Betanodavirus infections of teleost fish: a review

In the last decade betanodavirus infections have emerged as major constraints on the culture of marine fish in all parts of the world with the exception of the African continent. The occurrence of these infections appears to be a function of the number of species cultured and the intensity of culture. This has been further complicated by the promiscuous translocation of stock within and between countries. Great strides have been made in defining these agents and producing diagnostic techniques but much more remains to be done. Lack of knowledge of the epidemiology of the diseases caused by nodaviruses, except for vertical transmission of the pathogen in some species, has impeded the development of control measures but, even so, the measures identified to date have not been adequately implemented by producers with the result that catastrophic losses still occur on a regular basis.     

Current knowledge of nocardiosis in teleost fish

Nocardia sp. is the causative agent of nocardiosis, a lethal granulomatous disease of the skin, muscle, and various inner tissues affecting various teleost and shellfish. Four species of Nocardia have been isolated from diseased fish and shellfish, namely Nocardia asteroides, Nocardia seriolae, Nocardia salmonicida and Nocardia crassostreae. Therefore, in fish aquaculture, nocardiosis has caused severe economic losses, especially in the Asian region. Considerable research has been performed, since the first report of identified Nocardia sp. in fish, to characterize Nocardia sp. and identify rapid detection techniques, immune response against infection and prophylactic approaches. In this review, the current state of knowledge about nocardiosis in fish has been presented, including the pathogenesis, diagnosis, host immune response and vaccine development.     

鱼类趋化因子家族的研究进展

趋化因子(chemokines)是一类分子量仅 8~10 kD 的小分子细胞因子,由巨噬细胞、成纤维细胞、T 细胞和 B 细胞等表达分泌。按生理功能的差异可将趋化因子分为组成型和诱导型,前者通常参与淋巴细胞的迁移、定位以及免疫监视, 而后者仅在感染或炎性刺激时分泌并诱导白细胞迁移至炎症部位。根据氨基酸序列中前两个半胱氨酸(Cys)的排列方式, 可以将趋化因子分为 4 类: CXC 亚族、CC 亚族、C 亚族和 CX3C 亚族。鱼类至今尚未发现有关 CX3C 亚族趋化因子的研究报道, 但拥有一类特有的 CX 亚族。除了基本的募集和激活白细胞功能外, 鱼类趋化因子在免疫应答、应激反应、胚胎发育及血管生成等方面也发挥着重要作用。本文从鱼类趋化因子及其受体的分类、鉴定、结构、表达以及功能等方面进行了综述。     

The ovarian regulation of ovulation in teleost fish

Ovulation is the release of a mature oocyte from its follicle wall enclosure in the ovary. This process requires the separation of the oocyte from the granulosa layer, the rupture of the follicle wall and the active expulsion of the oocyte through the rupture site. Results of experiments on various vertebrates, including fish, have shown that the control of these processes may involve the cooperative action of a number of ovarian regulators including proteases, protease inhibitors, progestational steroids, eicosanoids, catecholamines and vasoactive peptides. We have used two teleost models, the brook trout (Salvelinus fontinalis) and the yellow perch (Perca flavescens) to study the mechanism and control of ovulation in fish. Using subtractive cDNA cloning, a family of ovarian and ovulation specific mRNAs (TOPs = trout ovulatory proteins) was isolated from the brook trout ovary. These mRNAs have not previously been observed in the ovary of any vertebrate species; however, the proteins they code for have significant sequence homology to a group of mammalian protease inhibitors called antileukoproteinases. These inhibitors have been isolated from several mammalian mucosal tissues and their function may be to protect the mucosal lining from nonspecific degradation by proteases released from infiltrating leukocytes. The ovarian proteins encoded by the TOP mRNAs have now been characterized by Western blotting using antibodies derived against recombinant TOPs. Given the similarity of TOPs to antileukoproteinases, one function of TOPs may be to regulate proteolysis at the time of ovulation. In yellow perch, the maturational steroid, 17,20-dihydroxy-4-pregnen-3-one (17,20-PG), stimulates both germinal vesicle breakdown and ovulation in vitro. The stimulation of ovulation can be blocked by indomethacin, a prostaglandin endoperoxide synthase inhibitor. Thus, it appears that 17,20-PG acts through the production of an eicosanoid that is most likely a primary prostaglandin. This hypothesis is further supported by the observations that (1) a direct correlation exists between indomethacin levels that block ovulation and those that block primary prostaglandin synthesis in the ovary; (2) ovulation can be restored in indomethacin-blocked incubates with primary prostaglandins; (3) PGF levels increase at the time of ovulation in incubations of yellow perch follicles stimulated with 17,20-PG; and (4) the stimulation of PGF by steroids in the ovary is specific for 17,20-PG. Finally, 17,20-PG-stimulated ovulation and follicular prostaglandin synthesis requires the close interaction of extrafollicular tissue and other follicle wall layers.     

硬骨鱼免疫相关microRNA研究进展

MicroRNA(miRNA)是一类长度为18～25 nt的非编码RNA,在硬骨鱼免疫应答中起重要作用.近年来,借助高通量测序与荧光定量PCR(RT-PCR)技术,筛选到大量免疫相关的差异表达miRNA(DE miRNA).通过比较多种硬骨鱼在不同病毒/细菌感染条件的miRNA表达谱,筛选出相同DE miRNA,并认为...     

Immunochemical relationships of cytochrome P4503A-like proteins in teleost fish

Multiple P450 proteins have been purified from several teleost species, including rainbow trout (Oncorhynchus mykiss), scup (Stenotomus chrysops) and Atlantic cod (Gadus morhua). Identity, relationships and/or functions have been established in these fish species for the cytochrome P4501 As. Information about the structure, function, regulation and relationships of other piscine cytochrome P450 (CYP) proteins is sparse. In the present study we have focused on constitutively expressed CYP forms, P450con and LMC5 isolated from rainbow trout, P450A from scup, and P450b from Atlantic cod, and we consider evidence for the relationship of these proteins to mammalian members of the CYP3A subfamily. Reciprocal western blot analysis shows that P450con and LMC5, isolated from rainbow trout in two different laboratories, are closely related and ostensibly identical proteins. These trout proteins show specific reciprocal cross-reactivity with scup P450A, and polyclonal antibodies (PAb) to the trout and scup proteins both recognize cod P450b, indicating that rainbow trout P450con/LMC5, scup P450A and cod P450b are immunochemically-related proteins. In analyses of liver microsomes of trout, scup and cod, PAb to trout P450con/LMC5 and scup P450A recognize only bands that are identical in migration to the CYP proteins purified from these species, and which were used as immunogens. These CYP proteins purified from fish are each immunochemically-related to mammalian CYP3A proteins, showing recognition by PAb to human CYP3A4 and to rat CYP3A1. PAb to the mammalian CYP3As also recognize the same bands in liver microsomes from these fish species as seen by PAb to the fish proteins. These results strongly suggest that these fish proteins are members of theCYP3 gene family and probably theCYP3A subfamily. Although sequence analysis is required before their designation in the CYP3A subfamily can be confirmed and specified, we refer to these as CYP3A-like. Immunoblot analyses of hepatic microsomes from other fish species with PAb to scup P450A and trout P450con show that multiple CYP3A-like proteins are expressed in liver of several species, including killifish (Fundulus heteroclitus) and winter flounder (Pleuronectes americanus). Important questions still remain to be addressed concerning CYP3A structure, multiplicity, physiological function, regulation and metabolism of endogenous as well as exogenous substrates in fish.Part of this study was presented at the 10th International Symposium on Microsomes and Drug Oxidations. Toronto, Canada, July 18–21, 1994.     

氯化钴在大菱鲆饲料中的安全性评价

不同鱼类的脂肪组织分布模式具有高度多样性。大菱鲆具有相对特殊的脂质储存模式;鳍条附近皮下脂肪组织是其重要的脂肪存储部位。为了更好地了解大菱鲆的脂质代谢生理,本实验初步研究了29个脂质代谢相关基因在大菱鲆体内的组织表达,这些基因参与了脂肪生成、脂肪酸β氧化、甘油酯的生物合成和水解、脂质运输以及相关的脂代谢转录调控过程。从30尾鱼(10尾混样作为一个重复)中采集眼、鳃、脑、皮肤、肌肉、肝脏、胃、肾脏、脾脏、心脏、前肠、幽门盲囊、后肠、盲肠和脂肪等共15个组织样本进行qRT-PCR分析。结果显示,肠和脑中脂肪生成基因表达量高,肝脏和肌肉脂肪生成基因表达量低。肠内大部分载脂蛋白和脂质代谢相关转录因子的基因表达水平也较高。肌肉中脂肪酸β氧化相关基因的表达水平较低。细胞内甘油酯酶在脑、眼和心脏中高表达。研究表明,在大菱鲆体内,肠道可能不仅是脂质摄取的场所,也是大菱鲆体内重要的脂肪代谢器官。本研究为进一步探讨大菱鲆脂代谢特征及探索不同储脂类型鱼类中脂肪代谢的多样性提供了基础数据,也将有助于在分子水平上进一步研究鱼类的脂质代谢调节。

     

Estrogen-induced and suppressed gene expression during XY sex reversal in a teleost fish, tilapia

To identify the down-stream gene products of estrogen during ovarian differentiation, we performed subtractive hybridization using mRNA derived from normal and estrogen treated XY gonads. The clones obtained after subtractive hybridization screening were re-screened by reverse-Northern analysis. Finally seven up-regulated and three down-regulated gene products were obtained.     

The role of phospholipids in nutrition and metabolism of teleost fish

It has been known for almost 25 years now that inclusion of intact phospholipids in the diet could improve culture performance of various freshwater and marine fish species. The primary beneficial effect was improved growth in both larvae and early juveniles, but also increased survival rates and decreased incidence of malformation in larvae, and perhaps increased stress resistance. Determination of absolute dietary requirements has been hampered by the use, in different dietary trials, of a wide range of phospholipid preparations that can vary greatly both in phospholipid content and class composition. Larval studies have been compromised further by the need on many occasions to supply phospholipid through enrichment of live feeds with subsequent re-modelling of the phospholipid and fatty acid compositions. Generally, the levels of phospholipid requirement are around 2–4% of diet for juvenile fish and probably higher in larval fish. The effects were restricted to young fish, as a requirement for dietary phospholipids has not been established for adult fish, although this has been virtually unstudied. As the majority of studies have used crude mixed phospholipid preparations, particularly soybean lecithin, but also other plant phospholipids and egg yolk lecithin, that are enriched in several phospholipids, it has been difficult to elucidate which specific phospholipid classes impart beneficial effects. Based on the few studies where single pure phospholipid species have been used, the rank order for efficacy appears to be phosphatidylcholine > phosphatidylinositol > phosphatidylethanolamine > phosphatidylserine. The efficacy of other phospholipid classes or sphingolipids is not known. The mechanism underpinning the role of the phospholipids in larval and early juvenile fish must also explain their lack of effect in adult fish. The role of phospholipids appears to be independent of fatty acid requirements although the presence of an unsaturated fatty acid at the sn-2 position may be important. Similarly, the phospholipid requirement is not related to the delivery of other essential dietary components such as the bases choline and inositol. Studies also suggested that the phospholipid effect was not due to generally enhanced emulsification and digestion of lipids. Rather the evidence led to the hypothesis that early developing stages of fish had impaired ability to transport dietary lipids away from the intestine possibly through limitations in lipoprotein synthesis. The current hypothesis is that the enzymic location of the limitation is actually in phospholipid biosynthesis, perhaps the production of the glycerophosphobase backbone and that dietary supplementation with intact phospholipids in larvae and juvenile fish compensated for this. Thus, dietary phospholipids increase the efficiency of transport of dietary fatty acids and lipids from the gut to the rest of the body possibly through enhanced lipoprotein synthesis.     

Total and free thyroid hormones in plasma of tropical marine teleost fish

Plasma levels of L-thyroxine (T₄) and 3,5,3-triiodo-L-thyronine (T₃) and the percentage of plasma T₄ and T₃ present in the free (dialyzable) form (%FT₄ and %FT₃) were measured in 16 species (11 families) of tropical marine teleosts from an inshore Barbados reef. Mean plasma T₄ varied from 0.2 ng/ml to 42 ng/ml; mean plasma T₃ varied from < 0.2 ng/ml to 50 ng/ml. The highest T₄ and T₃ levels were recorded in parrot-fish and the lowest levels in filefish. The %oFT₄ and %FT₃ varied from 0.05–3.41%. Estimated levels of plasma free T₄ and free T₃ levels ranged from 0.4–466 pg/ml. The extremely wide inter- and intra-species ranges in levels of free T₄ and T₃ do not support a previous suggestion, based on temperate freshwater salmonid species, that free T₄ and T₃ levels in fish may fall within a relatively range narrow comparable to that of homeothermic vertebrates.     

Administration of gulonolactone does not evoke ascorbic acid synthesis in teleost fish

Evidence is presented that for common carp (Cyprinus carpio L.), intraperitoneal injection of L-gulonolactone, a precursor of ascorbic acid synthesis in the D-glucuronic acid pathway, does not result in an increased concentration of ascorbate in tissue. Control fish injected with an equimolar amount of ascorbic acid have shown a significant increase in ascorbic acid concentration in the kidney, hepatopancreas, plasma and spleen. The ascorbate status in the carp body,i.e., the ascorbate nutritional history, produced significant differences in ascorbate withdrawal from circulation and probably in the catabolic rate. Acute fasting decreased ascorbate uptake into tissues as compared to fish fed a diet lacking ascorbate. Intraperitoneally injected ascorbate affects common carp being fed a diet containing 295 mg of total ascorbic acid kg^–1 by causing tissue to become saturated with vitamin C, similar to the tissues in the group undergoing acute fasting. There was no gulonolactone oxidase activity in the hepatopancreas of the common carp. These results suggest that the metabolic rate induced by feeding is the primary factor regulating ascorbate requirement.     

硬骨鱼类鳃黏膜免疫相关分子的研究进展

鳃为鱼类重要的呼吸器官,是鱼类进行离子交换、酸碱调节和含氮废物排泄的重要结构基础,也是鱼类重要的外周黏膜免疫器官之一,在抵御病原微生物侵染过程中发挥重要的免疫屏障作用。当前,硬骨鱼类鳃黏膜免疫反应是研究热点之一。本文首先对硬骨鱼类鳃的结构和特点进行分析,之后综述了抗菌肽、干扰素、白细胞介素、Toll样受体、补体等先天性免疫相关分子以及T细胞受体和免疫球蛋白等适应性免疫相关分子在硬骨鱼类鳃黏膜中的表达规律、分子功能,最后探讨了化学因素(重金属、杀虫剂等)、生物因素(细菌、病毒、真菌、和寄生虫等)以及营养物质和疫苗等对硬骨鱼类鳃黏膜结构的影响,以期为深入研究鳃在鱼类黏膜免疫反应中的角色和应答机制提供指导,为硬骨鱼类病原性疾病的免疫防控策略的制定提供理论基础。     

Natriuretic peptides and the control of catecholamine release in two freshwater teleost and a marine elasmobranch fish

Experiments were carried out in situ and in vivo to investigate the relationship between natriuretic peptides (NPs) and humoral catecholamine secretion in the American eel (Anguilla rostrata), rainbow trout (Oncorhynchus mykiss) and spiny dogfish (Squalus acanthias). In situ perfusion of the chromaffin tissue of A. rostrata with homologous atrial NP (ANP; 10–9 mol l–1) or ventricular NP (VNP; 10–9 mol l–1), or O. mykiss with either rat ANP (10–9 mol l–1), eel VNP (10–9 mol l–1), or trout VNP (10–9 mol l–1), did not significantly affect basal or carbachol-elicited (10–5 mol kg–1) catecholamine release. Bolus injections of homologous ANP (10–9 mol kg–1) or VNP (10–9 mol kg–1) in A. rostrata in vivo elicited a rapid and prolonged reduction in arterial blood pressure and an increase in heart rate (fH); circulating plasma catecholamine levels were unaffected. In O. mykiss, bolus injections of rat ANP (10–9 mol kg–1) or trout VNP (10–9 mol kg–1) elicited a significant bi-phasic pressor- depressor response and a marked increase in fH. Neither the acute pressor or the longer-term depressor effects of NPs in O. mykiss were associated with any change in circulating plasma catecholamine levels. In S. acanthias, bolus injections of homologous C-type natriuretic peptide (CNP; 10–9 mol kg–1) elicited a bi-phasic pressor-depressor response, an increase in systemic resistance, a decrease in cardiac output and stroke volume, but no change in fH. Plasma noradrenaline levels were elevated 15- fold after CNP injection while circulating adrenaline levels remained unchanged. These results show that NPs of systemic origin do not directly or indirectly affect basal or cholinergically-mediated catecholamine release in A. rostrata and O. mykiss and that the initial pressor response to NP's in trout cannot be attributed to an elevation of circulating catecholamines. Conversely, CNP appears to be a potent secretagogue (direct or indirect) of noradrenaline release in S. acanthias and thus there is likely to be a significant humoral adrenergic component to the cardiovascular effects of NPs in this species.     

Rapid response of fish and aquatic habitat to removal of a tidal barrier

1. River barrier removal is used increasingly as a conservation tool to restore lotic habitat and river connectivity, but evidence of its efficacy is incomplete. This study used a before–after methodology to determine the effects of removing a tidal-limit barrier on the fishes, macroinvertebrates, and habitats of an English coastal stream.
2. Following barrier removal, habitat diversity increased immediately upstream and remained similar downstream. Mobilized silt altered the substrate composition immediately downstream, but this was temporary as silt was flushed out the following winter. Changes to macroinvertebrate communities occurred upstream and downstream of the former barrier but these were transient.
3. A dramatic and sustained increase in fish density occurred immediately upstream of the barrier after its removal, but effects downstream were minor. The fish community upstream changed, largely due to rapid recruitment and dispersal of endangered European eel (Anguilla anguilla). Eel density in the formerly impounded zone increased from 0.5 per 100 m² before barrier removal to 32.5 per 100 m² 5 months after removal. By 17 months after barrier removal there was no difference in eel density across the six sections sampled.
4. Although resident stream fishes such as bullhead (Cottus gobio species complex, protected under the European Habitats Directive) were abundant in middle and upper-stream sections, brown trout (Salmo trutta, a listed species for biodiversity conservation in England and Wales) density remained low during the study and recruitment was poor. This suggests that although colonization access for anadromous trout was available, habitat upstream may have been unsuitable for reproduction, indicating that wider catchment management is required to complement the restoration of connectivity.
5. These findings suggest that tidal barrier removal is an effective method of restoring lotic habitats and connectivity, and can be beneficial for resident and migratory fishes including those of conservation importance (e.g. European eel) in coastal streams.

     

A novel technique for the separation of yolk from the developing embryonic tissue in a teleost fish, Oncorhynchus mykiss

We have developed and validated a novel technique that allows for a relatively quick and efficient isolation of embryonic tissue from the yolk in the rainbow trout, Oncorhynchus mykiss. After removal of the chorion, the yolk is first dissolved in an imidazole-KCl buffer and separated from the embryo proper and yolk sac (tissues) by centrifugation. SDS gel was used to identify proteins of 165, 48, 35 and 10 kDa unique to the embryonic tissue and a 6 kDa protein unique to the yolk. Using protein analysis and enzyme activity measurements (lactate dehydrogenase and glutamate dehydrogenase) of each fraction, we have determined that cross contamination is less than 10%. This new method will facilitate biochemical studies of the embryo and yolk that were previously very time consuming. This revised version was published online in August 2006 with corrections to the Cover Date.     

A comparison of sex steroid binding protein (SBP) in four species of teleost fish

A sex steroid binding protein (SBP) binding E₂ with high affinity has been detected in a pleuronectid (greenback flounder Rhombosolea tapirina), two sparids (black bream Acanthopagrus butcheri and snapper Pagrus auratus), and its presence has been confirmed in a salmonid (rainbow trout Oncorhynchus mykiss). SBP binding characteristics were measured using a hot saturation assay for trout, bream and snapper, and a cold saturation assay for flounder. Bound and unbound steroid were separated by incubation with dextran-coated charcoal (DCC). Affinity for E₂ was highest in trout (k_D=0.44 nM), followed by bream (k_D=3.39 nM) and snapper (k_D=10.7 nM). The lowest affinity was found in flounder (k_D=84.7 nM). Binding capacity, however, was greatest in flounder (B_max=164 nM), followed by trout (B_max=92 nM), and then bream and snapper (B_max=50 and 39 nM, respectively). Binding of E₂ to SBP had a very rapid rate of association, and most dissociation occurred within 5 min. To confirm that the plasma protein measured here was SBP, the relative binding affinities of SBP for a range of steroids were measured. In trout, bream and snapper, SBP bound E₂ with the highest affinity, followed by T. In contrast, the relative affinity of T for flounder SBP was more than twice that of E₂. The rank orders of affinity of binding indicate the importance of an unhindered 17-hydroxyl group, and a 3-hydroxyl or 3-ketone group for high affinity binding to SBP. These requirements for high affinity binding are present in most animals possessing SBP and indicate conservation of the SBP molecule through evolution.     

A new method for recording activities of spinal motoneurons during fictive swimming in paralyzed teleost fish

ABSTRACT: A new method for monitoring spinal motoneuron activities during fictive swimming in teleost fish was developed. Enamel-insulated copper wire electrodes were implanted in the trunk muscle of goldfish or carp. For each freely moving fish, an electromyogram (EMG) was recorded using the electrodes. In those fish paralyzed with curare, using the same electrode set, bursts of electrical activities consisted of spikes of smaller amplitude and of shorter duration compared with those recorded by EMG. Simultaneous recording of the extracellular activity and intracellular recording from the muscle revealed that the bursts of spikes recorded in the paralyzed fish were motor nerve impulses innervating the muscle and are considered to be fictive swimming activity. The method developed in the present study provides a useful tool with which to investigate neuronal mechanisms underlying swimming activity in teleost fish.     

Procedures for evaluating and prioritising the removal of fish passage barriers: a synthesis

Abstract Techniques for assessing the impact of structural barriers on fish passage and for prioritising restoration actions are reviewed. Current survey methodologies are biased towards specific structures, primarily culverts and economically significant fish. Assessment criteria are often based on swimming capabilities of upstream migrating adult salmonids, while ignoring other life‐stages, non‐salmonid species, downstream migration and behaviour. The development of comprehensive and centrally owned geospatial inventories of barriers is essential. The collection, maintenance and dissemination of pertinent structural and environmental data can be technically, logistically and financially challenging. Standardised procedures are needed to rapidly and cost‐effectively survey large numbers of barriers over wide geographic areas. The prioritisation of barrier repair and removal projects is most often based on simple cost‐benefit analysis, whereby individual barriers are scored based on a set of assessment criteria and then ranked in order of priority. The benefits of using scoring‐and‐ranking systems, however, are unacceptably low because they consider barriers independently, thereby ignoring the cumulative, non‐additive impacts produced by multiple, spatially interconnected structures. Optimisation modelling offers a more robust approach for efficiently prioritising decision making in river restoration planning, allowing decision makers to account for key uncertainties and effectively balance multiple, possibly competing, environmental and socioeconomic goals and constraints.