TRPV1 as a key determinant in ciguatera and neurotoxic shellfish poisoning

Ciguatera fish poisoning and neurotoxic shellfish poisoning are distinct clinical entities characterized by gastrointestinal and neurological disturbances, following the consumption of certain reef fish and shellfish containing toxic polyether compounds sporadically present in certain toxic marine dinoflagellates. The biotransformation and bioaccumulation of gambierol and brevetoxin, and their congeners, are believed to be involved in the pathogenesis of these "food-chain diseases", for which no effective treatments are available. Here, we describe for the first time the potent effect of gambierol and brevetoxin on TRPV1 channels, a key player in thermal and pain sensation. Our findings may lead to promising new therapeutic interventions.     

Determination of paralytic shellfish poisoning (PSP) toxins in UK shellfish

A study was conducted to aid the interpretation of data generated by parallel testing of the qualitative Jellett Rapid Test (JRT) and the mouse bioassay (MBA) for detection of paralytic shellfish poisoning (PSP) toxins within the UK statutory shellfish biotoxin monitoring programme. A selection of stored sample extracts subjected to testing by MBA and/or JRT were further analysed by liquid chromatography with fluorescence detection (LC–FLD) to provide additional information on the concentrations of PSP toxins and toxin profiles.Results, from this study, demonstrate the potential of the JRT to effectively screen out PSP toxin negative shellfish samples and samples containing low concentrations of toxins from UK monitoring programmes. Additionally, data generated using LC–FLD highlights the potential of introducing alternative analytical techniques to completely replace the requirement for the MBA.     

The Effect of Overnight Fasting on the Synthesis of Glycerolipids by Liver Slice

The uptake by liver slices of radioactive acetate, palmitate, stearate, linoleate and glycerol into glycerolipids was compared in fed and fasted (overnight, 16 hr) rats.

The incorporation of l-¹⁴C-acetate into long-chain fatty acids and glycerolipids was depressed by fasting. There was a considerable decrease in the incorporation of 1-¹⁴C-palmitate into triglyceride (TG) and that of l-¹⁴C-stearate into phosphatidylcholine (PC) in fasted liver slices. No such differences were observed with l-¹⁴C-linoleate. The incorporation of l-¹⁴C-glycerol into TG was slightly decreased, whereas that into PC and phosphatidylethanolamine (PE) was increased by fasting.

These observations, together with those with the incorporation of the precursors into molecular species of TG, PC and PE, suggested that the changes in the fatty acid composition of glycerolipids by fasting may be governed by the changes in the availability of acyl moieties as well as in the relative balance of the pathways participating to formation of TG and phospholipids.     

A novel type of kleptoplastidy in Dinophysis (Dinophyceae): presence of haptophyte-type plastid in Dinophysis mitra

Red-fluorescent, non-phycobilin-containing plastids were found in the heterotrophic dinoflagellate, Dinophysis mitra. Transmission electron microscopy showed that they contained a three-layer thylakoid, the absence of girdle lamella, and an embedded pyrenoid with thylakoid intrusions. These characteristics all coincide with haptophyte plastids. Phylogenetic analysis of the plastid small-subunit ribosomal DNA (SSU rDNA) revealed that the Dinophysis mitra sequences are distantly related to those of phycobilin-containing Dinophysis species and are positioned within a lineage of haptophytes belonging to Prymnesiophyceae. Because the plastid SSU rDNA sequences of Dinophysis mitra showed significant heterogeneity, despite being derived from a single species, it is highly likely that they were not established as plastids through an evolutionary process but are "kleptoplastids" (temporally stolen plastids) from multiple sources of haptophytes in the environment. We deduced that Dinophysis mitra takes up haptophytes myzocytotically and selectively retains the plastid with surrounding plastidal membranes, whereas other haptophyte cell components are degraded. This represents another type of kleptoplastidy in the Dinophysis species, which mostly harbor cryptophyte plastids, and is the first evidence of kleptoplastidy originating from haptophytes.     

Effect of storage on amnesic shellfish poisoning (ASP) toxins in king scallops (Pecten maximus)

Since 1998, king scallops (Pecten maximus) obtained from Scottish offshore sites have been monitored for domoic acid (DA) and epi-domoic acid (epi-DA), the principal toxic compounds associated with amnesic shellfish poisoning (ASP). The presence of these toxins in king scallops harvested from Scottish waters at concentrations exceeding the current regulatory limit (20 μg g⁻¹ shellfish flesh) is a recurrent event. However, little information was available to determine the effects that different storage conditions experienced during sample transportation to the monitoring laboratory may have on the toxin concentrations, which are subsequently detected. Furthermore, the stability of DA and epi-DA in the solvents (methanol:water (1:1, v/v) and citric acid buffer (0.5 M, pH 3.2)) routinely used for their extraction from shellfish has not previously been assessed. Results from this study demonstrate that when king scallop samples were stored for 2–3 days at 12 °C, a significantly higher toxin concentration was detected in the gonad than when samples were stored at 4 °C and analysed within 48 h. This implies that monitoring programmes must consider transport and storage conditions between harvest and analysis. Stability studies showed rapid decomposition of DA and epi-DA in aqueous methanol extracts while DA and epi-DA seem acceptably stable when stored refrigerated in citrate buffer.     

口蹄疫病毒非结构蛋白3A、3B和2C基因的表达及产物纯化与活性检测

[目的]口蹄疫病毒(FMDV)非结构蛋白(NSP)3A、3B和2C基因的表达及产物纯化与活性检测.[方法]利用原核表达系统表达了FMDV NSP 3A、3B和富含B细胞抗原位点序列的2C蛋白.利用高浓度尿素裂解包涵体,采用稀释法和氧化型、还原型谷胱甘肽系统相结合方法对2C蛋白进行复性.用金属鳌合亲合层析的方法对表达的FMDV NSP 3A、3B和2C进行纯化.采用ELISA方法对比检测了3种纯化蛋白在检测羊血清NSP抗体的效果.[结果]检测得知3A和3B为可溶性表达蛋白,2C以包涵体形式表达.通过Western-blot分析,表明纯化后蛋白能与FMDV感染动物血清发生特异性反应.纯化的3A、3B和复性后的2C融合蛋白与3ABC抗原的检测结果具有很高的符合性.[结论]该研究为建立鉴别FMDV自然感染动物和灭活疫苗免疫动物的酶联免疫电转移印迹技术(EITB)提供了所需的材料.     

Pre-ingestive selection efficiency in two populations of the razor clam Tagelus dombeii with different histories of exposure to paralytic shellfish poisoning (PSP)

We studied toxic effects of the dinoflagellate Alexandrium catenella on filtration activity and pre-ingestive selection efficiency in Tagelus dombeii (razor clam). Samples came from two populations with different histories of exposure to paralytic shellfish poisoning (PSP): Melinka, Aysén (with frequent exposure to PSP) and Corral, Valdivia (without previous exposure to PSP). Feeding activity of T. dombeii was affected by a diet containing A. catenella, showing a reduction in individuals from Corral, Valdivia and Melinka, Aysén. Furthermore, pre-ingestive selection efficiency was significantly higher in specimens from the population of Melinka, than those from Corral. Significantly higher values of clearance rate and pre-ingestive selection efficiency from the Melinka samples may reflect adaptation to specific environmental conditions where PSP events frequently occur.     

Development of Molecular Probes for Dinophysis (Dinophyceae) Plastid: A Tool to Predict Blooming and Explore Plastid Origin

Dinophysis are species of dinoflagellates that cause diarrhetic shellfish poisoning. We have previously reported that they probably acquire plastids from cryptophytes in the environment, after which they bloom. Thus monitoring the intracellular plastid density in Dinophysis and the source cryptophytes occurring in the field should allow prediction of Dinophysis blooming. In this study the nucleotide sequences of the plastid-encoded small subunit ribosomal RNA gene and rbcL (encoding the large subunit of RuBisCO) from Dinophysis spp. were compared with those of cryptophytes, and genetic probes specific for the Dinophysis plastid were designed. Fluorescent in situ hybridization (FISH) showed that the probes bound specifically to Dinophysis plastids. Also, FISH on collected nanoplankton showed the presence of probe-hybridized eukaryotes, possibly cryptophytes with plastids identical to those of Dinophysis. These probes are useful not only as markers for plastid density and activity of Dinophysis, but also as tools for monitoring cryptophytes that may be sources of Dinophysis plastids.