Abstract
The genus Crypthecodinium (Dinophyceae) currently consists of only one species: C. cohnii, a heterotrophic marine dinoflagellate widely known able to produce prolific amount of DHA. However, previous studies revealed that there are morphological and genetic differences among Crypthedodinium cohnii-like strains, indicating the potential of undiscovered diversity of this dinoflagellate. Attempts of isolating heterotrophic marine dinoflagellate strains were made from submerged mangrove leaves and seaweed. A total of nine strains were established, cultured and maintained using seawater-based GTY medium for taxonomic identification and phylogenetic analyses based on SSU, ITS 1, 5.8S, ITS2 and LSU rDNA. Morphological observation revealed all strains shared similar morphology, e.g.: motile cells were delicately thin armored with single or double flagella surrounding the cingulum and protruding along the sulcus posteriorly, while non-motile cell stage (cyst) may retain more than two daughter cells inside the transparent and thick cell wall. Both cell stages varied in sizes and shapes even within the same clonal culture. Numerous lipid granules can be observed in the cytoplasm. Two types of cingulum encirclement were observed: partially or completely surrounding the cell body. All strains indicated close relatedness (>95% sequence similarity) with previously reported Cryphtecodinium cohnii strains, except for C. cohnii CAAE-CL2 (87.2-87.4% similarity) in SSU and C. cohnii CCMP316 (79.9-80.4% similarity) in ITS. Moreover, molecular phylogenetic tree grouped the Okinawan isolates into two genetically distinct clades with high similarity (>98%) among members of each clade in SSU and LSU but only shared 80.5% similarity in ITS rDNA. The results of this study showed evidences of undiscovered diversity in the heterotrophic dinoflagellates currently regarded as Crypthecodinium.