Using RDNA sequences to define dinoflagellate species

Abstract

Dinoflagellate species are traditionally defined using morphological characters, but molecular evidence accumulated over the past several decades indicates many morphologically-based descriptions are inaccurate. This recognition led to an increasing reliance on DNA sequence data, particularly rDNA gene segments, in defining species. The validity of this approach assumes the divergence in rDNA or other selected genes parallels speciation events. Another concern is whether single gene rDNA phylogenies by themselves are adequate for delineating species or if multigene phylogenies are required instead. Currently, few studies have directly assessed the relative utility of multigene versus rDNA-based phylogenies for distinguishing species. To address this, the current study examined D1-D3 and ITS/5.8S rDNA gene regions, a multi-gene phylogeny, and morphological characters in Gambierdiscus and other related dinoflagellate genera to determine if they produce congruent phylogenies and identify the same species. Data for the analyses were obtained from previous sequencing efforts and publicly available dinoflagellate transcriptomic libraries as well from the additional nine well-characterized Gambierdiscus species transcriptomic libraries generated in this study. The D1-D3 and ITS/5.8S phylogenies successfully identified the described Gambierdiscus and Alexandrium species. Additionally, the data showed that the D1-D3 and multigene phylogenies were equally capable of identifying the same species. The multigene phylogenies, however, showed different relationships among species and are likely to prove more accurate at determining phylogenetic relationships above the species level. These data indicated that D1-D3 and ITS/5.8S rDNA region phylogenies are generally successful for identifying species of Gambierdiscus, and likely those of other dinoflagellates. To assess how broadly general this finding is likely to be, rDNA molecular phylogenies from over 473 manuscripts representing 232 genera and 863 described species of dinoflagellates were reviewed. Results showed the D1-D3 rDNA and ITS phylogenies in combination are capable of identifying 97% of dinoflagellate species including all the species belonging to the genera Alexandrium, Ostreopsis and Gambierdiscus, although it should be noted that multi-gene phylogenies are preferred for inferring relationships among these species. A protocol is presented for determining when D1-D3, confirmed by ITS/5.8S rDNA sequence data, would take precedence over morphological features when describing new dinoflagellate species. This protocol addresses situations such as: a) when a new species is both morphologically and molecularly distinct from other known species; b) when a new species and closely related species are morphologically indistinguishable, but genetically distinct; and c) how to handle potentially cryptic species and cases where morphotypes are clearly distinct but have the same rDNA sequence. The protocol also addresses other molecular, morphological, and genetic approaches required to resolve species boundaries in the small minority of species where the D1-D3/ITS region phylogenies fail.

Oleaginous Heterotrophic Dinoflagellates—Crypthecodiniaceae

“…Despite its importance in the certification for industrial licensing, there has been relatively little study of inter- and intra-specific variations between different OTUs of the genus. Ribotypes are well recognized to trace dinoflagellate evolution, with ITS region (ITS1-5.8S-ITS2) successfully tested to demarcate species between morphological similar taxons [ 40 , 41 , 42 , 43 ].…”

“…We report here phylogenetic analysis of Crypthecodiniaceae based on three ribosomal markers [ 39 , 40 ]. Cumulated with morphological characters, thecal plate formulas, AFLP and three molecular markers, as well as genome size differences, we report Crypthecodinium croucheri sp.…”

Genomic copy number variability at the genus, species and population levels impacts in situ ecological analyses of dinoflagellates and harmful algal blooms

“…Given the ubiquity of microbial eukaryotic ecological analyses routinely targeting rRNA genes for qPCR and meta-barcoding, it can appear as though such approaches are very well characterised [ 9 , 10 , 13 , 17 , 20 , 56 , 59 , 71 ]. Considerable and justifiable attention has focused on sources of bias that confound universal rRNA gene markers, particularly PCR primer bias, sequencing bias and statistical bias [ 3 , 72 , 73 ].…”

Gambierone and Sodium Channel Specific Bioactivity Are Associated with the Extracellular Metabolite Pool of the Marine Dinoflagellate Coolia palmyrensis

“…The isolate was maintained in a laboratory culture (see methods) and confirmed by 28S rRNA gene sequence analysis as C. palmyrensis (GenBank Accession # OP392325). The LSU D1-D2 was used to determine the species of our strain, as is common for benthic dinoflagellate identification [37]. Our sequence had 100% coverage in existing entries for C. palmyrensis and 98.61% identity with prior database entries, e.g., GenBank Accession # MT295362.1.…”