First Report of Paralytic Shellfish Toxins in Marine Invertebrates and Fish in Spain

A paralytic shellfish poisoning (PSP) episode developed in summer 2018 in the Rías Baixas (Galicia, NW Spain). The outbreak was associated with an unprecedentedly intense and long-lasting harmful algal bloom (HAB) (~one month) caused by the dinoflagellate Alexandrium minutum. Paralytic shellfish toxins (PSTs) were analyzed in extracts of 45 A. minutum strains isolated from the bloom by high-performance liquid chromatography with post-column oxidation and fluorescence detection (HPLC-PCOX-FLD). PSTs were also evaluated in tissues from marine fauna (invertebrates and fish) collected during the episode and in dolphin samples. The analysis of 45 A. minutum strains revealed a toxic profile including GTX1, GTX2, GTX3 and GTX4 toxins. With regard to the marine fauna samples, the highest PSTs levels were quantified in bivalve mollusks, but the toxins were also found in mullets, mackerels, starfish, squids and ascidians. This study reveals the potential accumulation of PSTs in marine invertebrates other than shellfish that could act as vectors in the trophic chain or pose a risk for human consumption. To our knowledge, this is the first time that PSTs are reported in ascidians and starfish from Spain. Moreover, it is the first time that evidence of PSTs in squids is described in Europe.En prens

Paretic syndrome in gulls from southern Portugal: searching for the causative agent

Between 2010 and 2019, 2.432 gulls (Larus michahellis and Larus fuscus) with paretic syndrome were received at RIAS Wildlife Rehabilitation and Research Centre. The clinical signs included weakness, anorexia, paralysis, diarrhoea (flaccid cloacae), dyspnoea and, in some cases, death. Several biotic contaminants are among the potential cause of this syndrome: marine biotoxins, Clostridium botulinum, cyanotoxins and virus. This presentation compiles the results of the Clostridium botulinum and marine biotoxins analysis conducted at the French National Reference Centre for anaerobic bacteria and botulism, Pasteur Institute (Paris) and the Vigo Centre of the Spanish Oceanographic Institute, respectively. C. botulinum analyses were conducted in livers and intestines from 5 gulls with paretic syndrome symptoms admitted at RIAS Wildlife Rehabilitation and Research Centre. Samples were pooled in two groups according to the tissue and analysed by targeted Real Time Polymerase Chain Reaction (PCR) on neurotoxin genes after sample enrichment culture under anaerobic conditions. The presence of botulinum toxin was confirmed by a lethality test on mice (mouse bioassay). Mice were intraperitoneally injected with filtered supernatant of the culture. Paralytic shellfish toxins (PSTs) were analysed by Liquid Chromatography with Fluorescence Detection and Post-column Oxidation in samples from ten gull kidneys and in the cloacae contents from another gull. Domoic acid (DA) analysis was conducted following a procedure that involved a methanolic extraction and analysis by Liquid Chromatography coupled to High Resolution Mass Spectrometry. DA was analysed in twenty three gull samples: ten livers, ten intestines and three cloacae contents. PSTs and DA were not detected in any of the samples tested. Results obtained so far point to C. botulimum type C/D as the causative agent of the paretic syndrome in gulls

Differences in the toxin profiles of Alexandrium ostenfeldii (Dinophyceae) strains isolated from different geographical origins: evidence of paralytic toxin, spirolide and gymnodimine

Among toxin-producing dinoflagellates of the genus Alexandrium, Alexandrium ostenfeldii is the only species able to produce paralytic shellfish poisoning (PSP) toxins, spirolides (SPXs) and gymnodimines (GYMs). In this study we characterized and compared three A. ostenfeldii strains isolated from the Baltic, Mediterranean, and southern Chile Seas with respect to their toxin profiles, morphology, and phylogeny. Toxin analyses by HPLCeFD and LCeHRMS revealed differences in the toxin profiles of the three strains. The PSP toxin profiles of the southern Chile and Baltic strains were largely the same and included gonyautoxin (GTX)-3, GTX-2, and saxitoxin (STX), although the total PSP toxin content of the Chilean strain (105.83 ± 72.15 pg cell 1) was much higher than that of the Baltic strain (4.04 ± 1.93 pg cell 1). However, the Baltic strain was the only strain that expressed detectable amounts of analogues of GYM-A and GYM-B/-C (48.27 ± 26.12 pg GYM-A equivalents cell 1). The only toxin expressed by the Mediterranean strain was 13-desmethyl SPX-C (13dMeC; 2.85 ± 4.76 pg cell 1). Phylogenetic analysis based on the LSU rRNA showed that the studied strains belonged to distinct molecular clades. The toxin profiles determined in this study provide further evidence of the taxonomic complexity of this species.Preprin

First record of the dinoflagellate Prorocentrum borbonicum in the continental coast of Colombian Caribbean: A new 42 hydroxi-palytoxin producer

The dinoflagellate genus Prorocentrum includes several harmful toxigenic species, predominantly benthic ones. In the past, fast-acting toxicity in mice has been observed in extracts of the epibenthic species P. borbonicum, with the finding of two compounds termed as borbotoxins-A and -B. The presence of palytoxin-like compounds was also suggested from electrophysiological experiments. In the present study, a strain of P. borbonicum was isolated in the continental coast of Colombian Caribbean, in seagrass beds of Thalassia testudinum in Bonito Gordo, Tayrona National Natural Park (PNNT). The morphological and molecular characteristics were consistent with the original and former descriptions for this species. Typical haemolytic activity due to palytoxin was confirmed in P. borbonicum extracts in presence of ouabain, the toxin contents being estimated as 1.9 pg palytoxin equivalents cell-1. HPLC-HRMS analyses of these extracts unambiguously identified the presence of borbotoxins and 42-hidroxy-palytoxin (42-OH-PLTX). This is the first report of palytoxin-like compounds in another dinoflagellate genus than Ostreopsis and the first record of P. borbonicum in Colombia and the Caribbean region.En prens

A Kinetic and Factorial Approach to Study the Effects of Temperature and Salinity on Growth and Toxin Production by the Dinoflagellate Alexandrium ostenfeldii from the Baltic Sea

19 páginas, 6 tablas, 4 figuras.-- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are creditedAlexandrium ostenfeldii is present in a wide variety of environments in coastal areas worldwide and is the only dinoflagellate known species that produces paralytic shellfish poisoning (PSP) toxins and two types of cyclic imines, spirolides (SPXs) and gymnodimines (GYMs). The increasing frequency of A. ostenfeldii blooms in the Baltic Sea has been attributed to the warming water in this region. To learn more about the optimal environmental conditions favoring the proliferation of A. ostenfeldii and its complex toxicity, the effects of temperature and salinity on the kinetics of both the growth and the net toxin production of this species were examined using a factorial design and a response-surface analysis (RSA). The results showed that the growth of Baltic A. ostenfeldii occurs over a wide range of temperatures and salinities (12.5–25.5°C and 5–21, respectively), with optimal growth conditions achieved at a temperature of 25.5°C and a salinity of 11.2. Together with the finding that a salinity > 21 was the only growth-limiting factor detected for this strain, this study provides important insights into the autecology and population distribution of this species in the Baltic Sea. The presence of PSP toxins, including gonyautoxin (GTX)-3, GTX-2, and saxitoxin (STX), and GYMs (GYM-A and GYM-B/-C analogues) was detected under all temperature and salinity conditions tested and in the majority of the cases was concomitant with both the exponential growth and stationary phases of the dinoflagellate’s growth cycle. Toxin concentrations were maximal at temperatures and salinities of 20.9°C and 17 for the GYM-A analogue and > 19°C and 15 for PSP toxins, respectively. The ecological implications of the optimal conditions for growth and toxin production of A. ostenfeldii in the Baltic Sea are discussedThis work is a contribution of the Unidad Asociada "Microalgas Nocivas" (CSIC-IEO) and was financially supported by the CCVIEO project and CICAN-2013-40671-R (Ministry of Economy and Competitiveness). P. Salgado is a researcher at IFOP, which has provided financial support for his doctoral stayPeer reviewe

19,19'-diacyloxy signature: an atypical level of structural evolution in carotenoid pigments.

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Interactive effects of temperature and salinity on the growth and cytotoxicity of the fish-killing microalgal species Heterosigma akashiwo and Pseudochattonella verruculosa

Fish-killing blooms of Heterosigma akashiwo and Pseudochattonella verruculosa have been devastating for the farmed salmon industry, but in Southern Chile the conditions that promote the growth and toxicity of these microalgae are poorly understood. This study examined the effects of different combinations of temperature (12, 15, 18 °C) and salinity (10, 20, 30 psu) on the growth of Chilean strains of these two species. The results showed that the optimal growth conditions for H. akashiwo and P. verruculosa differed, with a maximum rate of 0.99 day−1 obtained at 15 °C and a salinity of 20 psu for H. akashiwo, and a maximum rate of 1.06 day−1 obtained at 18 °C and a salinity of 30 psu for P. verruculosa. Cytotoxic assays (2 × 101 – 2 × 105 cell mL−1; cells, filtrates, and cell lysates) performed at salinities of 20 and 30 psu showed a 100% reduction in the viability of embryonic fish cells exposed to intact cells of H. akashiwo and a 39% reduction following exposure to culture filtrates of P. verruculosa. Differences in the fish-killing mechanisms (direct cell contact vs. extracellular substances) and physiological traits of H. akashiwo and P. verruculosa explain the recent occurrence of very large blooms under contrasting (cold-brackish vs. hot-salty) extreme climate conditions in Chile.Postprint2,35

Latitudinal Variation in the Toxicity and Sexual Compatibility of Alexandrium catenella Strains from Southern Chile

The bloom-forming toxic dinoflagellate Alexandrium catenella was first detected in southern Chile (39.5–55° S) 50 years ago and is responsible for most of the area’s cases of paralytic shellfish poisoning (PSP). Given the complex life history of A. catenella, which includes benthic sexual cysts, in this study, we examined the potential link between latitude, toxicity, and sexual compatibility. Nine clones isolated from Chilean Patagonia were used in self- and out-crosses in all possible combinations (n = 45). The effect of latitude on toxicity, reproductive success indexes, and cyst production was also determined. Using the toxin profiles for all strains, consisting of C1, C2, GTX4, GTX1, GTX3, and NeoSTX, a latitudinal gradient was determined for their proportions (%) and content per cell (pg cell−1), with the more toxic strains occurring in the north (−40.6° S). Reproductive success also showed a latitudinal tendency and was lower in the north. None of the self-crosses yielded resting cysts. Rather, the production of resting cysts was highest in pairings of clones separated by distances of 1000–1650 km. Our results contribute to a better understanding of PSP outbreaks in the region and demonstrate the importance of resting cysts in fueling new toxic events. They also provide additional evidence that the introduction of strains from neighboring regions is a cause for concern.En prens

HABs in coastal upwelling systems: Insights from an exceptional red tide of the toxigenic dinoflagellate Alexandrium minutum

Alexandrium minutum blooms generally occur in semi-enclosed sites such as estuaries, harbours and lagoons, where enhanced stratification, restricted circulation and accumulation of resting cysts in the sediment set suitable habitat conditions for the proliferation of this paralytic shellfish poisoning toxigenic species. In the Galician Rías Baixas (NW Iberian Peninsula), according to weekly time-series between 1994 and 2020, blooms of A. minutum were recurrent in small, shallow estuarine bays inside the Rías de Vigo and Pontevedra, but rarely detected, and if so at low concentrations, out of these environments. However, from May to July 2018 it developed as usual in the small inner bays but then spread over both Rías (Vigo and Pontevedra) causing discoloured waters during one month and prolonged harvesting closures. Meteorological conditions during that period (rains / runoff higher than climatological averages, sustained temperature increment and oscillating wind pattern –i.e., series of upwelling-relaxation cycles), fostered optimal circumstances for the development of that extensive and massive proliferation: strong vertical stratification and the alternation of retention and dispersion processes. Simulations from a particle tracking model portrayed the observed bloom development phases: onset and development inside a small inner bay; transport within the surface layer, from these sites towards the interior parts of the Ría; and dispersion all over the embayment. Seedbeds with high concentrations of resting cysts were detected several months after the bloom, which may have favoured flourishment of A. minutum in the following two years, markedly in 2020. The present work contributes to the general understanding of the dynamics of harmful algal blooms (HABs), from which surveillance indicators of the state of marine ecosystems and their evolution can be derived. We hypothesize that the intensity and frequency of A. minutum proliferations in the Galician Rías could increase under projected climate trends.Postprint2,69

Estudio de crecimiento y producción de toxinas en tres especies de Alexandrium: A. catenella, A. peruvianum y A. ostenfeldii.

Se estudió el crecimiento y el contenido en toxinas de 3 especies de dinoflagelados: Alexandrium catenella, A. peruvianum y A. ostenfeldii. A. catenella mostró tasas de crecimiento más elevadas que las otras dos especies. Las tres especies crecieron más rápidamente a 26ºC y menos a 15ºC aunque las tasas de crecimiento no mostraron grandes diferencias. Se observaron diferentes ventanas óptimas de crecimiento en relación a la salinidad en las tres especies. Las máximas tasas de crecimiento fueron a 32 y 37 de salinidad para A. catenella, a 25 de salinidad para A. peruvianum y a 10 de salinidad para A. ostenfeldii. Las tres especies produjeron toxinas PSP pero en muy diferentes cantidades, mostrando notables diferencias en el perfil de toxinas: (10 componentes A. catenella: C1, C2, GTX1-6, neoSTX y STX; 3 componentes A ostenfeldii y A. peruvianum: GTX2-3 y STX). La concentración de toxinas PSP en A. peruvianum fue muy baja. Se detectó también presencia de espirólidos (13-Desmetil-SPX C) en A. peruvianum y Gymnodimina-A en A. ostenfeldii