Elsevier

Protist

Volume 162, Issue 4, October 2011, Pages 650-667
Protist

Original Paper
Taxonomy and Phylogeny of a New Kleptoplastidal Dinoflagellate, Gymnodinium myriopyrenoides sp. nov. (Gymnodiniales, Dinophyceae), and its Cryptophyte Symbiont

https://doi.org/10.1016/j.protis.2011.01.002Get rights and content

A new kleptoplastidal dinoflagellate, Gymnodinium myriopyrenoides sp. nov., was described using light microscopy, electron microscopy and phylogengetic analysis based on partial LSU rDNA sequences. Cells were dorsiventrally flattened, elongate-elliptical in ventral view. There was no displacement of the cingulum encircling the anterior part of the cell. The cingulum was curved posteriorly at the terminal junction with the sulcus. The sulcus was generally narrow but expanded in the posterior end. The epicone possessed an apical groove made of one and one-half counterclockwise revolutions. Phylogenetic analysis based on LSU rDNA showed that the sequence of G. myriopyrenoides was included in the Gymnodiniales sensu stricto clade and had special affinities with the species Amphidinium poecilochroum and Gymnodinium acidotum, which also harbor kleptochloroplasts. Phylogenetic analysis based on plastid-encoded SSU rDNA and ultrastructural observations suggested that the symbionts of G. myriopyrenoides were cryptophytes of the genus Chroomonas or Hemiselmis. Organelles including the nucleus, the nucleomorph, mitochondria, Golgi bodies and large chloroplasts remained in the cytoplasm of the symbionts, but not the periplast, ejectosomes or flagellar apparatus. The symbiotic level of G. myriopyrenoides was estimated to be a relatively early stage in the unarmored kleptoplastidal dinoflagellates.

Introduction

The typical chloroplasts of dinoflagellates are bounded by three membranes and possess chlorophylls a and c2 as well as peridinin as major accessory pigment (e.g., Graham and Wilcox, 2000, Stoebe and Maier, 2000). This type of chloroplast is considered to be originated from a red alga via a secondary endosymbiosis (e.g., Durnford et al., 1999, Fast et al., 2001, Zhang et al., 1999). Molecular phylogenetic studies suggest that the common ancestor of the dinoflagellates obtained the peridinin-containing chloroplast only once, at a relatively early stage in their evolutionary history (Saldarriaga et al., 2001, Saunders et al., 1997). However, about half of the dinoflagellates are colorless, perhaps through the secondary loss of chloroplasts (e.g., Saldarriaga et al., 2001, Taylor and Pollingher, 1987). Furthermore, some dinoflagellates appear to have replaced the peridinin-containing chloroplast with another type of chloroplast via tertiary endosymbioses (Hackett et al., 2004, Horiguchi, 2006, Morden and Sherwood, 2002, Saldarriaga et al., 2001, Stoebe and Maier, 2000).

It is most likely that some groups of dinoflagellates acquired chloroplasts via tertiary endosymbiosis. Karenia, Karlodinium and Takayama possess chloroplasts containing 19’-hexanoylofucoxanthin and/or 19’-buthanoyloxyfucoxanthin, and their chloroplasts are considered to have originated from a haptophyte alga (Hansen et al., 2000, de Salas et al., 2003). In these species, the chloroplast is surrounded by three membranes and neither cytoplasm nor organelles from the symbiont remain. Another group of dinoflagellates originated via tertiary endosymbiosis and includes some armored dinoflagellates such as Durinskia, Kryptoperidinium and Peridinium quinquecorne Abé. These dinoflagellates possess fucoxanthin-containing chloroplasts derived from diatoms (Chesnick et al., 1997, Horiguchi and Takano, 2006). Surprisingly, the nucleus, mitochondria and complete chloroplasts of the diatom are retained and are surrounded by a single membrane (Horiguchi, 2006, Horiguchi and Pienaar, 1991). These dinoflagellates are considered to have acquired the fucoxanthin-containing chloroplast from a common single ancestor (Horiguchi and Takano 2006). However, P. quinquecorne and Peridiniopsis spp. appear to have experienced a further replacement of their symbionts by another diatom (Horiguchi and Takano, 2006, Takano et al., 2008).

Possible intermediate states between a simple predation and tertiary endosymbiosis are known in the dinoflagellates. Some dinoflagellates engulf free-living algae and temporarily retain their chloroplasts (sometimes with other organelles) in a functional state for a few weeks to a month. These temporary symbiotic relationships are called “kleptoplastidy” and the symbionts (chloroplasts) are termed “kleptochloroplasts” (or “kleptoplastids”) (Schnepf 1992). The origin and structure of the kleptochloroplasts in dinoflagellates shows great diversity.

Some species of armored toxic dinoflagellates such as Dinophysis acuminata Claparède et Lachmann, D. acuta Ehrenberg and D. norvegica Claparède et Lachmann temporarily possess chloroplasts originating from cryptophyte algae, in particular the cryptophycean species of Geminigera/Teleaulax (Hackett et al., 2003, Janson, 2004, Janson and Granéli, 2003, Takishita et al., 2002). These kleptochloroplasts are bounded by two membranes, and no other organelles from Geminigera/Teleaulax are retained (Larsen 1992). Surprisingly, Park et al. (2006) showed that D. acuminata plundered the kleptochloroplasts from Myrionecta rubra Jankowski [usually referred to as Mesodinium rubrum (Lohmann) Hamburger et Buddenbrock], a ciliate that ingests and harbors Teleaulax. However, this issue is debated and some authors consider that the chloroplasts of D. acuminata are not kleptochlotoplasts but permanent ones (e.g., Garcia-Cuetos et al. 2010). In addition, D. mitra (Schütt) Abé possesses kleptochloroplasts derived not from a cryptophyte but from a haptophyte alga (Koike et al. 2005). Pfiesteria piscicida Steidinger et Burkholder, Amylax buxus (Balech) Dodge and A. triacantha (Jörgensen) Sournia also possess kleptochloroplasts originating from cryptophyte algae (Koike and Takishita, 2008, Lewitus et al., 1999). It is clear that Dinophysis, Pfiesteria and Amylax are distantly related (Fensome et al., 1993, Saldarriaga et al., 2004, Steidinger et al., 1996) and that these three genera apparently gained the kleptoplastidy independently. Gast et al. (2007) reported a new dinoflagellate with a kleptochloroplast originating from a haptophyte alga, but no ultrastructural study has been carried out on this dinoflagellate.

Several unarmored dinoflagellates also possess kleptochloroplasts of cryptophyte origin: Amphidinium poecilochroum Larsen (Larsen 1988), A. latum Lebour (Horiguchi and Pienaar 1992), A. vigrense Woloszynska (Wilcox and Wedemayer 1985), Gymnodinium acidotum Nygaard (Farmer and Roberts, 1990, Fields and Rhodes, 1991), G. aeruginosum Stein (Schnepf et al. 1989) and G. gracilentum Campbell (Skovgaard 1998). The ingestion and retention of cryptophyte algae was confirmed in these species, except for the ingestion by A. vigrense (Fields and Rhodes, 1991, Horiguchi and Pienaar, 1992, Larsen, 1988, Schnepf et al., 1989, Skovgaard, 1998). The synchronized division of the symbiont (chloroplast with other organelles) and host was observed only in G. acidotum (Farmer and Roberts, 1990, Fields and Rhodes, 1991). Ultrastructural studies revealed that the remnant conditions of the symbionts differed depending on the species. Therefore, studies on unarmored kleptoplastidal dinoflagellates would be very useful for a better understanding of the process of endosymbioses and algal evolution.

We found a new unarmored dinoflagellate possessing a blue-green chloroplast from Isonoura Beach, Wakayama, Japan. This blue-green chloroplast seemed to be a kleptochloroplast. In this study, we propose Gymnodinium myriopyrenoides Yamaguchi, Nakayama, Kai et Inouye sp. nov. for this new unarmored kleptoplastidal dinoflagellate, based on light and electron microscopy and molecular phylogenetic analysis using the partial large subunit ribosomal RNA gene (LSU rDNA) sequences. We also determined the sequences of plastid-encoded small subunit ribosomal RNA gene (pl SSU rDNA) derived from its kleptochloroplast to identify the origin of symbionts.

Section snippets

Description

Gymnodinium myriopyrenoides Yamaguchi, Nakayama, Kai et Inouye sp. nov.

Dinoflagellata inarmata cum canalis apicalis antihelicta 1.5 versatilis. Cellulae dorsoventraliter complanata, longa elliptica, 48–79 μm longa, 33–65 μm lata. Cingulum non descendens. Kleptochloroplastus ab Chroomonas vel Hemiselmis cum myri pyrenoidis.

Unarmored unicellular dinoflagellates with an apical groove of one and a half counterclockwise revolutions. Cells dorsiventrally flattened and elongate-elliptical, 48–79 μm long,

Taxonomy of the Dinoflagellate

In the traditional sense, unarmored dinoflagellates were classified mainly based on the relative sizes of the epicone and hypocone, and they were divided into several genera such as Gymnodinium, Gyrodinium, Amphidinium and Katodinium (e.g., Kofoid and Swezy 1921). Although recent studies indicate that this classification does not reflect their phylogenetic relationships (see below), many unarmored species remain and are still treated in the traditional classification. In this classification,

Methods

Sampling: In April, May and July 2006 and June, July and September 2007, sand samples containing Gymnodinium myriopyrenoides were collected at Isonoura Beach, Wakayama, Japan. The salinity of the water was about 34 psu. After returning to the laboratory, these samples were washed in Daigo IMK medium (Wako, Osaka, Japan) inside plastic cups, and the cells of G. myriopyrenoides were collected. G. myriopyrenoides was maintained for about 10 days at 20 °C, under 50 μmol photon m–2 s–1 with a 14/10 h LD

Acknowledgements

We thank Dr. Masanobu Kawachi (National Institute for Environmental Studies) and Dr. Yuji Inagaki (University of Tsukuba) for their valuable advice and for their correction of our manuscript. This research was supported by the Japan Science Society “Sasagawa Science Research Grant (2006-18-207)” (to H. Y.) and the Japan Society for the Promotion of Science (JSPS) (to H. Y.).

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