The Death Mechanism Of Thalassiosira Pseudonana Induced By Bacterium Chitinimonas Prasina LY03

Diatoms are a kind of photoautotrophic single cell eukaryotic organisms,which widely distributed in the global ocean and in a variety of freshwater systems.As an important contributor to marine primary productivity,diatom plays an important role in biogeochemistry cycle of marine ecosystems.Diatom is also a bloom forming phytoplankton.The model diatom Thalassiosira pseudonana plays an important role in the global carbon and sulfur cycle.We studied the death mechanism of T.pseudonana induced by algicidal bacterium Chitinimonas prasina LY03 with direct attack.The genome of the C.prasina LY03 and transcriptome of T.pseudonana were analyzed.We have obtained a novel algicidal compound from strain LY03 and investigatet he characteristics of algicidal compound.We investigated algal lysis process and mechanism from morphological alteration,photosynthetic systems and the response of antioxidant enzymes to oxidative stress of the algal cells.1.The strain LY03 lysed T.pseudonana cells through direct attack.The bacterial chemotactic ability and direct contact between C.prasina LY03 and T.pseudonana were investigated.Apart from T.pseudonana,strain LY03 could also lysed other diatom species.2.The genome of the C.prasina LY03 was 4.61 Mb and G+C content was 60.95%.We obtained the chemotaxis genes of C.prasina LY03.We found the mutant strain QY03 of algicidal bacterium C.prasina LY03.Strain QY03 did not show any obvious algicidal activity against T.pseudonana.The unique genes of LY03 were obtained after comparative genomic analysis with QY03.It was speculated that the algicidal function of strain LY03 may be related to the secretion system and the chemotaxis of bacteria.3.The transcriptome analysis was performed after T.pseudonana cells treated with C.prasina LY03.It was found that these differentially expressed genes were enriched in photosynthesis,glutathione metabolism,ascorbic acid metabolism,plant-pathogen interaction,carbon fixation in photosynthetic organisms and other related metabolic pathways.The photosynthesis of T.pseudonana was inhibited after LY03 treatment.The expression of glutathione,ascorbate peroxidase,glutaredoxin and thioredoxin were up-regulated,indicating that strain LY03 caused severe oxidative stress against T.pseudonana cells.The expression level of chitin synthase genes in T.pseudonana cells were reduced significantly.The chitin synthesis of T.pseudonana was inhibited.Strain LY03 damaged the siliceous cell wall of T.pseudonana.The expression of key enzymes involved in glycolysis were down-regulated,indicating that glycolysis metabolism was inhibited by strain LY03.The results showed that C.prasina LY03 caused stress on algae cells,and the normal metabolism of algae cells such as photosynthesis,chitin,synthesis,glycolysis,reactive oxygen species and antioxidant system was seriously affected.4.We obtained a novel algicdal compound from C.prasina LY03 with molecular weight of 381 and the molecular formula was C24H35N3O.The compound showed algicidal activity against T.pseudonana with a LD50 of 0.12 ?g/mL.5.The photosynthesis system of T.pseudonana cells was impacted severely by algicdal compound.The photosynthesis pigments were destructed and Fv/Fm and the rETR decreased rapidly after algal cells were treated.We found that the algicidal process and structure changes of T.pseudonana cell with C.prasina LY03 by scanning electron microscopy and transmission electron micrograph observation.The cell organelles loosely arranged,cell contents flowed out and finally cell broke down.6.The ROS content of T.pseudonana increased rapidly in a short time and reached the maximum value with 1 h of treatment.The intracellular ROS content after 1 h of treatment with algicdal compound was 4.04 times of the control.The NAC,GSH and AsA could effectively scavenge excessive ROS in the T.pseudonana cells.The antioxidant system of T.pseudonana cells,including SOD,CAT,POD,GPX,GR,GSH and AsA,were triggered to different degrees to scavenge ROS.The accumulation of ROS in a short time caused lipid peroxidation.The cell membranes and walls ruptured,cell contents lost,and finally algal cell lysis.