An investigation on bloom dynamics of Alexandrium catenella and A. pacificum and toxin accumulation in shellfish along the coast of Qinhuangdao, China.

The dinoflagellate genus Alexandrium comprises most of the toxic bloom-forming species producing paralytic shellfish toxins (PSTs) in the sea. Recently, repeated paralytic shellfish poisoning episodes have been recorded in Qinhuangdao located at the west coast of the Bohai Sea. To elucidate the relationship between toxic Alexandrium blooms and the poisoning episodes, a year-round investigation was carried out in this region from July 2020 to July 2021. Two qPCR assays were used to detect A. catenella and A. pacificum, and LC-MS/MS was applied to analyze PSTs in phytoplankton and shellfish samples. The blooms of A. catenella and A. pacificum were found in April and July, respectively, and PST content in three bivalves exhibited notable increase following the bloom of A. catenella. The results revealed bloom dynamics of the two toxic Alexandrium species in the Bohai Sea for the first time, and further confirmed A. catenella as the causative agent of poisoning episodes.

The complete mitochondrial genome of Mactra quadrangularis (Mactridae).

The whole mitochondrial genome sequence of Mactra quadrangularis (Reeve, 1854) was determined. It had a total length of 16,848 bp and it contained 12 protein coding genes, 2 ribosome RNA genes, and 22 transfer RNA genes. The base composition was 25.75% A, 20.82% G, 11.53% C, and 41.90% T, respectively. Furthermore, state codon of ND4 was ATT; ND1 and CYTB were ATA; COX1 was GTG; ND5, COX2, ND4L, ND6, ND2, COX3, ATP6, and ND3 were ATG. Phylogenetic analysis demonstrated that M. quadrangularis was most closely related to Mactra chinensis. The mitochondrial genome will provide reference for the further investigation and research of M. quadrangularis.

Microalgal Microscale Model for Microalgal Growth Inhibition Evaluation of Marine Natural Products.

Marine organisms especially sessile invertebrates, such as soft corals, gorgonians and sponges, can survive in the competitive environment mainly relying on their second metabolites with chemoecological effects including allelopathy and algal growth inhibition. It is well known that the microscale models are urgently needed in marine chemoecology assessment to evaluate the algal growth inhibition activity of trace quantity natural products. In this work, a microalgal growth inhibition model was established for microalgal inhibition evaluation of marine natural products with 96-well microplate by automatic fluorescence observation using microplate reader. Subsequently, this model was applied to bioassay-guided isolation and preliminary bioactivity screening of the secondary metabolites from soft corals, gorgonians, sponges and their symbiotic microbes collected from the South China Sea. As a result, fifteen compounds (1‒15) were found to exhibit microalgal growth inhibition activities against at least one of marine microalgae, Karenia mikimotoi, Isochrysis galbana, and Heterosigma akashiwo. Specifically, altersolanol C (13) demonstrated potent activity against K. mikimotoi with the 96h-EC50 value of 1.16 µg/mL, more than four times stronger than that of the positive control K2Cr2O7. It was suggested that the microalgal growth inhibition microscale model is suitable for bioassay-guided isolation and preliminary bioactivity screening of marine natural products.

Description of a Zostera marina catalase gene involved in responses to temperature stress.

Catalase (CAT) is an antioxidant enzyme that plays a significant role in cellular protection against oxidative damage by degradation of hydrogen peroxide to oxygen and water. In the present study, the complete CAT cDNA sequence of Zostera marina was identified through expressed sequence tags (EST) analysis and the rapid amplification of cDNA ends (RACE) technique. The nucleotide sequence of ZmCAT cDNA consisted of 1,816 bp with a 1,434 bp open reading frame (ORF), encoding a polypeptide of 477 amino acid residues, which possessed significant homology to other known plant CATs. The molecular mass of the predicted protein was 55.3 kDa with an estimated isoelectric point of 6.40. Phylogenetic analysis showed that ZmCAT was closely related to CAT from gramineous species. In response to temperature stress, H2O2 and MDA contents in Z. marina increased significantly with cold stress (<10 °C) and heat stress (>25 °C). ZmCAT expression was significantly upregulated at temperatures from 5 to 10 °C and then gradually downregulated, reaching its lowest expression at 30 °C. Recombinant ZmCAT protein exhibited strong antioxidant activity over a wide temperature range, with the highest rZmCAT activity observed at 25 °C and a higher relative activity retained even with heat stress. All these results indicated that ZmCAT was a member of the plant CAT family and involved in minimizing oxidative damage effects in Z. marina under temperature stress.

Protective roles of cornuside in acute myocardial ischemia and reperfusion injury in rats.

Cornuside is a secoiridoid glucoside isolated from the fruit of Cornus officinalis SIEB. et ZUCC. In this study, we investigated the anti-myocardial ischemia and reperfusion (I/R) injury effects of cornuside in vivo and elucidated the potential mechanism. Rat models of myocardial I/R were induced by coronary occlusion followed by reperfusion or by Isoproterenol (ISO), treatment of rats with cornuside (20 and 40 mg/kg, i.v.) protected the animals from myocardial I/R injury as indicated by a decrease in infarct volume, improvement in hemodynamics and reduction of myocardial damage severity. Treatment with cornuside also attenuated polymorphonuclear leukocytes (PMNs) infiltration, decreased myeloperoxidase (MPO) activity in the heart, lowered serum levels of pro-inflammatory factors and reduced phosphorylated IκB-α and nuclear factor kappa B (NF-κB) levels in the heart. Additionally, cornuside was shown to have remarkable antioxidant activity and inhibited ISO-induced myocardial cell necrosis. Thus, cornuside appeared to protect the rat from myocardial I/R injury by acting as an anti-inflammatory agent. These findings suggested that cornuside may be used therapeutically in the setting of myocardial I/R where inflammation and oxidant injury are prominent.

[Effects of two organic pollutants on biomarker system of fish Lateolabrax japonicus and the pollution assessment].

Responses of a select suite of protective enzymes in Lateolabrax japonicus including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (Gpx), reduced glutathione (GSH) glutathione S-transferase (GST), nitric oxide synthase (iNOS) in liver tissue and brain acetylcholinesterase (AChE), which could work as sensitively biochemical biomarkers were analyzed when exposed to different concentrations of sodium dodecylbenzene sulfonate (SDBS) and benzo[a]pyrene (B[a]P) under controlled laboratory conditions. Results showed that: (1) The biochemical biomarkers included in this investigation, different responses occurred in the enzymatic activities when exposed to different pollutants: the activities of SOD, GST, Gpx and GSH content were significantly induced by B[a]P (p < 0.05) while others present little different as compared to the control. Differently, SDBS exposure greatly affected the activities of CAT, Gpx, iNOS and AChE while showed little effect on the other evaluated biomarkers. Gpx was the most sensitive biomarker that sensitively indicated the organic pollution stress. (2) The simultaneous assay on heat shock protein 70 (Hsp70) expression was conducted by flow cytometry (FCM) and result presented that Hsp 70 expression could be effectively induced by SDBS as compared to the control throughout the experiment; however, B[a]P obviously induced Hsp70 during the first 12 d and then decreased to the level of the control. (3) The evaluated method of biomarker system combined with principal component analysis (PCA) could effectively distinguish different pollutants under controlled laboratory conditions and might be a promising and warning method for the early assessment of environmental change exposed to different organic pollutions.

Apoptosis induced by yessotoxins in Hela human cervical cancer cells in vitro.

To investigate the apoptotic effects of an extract of red-tide algae, Protoceratium reticulatum, on HeLa cells, we used liquid chromatography-tandem mass spectrometry, optical microscopy, Hoechst 33342/propidine iodide staining, and DNA gel electrophoresis to analyze its constituents and toxicity, as well as rhodamine 123 staining to investigate changes in mitochondrial membrane potential. Analysis showed that the P. reticulatum extract contained the yessotoxins (YTXs) homo-YTX, 45-OH-YTX and 45-OH-homo-YTX. The results indicated that P. reticulatum extract negatively influences HeLa cells and induces their apoptosis.

[Effects of artificial seawater with different N/P ratio on early development of Laminaria japonica embryo spore].

The embryo spores of Laminaria japonica were cultured in artificial seawater with different N/P ratio to observe their early development. 80 percent of the spores cultivated in whole-ingredient artificial seawater developed normally, but the time of spore germination and gametophyte formation was lagged significantly and the amount of female gametes was significantly higher than that of male gametes (P<0.001), as compared to the control cultivated in natural seawater. Artificial seawater with different N/P ratio showed different effects on the spore germination rate. The artificial seawater with N/P > 15 decreased the spore germination rate, while that without N or P inhibited the spore germination significantly (P<0.001). It was concluded that L. japonica spores could develop normally in artificial seawater, but their germination rate was affected by the seawater N/P. The deficiency of N or P in artificial seawater would lag the time of gametophyte formation and affect the differentiation ratio of female-male gametes.

[Allelopathic effect of Corallina pilulifera on Heterosigma akashiwo and its responses to UV-B irradiation].

By the method of co-culture and using cell density as the main indicator, this paper studied the allelopathic effect of Corallina pilulifera on Heterosigma akashiwo and its responses to UV-B irradiation. Under normal condition, the fresh tissue and aqueous extracts of C. pilulifera had significant inhibitory effects on the growth of H. akashiwo (P < 0.05), indicating their allopathic effect on H. akashiwo, while the dry power and culture media filtrate of C. pilulifera had less effect (P > 0.05). After pre-treated with different dose UV-B radiation and then co-cultured with H. akashiwo, C. pilulifera had some changes in the allelopathic activity of its fresh tissue, dry powder, and aqueous extracts. High-dose UV-B radiation (3.0 J x m(-2)) induced the decrease of the allelopathic effect, whereas low-dose UV-B radiation (0.9 J x m(-2)) was in adverse (P < 0.05).

[Acute toxicity effects of three red tide algae on Brachionus plicatilis].

Acute toxicity testing method was used to study effects of different density of Prorocentrum donghaiense, Heterosigma akashiwo and Alexandrium tamarense on mortality rates and population growth parameter of Brachionus plicatilis under controlled experimental conditions. Results showed that 24 h LC50 values of Prorocentrum donghaiense, Heterosigma akashiwo and Alexandrium tamarense treatment to mortality rate of Brachionus plicatilis were 3.56, 1.21 and 0.49 (x 10(4) cells/mL) respectively. Marked density effects were presented when three species of red tide microalga showed their toxicity to Brachionus plicatilis. There were significant inhibitory effects on Brachionus plicatilis when it was exposed to cells of Prorocentrum donghaiense at the concentration of 10(4) cells/mL, filtrate and cell contents of Heterosigma akashiwo at the concentration of 10(5) cells/mL, and cells, filtrate and cell contents of Alexandrium tamarense at the concentration of 10(3) cells/mL respectively. Inhibitory effects of three species of microalga on Brachionus plicatilis were enhanced with increasing of microalgal density.

[Allelopathic effects of Corallina pilulifera on red tide microalgae Heterosigma akashiwo].

Different concentration methanol-, acetone-, ether-, and chloroform extracts of Corallina pilulifera were used to study their growth inhibitory effects on red tide microalgae Heterosigma akashiwo. The results showed that methanol extract at relatively higher concentrations had the highest growth inhibitory activity and killed all H. akashiwo cells, while the other three kinds of organic solvent extracts had no apparent inhibitory effects, suggesting that the growth inhibitory substances in C. pilulifera had relatively high polarity. The methanol extract was partitioned to petroleum ether phase, ethyl acetate phase, butanol phase, and distilled water phase by liquid-liquid fractionation, and the bioassays on the activity of each fraction were carried out on H. akashiwo. It was found that petroleum ether phase and ethyl acetate phase had strong algicidal effects on H. akashiwo, suggesting that the fatty acids in C. pilulifera tissues might be one of the main allelochemicals.

[Response of interspecific competition between Ulva pertusa and Grateloupia filicina to UV-B irradiation enhancement].

The interspecific competition between Ulva pertusa and Grateloupia filicina and it's response to the UV-B irradiation enhancement were analyzed using mono-culture and co-culture methods. The study adopted reasonable experimental design and took biomass as the main examined index. Results showed that the relation of interspecific competition included both allelopathy effect and nutrient competition. Specific growth rates of U. pertusa under treatment with abundant nutrition and limited nutrition was 2.54 and 2.47 times of those of G. filicina. Thus, compared to U. pertusa, G. filicina was in inferior position. UV-B irradiation could inhibit the growth of U. pertusa and G. filicina under the condition of mono-culture. The higher the dosage and the longer exposure of UV-B irradiation were, the more significant the inhibitive effect was. When they were cultured together, low dosage [1.6 kJ x (m2 x d)(-1)] and medium dosage [4.8 kJ x(m2 x d)(-1)] of UV-B irradiation reduced the competitive ability of U. pertusa, and weights of U. pertusa and G. filicina declined 6.81% and 3.88% in low dosage, and 10.47% and 6.98% in medium dosage, respectively. So the relation of interspecific competition tended to be at a balanced level even though U. pertusa was still the dominant algae. However, on the 12th day, weight of U. pertusa decreased by 13.09%, but the value of G. filicina was 14.72%, which was higher than that of U. pertusa. Therefore, high dosage [9.6 kJ x (m2 x d)(-1)] of UV-B irradiation had more serious inhibitive effect on G. filicina, and competitive dominant position of U. pertusa tended to be more obvious. Thus, UV-B changed the relation of competitive balance of U. pertusa and G. filicina, which changed along with the dosage of UV-B. Moreover, UV-B irradiation might influence the metabolism of the allelochemicals produced by U. pertusa and G. filicina in a long time.

Cryopreservation of gametophytes of Laminaria japonica (phaeophyta) with two-step cooling: interactions between variables related to post-thaw survival.

Little attention has been given to the effect of interactions between different cryogenic parameters on the viability of cryopreserved algae. Gametophytes of Laminaria japonica were cryopreserved in liquid nitrogen by two-step cooling, and interactions were tested for optimizing the cooling protocol and improving freeze-tolerance. UNOVA of a general linear model suggested that interactions between both cooling rate and holding time and between holding temperature and holding time significantly affected the survival of thawed gametophytes. Based on the magnitude of effect, the importance order of factors was found to be: holding temperature, holding time, cooling rate, cooling rate x holding temperature, holding temperature x holding time. UNOVA also suggested significant main effects of pre-culture conditions and sex on survival of thawed gametophytes. Under the optimal procedure, post-thaw survivals obtained were 84 percent for male and 69 percent for female gametophytes. Male gametophytes were observed to be more tolerant to the whole procedure of cryopreservation than females. Following thawing viable gametophytes could grow asexually or give rise to morphologically normal sporophytes.

[Effects of red tide microalgae Alexandrium tamarense on the life history of rotifer Brachionus plicatilis].

In this paper, life-table method was used to study the effects of different concentration Alexandrium tamarense on the durations of different development stages of Brachionus plicatilis and the characters of its population growth. The results showed that A. tamarense had significant effects on the growth and development of B. plicatilis via prolonging the durations of the rotifer' s pre-reproduction and generation succession, shortening the durations of its reproduction and post-reproduction and its mean lifespan, and reducing its laying eggs and fecundity. The net reproduction rate and intrinsic increasing rate of B. plicatilis decreased significantly, in comparison with those of the control. B. plicatilis could maintain definite population increase at the presence of different concentration A. tamarense.

3-(4-Amino-5-thioxo-4,5-dihydro-1H-1,2,4-triazol-3-yl)pyridinium chloride.

In the title compound, C(7)H(8)N(5)S(+)·Cl(-), the dihedral angle formed by the pyridine ring with the triazole ring is 10.0 (1)°. There are weak inter-molecular hydrogen-bond inter-actions in the crystal structure, involving the NH and NH(2) groups as donors, and the chloride anion, the S atom in the thio-ketone group and the unsubstituted ring N atom as acceptors.

Effect of alginic acid decomposing bacterium on the growth of Laminaria japonica (Phaeophyceae).

We collected the diseased blades of Laminaria japonica from Yantai Sea Farm from October to December 2002, and the alginic acid decomposing bacterium on the diseased blade was isolated and purified, and was identified as Alteromonas espejiana. This bacterium was applied as the causative pathogen to infect the blades of L. japonica under laboratory conditions. The aim of the present study was to identify the effects of the bacterium on the growth of L. japonica, and to find the possibly effective mechanism. Results showed that: (1) The blades of L. japonica exhibited symptoms of lesion, bleaching and deterioration when infected by the bacterium, and their growth and photosynthesis were dramatically suppressed. At the same time, the reactive oxygen species (ROS) generation enhanced obviously, and the relative membrane permeability increased significantly. The contents of malonaldehyde (MDA) and free fatty acid in the microsomol membrane greatly elevated, but the phospholipid content decreased. Result suggested an obvious peroxidation and deesterrification in the blades of L. japonica when infected by the bacterium. (2) The simultaneous assay on the antioxidant enzyme activities demonstrated that superoxide dismutase (SOD) and catalase (CAT) increased greatly when infected by the bacterium, but glutathione peroxidase (Gpx) and ascorbate peroxidase (APX) did not exhibit active responses to the bacterium throughout the experiment. (3) The histomorphological observations gave a distinctive evidence of the severity of the lesions as well as the relative abundance in the bacterial population on the blades after infection. The bacterium firstly invaded into the endodermis of L. japonica and gathered around there, and then resulted in the membrane damage, cells corruption and ultimately, the death of L. japonica.

Mass transport model of ions within biofilms under the effect of external field.

A mass transport model was developed to predict the transport rate of ions within biofilms, which was experimentally verified using the fluxes of NH4+ and Ca2+ through the heterotrophic biofilms with the thickness varying from 230 to 1430 microm under the effect of external field in the range of -20 V/m to 60 V/m. It is found that the result predicted by the model is in agreement with the experimentally obtained one, with the error less than 5 percent for the thin biofilms. The error increases with the increase of the biofilm thickness. The transport rate of ions caused by electric migration is affected by the charges, field strength, and biofilm thickness and so on.

Effects of UV-B radiation on the growth interaction of Ulva pertusa and Alexandrium tamarense.

Enhanced UV-B (280 - 320 nm) radiation resulting from ozone depletion is one of global environmental problems. Not only marine organisms but also marine ecosystems can be affected by enhanced UV-B radiation. The effects of UV-B radiation on interaction of macro-algae and micro-algae were investigated using Ulva pertusa Kjellman and Alexandrium tamarense as the materials in this study. The results demonstrated that UV-B radiation could inhibit the growth of Ulva pertusa and Alexandrium tamarense when they were both mono-cultured, and the growth inhibition of algae was more significant with increasing doses of UV-B radiation. Alexandrium tamarense could inhibit the growth of Ulva pertusa in mixed culture, and the growth inhibition was more significant when increasing the initial cell density. However, Ulva pertusa could inhibit the growth of Alexandrium tamarense in early phase and stimulate the growth in latter phase when they were grown in mixed culture. Lower initial cell density (10(2) cell/ml) of Alexandrium tamarense could inhibit the growth of Ulva pertusa under UV-B radiation treatment, however, with the initial cell density increasing (10(3) and 10(4) cell/ml), the growth of Ulva pertusa was stimulated under lower dose of UV-B radiation and inhibited under higher dose of UV-B radiation by Alexandrium tamarense. Compared with that in mixed culture, Ulva pertusa showed more positive inhibition to the growth of Alexandrium tamarense under UV-B radiation treatment.

Effects of scorpion venom bioactive polypolypeptides on platelet aggregation and thrombosis and plasma 6-keto-PG F1alpha and TXB2 in rabbits and rats.

Effects of scorpion venom active polypeptide (SVAP) from scorpion venom of Buthus Martensii Karsch of Chinese on platelet aggregation in ex vivo and vitro in rabbits, thrombosis in carotid artery of rats and plasma 6-keto-PG F1alpha and TXB2 in rats were studied by the turbidimetry, the duplicated thrombosis model by electrostimulation and RIA, respectively. The results showed that SVAP 0.125, 0.25, 0.5 mg/ml inhibited significantly the rabbit platelet aggregation triggered by 0.3 U/ml thrombin, 10 microM ADP in vitro (P<0.05 or 0.01) and SVAP at the dose of 0.32, 0.64 mg/kg iv prolonged distinctively the occlusion time of thrombosis that were induced by electrical stimulation. Increased% of 0.16, 0.32 and 0.64 mg/kg were 30.16, 71.74, 98.27%, respectively, which showed a good dose-effect relationship. SVAP 0.22 mg/ml (in vitro) or 0.2, 0.4 mg/kg (in ex vivo) could obviously increase the plasma concentration of 6-keto-PG F1alpha, but slightly effect rats plasma concentration of TXB2 in vitro and in ex vivo and significantly increase of value of PG I2/TXA2, which suggested that the mechanism of the antithrombotic action of SVAP is related to the resistance against platelet aggregation, increase of the concentration of PG I2 in plasma.