁:the algae dissolving R₁ was screened out from the snails growing in the blue-green algae outbreak waters of Taihu Lake through a series of ways.The experimental results show that the R₁(Bacillus)has high algae dissolving ability,and the algae solution after algae dissolving is non-toxic and will not cause secondary pollution to the water body,indicating that it is a high efficiency algae dissolving bacteria with engineering application value.When R₁was in the logarithmic growth period,at the ratio of bacteria to algae of 1:10 and the initial algae concentration of 3338.26 mg·m^-3,R₁ could reach 89.37%algae dissolving ability in 7 days.The algae dissolving experiment proved that R₁ could exert its algae dissolving ability to the maximum in the suitable algae dissolving environment,but only when the concentration of algae dissolving bacteria reached a certain value could it produce algae dissolving effect.Different initial algae concentrations would affect the final algae removal rate.Generally,the higher the initial algae concentration was,the more obvious the algae dissolving effect was,and the shorter the time to achieve the best algae dissolving effect was.In terms of the the construction of algae dissolution kinetics,the growth curve of R₁ conformed to the Logistic model,and the degradation curve of chlorophyll-a was in accordance with the first-order kinetic model.In the process of algae dissolving,the strain in logarithmic growth stage had the strongest ability of algae dissolving,and there was an inverse relationship between chlorophyll-a content and algae dissolving time.Among them,the chlorophyll-a degradation kinetic model[Chla]=4831.82071×e^-0.0241t can be used to predict the degradation effect of Microcystis aeruginosa by dissolving bacteria.By studying the algae dissolving mechanism of R₁ strain,it was found that R₁ strain not only secretes some high temperature resistant algae dissolving substances for indirect algae dissolving,but also directly attacks algae cells or feeds on Microcystis to directly dissolve algae.When Microcystis cells break down,it will decompose aromatic amino acids,amides and other substances.In terms of the construction of compound microbial agent XR₁:the protoplasts of alginolytic bacteria R₁ and denitrifying phosphorus accumulating bacteria B8 were successfully fused by cell fusion technology,so as to construct a double effect engineering bacteria XR₁with the characteristics of both parents,which has the functions of alginolytic,nitrogen and phosphorus removal.By response surface analysis,the optimal algicidal conditions of XR₁ were found(when temperature is 27.17?,pH is 7.66 and shaking speed is 152.81 r·min^-1).It was found that there was interaction among the three factors,and the order of affecting algicidal rate was temperature>pH>shaking speed.Within 7 days,the algae dissolving,nitrogen and phosphorus removal efficiency of XR₁ reached 93.95%,58.39%and 61.7%respectively.The results showed that XR₁ not only directly dissolved Microcystis aeruginosa as carbon source,but also indirectly dissolved algae by absorbing nitrogen,phosphorus and other nutrients in water.In conclusion,the double effect engineering bacteria XR₁ has high algae dissolving efficiency,nitrogen and phosphorus removal function,strong adaptability to the environment and non-toxic products,which can be applied to the subsequent engineering application of water bloom control.In terms of microbial agent combined with subsurface flow wetland efficiency:the vertical subsurface flow wetland system has good degradation ability to TN,TP,CODMn and Chla in the tail water of crab pond culture within 50 days,and the average degradation rates of TN,TP,COD_Mn and Chla are 86.29%±2.77%,91.19%±1.18%,70.88%±3.10%and 90.95%±1.64%,respectively.After purification,the aquaculture tail water could meet the discharge standard.The improved subsurface flow wetland combines the characteristics of trickle filtration and swamp filtration,and its purification is mainly based on the physical adsorption of filter stone layer,supplemented by artificial dosing of bacteria.Among them,the volcanic stone filter material layer has made a great contribution to the purification of nitrogen and phosphorus in the system,but the filter material needs to be replaced regularly to maintain the continuous purification effect.The investment of composite bacteria can improve and enhance the overall purification ability of the system in a short time.With the enrichment of microorganisms on the biofilm,the efficiency of composite bacteria is gradually replaced.The results of biological microscopic examination confirmed that with the continuous operation of the subsurface flow wetland,the changes of indicator organisms in the system before and after can prove the improvement of water quality,and the enriched microorganisms show that the improved water quality is more conducive to the growth of microorganisms.In terms of the actual purification efficiency of the submerged wetland:the improved version of subsurface flow wetland was successfully applied to the purification of crab pond tail water,achieving the purification effect of zero discharge of aquaculture tail water.Adding double effect engineering bacteria in advance in the wetland will accelerate the construction of colonies in the system,strengthen the overall self purification ability of the wetland,and the emergent plants and microorganisms in the sediment in the system can play a synergistic role to achieve stable and rapid degradation of various pollutants.The results showed that the removal efficiency of NH₃-N,TN,NO₃-N and TP in crab pond tailwater was 92.3%,77.5%,88.6%and 87.1%,respectively.The system had a remarkable effect on the degradation of NH₃-N in tailwater,but it was urgent to further improve the overall removal efficiency of COD.The degradation trend of pollutants in each experimental cycle was in line with the first-order kinetic equation,and the model could be used to guide the follow-up wetland efficiency The optimization of the system.With the continuous operation of the wetland,the complex microbial agent is gradually replaced by the enriched colonies in the system.The dominant bacteria in the sediment is Proteus,which contains a variety of bacteria with the functions of ammoniation,nitrification and denitrification.Chlorella appeared in every cycle,and indicator algae such as crescent algae and Chaetoceros appeared in the middle and late period of system operation,which indicated that pollutants in subsurface flow wetland were effectively degraded and water quality was significantly improved.To sum up,this study successfully improved the traditional subsurface flow wetland and applied it to the crab pond aquaculture demonstration base.Combined with the use of adaptive bacteria,the wetland can establish a fast and stable purification system and achieve the goal of zero discharge of aquaculture tail water.During the operation of the system,observing the changes of the biofacies can understand the operation of the system,and at the same time,artificial dosing of bacteria can be used to deal with the local algae explosion.The kinetic equation of Microcystis degradation established in this experiment can also be used to predict the algae removal process.