Allelopathic eff ects of mixotrophic dinofl agellate Akashiwo sanguinea on co-occurring phytoplankton:the signifi cance of nutritional ecology

Blooms of Akashiwo sanguinea frequently break out around the world,causing huge economic losses to the aquaculture industry and seriously damaging coastal ecosystems.However,the formation mechanisms of A.sanguinea blooms remain unclear.We investigated the allelopathic effects of A.sanguinea on multiple phytoplankton species,explored the mode of allelochemicals action and the way of nutrient factors regulation of the allelopathic activity.Results show that strains of A.sanguinea could inhibit the growth of co-occurring phytoplankton including Scrippsiella trochoidea,Phaeocystis globosa,and Rhodomonas salina,but inhibition of Prorocentrum micans was not obvious.The inhibition rates on phytoplankton were positively correlated with the cell densities of A.sanguinea.The highest inhibition rate of 94%on R.salina was for A.sanguinea CCMA256 culture of 2000 cells/mL at 72 h.We observed that cells of S.trochoidea,Ph.g lobosa,and R.sali na were lysed when co-cultured with A.sanguinea,with the shortest time for S.trochoidea.Additionally,the growth rates of A.sanguinea were promoted by coculturing with S.trochoidea,Ph.globosa,and R.salina.Four components of A.sanguinea culture were all able to inhibit growth of R.salina:the strongest inhibitory effect was found in the sonicated culture,followed by whole-cell culture,filtrates of sonicated culture,and filtrate culture.The crude extract of A.sanguinea culture also lysed cells of R.salina,and the inhibition rates on R.salina increased with the increasing dose of crude extract.It was shown that both nutrient enrichment and nitrogen:phosphorus ratio imbalance enhanced remarkably the allelopathic activity of A.sanguinea.The highest inhibition rate on R.salina of 70%occurred in A.sanguinea JX13 treatment at 2000 cells/mL under high nutrient condition in 48 h.In JX14 treatment at 2000 cells/mL for N:P of 10:1,the inhibition rate increased by 1.7 times of that for N:P of 20:1.In addition,the allelopathy of A.sanguinea could not only be a competitive strategy but also a nutrition strategy,playing an important role in formation and/or maintenance of blooms of the mixotrophic dinoflagellate A.sanguinea.

Reviving and characterizing three species of dinoflagellate cysts dormant for about 70 years in the East China Sea:Biecheleria brevisulcata,Biecheleriopsis adriatica,and Scrippsiella donghaienis

Many marine dinoflagellates can form resting cysts as a part of their life cycle,and the cysts could be buried in sediment and remained viable for as long as over 150 years.However,only a very limited number of cyst species have been revived from long-buried sediments and investigated in regard to a possible shift in the intra-specific genetic structure of a species detected from the historical record at a particular location.Here,we report a successful germination of three species of resting cysts that were sampled from the depth dated back to 1941±18 AD from a 44-cm sediment core from the East China Sea.Seven isolates were established from germination of single cyst isolation or multi-cyst germinations.LSU rRNA gene or ITS sequences of these strains were obtained,then they were identified to be Biecheleria brevisulcata(five strains),Biecheleriopsis adriatica(one strain),and Scrippsiella donghaienis(one strain)in terms of morphology and rRNA gene sequence.Biecheleria brevisulcata strain 1,Bps.adriatica strain 21,and S.donghaienis strain 23 were examined in detail with light microscope(LM)and scanning electron microscope(SEM),and analyzed with high performance liquid chromatography(HPLC)for their pigment compositions,and genetic diversity.We also confirmed the presence of a resting cyst of Bps.adriatica in the field for the first time.The LSU rRNA gene-based genetic distances of Bps.adriatica from that obtained from water sample,single-cell PCR sequencing for the cysts isolated from the surface sediment of the same sea area and that reported from other regions during the recent years,and ITS-based genetic distances of S.donghaienis from that obtained from cysts isolated from the surface sediment of the same location and that reported from other regions during the recent years indicated that the intra-specific genetic structure of each species in the sampling area may have shifted during the last 70 years.Our work confirms that B.brevisulcata,Bps.adriatica,and S.donghaienis,all described as new species around 2010,have inhabited the East China Sea for about 70 years.The present work reports for the first time the revival of dinoflagellate resting cysts long-buried in the coastal sediments of China,which facilitates further study on the historical occurrences of other harmful dinoflagellates and their relevance to the regional climate and environmental changes in China.