Effect On Paralytic Shellfish Toxins In Flocculating Alexandrium Pacificum By Modified Clay

As a common harmful algal blooms(HABs)organism of China offshore,Alexandrium pacificum,which can produce paralytic shellfish toxins(PSTs),poses a serious threat to marine aquaculture and human health.Therefore,in the mitigation of A.pacificum algal blooms,attention should be paid to the dynamic changes of algal toxins.The rapid flocculation of modified clay(MC)to eliminate algal cells in water column is a widely recognized method for controlling algal blooms.However,the effects of A.pacificum flocculation by MC on PSTs toxins have not been studied systematically.In this study,the variation characteristic of intracellular and extracellular PSTs of A.pacificum associated with growth phases were systematically summarized.On this basis,the poly aluminum chloride MC(PAC-MC),the frequently-used modified clay in the field,was selected to investigate the removal effect on both intracellular and extracellular PSTs from water column while removing A.pacificum,and the effect on the PSTs biosynthesis ability and characteristics of algal cells of the whole system,were revealed according to algal cells state,gene transcription expression,algal-bacteria relationship,water quality parameters.In order to find a more effective method to remove algae and reduce toxicity of A.pacificum algal bloom,the degradation of PSTs toxins by oxidized modified clay in the flocculation process was further studied,and the mechanism on PSTs toxins biosynthesis and secretion was further elaborated combined with algal cells state,bacterial biomass,algal-bacteria relationships and water quality parameters changes.The main results are as follows:(1)Toxins production characteristic in A.pacificum of different growth stage.Based on the LC-MS method,the contents and components of intracellular and extracellular PSTs of A.pacificum at different growth stages were determined,and the variation characteristics of the toxic production of A.pacificum at different growth stages were described in detail.During exponential growth phase,both intracellular and extracellular PSTs content and toxicity of A.pacificum increased sharply.On 10 th day,the intracellular toxin content and toxicity were 7.12 times and 6.83 times higher than that on 2th day,and the extracellular toxin content and toxicity were 10.18 times and 8.61 times higher than that on 2th day,respectively.Since the increase of extracellular PSTs content mainly occurred at exponential phase,when most cells were living,we speculated that active PSTs secretion of living cells might be an important producing way of extracellular toxins.In addition,the study also found that the single-cell toxin content and toxicity of A.pacificum increased first and then decreased in the exponential growth period,reaching the highest on the 6th day.The results showed that the toxin content and toxicity of A.pacificum were significantly different in different growth stages,and the toxin production was stronger in the exponential growth stage.(2)The effect on PSTs of A.pacificum flocculation by PAC modified clay.In this study,the removal effect of PAC-MC on A.pacificum cells and total PSTs in water column were investigated.The results showed that 0.2 g/L PAC-MC had a good removal effect on A.pacificum cells and total PSTs in water column.The decrease of intracellular PSTs content accounted for more than 90% of the total.And there was no significant difference in the single-cell PSTs content and component between the residual algal cells in PAC-MC treatment group and the algal cells in control group.It has been also showed that PAC-MC could effectively inhibit the increase in the PSTs content and toxicity in the whole system in 12 days.Mechanistic analysis results showed that the restriction of the PSTs content and toxicity by PAC-MC was mainly achieved by affecting A.pacificum physiological processes and destroying algae-bacteria interactions.In addition,there was no significant increase in single-cell toxicity in PAC-MC treatment groups throughout the experiment.Algal cells after 0.2g/L PAC-MC flocculation tended to synthesize low-toxicity sulfated PSTs components such as GTX2&3,C1&2,GTX5,as a result of the up-regulation of sxtN and sxtO genes related to the sulfation of PSTs in Alexandrium cells.(3)The effect on PSTs of A.pacificum flocculation by oxidized modified clay.Considering the fact that PSTs are easy to be oxidized and degraded in seawater,this study adopted oxidized modified clay,to explore the effect of oxidized MC on PSTs during the flocculation to A.pacificum.The results showed that 0.1 g/L oxidized modified clay could degrade PSTs dissolved in water by oxidation while removing A.pacificum cells by flocculation.The extracellular PSTs toxicity in oxidized MC treatment group decreased by more than 92% within 15 min compared with that in control group.In addition,0.1 g/L oxidized MC might induce the transformation of more toxic GTX1&4 to less toxic GTX2&3 and C1&2,resulting in intracellular PSTs toxicity reduced within 15 min.And the intracellular PSTs toxicity was effectively inhibited by oxidized MC,possibly by inhibiting the A.pacificum cells growth and destroying algae-bacteria interactions.Intracellular and extracellular PSTs toxicity in oxidized MC treatment groups were consistently lower than the control group within48 h.The results provided a theoretical basis for evaluating the effectiveness and safety of modified clay method for controlling toxigenic Alexandrium blooms,and provided a new idea for solving the toxigenic Alexandrium blooms and paralytic shellfish toxins pollution.