Gonyaulax species (Dinophyceae) from Korean coastal waters: Morphology, phylogeny, and growth responses to changes in combinations of temperature and salinity

Abstract

In Korean coastal waters, morphological and molecular phylogeny of Gonyaulax species have beern rarely reported. In addition, the growth responses to changes in combinations of temperature and salinity was not reported. To investigate Gonyaulax species from Korean coastal waters, plankton samples were collected, and Gonyaulax species ware isolated from the samples and its cultures were successfully established. Morphological features of Gonyaulax species were examined using light and fluorescence microscopy, and scanning electron microscopy, and molecular phylogeny was analyzed. Based on obtained results, G. baltica, G. cf. digitale, G. cf. spinifera, G. hyalina, two clades of G. polygramma and G. whaseongensis are recorded, and four new Gonyaulax species, Gonyaulax sp. 1, Gonyaulax sp. 2, Gonyaulax sp. 3 and Gonyaulax sp. 4 are proposed in this study. Gonyaulax species identified in this study were divided two groups, based on the species characterized by dextral torsion (Group I) or neutral tortion (Group II). Group I had the plate formula of 2 pr, 4ʹ, (1a), 6ʹʹ, 6C, 5s, 6ʹʹʹ, 1p, 1ʹʹʹʹ, with S-type ventral organization, and the relative positions of the 3''/4'' and 4'''/5''' boundaries where 3''/4'' is either shifted to the right (dextral torsion), whereas Group II had the plate formula of 2 pr, 4ʹ, 6ʹʹ, 6C, 5s, 6ʹʹʹ, 1p, 1ʹʹʹʹ, with S-type ventral organization, and the relative positions of the 3''/4'' and 4'''/5''' boundaries where 3''/4'' is aligned (neutral torsion). G. hyalina, G. polygramma, G. whaseongensis and Gonyaulax sp. 2 belong to Group I, and G. batica, G. cf. digitale, G. cf. spinifera, Gonyaulax sp. 1, Gonyaulax sp. 3 and Gonaulax sp. 4 are assigned to Group II. G. baltica was characterized variably recticulated ornamentation on plates, short apical horn with slight shoulders, a rounded hypotheca, 2 cingulum width and short acuminate antapical spines. G. cf. digitale was characterized equilateral triangle shape of the 6ʹʹ plate, distinctive shape of the apical horn, idented shape of apical plate series. G. cf. spinifera was characterized vyeelow, symmetrical, with flagellum, singulum and sulcus concave and cingulum overhang and the centrally located nucleus. G. hyalina was characterized almost symmetrical in transversal axes, and to very delicate membrane and marked longitudinal striae with small ridge between them, G. hyalina appears as a rugged topographic map with many close contour lines. G. polygramma was characterized by remarkable apical horn and shoulder, longer vertically than horizontally, spine prominent on the left side of the cell and vertical stripes on the surface. In Korean specime of G. polygramma, the obsence and presenc of the 1a plate was found. G. polygramma was divided into two clades in the phylogenetic tree. G. whaseongensis was characterized small displacement of the cingulum, slight overhang and steep angle between the ends of the cungulum, and one or two antalical spines. In Korean specimen, two posterioi intercalary plate was occasionally observed. Gonyaulax sp. 1 was characterized by numerous minute antapical spines, large displacement of cingulum, and reticulated cell surface. Gonyaulax sp. 2 was distinguished by heavily recticulated plate pattern, having antapical spine or not, overhang of cingulum ends or not, displacement of cingulum, and square of sixth precingular plate. Gonyaulax sp. 3 was characterized by remarkable connected two antapical spines, small displacement of cingulum, and vermiculated cell surface. Gonyaulax sp. 4 was characterized by remarkable two antapical spines, large cingulum overhang, and randomly cuvated cell surface. These morphological characteristics allow us to distinguished Korean isolate from previously reported Gonyaulax species. The ML and BI based on SSU, ITS and LSU rDNA generated similar phylogenetic trees that differed at only a few topological features. The molecular tree based on concanated SSU-ITS-LSU rDNA formed a clade consisting of mainly Gonyaulax species, and those of genera Ataxiodinium, Spiniferites and Impagidinium. The Gonyaulax sp. 2 was clearly divergent from other Gonyaulax species, however clustered with G. whaseongensis, G. fragilis, G. hyalina and G. polygramma, with weak support (100/1 for SSU-ITS-LSU, 68/0.91 for SSU, 99/1 for ITS, 100/1 for LSU). The isolate of G. whaseongensis shared identical sequences with previously recorded G. whaseongensis (LS481152). G. hyalina was divergent from G. fragilis, with week support. G. polygramma was divided into two clades. Gonyaulax sp. 3 and Gonyaulax sp. 4 were clustered with G. cf. digitale, and G. cf. spinifera, however were clearly divergent from other Gonyaulax species. G. cf. digitale was divergent from G. digitale. Korean isolate of Gonyaulax cf. spinifera shared identical sequences of G. cf. spinifera and Spiniferites ramosus. Gonyaulax sp. 1 was clustered with G. baltica, and was divided into two clades. Identified Gonyaulax species were examined the specific growth rate under different water temperatures (5, 10, 15, 20, 25, 30℃) and salinity (15, 20, 25, 30, 35, 40). The maximum growth rates and maximum cell densities also differed depending on the species. Gonyaulax sp. 2 was only survived in 25℃, salinity level 25, 30. This result indicates Gonyaulax sp. 2 was stenothermal and stenohaline species. Gonyaulax baltica, Gonyaulax sp. 1, Gonyaulax sp. 3, Gonyaulax sp. 4, G. whaseongensis were eurythermal and euryhaline species, but the salinity of maximum growth rate was different. The species in two clades of Gonyaulax polygramma showed a similar growth response. In addition, maximum growth rates and maximum cell densities differed depending on the species. Therefore, growth responses to changes in combinations of temperature and salinity can be good data for further classification.