Representatives of the marine dinophyte family Amphidomataceae produce lipophilic phycotoxins called azaspiracids (AZA) which may cause azaspiracid shellfish poisoning (AZP) in humans after consumption of contaminated seafood. Three of the four known toxigenic species are observed frequently in the eastern North Atlantic. In 2018, a research survey was performed to strengthen knowledge on the distribution and abundance of toxigenic Amphidomataceae and their respective toxins in Irish coastal waters and in the North Sea. Species-specific quantification of the three toxigenic species (Azadinium spinosum, Azadinium poporum and Amphidoma languida) was based on recently developed qPCR assays, whose performance was successfully validated and tested with specificity tests and spike experiments. The multi-method approach of on-board live microscopy, qPCR assays and chemical AZA-analysis revealed the presence of Amphidomataceae in the North Atlantic including the three targeted toxigenic species and their respective AZA analogues (AZA-1,-2,-33,-38,-39). Azadinium spinosum was detected at the majority of Irish stations with a peak density of 8.3 x 10 4 cells L-1 and AZA (AZA-1,-2,-33) abundances up to 1,274 pg L-1. Amphidoma languida was also present at most Irish stations but appeared in highest abundance in a bloom at a central North Sea station with a density of 1.2 x 10 5 cells L-1 and an AZA (AZA-38,-39) abundances of 618 pg L-1. Azadinium poporum was detected sporadically at the Irish south coast and North Sea and was rather low in abundance during this study. The results confirmed the wide distribution and frequent occurrence of the target species in the North Atlantic area and revealed, for the first time, bloom abundances of toxigenic Amphidomataceae in this area. This emphasizes the importance of future studies and monitoring of amphidomatacean species and their respective AZA analogues in the North Atlantic.
Azaspiracids (AZA) are lipophilic marine biotoxins associated with shellfish poisoning which are produced by some species of Amphidomataceae. Diversity and global biogeography of this family are still poorly known. In summer 2017 plankton samples were collected from the central Labrador Sea and western Greenland coast from 64°N (Gothaab Fjord) to 75°N for the presence of Amphidomataceae and AZA. In the central Labrador Sea, light microscopy revealed small Azadinium-like cells (9200 cells l −1 ). Clonal strains established from plankton samples and scanning electron microscopy of fixed plankton samples revealed at least eight species of Amphidomataceae: Azadinium obesum, Az. trinitatum, Az. dexteroporum, Az. spinosum, Az. polongum, Amphidoma languida, Azadinium spec., and a new species described here as Azadinium perforatum sp. nov. The new species differed from other Azadinium species by the presence of thecal pores on the pore plate. All samples, including cultured strains, filtered seawater samples, and solid phase adsorption toxin tracking (SPATT) samplers deployed during the expedition in a continuous water-sampling system (FerryBox), were negative for AZA. DNA samples and PCR assays were positive for Amphidomataceae from most stations, whereas species-specific assays for three toxigenic species were rarely positive (two stations for Az. poporum, one station for Am. languida). The results highlight the presence of Amphidomataceae in the area but the lack of toxins and low abundance of toxigenic species currently indicate a low risk of toxic Amphidomataceae blooms in Arctic coastal waters.
SUMMARY Prorocentrum comprises a unique group of dinophytes with several apomorphic traits, such as an apical insertion of flagella and the presence of two major, large thecal plates. Species delimitation is challenging, especially for morphologically very similar, small planktonic species. Contemporary analyses, including SEM studies and molecular phylogenetics of type material, are not available for many described species. Based on six strains isolated from Antarctic, subarctic and North Atlantic waters, Prorocentrum pervagatum sp. nov. is described. Prorocentrum pervagatum was small (12–16 μm long and deep), oval to round in outline, and moderately compressed. One small, pyrenoid‐like structure was faintly visible in some cells. Rod‐like, long trichocysts were present. Cells had one distinct apical spine (1.1–1.7 μm in length) visible in light microscopy. The plate surface appeared smooth in light microscopy with few pores located close to the plate margin visible in empty thecae. Electron microscopy revealed plates to be densely covered by small projections and two size classes of thecal pores. Cells had a row of mostly four large pores in apical‐ventral position on the right thecal plate. The periflagellar area consisted of eight small platelets. The apical spine was formed by platelet six. In molecular phylogenetics, P. pervagatum was part of a species group generally exhibiting small size and spiny thecal ornamentation, together with Prorocentrum cordatum and Prorocentrum obtusidens. The new species is distinct in DNA trees and differs from the protologues of other small species of Prorocentrum by the unique combination of size, shape (i.e. only moderately compressed or round), presence of a distinct apical spine, and position of thecal pores (i.e. located at the plate margins only). Its clear description and illustration may stimulate similar work of other small species of Prorocentrum, particularly including the re‐investigation of taxa with historical names collected at the corresponding type localities.
Species of the planktonic dinoflagellates Azadinium and Amphidoma are small, inconspicuous and difficult, if not impossible to be identified and differentiated by light microscopy. Within this group, there are some species that produce the marine biotoxin azaspiracid (AZA) while others are non-toxigenic, therefore a requirement exists for precise species identification. A quantitative polymerase chain reaction (qPCR) assay for molecular detection and quantification of one of the toxigenic species, Amphidoma languida, was designed and extensively tested. The assay was highly specific and sensitive to detect and quantify down to 10 target gene copies (corresponding to ca. 0.05 cells) per reaction. DNA cell quota and copy number cell −1 were constant for four different Am. languida strains, and for one strain they were shown to be stable at various time points throughout the growth cycle. Recovery of known cell numbers of Am. languida spiked into natural samples was 95-103%, and the assay was successfully tested on field samples collected from Irish coastal waters. This new qPCR assay is a valuable tool for routine monitoring for the prevention of AZA-shellfish-poisoning caused by the consumption of contaminated shellfish and is a supportive tool for studies on the biogeography of this AZA-producing species.
Species of the dinophyte genus Alexandrium are widely distributed and are notorious bloom formers and producers of various potent phycotoxins. The species Alexandrium taylorii is known to form recurrent and dense blooms in the Mediterranean, but its toxin production potential is poorly studied. Here we investigated toxin production potential of a Mediterranean A. taylorii clonal strain by combining state-of-the-art screening for various toxins known to be produced within Alexandrium with a sound morphological and molecular designation of the studied strain. As shown by a detailed thecal plate analysis, morphology of the A. taylorii strain AY7T from the Adriatic Sea conformed with the original species description. Moreover, newly obtained Large Subunit (LSU) and Internal Transcribed Spacers (ITS) rDNA sequences perfectly matched with the majority of other Mediterranean A. taylorii strains from the databases. Based on both ion pair chromatography coupled to post-column derivatization and fluorescence detection (LC-FLD) and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis it is shown that A. taylorii AY7T does not produce paralytic shellfish toxins (PST) above a detection limit of ca. 1 fg cell−1, and also lacks any traces of spirolides and gymnodimines. The strain caused cell lysis of protistan species due to poorly characterized lytic compounds, with a density of 185 cells mL−1 causing 50% cell lysis of cryptophyte bioassay target cells (EC50). As shown here for the first time A. taylorii AY7T produced goniodomin A (GDA) at a cellular level of 11.7 pg cell−1. This first report of goniodomin (GD) production of A. taylorii supports the close evolutionary relationship of A. taylorii to other identified GD-producing Alexandrium species. As GD have been causatively linked to fish kills, future studies of Mediterranean A. taylorii blooms should include analysis of GD and should draw attention to potential links to fish kills or other environmental damage.
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