Studies On Photosynthetic Damage And Apoptosis Induced By Linoleic Acid In Karenia Mikimotoi

The harmful algal bloom(HAB)is a kind of marine ecological anomaly and one of the main environmental problems of global offshore water pollution.In recent years,eutrophication in coastal areas around the world have led to frequent outbreaks of HAB,causing a serious threat to the sustainable development of aquaculture,marine ecological security and human health in coastal areas.At present,the outbreak,ecological effect and the prevention of HAB have attracted extensive attention,and the search for effective methods of preventing and controlling HAB has become an important research content in the field of water environment.Physical and chemical methods inevitably cause secondary environmental pollution problems.Biological methods are to use organisms themselves and their secretions to prevent and control HAB.Currently,they mainly include cultivation of macroalgae,cultivation of filter-feeding animals and introduction of bacteria and viruses that can infect microalgae.Among them,the use of macroalgae aimed to control HAB has the advantages of ecological friendliness,low cost,high efficiency and long control time,and has a good application prospect as well.We chose Karenia mikimotoi,a preponderant harmful algal bloom,as the research object.We did this research from the perspective of growth inhibition of apoptosis,and photosynthetic mechanism,aimed to explore the allelopathy mechanism of linoleic acid(LA)secreted by Karenia mikimotoi.And we put emphasize on the action of photosynthetic algae cell damage mechanis,intracellular reactive oxygen species level,and the relationship between the cell cycle and cell apoptosis under the pressure of LA.This study provides a theoretical basis for the use of allelochemicals secreted by macroalgae to controlHAB.The main research results are as follows: 1.Growth inhibition level: low concentration(<100?g/L)of LA promoted the growth of Karenia mikimotoi,while high concentration(>500?g/L)of LA inhibited the growth of Karenia mikimotoi.In the low LA treatment group,the number of cells wasincreased,while in the LA treatment group,the number of cells was less,the cells presented irregular shape,and the cells became lighter in color,and the number of cells in the same field gradually decreased with the increase of LA concentration.Flow cytometry results showed that under high concentration of LA,the normal cycle of algal cells was blocked at S phase.Moreover,the addition of antioxidant(NAC)and caspase-3 inhibitor(Ac-DEVD-CHO)did not alleviate the periodic retardation effect of LA on the Karenia mikimotoi.2.Photosynthetic mechanism: high concentration of LA reduced the content of photosynthetic pigments(chlorophyll a,chlorophyll c and carotenoids)in Karenia mikimotoi.Transmission electron microscopy showed thatLA could make thechloroplast of Karenia mikimotoi swell and deform,and the granule lamellar structure was loose and fractured.The addition of antioxidant(NAC)and caspase-3 inhibitor(Ac-DEVD-CHO)alleviated the damage of LA to the chloroplast structure of microalgae.Plant efficiency test showed that LA effect,chlorophyll fluorescence parameters of Karenia mikimotoi-PS? maximum photochemical efficiency(Fv/Fm),based on the absorption of light energy performance index(PI)andenergy for electron transport(ETo/RC)shows the tendency of lowerexposure to LA;the energy absorbed by antenna pigment(ABS/RC)and the energy used for heat dissipation(DIo/RC)showed an increasing trend;the energy used to reduce QA(TRo/RC)increased gradually with the increase of LA concentration,and decreased significantly when LA concentration was 900?g/L.The resultsabove showed that LA damaged the photosynthetic system of Karenia mikimotoi.3.Apoptosis mechanism: according to the observation with inverted phase contrast fluorescence microscope,LA marginalized the nucleus of Karenia mikimotoi,making the nucleus smaller and condensed.The activity of caspase-3 and caspase-9 related to apoptosis was increased,and the proportion of apoptotic cells was increased detected by flow cytometry under the action of LA,indicating that LA could promote the cell apoptosis of Karenia mikimotoi.After the addition of antioxidant(NAC)and caspase-3 inhibitor(Ac-DEVD-CHO),the proportion of apoptotic cells decreased under the action of LA,indicating that the two inhibitors could slow down the apoptotic effect of LA on Karenia mikimotoi.4.Relationship between cycle-oxidative stress–apoptosis: under the action of LA,the cell cycle process of Karenia mikimotoi was less correlated with the intracellular reactive oxygen level,and the oxidative stress caused by LA in Karenia mikimotoi could cause cell apoptosis.