New morphological data and molecular phylogeny of the benthic dinoflagellate Pseudothecadinium campbellii (Dinophyceae, Gonyaulacales)

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Abstract

Pseudothecadinium campbellii, a phototrophic, thecate, marine benthic species, has been found in the Sea of Japan, Russia. The morphological description of the species has been emended, and the thecal tabulation pattern is now APC 4′ 2a 4″ 6c 6 s 5‴ 1⁗. Our study indicates that P. campbellii is related to Thecadinium kofoidii and Thecadiniopsis tasmanica, based on a unique morphological feature: incomplete precingular plate series. Previously, molecular data was not available for the taxa, and thus the phylogenetic position of P. campbellii within the Dinophyceae remained obscure. In this study, analyses of the rRNA gene sequences (partial 18S and 28S) revealed that unexpectedly, P. campbellii is most closely related to Halostylodinium arenarium. It formed a common clade with the Thecadinium sensu stricto clade comprising T. kofoidii and T. pseudokofoidii. This clade was placed within the order Gonyaulacales. However, almost no similarity in morphology was found between the two genetically closest species. In addition, they have different lifestyles: unlike P. campbellii, the nonmotile stage is dominant in the life cycle of H. arenarium. It has been shown that other genetically similar species (T. kofoidii, T. pseudokofoidii, T. yashimaense) exhibited some morphological features that unite them with gonyaulacoids.

Introduction

The monotypic genus Pseudothecadinium Hoppenrath and Selina, 2006, with P. campbellii as type species, was described from plankton of shallow-water from the Lunskiy Bay in the Sea of Okhotsk (Hoppenrath and Selina 2006). Earlier, this species has been recorded from plankton of the estuarine region of Gales Greek, North Carolina, under the invalidly published name Thecadinium aureum (nomen nudum; Campbell 1973) and from the Gulf of Mexico as ‘unidentified thecate #2’ (Steidinger and Williams 1970). Pseudothecadinium campbellii showed some similarity in cell morphology (incomplete precingular plate series) to Thecadinium kofoidii (Herdman) Larsen and Thecadiniopsis tasmanica Croome, Hallegraeff and Tyler, and hence, together with the latter species, it was tentatively assigned to the family Thecadiniaceae of the order Gonyaulacales (Hoppenrath and Selina 2006). Apart from morphological differences, the above species differed in their ecology: Thecadiniopsis tasmanica is a freshwater planktonic species, while T. kofoidii is marine sand-dwelling (Croome et al. 1987; Hoppenrath 2000). Pseudothecadinium campbellii was first recorded in coastal plankton (Campbell 1973; Steidinger and Williams 1970) and described from plankton of a shallow lagoon (Hoppenrath and Selina 2006). Long-term studies of benthic dinoflagellates in Peter the Great Bay, Sea of Japan, showed that P. campbellii is a permanent component of the epiphytic community and sometimes reaches high abundances in sublittoral sands (Selina and Levchenko 2011; Selina et al. 2014).

In the classification by Fensome et al. (1993), T. kofoidii and Thecadiniopsis tasmanica were placed in the order Gonyaulacales, but family uncertain. Later, the three genera were separated into a distinct group, the Thecadinioide (Hoppenrath 2017). The phylogenetic relationships and position within the group Thecadinioide remain obscure. In Adl et al. (2019) these genera were classified as “incertae sedis Dinoflagellate”. However, so far, no molecular data are available for Pseudothecadinium. In this study, we determined and analyzed the small subunit ribosomal RNA gene sequences (18S rRNA), and the D1/D2 and D8/D10 regions of the large subunit (28S rRNA) of P. campbellii. Data on the morphology of the apical pore complex, cingular and sulcal areas of P. campbellii were specified and added.

Section snippets

Cell isolation and culturing

Investigations were carried out using clonal cultures of Pseudothecadinium campbellii (strains MBRU_Ps1, MBRU_Ps2, MBRU_Ps3) and field samples. Cells of P. campbellii for establishment of cultures were isolated from the green alga Codium fragile (Suringar) Hariot that was collected on 27 November 2014 in the Bosfor Vostochny Strait (43°04′24″N, 131°57′34″E) of Peter the Great Bay, Sea of Japan at about 1 m depth. In addition, field samples from epiphytic assemblage at same station on 27

Morphology of Russian population of Pseudothecadinium campbellii (Figs. 1–3)

Cells laterally flattened, oval or obovate in lateral view (Figs. 1, 2a–c), 33.6–54.5 μm (43.4 ± 5.3, n = 50) long, 26.1–45.8 μm (34.6 ± 4.4, n = 50) deep, with a length/depth ratio of 1.1–1.4 (1.25 ± 0.06, n = 50). Nucleus oval, at dorsal side of hypotheca (Fig. 1a, b). Chloroplasts numerous, yellow-brown, radiating from cell center. In some cells small rounded greenish granules located in parietal part of epitheca and hypotheca (Fig. 1a). Left part of epitheca almost two times smaller

Discussion

Light and scanning electron microscopic observations of cultured cells revealed all species-specific characteristics of Pseudothecadinium campbellii: (1) incomplete precingular plate series, (2) apical intercalary plates separated from each other, and (3) ventral intercalary plate with a pore (Hoppenrath and Selina 2006). One more characteristic feature was specified — the antapical plate was not entirely on the right lateral side of the hypotheca, a small part continued onto the left side. The

Author contributions

K.V.E. and M.S.S. conceptualised the project; M.S.S. collected samples; K.V.E. and M.S.S. carried out the research; K.V.E. performed molecular-phylogenetic analysis; M.S.S. performed electron microscopy analysis; K.V.E., M.S.S. and M.H. wrote the manuscript. All authors read and approved the final manuscript.

Funding

Financial support was provided by the Dalnii Vostok program (project 18-4-050); and Ministry of Science and High Education Program for support of bioresource collections for maintenance of microalgae cultures.

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