Study On The Aggregation Characteristics Of Cyanobacterial Cells Under Allelopathic Stress And The Mechanism Of EPS Alleviating Stress

Serious water eutrophication increasing rapidly leads to the excessive proliferation of algae blooms and frequent water blooms,damaging water functions and aquatic ecosystems.The removal technology of cyanobacteria in eutrophic water has become a research focus in the field of environmental science and protection.Using allelochemicals secreted by organisms to inhibit the growth of algae provided a great idea for the control of algal blooms,motivating this study.Further study on the allelopathic cyanobacteria inhibition is helpful to evaluate the mechanism of allelopathy on cyanobacteria in natural system.However,the aggregation of cyanobacteria and the mechanism of alleviating allelopathy stress by cyanobacteria are still unclear.To explore the environmental behavior of cyanobacteria under allelopathy stress,this study selected Pyrogallic acid(PA),Linoleic acid(LA),and Tannic acid(TA)as the representative allelochemicals of phenolic acids,fatty acids and tannins.Microcystis aeruginosa was used as the experimental cyanobacteria to study the interaction between typical allelochemicals with cyanobacteria cells and the extracellular polymeric substances(EPS)released by them.The effects of typical allelochemicals on the aggregation of Microcystis aeruginosa were studied by measuring the aggregation rate and its surface properties(zeta potential,hydrophilicity,and the composition of EPS)under allelopathy stress;the effect of EPS on the growth inhibition of Microcystis aeruginosa by allelochemicals was explored by comparing the changes of cell growth inhibition rate,esterase activity,and photosynthetic activity indexes before and after removal of EPS;the mechanism of the interaction between typical allelochemicals with EPS of Microcystis aeruginosa were analyzed by three-dimensional fluorescence spectroscopy,synchronous fluorescence spectroscopy,infrared spectroscopy,and other characterization methods.The results are not only shed light on the understanding of the role of EPS in the process of algal inhibition by allelochemicals,but also provide new ideas for screening high-efficiency allelopathic anti-cyanobacteria materials.The main conclusions are as follows:1.Under the short-term(24h)effect of allelopathic stress,the typical allelochemicals reduce the aggregation rate of Microcystis aeruginosa by affecting the surface charge of cells.However,this downward trend will be eased under the long-term effect of allelopathic stress because of the hydrophobicity of cyanobacteria cells increased after the increase of EPS secretion.Under LA stress,more polysaccharide of EPS were secreted,while under PA or TA stress,more protein and humic acid of EPS were secreted.The ability to enhance the hydrophobicity and aggregation rate of cyanobacteria cells was LA>PA>TA.2.The EPS secreted by Microcystis aeruginosa could alleviate the toxic effects of typical allelochemicals.Under the exposure concentrations of LA 0.1-4 mg L^-1,TA2.5-80 mg L^-1and PA 0.5-16 mg L^-1,the metabolism and photosynthetic activity of Microcystis aeruginosa were inhibited by allelopathic stress.Compared with intact cells,the toxicity of typical allelochemicals to Microcystis aeruginosa removed EPS was enhanced,the 96h IC₅₀of typical allelochemicals decreased from 0.80 mg L^-1to0.72 mg L^-1(LA),from 19.22 mg L^-1to 12.48 mg L^-1(PA),and from 55.36 mg L^-1to20.59 mg L^-1(TA),respectively.EPS can effectively alleviate the toxic effect of allelochemicals on Microcystis aeruginosa and stabilize the metabolism and photosynthetic activity of Microcystis aeruginosa.Besides,the ability of EPS to alleviate the cytotoxicity effect of Microcystis aeruginosa was TA>PA>LA.3.EPS alleviated the toxic effect of allelochemicals on cyanobacteria cells through chemical combination.Tryptophan-like proteins and humic acid-like substances in EPS provided binding sites and had a high binding constant to TA,LA,and PA.Among them,TA(or LA)had the highest binding capacity to proteins(or the humic acid)in EPS.After binding,the tryptophan type peptide chain in EPS extended,increasing the hydrodynamic diameter of macromolecules in EPS.Generally,bound extracellular polymeric substances(B-EPS)played a more crucial role during the binding process than soluble extracellular polymeric substances(S-EPS).The binding reaction between EPS and allelochemicals was mainly hydrophobic,and the reaction was endothermic,revealing that the increase in temperature was conducive to to reduce the toxic effect of allelochemicals on cyanobacteria.