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

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

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

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

The toxicity and intraspecific variability of Alexandrium andersonii Balech

The toxicity of Alexandrium andersonii Balech is unclear and its intraspecific variability has yet to be studied. To address these gaps in our knowledge, in the present work five strains of A. andersonii from four different localities were characterized. The results showed that despite genetic homogeneity in the 5.8-ITS (internal transcribed spacer) and large subunit (LSU) regions and similar growth rates, strains originating from different locations varied with respect to cell size, the ratios of certain pigments, and their growth patterns. Cultures of the strains grown at 20 8C were analyzed for toxicity using four different methodologies. The two officially established methods, mouse bioassay and high-performance liquid chromatography with fluorescence detection (HPLC-FLD) and post-column reaction analysis of PSP toxins, failed to show the toxicity of any strain. Strains grown at 14 8C were also negative for PSP toxins by HPLC-FLD. However, strains grown at 20 8C exhibited both a response characteristic of the presence of toxin-inhibiting voltage-gated sodium channels, as demonstrated in a neuroblastoma neuro- 2a cell-based assay, as well as hemolytic activity in a sheep red blood cell assay

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

Biphasic toxicodynamic features of some antimicrobial agents on microbial growth: a dynamic mathematical model and its implications on hormesis

<p>Abstract</p> <p>Background</p> <p>In the present work, we describe a group of anomalous dose-response (DR) profiles and develop a dynamic model that is able to explain them. Responses were obtained from conventional assays of three antimicrobial agents (nisin, pediocin and phenol) against two microorganisms (<it>Carnobacterium piscicola </it>and <it>Leuconostoc mesenteroides</it>).</p> <p>Results</p> <p>Some of these anomalous profiles show biphasic trends which are usually attributed to hormetic responses. But they can also be explained as the result of the time-course of the response from a microbial population with a bimodal distribution of sensitivity to an effector, and there is evidence suggesting this last origin. In light of interest in the hormetic phenomenology and the possibility of confusing it with other phenomena, especially in the bioassay of complex materials we try to define some criteria which allow us to distinguish between <it>sensu stricto </it>hormesis and biphasic responses due to other causes. Finally, we discuss some problems concerning the metric of the dose in connection with the exposure time, and we make a cautionary suggestion about the use of bacteriocins as antimicrobial agents.</p> <p>Conclusions</p> <p>The mathematical model proposed, which combines the basis of DR theory with microbial growth kinetics, can generate and explain all types of anomalous experimental profiles. These profiles could also be described in a simpler way by means of bisigmoidal equations. Such equations could be successfully used in a microbiology and toxicology context to discriminate between hormesis and other biphasic phenomena.</p