Study On The Discovery Of Novel Algicldal Bacteria And The Mechanisms Of Algicidal Bacteria Against Toxic Alexandrium Sp.

Compared to decades ago,harmfula algal blooms(HABs),as one of the serious disarsters,have broken out more frequently.As a result,there are more toxic algal species,algal toxins,areas affected,fisheries resources impacted,and higher economic losses.It is known that bacteria in phycosphere can regulate the growth of algae,stimulating or inhibiting the algal growth.Thus,as a potencial HABs regulator,the interaction between alga and bacteria has been attracted increasing attention.In this study,the bacterial diversity of associated' bacteria of a toxic dinoflagellate Alexandrium fundyense was stuedies.Two algicidal bacteteria against A.fundyense were isolated from its phycosphere,one of which,strain B201 was conducted for further analysis.What's more,the physiological and biochemical responses and protein expression profile of Alexandrium tamarense were studied when algicial substance from algicidal bacteria Pseudoalteromonas sp.SP48 was induced into A.tamarense culture.The results are listed as below:(1)The bacterial diversity of associated bacteria of A.fundyense was characterized by analyzing the 16S rRNA gene library.Clones with bacterial 16S rRNA genes were sequenced and phylogenetic analysis was conducted.Three groups of bacteria were found in the phycosphere of A.fundyense:a-Proteobacteria,y-Proteobacteria and Flavobacteriaceae.(2)Two algicidal bacteria against A.fundyense were isolated from the phycosphere.They belong to y-Proteobacteria and Flavobacteriaceae,and were identified as Marinobacteria sp.strain B82 and Croceibacter sp.strain B201,respectively.This is the first report about algicidal bacteria against A.fundyense and also the first record that Croceibacter sp.was isolated with algicidal activity.(3)It is showed that Croceibacter sp.B201 could inhibit the growth of A.fundyense only with additional nutrients exist(5%or even 1%peptone or yeast exact).Algicidal activity was detected in cell-free supernatant when B201 was cultured with 2216E medium,which indicated that the bacterial isolate B201 could inhibit growth of A.fundyense via indirect attack,and the additional nutrients were obligated for synthesization of unidentified algicides.Except for A.fundyense,strain B201 could also lyse 4 other species:Alexandrium minitum TW01,Scrippsiella trochoidea XM01,Thalassiosira pseudonana and Nitzschia closterium.(4)The physiological and biochemical responses of A.tamarense were studied when treated with algicidal substance from Pseudoalteromonas sp.SP48.ROS concentration hoiked and oxidative burst was induced in a short time.As treated time increasing,ROS was accumulated in algal cells,which cuased lipid peroxidation and MDA content increased.SOD and CAT,as key oxidases in cells,were also triggered by ROS accumulation and their activities increased.At the same time,algal Fv/Fm was inhibited as chl a content in algal cells decreased.(5)The protein expression profiles were also analyzed using 2-DE combined with MALDI-TOF/TOF MS when A.tamarense were exposured to algicidal substance.Proteins or key enzymes associated with carbohydrate metabolism,photosynthesis,respiration and other metabolisms were found affected by algicidal substance.It is indicated that the important physiological processes of A.tamarense were disturbed by algicidal substance from SP48.The novel isolated algicidal bacteria could enrich the resource of algicidal bacteria.The studies on bacterial diversity and mechanisms of algicidal bacteria killing algae would deepen our understanding of bacteria-alga interaction and finally provide potencial powerful tools for controlling HABs.