Kleptoplasty in Dinophysis spp. Ecological role and evolutionary implications

Transcription

1 Kleptoplasty in Dinophysis spp Ecological role and evolutionary implications

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3 Linnaeus University Dissertations No 19/2010 KLEPTOPLASTY IN DINOPHYSIS SPP Ecological role and evolutionary implications SUSANNA MINNHAGEN LINNAEUS UNIVERSITY PRESS

4 KLEPTOPLASTY IN DINOPHYSIS. Ecological role and evolutionary implications. Doctoral dissertation, School of Natural Sciences, Linnaeus University Series editor: Kerstin Brodén Cover: Illustrations by Mats Minnhagen ISBN: Printed by: Intellecta Infolog, Gothenburg, 2010

5 Abstract Minnhagen, Susanna (2010). Kleptoplasty in Dinophysis spp. Ecological role and evolutionary implications. Linnaeus University Dissertations No 19/2010. ISBN: Written in English. This thesis deals with the question of whether planktonic protits of the genus Dinophysis have permanent plastids (=chloroplasts) or practice kleptoplasty, i.e. acquire plastids via predation on other microorganisms. Sequencing the plastid 16S rdna of Dinophysis spp. collected from 4 different geographical regions unveiled two different plastid genotypes within this genera: one that was found at all locations investigated, identical to that of the free-living cryptophyte Teleaulax amphioxeia, and another found only in the Greenland Sea, closely related to that of the cryptophyte Geminigera cryophila. Both types were found within the species D. acuminata. These findings imply that the plastids in Dinophysis spp. were not inherited from a common ancestor, but acquired from feeding. By using flow cytometry in combination with an acidotrophic probe, it was shown that 71 % of the cells in a D. norvegica population in the aphotic zone of the Baltic Sea had food-vacuoles. Dinophysis used to be regarded as a primarily phototrophic organism, and this was a higher proportion of cells with food-vacuoles than reported earlier. To further study if Dinophysis needs constant refill of new plastids from the environment, a new method combining flow-cytometry and quantitative real-time PCR was developed to compare the levels of nuclear and plastid DNA in different phases of the cell-cycle. Results showed that plastid acquisition in Dinophysis was uncoupled with the cell-cycle, which is different than the pattern seen in microalgal species with permanent plastids. Furthermore, when quantitative real-time PCR combined with flowcytometry was used to follow D. caudata cultures during a 65 days starvation/feeding experiment, the cells first went through a steady decrease in plastid DNA during starvation. In contrast, after feeding on the ciliate Myrionecta rubra, plastid DNA in starved cells increased 7-fold, thereby directly revealing the kleptoplastic behavior. The main conclusion from this thesis is that Dinophysis cells are actively taking up kleptoplastids from the ciliates on which they feed, and that kleptoplasty is an important key to understand Dinophysis ecology. Part of this thesis work has also been dedicated to the application and optimization of new methods, and it shows how quantitative real-time PCR, flow cytometry and molecular methods in different combinations can be used as powerful tools for the study of plankton ecology. Keywords: Dinophysis, kleptoplastid, real-time PCR, 16S rrna, plastid evolution, Baltic Sea, flow cytometry, aquatic ecology

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21 (B) (C) (A) WF Carvalho 10 µm (E) (F) 10 µm MG Park (G) (H) (A) 10 µm (I) 10 µm

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25 Amplification plot: Standard curve: Fluorescence C T Threshold No. PCR cycles Log. DNA copies (or cells) Figure 3

26 Orange fluorescence Particle size Figure 4

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30 Log (cells L -1 ) 6 4 Teleaulax M. rubra D. acuminata Stratification Chl a Year

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33 (A) Hemiselmis AB Plagioselmis AB D. acuminata Greenland Sea 2002 (paper 1) D. acuminata Kalmar harbour 2004 (paper 1) D. acuminata BY (manus. in prep.) D. norvegica BY (manus. in prep.) M.rurbra BY (manus. in prep.) T. amphioxeia AY D. acuminata Kalmar harbour 2003 (manus. in prep) M. rubra Kalmar harbour 2007 (manus. in prep) D. acuminata DQ Greenland Sea 2002 (paper 1) M. rubra CCMP 2563 McMurdo Sound 2000 (Kindly provided by Matthew Johnson) Geminigera cryophila AB (Takishita et al. 2002) (B) Mesodinium pulex AY M. rubra AY Antarctica M. rubra EF Korea M. rubra AY Chesapeake bay 0,01 M. rubra Baltic Sea Figure 7. ab

34 Cyanobacterial symbionts Cryptophyte plastids Haptophyte plastids Other bacterial symbionts Eukaryotic endosymbionts Figure 8

35 Kleptoplastid stage: -Digestion prevented -Repeated uptake -Unsynchronized division I P u N P n Persistent symbiont: -Nucleus retained -Division synchronized 2 N n P n P N Gene transfer: -DNA from symbiont transferred to the host nucleus -Protein transport system established Permanent plastid: -Symbiont nucleus erased 3 4 Nnp P PP u Nnp P Nnp Nnp New plastid? P Dead end? n 5 6 Plastid lost: -Plastid genes and plastid protein import system remains. New uptake: -Prey nuclei immediately erased -Only plastid retained

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