Effects of turbulence on diatoms of the genus Pseudo-nitzschia spp. and associated bacteria.

Turbulence is one of the least investigated environmental factors impacting the ecophysiology of phytoplankton, both at the community and individual species level. Here, we investigated, for the first time, the effect of a turbulence gradient (Reynolds number, from Reλ = 0 to Reλ = 360) on two species of the marine diatom Pseudo-nitzschia and their associated bacterial communities under laboratory conditions. Cell abundance, domoic acid (DA) production, chain formation, and Chl a content of P. fraudulenta and P. multiseries were higher for intermediate turbulence (Reλ = 160 or 240). DA was detectable only in P. multiseries samples. These observations were supported by transcriptomic analyses results, which suggested the turbulence related induction of the expression of the DA production locus, with a linkage to an increased photosynthetic activity of the total metatranscriptome. This study also highlighted a higher richness of the bacterial community associated with the nontoxic strain of P. fraudulenta in comparison to the toxic strain of P. multiseries. Bacillus was an important genus in P. multiseries cultures (relative abundance 15.5%) and its highest abundances coincided with the highest DA levels. However, associated bacterial communities of both Pseudo-nitzschia species did not show clear patterns relative to turbulence intensity.

Estimating the risks of exposure to harmful algal toxins among Scottish harbour seals.

Harmful algal bloom (HAB) toxins consumed by marine predators through fish prey can be lethal but studies on the resulting population consequences are lacking. Over the past approximately 20 years there have been large regional declines in some harbour seal populations around Scotland. Analyses of excreta (faeces and urine from live and dead seals and faecal samples from seal haulout sites) suggest widespread exposure to toxins through the ingestion of contaminated prey. A risk assessment model, incorporating concentrations of the two major HAB toxins found in seal prey around Scotland (domoic acid (DA), and saxitoxins (STX)), the seasonal persistence of the toxins in the fish and the foraging patterns of harbour seals were used to estimate the proportion of adults and juveniles likely to have ingested doses above various estimated toxicity thresholds. The results were highly dependent on toxin type, persistence, and foraging regime as well as age class, all of which affected the proportion of exposed animals exceeding toxicity thresholds. In this preliminary model STX exposure was unlikely to result in mortalities. Modelled DA exposure resulted in doses above an estimated lethal threshold of 1900 µg/kg body mass affecting up to 3.8 % of exposed juveniles and 5.3 % of exposed adults. Given the uncertainty in the model parameters and the limitations of the data these conclusions should be treated with caution, but they indicate that DA remains a potential factor involved in the regional declines of harbour seals. Similar risks may be experienced by other top predators, including small cetaceans and seabirds that feed on similar prey in Scottish waters.

First evidence of the induction of domoic acid production in Pseudo-nitzschia australis by the copepod Temora longicornis from the French coast.

Diatoms of the genus Pseudo-nitzschia are widespread in marine waters. Some of them can produce the toxin domoic acid (DA) which can be responsible for amnesic shellfish poisoning (ASP) when transferred into the food web. These ASP events are of major concern, due to their ecological and socio-economic repercussions, particularly on the shellfish industry. Many studies have focused on the influence of abiotic factors on DA induction, less on the role of biotic interactions. Recently, the presence of predators has been shown to increase DA production in several Pseudo-nitzschia species, in particular in Arctic areas. In order to investigate the relationship between Pseudo-nitzschia species and grazers from the French coast, exposures between one strain of three species (P. australis, P. pungens, P. fraudulenta) and the copepod Temora longicornis were conducted for 5 days. Cellular and dissolved DA content were enhanced by 1,203 % and 1,556 % respectively after the 5-days exposure of P.australis whereas no DA induction was observed in P. pungens and P. fraudulenta. T. longicornis consumed all three Pseudo-nitzschia species. The copepod survival was not related to DA content. This study is an essential first step to better understanding the interactions between planktonic species from the French coast and highlights the potential key role of copepods in the Pseudo-nitzschia bloom events in the temperate ecosystems.

Simultaneous determination of trace marine lipophilic and hydrophilic phycotoxins in various environmental and biota matrices.

An efficient and sensitivity approach, which combines solid-phase extraction or ultrasonic extraction for pretreatment, followed by ultra-performance liquid chromatography-tandem mass spectrometry, has been established to simultaneously determine eight lipophilic phycotoxins and one hydrophilic phycotoxin in seawater, sediment and biota samples. The recoveries and matrix effects of target analytes were in the range of 61.6-117.3 %, 55.7-121.3 %, 57.5-139.9 % and 82.6 %-95.0 %, 85.8-106.8 %, 80.7 %-103.3 % in seawater, sediment, and biota samples, respectively. This established method revealed that seven, six and six phycotoxins were respectively detected in the Beibu Gulf, with concentrations ranging from 0.14 ng/L (okadaic acid, OA) to 26.83 ng/L (domoic acid, DA) in seawater, 0.04 ng/g (gymnodimine-A, GYM-A) to 2.75 ng/g (DA) in sediment and 0.01 ng/g (GYM-A) to 2.64 ng/g (domoic acid) in biota samples. These results suggest that the presented method is applicable for the simultaneous determination of trace marine lipophilic and hydrophilic phycotoxins in real samples.

Identification of Candidate Protein Biomarkers Associated with Domoic Acid Toxicosis in Cerebrospinal Fluid of California Sea Lions (Zalophus californianus).

Since 1998, California sea lion (Zalophus californianus) stranding events associated with domoic acid toxicosis (DAT) have consistently increased. Outside of direct measurement of domoic acid in bodily fluids at the time of stranding, there are no practical nonlethal clinical tests for the diagnosis of DAT that can be utilized in a rehabilitation facility. Proteomics analysis was conducted to discover candidate protein markers of DAT using cerebrospinal fluid from stranded California sea lions with acute DAT (n = 8), chronic DAT (n = 19), or without DAT (n = 13). A total of 2005 protein families were identified experiment-wide. A total of 83 proteins were significantly different in abundance across the three groups (adj. p < 0.05). MDH1, PLD3, ADAM22, YWHAG, VGF, and CLSTN1 could discriminate California sea lions with or without DAT (AuROC > 0.75). IGKV2D-28, PTRPF, KNG1, F2, and SNCB were able to discriminate acute DAT from chronic DAT (AuROC > 0.75). Proteins involved in alpha synuclein deposition were over-represented as classifiers of DAT, and many of these proteins have been implicated in a variety of neurodegenerative diseases. These proteins should be considered potential markers for DAT in California sea lions and should be prioritized for future validation studies as biomarkers.

Applying metabolic modeling and multi-omics to elucidate the biotransformation mechanisms of marine algal toxin domoic acid (DA) in sediments.

Domoic acid (DA)-producing algal blooms are a global marine environmental issue. However, there has been no previous research addressing the question regarding the fate of DA in marine benthic environments. In this work, we investigated the DA fate in the water-sediment microcosm via the integrative analysis of a top-down metabolic model, metagenome, and metabolome. Results demonstrated that biodegradation is the leading mechanism for the nonconservative attenuation of DA. Specifically, DA degradation was prominently completed by the sediment aerobic community, with a degradation rate of 0.0681 ± 0.00954 d-1. The DA degradation pathway included hydration, dehydrogenation, hydrolysis, decarboxylation, automatic ring opening of hydration, and ß oxidation reactions. Moreover, the reverse ecological analysis demonstrated that the microbial community transitioned from nutrient competition to metabolic cross-feeding during DA degradation, further enhancing the cooperation between DA degraders and other taxa. Finally, we reconstructed the metabolic process of microbial communities during DA degradation and confirmed that the metabolism of amino acid and organic acid drove the degradation of DA. Overall, our work not only elucidated the fate of DA in marine environments but also provided crucial insights for applying metabolic models and multi-omics to investigate the biotransformation of other contaminants.

Temporal and spatial variation of domoic acid along Canada's coast.

Shellfish poisonings have posed severe risks to human health globally. The Canadian Shellfish Sanitation Program was established in 1948 to monitor the toxin levels at shellfish harvesting sites along the coast of six provinces in Canada. Domoic acid has been a causal toxin for amnesic shellfish poisoning, and a macro-scale analysis of the temporal and spatial variation of domoic acid along Canada's coast was conducted in this study. We aggregated the toxin levels by week in blue mussel (Mytilus edulis) and soft-shell clam (Mya arenaria) samples, respectively, over a one-year scale. The subsequent application of Functional Principal Component Analysis unveiled that magnitudes of seasonal variation and peaked DA levels around early summer, spring, or mid-fall formed the largest variation in the toxin levels in blue mussels along the coastlines of British Columbia and Prince Edward Island and in soft-shell calms along those of New Brunswick and Nova Scotia. In Quebec, the DA levels were low and varied mostly in terms of the overall magnitude from spring to fall. Downstream correlation analyses in British Columbia further discovered that, at most sites, the strongest correlations were negative between precipitation as well as inorganic nutrients (including nitrate, nitrite, phosphate, and silicate) on one side and DA a few weeks afterward on the other. These findings indicated associations between amnesic shellfish poisoning and environmental stresses.

Understanding the risks of co-exposures in a changing world: a case study of dual monitoring of the biotoxin domoic acid and Vibrio spp. in Pacific oyster.

Assessing the co-occurrence of multiple health risk factors in coastal ecosystems is challenging due to the complexity of multi-factor interactions and limited availability of simultaneously collected data. Understanding co-occurrence is particularly important for risk factors that may be associated with, or occur in similar environmental conditions. In marine ecosystems, the co-occurrence of harmful algal bloom toxins and bacterial pathogens within the genus Vibrio may impact both ecosystem and human health. This study examined the co-occurrence of Vibrio spp. and domoic acid (DA) produced by the harmful algae Pseudo-nitzschia by (1) analyzing existing California Department of Public Health monitoring data for V. parahaemolyticus and DA in oysters; and (2) conducting a 1-year seasonal monitoring of these risk factors across two Southern California embayments. Existing public health monitoring efforts in the state were robust for individual risk factors; however, it was difficult to evaluate the co-occurrence of these risk factors in oysters due to low number of co-monitoring instances between 2015 and 2020. Seasonal co-monitoring of DA and Vibrio spp. (V. vulnificus or V. parahaemolyticus) at two embayments revealed the co-occurrence of these health risk factors in 35% of sampled oysters in most seasons. Interestingly, both the overall detection frequency and co-occurrence of these risk factors were considerably less frequent in water samples. These findings may in part suggest the slow depuration of Vibrio spp. and DA in oysters as residual levels may be retained. This study expanded our understanding of the simultaneous presence of DA and Vibrio spp. in bivalves and demonstrates the feasibility of co-monitoring different risk factors from the same sample. Individual programs monitoring for different risk factors from the same sample matrix may consider combining efforts to reduce cost, streamline the process, and better understand the prevalence of co-occurring health risk factors.

Room-temperature fabrication of fluorinated covalent organic polymer @ Attapulgite composite for in-syringe membrane solid-phase extraction and analysis of domoic acid in aquatic products.

A novel fluorinated covalent organic polymer @ attapulgite composite (F-COP@ATP) was prepared at room temperature for in-syringe membrane solid-phase extraction (SM-SPE) of domoic acid (DA) in aquatic products. Natural ore ATP has the advantages of low cost, good mechanical strength and abundant hydroxyl group on its surface, and in-situ modified F-COP layer can provide abundant adsorption sites. F-COP@ATP combining the advantages of F-COP and ATP, becomes an ideal adsorbent for DA extracting. Moreover, a high-throughput sample preparation strategy was carried out by using the F-COP@ATP membrane as syringe filter and assembling syringes with a ten-channel injection pump. In addition, the experimental factors were optimized, such as pH of extract, amount of adsorbent, velocity of extraction and desorption, type and volume of desorption solvent. The DA analytical method was established by SM-SPE-HPLC/tandem mass spectrometry. The method had a wide linear range with low limit of detection (0.344 ng/kg) and low limit of quantification (1.14 ng/kg). F-COP@ATP membrane can be reused more than five times. The method realized the analysis of DA in scallop and razor clam samples, which shows its application prospect in practical analysis. This study provided an efficient, low-energy and mild idea for preparing other reusable natural mineral ATP-based composite materials for separation and enrichment, which reduces the experimental cost and is closer to environmental protection and green chemistry to a certain extent.

Determination of lipophilic marine biotoxins (azaspiracids, brevetoxins, and okadaic acid group) and domoic acid in mussels by solid-phase extraction and reversed-phase liquid chromatography with tandem mass spectrometry.

An accurate and efficient method was developed for the determination of azaspiracid shellfish toxins (azaspiracids-1, -2, and -3), neurotoxic shellfish toxins (brevetoxins-2 and -3), diarrhetic shellfish toxins (okadaic acid and dinophysistoxins-1 and -2), and the amnesic shellfish toxin (domoic acid) in mussels (Mytilus galloprovincialis). Lipophilic marine biotoxins (azaspiracids, brevetoxins, and okadaic acid group) were extracted with 0.5 % acetic acid in methanol under heating at 60°C to improve the extraction efficiency of okadaic acid group toxins and then cleaned up with a C18 solid-phase extraction cartridge. Domoic acid was extracted with 50 % aqueous methanol and then cleaned up with a graphitized carbon solid-phase extraction cartridge. Lipophilic marine biotoxins and domoic acid were quantified by reversed-phase liquid chromatography coupled to electrospray ionization tandem mass spectrometry. The developed method had insignificant matrix effects for the nine analytes and good recoveries in the range of 79.0 % to 97.6 % at three spiking levels for all analytes except brevetoxin-2 (43.8-49.8 %). The developed method was further validated by analyzing mussel tissue certified reference materials, and good agreement was observed between certified and determined values.

The amnesic shellfish poisoning toxin, domoic acid: The tattoo of the king scallop Pecten maximus.

Domoic acid (DA) is a potent neurotoxin produced by diatoms of the genus Pseudo-nitzschia and is responsible for Amnesic Shellfish Poisoning (ASP) in humans. Some fishery resources of high commercial value, such as the king scallop Pecten maximus, are frequently exposed to toxic Pseudo-nitzschia blooms and are capable of accumulating high amounts of DA, retaining it for months or even a few years. This poses a serious threat to public health and a continuous economical risk due to fishing closures of this resource in the affected areas. Recently, it was hypothesized that trapping of DA within autophagosomic-vesicles could be one reason explaining the long retention of the remaining toxin in P. maximus digestive gland. To test this idea, we follow the kinetics of the subcellular localization of DA in the digestive glands of P. maximus during (a) the contamination process - with sequential samplings of scallops reared in the field during 234 days and naturally exposed to blooms of DA-producing Pseudo-nitzschia australis, and (b) the decontamination process - where highly contaminated scallops were collected after a natural bloom of toxic P. australis and subjected to DA-depuration in the laboratory for 60 days. In the digestive gland, DA-depuration rate (0.001 day-1) was much slower than contamination kinetics. The subcellular analyses revealed a direct implication of early autophagy in DA sequestration throughout contamination (r = 0.8, P < 0.05), while the presence of DA-labeled residual bodies (late autophagy) appeared to be strongly and significantly related to slow DA-depuration (r = -0.5) resembling an analogous DA-tattooing in the digestive glands of P. maximus. This work provides new evidence about the potential physiological mechanisms involved in the long retention of DA in P. maximus and represents the baseline to explore procedures to accelerate decontamination in this species.

Development, calibration, and evaluation of a model of Pseudo-nitzschia and domoic acid production for regional ocean modeling studies.

Pseudo-nitzschia species are one of the leading causes of harmful algal blooms (HABs) along the western coast of the United States. Approximately half of known Pseudo-nitzschia strains can produce domoic acid (DA), a neurotoxin that can negatively impact wildlife and fisheries and put human life at risk through amnesic shellfish poisoning. Production and accumulation of DA, a secondary metabolite synthesized during periods of low primary metabolism, is triggered by environmental stressors such as nutrient limitation. To quantify and estimate the feedbacks between DA production and environmental conditions, we designed a simple mechanistic model of Pseudo-nitzschia and domoic acid dynamics, which we validate against batch and chemostat experiments. Our results suggest that, as nutrients other than nitrogen (i.e., silicon, phosphorus, and potentially iron) become limiting, DA production increases. Under Si limitation, we found an approximate doubling in DA production relative to N limitation. Additionally, our model indicates a positive relationship between light and DA production. These results support the idea that the relationship with nutrient limitation and light is based on direct impacts on Pseudo-nitzschia biosynthesis and biomass accumulation. Because it can easily be embedded within existing coupled physical-ecosystem models, our model represents a step forward toward modeling the occurrence of Pseudo-nitzschia HABs and DA across the U.S. West Coast.

Prolonged, Low-Level Exposure to the Marine Toxin, Domoic Acid, and Measures of Neurotoxicity in Nonhuman Primates.

BACKGROUND: The excitotoxic molecule, domoic acid (DA), is a marine algal toxin known to induce overt hippocampal neurotoxicity. Recent experimental and epidemiological studies suggest adverse neurological effects at exposure levels near the current regulatory limit (20 ppm, ∼0.075-0.1mg/kg). At these levels, cognitive effects occur in the absence of acute symptoms or evidence of neuronal death. OBJECTIVES: This study aimed to identify adverse effects on the nervous system from prolonged, dietary DA exposure in adult, female Macaca fascicularis monkeys. METHODS: Monkeys were orally exposed to 0, 0.075, and 0.15mg/kg per day for an average of 14 months. Clinical blood counts, chemistry, and cytokine levels were analyzed in the blood. In-life magnetic resonance (MR) imaging assessed volumetric and tractography differences in and between the hippocampus and thalamus. Histology of neurons and glia in the fornix, fimbria, internal capsule, thalamus, and hippocampus was evaluated. Hippocampal RNA sequencing was used to identify differentially expressed genes. Enrichment of gene networks for neuronal health, excitotoxicity, inflammation/glia, and myelin were assessed with Gene Set Enrichment Analysis. RESULTS: Clinical blood counts, chemistry, and cytokine levels were not altered with DA exposure in nonhuman primates. Transcriptome analysis of the hippocampus yielded 748 differentially expressed genes (fold change≥1.5; p≤0.05), reflecting differences in a broad molecular profile of intermediate early genes (e.g., FOS, EGR) and genes related to myelin networks in DA animals. Between exposed and control animals, MR imaging showed comparable connectivity of the hippocampus and thalamus and histology showed no evidence of hypomyelination. Histological examination of the thalamus showed a larger microglia soma size and an extension of cell processes, but suggestions of a GFAP+astrocyte response showed no indication of astrocyte hypertrophy. DISCUSSION: In the absence of overt hippocampal excitotoxicity, chronic exposure of Macaca fascicularis monkeys to environmentally relevant levels of DA suggested a subtle shift in the molecular profile of the hippocampus and the microglia phenotype in the thalamus that was possibly reflective of an adaptive response due to prolonged DA exposure. https://doi.org/10.1289/EHP10923.

Response mechanism of microbial community during anaerobic biotransformation of marine toxin domoic acid.

Domoic acid (DA) is a potent neurotoxin produced by toxigenic Pseudo-nitzschia blooms and quickly transfers to the benthic anaerobic environment by marine snow particles. DA anaerobic biotransformation is driven by microbial interactions, in which trace amounts of DA can cause physiological stress in marine microorganisms. However, the underlying response mechanisms of microbial community to DA stress remain unclear. In this study, we utilized an anaerobic marine DA-degrading consortium GLY (using glycine as co-substrate) to systematically investigate the global response mechanisms of microbial community during DA anaerobic biotransformation.16S rRNA gene sequencing and metatranscriptomic analyses were applied to measure microbial community structure, function and metabolic responses. Results showed that DA stress markedly changed the composition of main species, with increased levels of Firmicutes and decreased levels of Proteobacteria, Cyanobacteria, Bacteroidetes and Actinobacteria. Several genera of tolerated bacteria (Bacillus and Solibacillus) were increased, while, Stenotrophomonas, Sphingomonas and Acinetobacter were decreased. Metatranscriptomic analyses indicated that DA stimulated the expression of quorum sensing, extracellular polymeric substance (EPS) production, sporulation, membrane transporters, bacterial chemotaxis, flagellar assembly and ribosome protection in community, promoting bacterial adaptation ability under DA stress. Moreover, amino acid metabolism, carbohydrate metabolism and lipid metabolism were modulated during DA anaerobic biotransformation to reduce metabolic burden, increase metabolic demands for EPS production and DA degradation. This study provides the new insights into response of microbial community to DA stress and its potential impact on benthic microorganisms in marine environments.

Rapid detection of phycotoxin domoic acid in seawater and seafood based on the developed lateral flow immunoassay.

A lateral flow immunoassay (LFIA) of phycotoxin domoic acid (DA) contaminating seawater and marine organisms was developed in this investigation. Nine clones of monoclonal antibodies against DA were produced and characterized. The test system was implemented in the indirect competitive format, where gold nanoparticles as a marker were conjugated with secondary antibodies. The developed test system allows for the detection of DA with a cutoff of 60 ng mL-1 and an instrumental detection limit of 1.4 ng mL-1 within 15 min. The LFIA was applied to detect DA in seawater, mussels, shrimps, and octopuses. A simple method of seafood sample preparation was proposed. The entire analytical cycle, from obtaining a sample to the estimation of final results, takes only 30 min. The assay recoveries ranged from 88.5% to 124%. The developed analytical method is a promising solution for rapid on-site monitoring of marine toxicants in water and food throughout the farm-to-fork chain.

First subcellular localization of the amnesic shellfish toxin, domoic acid, in bivalve tissues: Deciphering the physiological mechanisms involved in its long-retention in the king scallop Pecten maximus.

Domoic acid (DA), the phycotoxin responsible for amnesic shellfish poisoning (ASP), is an excitatory amino acid naturally produced by at least twenty-eight species of the bloom-forming marine diatoms Pseudo-nitzschia spp. Suspension feeders, such as bivalve mollusks, can accumulate and lengthy retain high amounts of DA in their tissues, threatening human health and leading to extensive-prolonged fishery closures, and severe economic losses. This is particularly problematic for the king scallop Pecten maximus, which retains high burdens of DA from months to years compared to other fast-depurator bivalves. Nonetheless, the physiological and cellular processes responsible for this retention are still unknown. In this work, for the first time, a novel immunohistochemical techniques based on the use of an anti-DA antibody was successfully developed and applied for DA-detection in bivalve tissues at a subcellular level. Our results show that in naturally contaminated P. maximus following a Pseudo-nitzschia australis outbreak, DA is visualized mainly within small membrane-bounded vesicles (1 - 2.5 µm) within the digestive gland cells, identified as autophagosomic structures by means of immune-electron microscopy, as well as in the mucus-producing cells, particularly those from gonad ducts and digestive tract. Trapping of DA in autophagososomes may be a key mechanism in the long retention of DA in scallops. These results and the development of DA-immunodetection are essential to provide a better understanding of the fate of DA, and further characterize DA contamination-decontamination kinetics in marine bivalves, as well as the main mechanisms involved in the long retention of this toxin in P. maximus.

Construction of ratiometric fluorescence sensor and test strip with smartphone based on molecularly imprinted dual-emission quantum dots for the selective and sensitive detection of domoic acid.

Domoic acid (DA), a highly neurotoxic metabolite produced by phytoplankton, contaminates seafood products and threats humankind. Herein, we have proposed a molecular imprinting fluorescence sensor with internal standard ratiometric mode for sensing of DA in seafood and seawater. In this study, the silicon-coated blue luminous carbon dots (B-CDs@SiO2) and CdTe acted as reference probe (430 nm) and response probe (610 nm), respectively. Subsequently, the two probes were assembled and the molecularly imprinted polymer (MIP) was introduced as the recognition element to construct the core component of the sensor (B-CDs@SiO2/CdTe MIP). When DA exists, it can be specifically adsorbed by the amino-rich imprinted sites on surface of B-CDs@SiO2/CdTe MIP and further assembled into the hydrogen-bonds complex, which can lead to the decrease in the fluorescence signal of MIP at 610 nm owing to the electron transfer from CdTe to DA. However, the fluorescence signal of MIP at 430 nm is not affected because of the protection of silica layer. Based on this principle, the designed internal standard ratiometric fluorescence sensor reveals high sensitivity, excellent selectivity, and wide linear range of 0.03-1 µM with a detection limit of 18 nM. Further, the portable fluorescent test strip with smartphone has been designed for semi-quantitative sensing of DA, which has potential application prospects for field analysis.

Marine toxin domoic acid induces moderate toxicological response in non-target HepG2 cells.

Domoic acid (DA) is a marine neurotoxin produced as a defence compound by diatom Pseudo-nitzschia. Although its toxicity is well known in marine mammals and fish, data on DA cyto/genotoxicity in human non-target cells is still limited. Hence, we aimed to study the effect of DA (0.001-10 µg/mL) on cell viability and proliferation kinetics of human hepatocellular carcinoma (HepG2) cells as well as DNA damage induction after 4, 24 and 72 h of exposure. The results revealed that DA up to 10 µg/mL did not elicit significant changes in HepG2 cell viability, proliferation and cell cycle at applied conditions. DA did not generate DNA double-strand breaks, while it exhibited significant dose- and time-dependent increase of DNA damage in the form of either DNA single-strand breaks or alkali labile sites. Additionally, increased malondialdehyde level after DA treatment indicated oxidative damage to lipids. Altogether, the results showed that neurotoxin DA induced only minor adverse genotoxic effects in non-target HepG2 cells that most probably occurred resulting from the oxidative stress. However, additional research is needed to further elucidate the mechanisms of DA toxicity, particularly in terms of chronic exposure, as well as to understand its potential influence on human non-target cells.

Adaptogenic potential of ginsenosides against domoic acid-induced toxicity by regulating neuronal stress and kinate receptors: Ex vivo and in silico studies.

This study is focused on potential effects of ginsenosides from Panax ginseng (PG) against amnesic shell fish poison, that is, domoic acid-induced excitotoxicity. Mice received PG at two different dosages by oral feeding for a period of 28 days (50 and 100 mg kg-1 bwt.-1 ). Domoic acid was injected to the mice to induce excitotoxicity (DA; 3 mg kg-1  bwt.-1 ) and piracetam-injected animals (PIR; 100 mg kg-1  bwt.-1 ) were treated as positive control. DA-induced cognitive impairment was reverted by PG supplementation, which was observed in Morris water maze and novel object task. Moreover, PG supplementation restored levels of GABA and antioxidant enzymes. Our results further elucidated ameliorative effects of PG supplementation on DA-induced changes in the expression of synaptic plasticity (BDNF), inflammation (NFkB), and apoptotic (Bcl2, Bax, and Caspase 3) markers. Hence, this study elucidates potential nootropic effects of ginsenosides from P. ginseng extract against DA-induced neuronal impairments via, modulation of behavioral and biochemical mechanisms involved in excitotoxicity, oxidative stress, neuro-inflammation, and apoptosis. PRACTICAL APPLICATIONS: This study highlights potential effects of ginsenosides from Panax ginseng against amnesic shell fish poison, that is, domoic acid-induced excitotoxicity for the first time. This study confirms that ginsenosides have the beneficial effects against amelioration of DA-induced toxicity. This study elucidates the potential nootropic effects of P. ginseng extract against DA-induced neuronal impairments via, modulation of synaptic plasticity markers and oxido-inflammatory responses leading to apoptosis. This study will be helpful in offering various mechanisms involved in pharmacological applications of P. ginseng in the management of DA-induced excitotoxicity.