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Taxonomy and phylogeny of
marine unarmored dinoflagellates
(and others)
Mitsunori Iwataki
Asian Natural Environmental Science Center,
The University of Tokyo
Gonyaulacales Peridiniales Prorocentrales Gymnodiniales Dinophysiales
Noctilucales
Morphology of dinoflagellates
unarmored
(athecate, naked)
armored
(thecate) Woloszynskioids
(Taylor 1987) Amphiesmal vesicle
thecal
plates
Armored and unarmored dinoflagellates
Armored dinoflagellates are classified based on number and
arrangement of the thecal plates.
Scrippsiella Po, X, 4', 3a, 7", 6c, 4s, 5"', 2"”
Heterocapsa Po, X, 5', 3a, 7", 6c, 4s, 5"', 2””
Taxonomy of armored dinoflagellates
Gymnodinium Gyrodinium Cochlodinium Amphidinium Katodinium
Since the cells are not covered by thecal plates, position and torsion of the cingulum was used for he crassic taxonomic criterion for genus of unarmored dinoflagellates.
Previous taxonomy of unarmored dinoflagellates
Cingular displacement <1/5 of cell length Cingular displacement >1/5 of cell length
Gymnodinium Gyrodinium
Previous taxonomy of unarmored dinoflagellates
Gonyaulacales
Peridiniales
Peridiniales (Freshwater)
Gymnodiniales
Peridiniales (Maine)
Prorocentrales
Suessiales
Gymnodiniales
Gymnodiniales
Prorocentrales
GP
P c
om
ple
x
Peridiniales (Marine)
Akashiwo Gymnodinium
Karenia Karlodinium
Phylogenetic positions of unarmored dinoflagellates
Generic diagnoses of unaremod
dinoflagellates were emended
and several new genera were
newly proposed.
(Daugbjerg et al. 2000)
straight straight
Horseshoe -shaped
Horseshoe -shaped
sigmoid elliptical ?
curves in clockwise
Apical groove Apical groove is visible under scanning electron microscope.
Stable character in each genus, and used for taxonomic criterion of present
taxonomy of unarmored dinoflagellates.
C. polykrikoides
Apical groove of Cochlodinium fulvescens
C. fulvescens
C. fulvescens (Iwataki et al. 2010) (Iwataki et al. in press)
Togula britannica
(Flø Jørgensen et al. 2004)
Ankistrodinium semilunatum
(Hoppenrath et al. 2012)
Bispinodinium angelaceum
(Yamada et al. 2013)
Recently established genera (from Amphidinium) Amphidinium sp.
Apicoporus glaber
(Sparmann et al. 2008)
Testudodinium corrugatum
(Horiguchi et al. 2012)
(Sanpedro et al. 2011)
Barrufeta bravensis
Levanderina fissa
(Moestrup et al. 2014)
Ceratoperidinium falcatum
(Rene et al. 2013)
Moestrupia oblonga
(Hansen et al. 2011)
Recently established genera (Gymnodinium and Gyrodinium)
Paragymnodinium shiwhaense Gyrodiniellum shiwhaense
(Kang et al. 2010) (Kang et al. 2011)
Phylogenetic position of unarmored dinoflagellates
based on SSU rDNA sequences
Phaeobalechina catenata
nom. nud. (Omura Bay)
Cochlodinium polykrikoides
(Isahaya Bay, Malaysia)
Gyrodinium falcatum
(Inokushi Bay, Tsushima)
Gymnodinium nolleri
(Hiroshima Bay)
G. microreticulatum
(Sasebo, Obama)
Gyrodinium instriatum
(Isahaya Bay)
Torodinium teredo
(Nagasaki)
Gymnodinium sp.
(East China Sea)
G. cf. chlorophorum
(Tokyo Bay)
Gymnodinium sp.
(Yatsushiro Bay)
C. convolutum
(Omura Bay)
Gymnodinium sp.
(East China Sea)
Gymnodinium catenatum
(Philippines)
Spatulodinium pseudonoctiluca
(Omura Bay)
Karlodinium sp.
(Ariake Sound)
Takayama sp.
(Ariake Sound)
D. pseudolunula
(Omura Bay)
Cochlodinium fulvescens
(Tachibana Bay, Indonesia)
Photosynthetic Species 55 samples belonging to 15 genera
Erythropsidinium agile
(Nagasaki)
Achradina pulchra
(East China Sea)
Gyrodinium sp.
(East China Sea)
Gymnodinium gracile
(Nagasaki)
Nematodinium sp.
(Nagasaki)
Nematodinium sp.
(Nagasaki)
Balechina coerulea
(Nagasaki)
Gyrodinium spirale
(Nagasaki)
Katodinium glaucum
(Omura Bay, Ariake Sound)
Polykrikos cf. kofoidii
(Nagasaki)
Gymnodinium aureum
(Nagasaki)
Gyrodinium rubrum
(Nagasaki)
Gymnodinium cf. puniceum
(Nagasaki)
Nematodinium sp.
(Nagasaki)
Heterotrophic Species
Perkinsus sp. Perkinsus marinus
Noctiluca scintillans Spatulodinium pseudonoctiluca Spatulodinium pseudonoctiluca
Phaeobalechina catenata Phaeobalechina catenata
Balechina coerulea Balechina coerulea
Gyrodinium dorsum Gyrodinium uncatenum Gyrodinium instriatum
Alexandrium tamarense Alexandrium minutum
Pyrocystis noctiluca Pyrocystis lunula
Symbiodinium corculorum Symbiodinium microadriaticum
Gymnodinium gracile Gymnodinium gracile Gymnodinium gracile
Achradina pulchra Achradina pulchra Achradina pulchra Gymnodinium sp. Gymnodinium sp. Gyrodinium galatheanum Gymnodinium galatheanum Karlodinium micrum Takayama sp.
Gymnodinium breve Karenia brevis Karenia mikimotoi Karenia mikimotoi
Amphidinium carterae Amphidinium massartii
Akashiwo sanguinea Akashiwo sanguinea
Gymnodinium cf. puniceum Erythropsidinium agile
Gyrodinium sp. Polykrikos kofoidii
Pheopolykrikos hartmannii Gymnodinium fuscum
Nematodinium sp. Nematodinium sp. Nematodinium sp. Gyrodinium impudicum
Dissodinium pseudolunula Lepidodinium viride Gymnodinium chlorophorum Gymnodinium catenatum Gymnodinium catenatum
Gymnodinium nolleri Gymnodinium microreticulatum Gymnodinium microreticulatum
Gyrodinium falcatum Gyrodinium falcatum Gyrodinium falcatum Gymnodinium sp. Cochlodinium convolutum Cochlodinium convolutum Gymnodinium sp. Gyrodinium spirale Gyrodinium fusiforme Gyrodinium spirale Gyrodinium sp. Gyrodinium rubrum Gymnodinium rubrum Gyrodinium aureum Gymnodinium helveticum
Katodinium glaucum Katodinium glaucum
Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides Cochlodinium polykrikoides
Cochlodinium sp. Cochlodinium sp.
Torodinium teredo
Gamma corrected NJ tree, evolutionary model: Tamura & Nei (1993),
Proportion of invariable site: 0.2608, Shape parameter: 0.6499
N. scintillans
C. polykrikoides + C. fulvescens
Karenia + Karlodinium + Takayama
Gyrodinium sensu stricto
C. convolutum + G. falcatum + Gymnodinium spp.
Spatulodinium pseudonoctiluca
Erythropsidinium + Nematodinium + Polykrikos etc.
Gymnodinium sensu stricto + Lepidodinium
Phylogenetic Tree Based on SSU rDNA Sequences
Levanderina fissa
(Isahaya Bay)
Akashiwo sanguinea
Gymnodinium sp.
(East China Sea)
Torodinium teredo
(Nagasaki)
Katodinium glaucum
(Omura Bay, Ariake Sound)
Balechina coerulea
(Nagasaki)
Balechina sp.
(Omura Bay)
Gymnodinium gracile
(Nagasaki, Omura Bay)
Achradina pulchra
(East China Sea)
Related Species Unknown
88 Gymnodinium cf. puniceum Nagasaki
Lepidodinium viride
Erythropsidinium agile Nagasaki
Gyrodinium sp.
Gymnodinium cf. chlorophorum Tokyo Bay
Gymnodinium catenatum Philippines
Gymnodinium catenatum
Gymnodinium cf. nolleri Hiroshima Bay
Gymnodinium microreticulatum Sasebo Bay
Gymnodinium microreticulatum Obama Bay
Gymnodinium impudicum
Dissodinium pseudolunula Omura Bay
Nematodinium sp. Nagasaki
Nematodinium sp. Ariake Sound
Nematodinium sp. East China Sea
Polykrikos cf. kofoidii Nagasaki
Polykrikos hartmannii
Gymnodinium fuscum
100
100
80
100
100 100
95
- Heterotrophic species possessing nematocyst, ocellus eyespot and piston are
related to Gymnodinium sensu stricto.
Gymnodinium sensu stricto
77
100
90
100
100
Gyrodinium spirale
Gyrodinium fusiforme
Gyrodinium spirale
(Nagasaki)
Gyrodinium sp.
(East China Sea)
Gyrodinium cf. lineatum
(Nagasaki)
100
Gyrodinium helveticum
Gyrodinium cf. nasutum
(Nagasaki)
Gyrodinium rubrum
100
- Gyrodinium species characterized by possessing an elliptical apical groove and
cell surface striations are monophyletic.
Gyrodinium sensu stricto
98
Karlodinium micrum
(as Gyrodinium galatheanum)
Karlodinium sp. Ariake Sound
Takayama sp. Omura Bay
Karenia mikimotoi
（as Gymnodinium mikimotoi）
59
Karenia mikimotoi
(as Gyrodinium aureolum)
Karenia brevis
（as Gymnodinium breve）
Karlodinium micrum
(as Gymnodinium galatheanum)
Karenia brevis
（as Gymnodinium breve）
72
100
93
95
- Apical groove is strait or sigmoid, containing chloroplast derived from a haptophyte.
- Relationship between Karenia and Karlodinium/Takayama is not highly supported in
phylogenetic tree SSU rDNA.
Karenia/Karlodinium/Takayama
<50
100
100
Noctiluca scintillans
Spatulodinium pseudonoctiluca
(Omura Bay)
Spatulodinium pseudonoctiluca
(Omura Bay)
- Noctiluca scintillans and Spatulodinium pseudonoctiluca formed a monophyletic
group, which branched out the basal position of all dinoflagellates.
- Two forms of Spatulodinium pseudonoctiluca, Noctiluca-form (S. pseudonoctiluca)
and Gymnodinium-form (Gymnodinium lebourae) are the same species.
Noctiluca scintillans/Spatulodinium pseudonoctiluca
77
87
71
98
79 100
Ceratoperidinium falcatum
(Tsushima Is)
Ceratoperidinium falcatum
(Inokushi Bay)
Ceratoperidinium falcatum
(Indonesia)
Gymnodinium sp.
(Ariake Sound)
Cochlodinium convolutum
(Omura Bay)
Cochlodinium convolutum
(Omura Bay)
Gymnodinium sp.
(Yatsushiro Bay)
Consisted of Cochlodinium convolutum, Gyrodinium falcatum and Gymnodinium spp.
Photosynthetic, with a circle shaped apical groove (?) and forming hyaline cyst.
The clade related to Ceratoperidinium
Recent our works on taxonomy and phylogeny
of small species including
unarmored, woloszynskioid and
armored dinoflagellates
Small dinoflagellates occurred in the Western Pacific
Amphidinium
Woloszynskioids
Armored dinoflagellates
Amphidinium carterae
Amphidinium cupulatisquama
Scale bars = 10 µm (LM), 100 nm (TEM)
Amphidinium mootonorum
Amphidinium massarti
Amphidinium gibbosum
Identified Amphidinium spp.
Species Locality Length
（μm） Cell shape Chloroplast
Other
characters
A. carterae
Turtle Cove,
Palau 12.0-18.0 Ellipsoid Membranous
Ngeruktabel,
Palau 12.5-22.5 Ellipsoid Membranous
A. cupulatisquama
Tsuruoka,
Japan 32.5-45.0 Ellipsoid Rod-like
Eye spot,
Body scale
Tsuruoka,
Japan 23.0-37.0 Ellipsoid Rod-like
Eye spot,
Body scale
A. gibbosum
Pari Island,
Indonesia 25.0-37.5
Asymmetrical,
Heart-shaped Rod-like
Turtle Cove,
Palau 18.0-32.0
Asymmetrical,
Heart-shaped Rod-like
A. massartii Okinawa,
Japan 13.5-19.5 Ellipsoid Rod-like
A. mootonorum Yamagata,
Japan 32.5-50.0 Ellipsoid Rod-like
Nucleus located
in the middle
Cell shape, chloroplasts, presence of eyespot and body scales,
and position of nucleus are recognized as reliable character for
species identification of some species.
Identified Amphidinium spp.
Species Length（µm） Shape Pyrenoid Chloroplast Pusule Immotile cell
A. steinii 20-38 ellipsoid spherical rod-like 2 present
A. trulla 18-31 ellipsoid spherical rod-like 0 absent
(Murray et al. 2004)
A. operculatum
A. carteriae A. massarti
A. gibbosum A. steinii A. trulla
Species in middle cell size range are
difficult to identify…
Difficulty of Amphidinium identification
Amphidinium sp. 1 (Indonesia) Amphidinium sp. 2 (Japan)
Amphidinium sp. 3 (Australia) Amphidinium sp. 4 (Japan)
Amphidinium sp. 5 (Vietnam) Amphidinium sp. 6 (Malaysia)
Scale bars = 10 µm Unidentified Amphidinium spp.
Locality Length (µm) Cell shape Pyrenoid Chloroplast Pusule Immotile cell
Amphidinium
sp. 1
Pari Island, Indonesia 17.5-27.5 ellipsoid 1 rod-like - absent
Pari Island, Indonesia 15.0-25.0 ellipsoid 1 rod-like - present
Pari Island, Indonesia 17.5-30.0 ellipsoid 1 rod-like - -
Amphidinium
sp. 2
Sendai, Japan 17.5-27.5 ellipsoid 1 rod-like - present
Sendai, Japan 17.5-27.5 ellipsoid 1 rod-like - -
Amphidinium
sp. 3 Melbourne, Australia 17.5-32.5 ellipsoid 1 reticulate - -
Amphidinium
sp. 4
Tsuruoka, Japan 17.5-30.0 ellipsoid 1 rod-like - -
Tsuruoka, Japan 17.5-32.5 ellipsoid 1 rod-like - -
Tsuruoka, Japan 17.5-28.0 ellipsoid 1 rod-like 2 absent
Amphidinium sp. 5
Nha Trang, Vietnam 20.0-35.5 ellipsoid >10 rod-like 2 absent
Nha Trang, Vietnam 17.0-31.0 ellipsoid 1-4 rod-like 2 absent
Amphidinium sp. 6
Port Dickson, Malaysia 15.0-25.0 ellipsoid 1 reticulate 1 - 2 present
Port Dickson, Malaysia 13.0-23.0 ellipsoid 1 reticulate 2 present
Port Dickson, Malaysia 14.0-22.0 ellipsoid 1 reticulate 2 present
Unidentified Amphidinium spp.
Amphidinium sp. 1 (Indonesia)
Amphidinium sp. 2 (Japan)
Amphidinium sp. 3 (Australia)
Amphidinium sp. 4 (Japan)
Amphidinium sp. 5 (Vietnam)
Amphidinium sp. 6 (Malaysia)
A. carterae
A. massartii
A. gibbosum
Not related
Not related
A. trulla
A. thermaeum
A. herdmanii / mootonorum
A. steinii
Gamma corrected maximum likelihood (ML) tree,
evolutionary model: GTR+I+G
Bootstrap support values: ML/MP/NJ,
>50%, ●=100%
Molecular phylogeny based on LSU rDNA
Distribution of Amphidinium species in the East and Southeast Asia
Amphidinium carterae
Amphidinium cupulatisquama
Amphidinium gibbosum
Amphidinium massartii
Amphidinium mootonorum
Amphidinium thermaeum
Amphidinium sp. 2
Amphidinium sp. 6
A. thermaeum is widely distributed in East and SE Asia.
Distribution of Amphidinium in East and Southeast Asia
unarmored
(athecate, naked)
armored
(thecate) Woloszynskioids
(Taylor 1987) Amphiesmal vesicle
thecal
plates
Woloszynskioid dinoflagellates
Biecheleriopsis adriatica (Japan and Palau)
E1
E2
E3
E4
2μm
Original （Moestrup et al. 2009b） Present study
10μm
Biecheleriopsis adriatica
Original （Siano et al. 2009） Present study
10μm
2μm
1-1 1-2
1-3
1-4 1-5
1-6
2-2
Pelagodinium bei (Japan and Vietnam)
Pelagodinium bei
10μm
2μm
Biecheleria brevisulcata (Japan)
Biecheleria brevisulcata Takahashi et Iwataki 2014
Peridinioid sp.1 Peridinioid sp.2
・7 cultures of small armored dinoflagellates from
the western Pacific Peridinioid sp.1
From Palau, Guam, Philippines, Malaysia,
Peridinioid sp.2
From Indonesia, Philippines
Small armored dinoflagellates
Location of organelles change in motile and immotile
Motile cell Immotile cell
Mushroom-shaped Outline Mushroom-shaped
Posteriorly Nucleus (N) Anteriorly
Posteriorly Eyespot (E) Anteriorly
Anteriorly Pyrenoid
(Py) Posteriorly
17.1 μm (14.0–21.5)
Cell length n=31
17.5 μm (15.0–20.0)
N
E
Py
N
Py
E
Scale bar=10 μm
N
Peridinioid sp.1
2μm
Thecal plates: Po,cp,4’,3a,7”,6c,5’’’,2””
A conspicuous sulcal list in the sulcus
Scale bars=2 μm (SEM), 10 μm (LM)
Peridinioid sp.1
Motile cell Immotile cell
Ellipsoidal, conical epitheca Outline Spherical
Posterior Nucleus
(N) Central
Posterior Eyespot
(E) Peripheral
Anterior Pyrenoid
(Py) Central
Absent Red body Central
17.5 μm (16.5‐19.0)
Cell length n=31
18.7 μm (12.5‐28.8)
Scale bars=10 μm
Different cell shape in motile and immotile cells
N
Py
E
N E
Peridinioid sp.2
Scale bar=10μm
Thecal plates: Po,cp,4’,2a,6”,6c,5’’’,2””
Two anterior intercalary plate (2a)
(Peridinioids have 3a typically)
Peridinioid sp.2
ML Tree（D1-D3, 579 bp）
BS values>50%: ML/NJ
Peridinioid sp.1
Peridinioid sp.2
Peridinioid sp.1
formed a monophyletic group
related species was uncertain
Peridinioid sp.2
formed a monophyletic group
related to Peridiniopsis borgei,
Scrippsiella hexapraecingula
Phylogenetic tree
Summary
New genera have being newly proposed based on the phylogenetic
position and combination of morphological characters different from
those of previouly reported genera.
Levanderina fissa (Levander) Moestrup et al. 2014
= Gyrodinium instriatum
= Gyrodinium uncatenum
= Gyrodinium fissum
Many of small undescribed dinoflagellates are commonly found in the
Western Pacific, these species can be be distinguished from unarmored
dinoflagellates,
- Woloszynskioid: an eyespot (difficult to observe by light microscopy),
sigmoid sulcus in the ventral
- Small peridinioids: thecal plates and an eyespot (difficult to observe by
light microscopy), mushroom-shaped or coccoid immotile stage.
