Saxitoxin (STX), an exceptionally potent marine toxin for which no antidote is currently available, is produced by methanogens and cyanobacteria. This poses a significant threat to both shellfish aquaculture and human health. Consequently, the development of a rapid, highly sensitive STX detection method is of great significance. The objective of this research is to create a novel approach for identifying STX. Therefore, amplified luminescent proximity homogeneous assay (AlphaLISA) was established using a direct competition method based on the principles of fluorescence resonance energy transfer and antigen-antibody specific binding. This method is sensitive, rapid, performed without washing, easy to operate, and can detect 8-128 ng/mL of STX in only 10 min. The limit of detection achieved by this method is as low as 4.29 ng/mL with coefficients of variation for the intra-batch and inter-batch analyses ranging from 2.61% to 3.63% and from 7.67% to 8.30%, respectively. In conclusion, our study successfully establishes a simple yet sensitive, rapid, and accurate AlphaLISA method for the detection of STX which holds great potential in advancing research on marine biotoxins.

Investigating organic carriers\' utilization efficiency and bioactivity within organic-inorganic hybrid nanoflowers is critical to constructing sensitive immunosensors. Nevertheless, the sensitivity of immunosensors is interactively regulated by different classes of biomolecules such as antibodies and enzymes. In this work, we introduced a new alkaline phosphatase-antibody-CaHPO4 hybrid nanoflowers (AAHNFs) microreactor based colorimetric immunoprobe. This system integrates a biometric unit (antibody) with a signal amplification element (enzyme) through the biomineralization process. Specifically, the critical factors affecting antibody recognition activity in the formation mechanism of AAHNFs are investigated. The designed AAHNFs retain antibody recognition ability with enhanced protection for encapsulated proteins against high temperature, organic solvents, and long-term storage, facilitating the selective construction of lock structures against antigens. Additionally, a colorimetric immunosensor based on AAHNFs was developed. After ascorbic acid 2-phosphate hydrolysis by alkaline phosphatase (ALP), the generated ascorbic acid decomposes I2 to I-, inducing the localized surface plasmon resonance in the silver nanoplate, which is effectively tuned through shape conversion to develop the sensor. Further, a 3D-printed portable device is fabricated, integrated with a smartphone sensing platform, and applied to the data of collection and analysis. Notably, the immunosensor exhibits improved analytical performance with a 0.1-6.25 ng·mL-1 detection range and a 0.06 ng·mL-1 detection limit for quantitative saxitoxin (STX) analysis. The average recoveries of STX in real samples ranged from 85.9% to 105.9%. This study presents a more in-depth investigation of the recognition element performance, providing insights for improved antibody performance in practical applications.

Saxitoxin (STX) is a cyanotoxin with high toxicity, and therefore, there is an urgent need to develop a facile detection method for STX. In this study, an ordered nanopillar array-based electrochemical aptasensor was fabricated for the high-performance detection of STX. The anti-STX aptamer with methylene blue (MB) incorporated at the 3\'-end (MB-Apt) was immobilized at the surface of an Au@PAN nanopillar array electrode and used as the recognition element. The proposed aptasensor demonstrated highly sensitive and selective STX detection because of synergistic catalysis effects of MB and ordered nanopillar arrays along with the selection of MB-Apt. The nanopillar array-based electrochemical aptasensor exhibited high sensitivity over a wide linear concentration range of 1 pM-3 nM with a linear regression equation of ΔI (μA) = 28.0 + 6.9 × log[STX] (R2 = 0.98079) and 3-100 nM with a linear regression equation of ΔI (μA) = 10.7 + 43.4 × log[STX] (R2 = 0.98772), where R is the correlation coefficient. In addition, the limit of detection (LOD) was as low as 1 pM. Furthermore, the designed aptasensor demonstrated excellent selectivity toward STX, preventing interference from neo-STX, okadaic acid, and common metal ions. The presented orderly nanopillar array-based strategy to develop an electrochemical aptasensor for STX detection offers a promising method for developing high-performance electrochemical sensors, and the presented aptasensor should find useful application in the detection of shellfish poison.

Paralytic shellfish toxins (PSTs) can pose a serious threat to human health. Among them, saxitoxin (STX) is one of the most potent natural neurotoxins. Here, the copepod Tigriopus japonicus, was exposed to environmentally relevant concentrations (2.5 and 25 μg/L) STX for 48 h. Although no lethal effects were observed at both concentrations, the transcriptome was significantly altered, and displayed a concentration-dependent response. STX exposure decreased the copepod\'s metabolism and compromised immune defense and detoxification. Additionally, STX disturbed signal transduction, which might affect other cellular processes. STX exposure could inhibit the copepod\'s chitin metabolism, disrupting its molting process. Also, the processes related to damage repair and protection were up-regulated to fight against high concentration exposure. Collectively, this study has provided an early warning of PSTs for coastal ecosystem not only because of their potent toxicity effect but also their bioaccumulation that can transfer up the food chain after ingestion by copepods.

Marine algal toxin contamination is a major threat to human health. Thus, it is crucial to develop rapid and on-site techniques for detecting algal toxins. In this work, we developed colorimetric cloth and paper hybrid microfluidic devices (μCPADs) for rapid detection of gonyautoxin (GTX1/4) combined with molecularly imprinted polymers. In addition, the metal-organic frameworks (MOFs) composites were applied for this approach by their unique features. Guanosine serves as a dummy template for surface imprinting and has certain structural advantages in recognizing gonyautoxin. MOF@MIPs composites were able to perform a catalytic color reaction using hydrogen peroxide-tetramethylbenzidine for the detection of GTX1/4. The cloth-based sensing substrates were assembled on origami μPADs to form user-friendly, miniaturized colorimetric μCPADs. Combined with a smartphone, the proposed colorimetric μCPADs successfully achieved a low limit of detection of 0.65 μg/L within the range of 1-200 μg/L for rapid visual detection of GTX1/4. Moreover, the GTX1/4 of real shellfish and seawater samples were satisfactorily detected to indicate the application prospect of the μCPADs. The proposed method shows good potential in the low-cost, stable establishment of assays for the rapid detection of environmental biotoxins.

In this work, on the basis of Fe3O4@Au-Pt nanozymes (MAP NZs) and aptamer recognition, a magnetic fluorescent aptasensor (MFA) was developed for sensitive and accurate detection of saxitoxin (STX). With the bridge of STX aptamer (AptSTX) and complementary DNA (cDNA), AptSTX decorated MAP NZs (MAP/Apt) and cDNA modified green quantum dots (cDNA@g-QDs) were connected to form MAP/Apt-cDNA@g-QDs complex. As STX behaves a strong binding ability towards AptSTX, it will compete with cDNA and hybridize with Apt to release cDNA@g-QDs. With the addition of TMB, MAP will catalyze TMB to the oxidized TMB (ox-TMB), thereby quenching the fluorescence of g-QDs due to the inner filter effect. Based on this finding, the quantitative relationship between the change in fluorescence of gQDs and STX concentration was explored with a limit of detection (LOD, S/N = 3) of 0.6 nM. An internal standard signal of oxTMB was adopted and reduced the fluctuation of fluorescence signal output. Besides, the fluorescence probe can selectively recognize and detect STX among five marine toxins. Eventually, the MFA method behaved good performance in detecting seafood samples with recoveries of 82.0 % ∼ 102.6 % as well as coefficient of variations (CV) of 7.2 % ∼ 10.3 %. Therefore, the method with internal signal is hopeful to be a potential candidate for sensitive and accurate detection of STX in seafood.

BACKGROUND: Saxitoxin (STX) is the most toxic marine toxin, which can pose several adverse effects on human health. High sensitivity, fast response, and low-cost detection of STX contamination are of significance to reducing the fishery and seafood industries\' loss. Among the various types of biosensors, the electrochemical biosensors have been extensively studied in the detection of STX, but the electrode surface modification material is easy to fall off, resulting in unstable electrochemical signals and poor reproducibility. It is imperative to have a ratiometric electrochemical biosensor for STX.
RESULTS: In this study, we developed a novel aptamer-based electrochemical sensor (AECs) for the sensitive detection of STX based on a K3Fe(CN)6 regulated silver nanoparticles (Ag NPs) modified with aptamer. The AECs was constructed by immobilizing aptamer on Ag NPs surfaces. Under optimized conditions, the AECs showed a linear response towards STX in the range from 0.04 to 0.15 μM with the regression equation of Y = -8.0 + 233.7 X (R2 = 0.9956). The limit of detection (LOD) was calculated to be 1 nM (based on 3 N/S), which is significantly lower than the regulatory limits for STX in seafood. Moreover, the AECs showed excellent sensitivity, reproducibility and stability, as well as the detection in samples with acceptable recovery ranged from 71.2 % to 93.8 %, demonstrating its broad application prospects in detection of STX in seafood samples.
CONCLUSIONS: This work proposed an AECs to achieve sensitive detection of STX. A reaction system of K3Fe(CN)6 etched Ag NPs was introduced and used as the signal source to avoid the instability of the electrochemical signal, which can produce a ratiometric electrochemical signal output mode, improving the stability and sensitivity of electrochemical detection of STX.

Gonyautoxins (GTXs), a group of potent neurotoxins belonging to paralytic shellfish toxins (PSTs), are often associated with harmful algal blooms of toxic dinoflagellates in the sea and represent serious health and ecological concerns worldwide. In the study, a highly selective and sensitive fluorescence nanoprobe was constructed based on photoinduced electron transfer recognition mechanism to rapidly detect GTXs in seawater, using specific entrapment of molecularly imprinted polymers (MIPs) combined with fluorescence analyses. The green emissive fluorescein isothiocyanate was grafted in a silicate matrix as a signal transducer and fluorescence intensity of the nanoprobe with a core-shell structure exhibited a strong enhancement due to efficient analyte blockage in a short response time. Under optimal conditions, the developed MIPs nanoprobe presented an excellent analytical performance for spiked seawater samples including a recovery from 94.44 % to 98.23 %, a linear range between 0.018 nmol L-1 and 0.36 nmol L-1, as well as good accuracy. Furthermore, the method had extremely high sensitivity, with limit of detection obtained as 0.005 nmol L-1 for GTXs and GTX2/3. Finally, the nanoprobe was applied for the determination of GTXs in seven natural seawater samples with GTXs mixture (0.035-0.058 nmol L-1) or single GTX2/3 (0.033-0.050 nmol L-1), and the results agreed well with those of a UPLC-MS/MS method. The findings of our study suggest that the constructed MIPs-based fluorescence enhancement nanoprobe was suitable for rapid, selective and ultrasensitive detection of GTXs, particular GTX2/3, in natural seawater samples.

The intake of paralytic shellfish toxins (PSTs) may adversely affect human health. Therefore, this study aimed to show the prevalence of PSTs from commercially available shellfish in Zhejiang Province, China, during the period of frequent red tides, investigate the factors affecting the distribution of PSTs, and assess the risk of PST intake following the consumption of bivalve shellfish among the Zhejiang population. A total of 546 shellfish samples were collected, 7.0% of which had detectable PSTs at concentrations below the regulatory limit. Temporal, spatial, and interspecific variations in the occurrence of PSTs were observed in some cases. The dietary exposure to PSTs among the general population of consumers only was low. However, young children in the extreme scenario (the 95th percentile of daily shellfish consumption combined with the maximum PST concentration), defined as 89-194% of the recommended acute reference doses, were possibly at risk of exposure. Notably, Arcidae and mussels were the major sources of exposure to toxins. From the public health perspective, PSTs from commercially available shellfish do not pose a serious health risk; however, more attention should be paid to acute health risks, especially for young children, during periods of frequent red tides.

Saxitoxin (STX) causes high toxicity by blocking voltage-gated sodium channels, and it poses a major threat to marine ecosystems and human health worldwide. Our work evaluated the neurotoxicity and chronic toxicology of STX to Caenorhabditis elegans by an analysis of lifespan, brood size, growth ability, reactive oxygen species (ROS) and adenosine triphosphate (ATP) levels, and the overexpression of green fluorescent protein (GFP). After exposure to a series of concentrations of STX for 24 h, worms showed paralysis symptoms and fully recovered within 6 h; less than 5% of worms died at the highest concentration of 1000 ng/mL for first larval stage (L1) worms and 10,000 ng/mL for fourth larval stage (L4) worms. Declines in lifespan, productivity, and body size of C. elegans were observed under the stress of 1, 10, and 100 ng/mL STX, and the lifespan was shorter than that in controls. With STX exposure, the productivity declined by 32-49%; the body size, including body length and body area, declined by 13-18% and 25-27%, respectively. The levels of ROS exhibited a gradual increase over time, accompanied by a positive concentration effect of STX resulting in 1.14-1.86 times higher levels compared to the control group in L4 worms. Conversely, no statistically significant differences were observed between L1 worms. Finally, after exposure to STX for 48 h, ATP levels and GFP expression in C. elegans showed a significant dose-dependent increase. Our study reports the first evidence that STX is not lethal but imposes substantial oxidative stress on C. elegans, with a dose-responsive relationship. Our results indicated that C. elegans is an ideal model to further study the mechanisms underlying the fitness of organisms under the stress caused by paralytic shellfish toxins including STX.