深圳海域麻痹性贝类毒素含量特征

根据 1996～ 1997年间深圳海域 (大亚湾和大鹏湾 ) 13种贝类 (华贵栉孔扇贝、马氏珍珠贝、翡翠贻贝、波纹巴非蛤、毛蚶、栉江瑶、联珠蚶、文蛤、棕带仙女蛤、僧帽牡蛎、菲律宾蛤仔、习见蛙螺和近江牡蛎 )体中的麻痹性毒素的检测数据 ,阐述了贝类麻痹性毒素的含量水平、年际变化、PSP中毒事件以及与其它海区贝类 PSP的比较     

A new simple screening method for the detection of paralytic shellfish poisoning toxins

The current testing for paralytic shellfish poisoning(PSP) in shellfish is based on the mouse bioassay(MBA).To alleviate animal welfare concerns,we evaluated the utility of using sublethal indicators of toxicity as an alternative to measuring time to death.Live mice were injected with a PSP congener and the changes in neurotransmitter levels were measured 60,90,and 120 min after injection.Acetylcholine(ACh) was the most sensitive marker for PSP toxicity.The changes in neurotransmitter levels were most pronounced in the blood.Thus,measurement of Ach levels in the blood may serve as a sensitive predictor for PSP that would not require sacrifice of the mice.This method was relatively simple,sensitive(less than 1 μg/kg weight,equivalent to 20 ng/mL),low maintenance,and rapid(less than 60 min).     

Identification of paralytic shellfish toxin-producing microalgae using machine learning and deep learning methods

Journal of Oceanology and Limnology - Paralytic shellfish poisoning (PSP) microalgae, as one of the harmful algal blooms, causes great damage to the offshore fishery, marine culture, and marine...     

A sulfotransferase specific to N-21 of gonyautoxin 2/3 from crude enzyme extraction of toxic dinoflagellate Alexandrium tamarense CI01

Sulfotransferase (ST) is the first enzyme discovered in association with paralytic shellfish poisoning (PSP) toxin biosynthesis in toxic dinoflagellates. This study investigates the ST activity in crude enzyme extraction of a toxic dinoflagellate species, Alexandrium tamarense CI01. The results show that crude enzyme can transfer a sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to N-21 in the carbamoyl group of gonyautoxin 2/3 (GTX2/3) to produce C1/C2, but is inactive toward STX to produce GTX5. The crude enzyme is optimally active at pH 6.0 and 15°C. The activity is enhanced by Co2 , Mg2 , Mn2 and Ca2 individually, but is inhibited by Cu2 . Moreover, the activity shows no difference when various sulfur compounds are used as sulfate donors. These results demonstrate that the ST specific to GTX2/3 is present in the cells of A. tamarense CI01 and is involved in PSP toxin biosynthesis. In addition, the ST from different dinoflagellates is species-specific, which explains well the various biosynthesis pathways of the PSP toxins in toxic dinoflagellates.     

Molecular Identification of Dried Shellfish Products Sold on the Market Using DNA Barcoding

The dried shellfish products with rich nutrients and low-calorie content are favorite food in China, especially in coastal areas. However, the species of dried shellfish products in the market are usually unknown, as the taxonomic features were removed during the production process. This study described the application of DNA barcoding technique to the identification of 100 dried shellfish(scallop, squid, octopus and cuttlefish) products in markets. Samples were authenticated by comparing mitochondrial cytochrome oxidase subunit I(COI) gene and 16 S ribosomal RNA(16 S rRNA) gene sequences with public reference taxonomic databases. The results showed that all the 100 products can be identified at species level. Sixty four scallop adductor products were processed using the bay scallop, Argopecten irradians, and one was from Portuguese oyster, Crassostrea angulata. All the 27 squid, 2 cuttlefish and 6 octopus products were produced by the Jumbo flying squid, Dosidicus gigas. The neighbour-joining tree is in agreement with the results of DNA barcoding analysis. The 64 scallop samples formed one A. irradians cluster, leaving Sca65 clustered with the reference oyster sequence C. angulata(MH997922). All the 35 cephalopod(squid, octopus and cuttlefish) samples formed a D. gigas cluster. This investigation revealed a low variety of dried shellfish products sold on the market, and highlighted the high rate of mislabeling and species substitution. Our present work provides a practical method for tracing and authenticating shellfish products.     

方斑东风螺两种养殖模式的比较

在室内水泥池,利用沙层自净养殖模式和直接铺沙养殖模式对不同规格的方斑东风螺(Babylonia areolataLink)进行了高密度养殖的研究。结果表明,沙层自净养殖模式养殖小螺、中螺、大螺组日均增重分别为0.031、0.088、0.098 g/d,沙层NH4+-N最高含量w分别为1.3、2.1、3.1 mg/L,H2S最高含量w分别为0.03、0.07、0.14mg/L,各规格组东风螺保持正常生长和活动,成活率92.9%以上;直接铺沙养殖模式养殖小螺、中螺、大螺日均增重分别为:0.023、0.051、0.068 g/d,成活率分别为95.2%、86.7%、84.9%,沙层NH4+-N最高含量w达到13.7mg/L,H2S最高含量w达到0.47 mg/L,沙层底质恶化,东风螺活动异常、不摄食。可见,沙层自净养殖模式对方斑东风螺的生长、成活率、沙层水质控制效果显著,在一定程度上克服了直接铺沙养殖底质恶化问题。     

First observation of domoic acid and its isomers in shellfish samples from Shandong Province,China

Journal of Oceanology and Limnology - A total of 133 shellfish samples were collected in seven cities of Shandong Province, China, from May to October, 2019. The domoic acid (DA) concentrations...     

Interactions of aquaculture and waste disposal in the coastal zone

Throughout the world, the coastal zones of many countries are used increasingly for aquaculture in addition to other activities such as waste disposal. These activities can cause environmental problems and health problems where they overlap. The interaction between aquaculture and waste disposal, and their relationship with eutrophication are the subjects of this paper. Sewage discharge without adequate dispersion can lead to nutrient elevation and hence eutrophication which has clearly negative effects on aquaculture with the potential for toxic blooms. Blooms may be either toxic or anoxia-causing through the decay process or simply clog the gills of filter-feeding animals in some cases. With the development of aquaculture, especially intensive aquaculture, many environmental problems appeared, and have resulted in eutrophication in some areas. Eutrophication may destroy the health of whole ecosystem which is important for sustainable aquaculture. Sewage discharge may also cause serious public health problems. Filter-feeding shellfish growing in sewage-polluted waters accumulate micro-organims, including human pathogenic bacteria and viruses, and heavy metal ion, presenting a significant health risk. Some farmed animals may also accumulate heavy metals from sewage. Bivalves growing in areas affected by toxic algae blooms may accumulate toxins (such as PSP, DSP) which can be harmful to human beings.     

Application of rRNA probes and fluorescence in situ hybridization for rapid detection of the toxic dinoflagellate Alexandrium minutum

The dinoflagellate Alexandrium minutum is often associated with harmful algal blooms (HABs). This species consists of many strains that differ in their ability to produce toxins but have similar morphology, making identification difficult. In this study, species-specific rRNA probes were designed for whole-cell fluorescence in situ hybridization (FISH) to distinguish A. minutum from two phylogenetic clades. We acquired the complete SSU to LSU rDNA sequences (GenBank accession numbers JF906989-JF906999) of 11 Alexandrium strains and used these to design rRNA targeted oligonucleotide probes. Three ribotype-specific probes, M-GC-1, M-PC-2, and M-PC-3, were designed. The former is specific for the GC clade ("Global clade") of A. minutum, the majority of which have been found non-toxic, and the latter two are specific for the PSP (paralytic shellfish poisoning)-producing PC clade ("Pacific clade"). The specificity of these three probes was confirmed by FISH. All cells in observed fields of view were fluorescently labeled when probes and target species were incubated under optimized FISH conditions. However, the accessibility of rRNA molecules in ribosomes varied among the probe binding positions. Thus, there was variation in the distribution of positive signals in labeled cells within nucleolus and cytosol (M-GC-1, M-PC-3), or just nucleolus (M-PC-2). Our results provide a methodological basis for studying the biogeography and population dynamics of A. minutum, and providing an early warning of toxic HABs.     

Screening for lipophilic marine toxins and their potential producers in coastal waters of Weihai in autumn, 2020

Lipophilic marine toxins (LMTs) produced by some microalgae in the sea could accumulate in shellfish and pose potential threats to the health of seafood consumers. Phytoplankton and shellfish samples were collected from coastal waters of Weihai in Shandong Peninsula, China in autumn, 2020, and screened for lipophilic marine toxins and their potential producers using liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis and high throughput sequencing of partial DNA (V4 region of the 18S rRNA gene) extracted from phytoplankton. Pectenotoxin-2 (PTX2), trace amounts of azaspiracid (AZA1 or AZA40), and 13-desmethyl spirolide C (13-DesMe-C) were detected in phytoplankton samples, while PTX2 and gymnodimine (GYM) were detected in shellfish samples. The toxin content in shellfish samples was much lower than the regulatory limit or values reported previously. Results suggest that lipophilic marine toxins should have low risk in coastal waters of Weihai in autumn. Based on the data of high throughput sequencing, the OTUs were assigned to 5 identified species of Alexandrium, including A. ostenfeldii capable of producing 13-DesMe-C and GYM. Two OTUs were found closely related to the toxic species in genus Dinophysis, but it is impossible to assign them to any identified species due to the low resolving power of the V4 region for Dinophysis. The OTUs could not be assigned to any identified species in the genus Azadinium, suggesting the existence of unidentified species in this region.

     

Application and comparison of two symptom-based eutrophication-assessment methods in Jiaozhou Bay,China

Increasing numbers of approaches to assess eutrophication,such as estuarine trophic status,the Oslo-Paris Commission Common Procedure,and the Water Framework Directive,focus on the symptoms of eutrophication.In China,however,nutrient index methods dominate the assessment of coastal waters.In this study,an integrated method that includes both water quality and ecological response was compared with the Northwest Pacific Action Plan(NOWPAP)Common Procedure.Observation data from Jiaozhou Bay,Shandong,China,were used in a comparison of the two methods in a trophic status study.Overall,both clearly revealed a high level of nutrient enrichment in the bay,indicated by high nutrient concentrations.Though the two methods differed in their methodological design in the assessment of the ecological effects of nutrient enrichment,they have acquired similar results:the integrated method suggested that the status was good,and the NOWPAP Common Procedure suggested that the status was low(indicating that the bay had no serious eutrophication problem).The introduction of filter feeders(shellfish aquaculture)into the bay on a reasonable scale may have been effective in mitigating eutrophic conditions,and perhaps explains the low ecological impacts there.Our results will be useful to ecosystem-based eutrophication management in the bay and in similar areas.     

Development of a Label-Free Colorimetric Aptasensor with Rationally Utilized Aptamer for Rapid Detection of Okadaic Acid

Okadaic acid(OA)is a typical marine toxin with strong toxicity causing diarrheic shellfish poisoning(DSP).Aptamers show great advantages in toxin detection and attract increasing attentions in the field of food analysis.In this study,a label-free col-orimetric aptasensor was constructed for visual and rapid detection of OA in shellfish.To exploit the binding capability of the anti-OA aptamer,the inherent molecular recognition mechanism of aptamer and OA was studied,based on molecular docking,fluorescent assay,and biolayer interferometry.Consistent results showed that the stem-loop near the 3’terminal of the aptamer exhibit dominate binding capacity.Based on the revealed recognition information,the aptamer was thus rationally utilized and combined with AuNPs and cationic polymer polydiallyl dimethyl ammonium chloride(PDDA)for the development of the label-free colorimetric aptasensor,in which the 3’terminal was thoroughly exposed to OA.The aptasensor provided robust performance with a linear detection range of 100-1200 nmol L-1,a limit of detection of 41.30 nmol L-1,recovery rates of 91.6%-106.2%,as well as a high selectivity towards OA in shellfish samples.The whole detection process can be completed within 1 h.To our best knowledge,this is the first time that the anti-OA aptamer was thoroughly studied,and a label-free colorimetric aptasensor was rationally designed in this way.This study not only provides a rapid detection method for highly sensitive and specific detection of OA,but also serves as a reference for the design of efficient aptasensors in the future.     

Concentrations and distribution of organochlorine pesticides in shellfish from Changjiang estuary

To analyze and evaluate the status of organochlorine pollutants in the Changjiang (Yangtze River) estuary and adjacent waters, the concentrations of hexachlorocyclohexane (HCHs) and dichlorodiphenyltrichloroethane (DDTs) in shellfish collected in study area from 2006 to 2007 were determined with gas chromatography (GC). The concentration range of HCHs was (ND-12.13)×10-3 mg/kg wet weight and averaged at 0.54×10-3 mg/kg while the concentration of DDTs was in the range of (4.06-281.73) × 10-3 mg/kg with a mean of 57.52×10-3 mg/kg in the survey areas. The concentrations of DDTs in the shellfish were higher than HCHs', so that DDTs could be considered as typical organochlorine pollutants in the areas. The concentrations of DDTs in the shellfish were higher than HCHs', so that DDTs could be considered as typical organochlorines pollutants. The HCHs in all the shellfish conformed to the first level of criterion (0.02 mg/kg) of the Marion Biology Quality (GB 18421-2001), and that of DDTs in most samples were beyond the first level (0.01 mg/kg) but conformed to the second level (0.10 mg/kg). On average, α-HCH and δ-HCH occupied the most part of HCHs, while O,P'-DDT and P,P'-DDT occupied the most part of DDTs. The concentrations of organocholorine pesticides in shellfish samples varied in site and in species. The highest level occurred at the Shengsi (SS), followed by Yangkougang (YKG), Lvsi (LS), Dongyuan (DY) and Beibayao (BBY), low concentrations were observed at Changsha (CS), Beidaodi (BDD), and Gouqi (GQ). The concentration of HCHs and DDTs in most sites decreased clearly from 2006 to 2007 except for YKG, DY, BDD, LYS, and SS. All of above results suggested that the study area was slightly affected by organochlorine pesticide, special by DDTs.     

Historical Evolution of Mariculture in China During Past 40 Years and Its Impacts on Eco-environment

The rapid development of mariculture in China in recent decades has satisfied people's demand of seafood, and has made a great contribution to economic development. However, mariculture has also caused some negative impacts on the eco-environment. By statistically analyzing national data(as well as data for four main provinces) regarding the mariculture area and cultivation species in China since reform and opening up, the historical evolution of mariculture during the past 40 years was analyzed, the driving factors related to policy, market and technology innovation were discussed, and the potential impacts of mariculture on the coastal eco-environment were also illustrated. The statistical results indicated that the maricultural area increased radically during the past 40 years, from 1979(116.47 × 10~3 ha) to 2012(2205.65 × 10~3 ha), with an increase of nearly 20-fold. Shandong Province represented the fastest-growing region, with an increase of 31-fold. Moreover, the cultivated species had gradually become diversified. Initially, shellfish and algae were major species, and then it developed to various species including fish, and currently shellfish and crustaceans were the major species. The development of mariculture was driven by government policies, market economy and technology innovation. Rapid development of mariculture also caused significantly adverse impacts on the eco-environment of the coastal regions. For example, the sea reclamation for mariculture decreased the area and function of coastal wetlands and the contaminants originating from mariculture destroyed the coastal aquatic environment. Fortunately, the implementation of ‘13 th Five-Year Fisheries Planning' is expected to help both improve the seafood quality and reduce the contamination in coastal aquatic environment. This current study will provide reference for management and structure adjustment of mariculture in the future.     

An efficient method of noroviruses recovery from oysters and clams

Noroviruses (NoVs) are widespread causes of nonbacterial gastroenteritis. Outbreaks of NoVs caused diseases are commonly ascribed to the consumption of contaminated shellfish. The concentration and RNA extraction of NoVs are crucial steps of detecting NoVs in shellfish. This study aimed to select a simple, rapid and highly efficient recovery method of NoVs detection with real-time RT-PCR. Four methods of recovering GI.3 and GII.4 NoVs from spiked digestive tissues of oysters and clams, respectively, were compared, of them, the method involving proteinase K and PEG 8000 was found the most efficient. With this method, 9.3% and 13.1% of GI.3 and GII.4 NoVs were recovered from oysters and 9.6% and 12.3% of GI.3 and GII.4 NoVs were recovered from clams, respectively. This method was further used to detect NoVs in 84 oysters (Crassostrea gigas) and 86 clams (Ruditapes philippinarum) collected from 10 coastal cities in China from Jan. 2011 to Feb. 2012. The NoVs isolation rates were 10.47% of clams (9/86) and 7.14% of oysters (6/84). All the detected NoVs belonged to genotype GII. The NoVs recovery method selected is efficient for NoVs detection in oysters and clams.     

流沙湾贝类资源调查

对流沙湾潮间带及湾内浅水地带(20°22'103″～20°28'526″N,109°55'341″～109°59'185″E)的贝类资源进行周年调查,定性、定量分析该海湾贝类种类、生态分布、区系性质、群落结构及生物量的季节变化规律。结果表明:在流沙湾采集到的贝类有3纲7亚纲13目42科79属104种,其中帘蛤科24种,牡蛎科、贻贝科各6种,汇螺科和蜒螺科各5种,其他各科均小于5种;贝类组成表现出温带性质,温带种类占绝对优势,该海湾贝类区系属印度—西太平洋的中国-日本亚区,生物量的季节变化明显。     

Morphological and toxicity characteristics of Pseudo-nitzschia pungens strain PP0201-01 isolated from the East China Sea

The morphology and toxicity of Pseudo-nitzschia strain PP0201-01, isolated near the estuary of Changjiang （Yangtze） River in Year 2002 and tentatively identified as Pseudo-nitzschia pungens, were studied. The strain was identified morphologically as P. pungens using scanning electronic microscope （SEM）. The analytical results of LC-PDA （liquid chromatography with photodiode array detector）, LC-MS （liquid chromatography with mass detector）, and the toxicity data from mouse bioassay, showed that no domoic acid was produced by the strain. However, the mechanism of domoic acid production by Pseudo-nitzschia species is complicated. More works are needed to assess the toxic species and to evaluate the potential risk of amnesic shellfish poisoning （ASP） in China.     

Clean and safe supply of fish and shellfish to clear the HACCP regulation by use of clean and cold deep ocean water in Rausu, Hokkaido, Japan

For the supply of fish and shellfish to consumers in fresh condition, clean handling after catch from the sea is essential. According to HACCP （Hazard Analysis and Critical Control Points）, it is important to meet such requirement by keeping fish and shellfish under a certain low temperature and clean conditions after catching. The deep ocean water （DOW） characterized by low temperature and cleanliness has been chosen for fish and shellfish handlings, particularly for salmon, cod, and sea urchin in Town ＇Rausu＇ in Hokkaido, Japan. DOW below 2.9℃ of an amount of nearly 5 000 m^3 is planned to be pumped up every day from a depth of about 350 m, and temporarily stored in a large simulated tank on land. DOW is then supplied to fish boats through hydrants distributed throughout the harbor and used for keeping salmon in clean and cold conditions. Ice made from DOW is also used for lowering temperature if necessary. DOW and ice made from DOW are also used during the transportation of fish and shellfish. The entire system is scheduled to be completed by the summer of 2005.     

Cloning, expressing, and hemolysis of tdh, trh and tlh genes of Vibrio parahaemolyticus

Vibrio parahaemolyticus (VP) is one of the pathogenic vibrios endangering net-cage cultured Pseudosciaena crocea,Fennerpenaeus chinensis, and shellfish in coastal areas of China. Several types of hemolysins produced by Vp have been characterized as major virulence factors.They are thermostable direct hemolysin (TDH),TDH-related hemolysin (TRH) and thermolabile hemolysin (TLH). In this study, we cloned tdh, trh, and tlh genes from the genome DNA of VP by polymerase chain reaction (PCR).We ligated the three genes into prokaryotic expression vector pET-28a (+),and transformed the recombinant plasmids into Es-cherichia coli BL21 (DE3). The expression of recombinant proteins was induced by isopropyl-β-D-thiogalacto-pyranoside (IPTG). The recombinant proteins were expressed in a form of inclusion bodies and thus purified with Ni-NTA affinity chromatography. Western blotting results showed that recombinant proteins,TDH, TRH and TLH, could be recognized by rabbit anti-VP serum. The three purified proteins were renatured by gradient dialysis.The renatured proteins exhibited hemolytic activity except for TLH in the presence of phosphatidylcholine. These results not only are helpful for better understanding these genes’ functions under a single factor level, but also provide evidence for VP vaccine engineering.     

施氏獭蛤的生态观察

施氏獭蛤(LutrariasieboldiiReeve)属暖水性贝类,分布于热带—亚热带海区,在我国东海和南海都有。据在广西和海南沿海的调查,它生活在潮下带至水深78m的沙或沙泥底质中。营埋栖生活,深度达21cm。生活海区的水温为8~32℃,盐度11 .50~33 00,pH值8. 2~8 .4,透明度4~7m。食物以底栖硅藻和有机碎屑为主。繁殖期在5~9月。其肉供食用,是较珍贵的食品,可以发展人工增养殖。