Use of an optical biosensor with a silicone-immobilized enzyme to determine plasma total cholesterol concentrations in fish

Decreased plasma total cholesterol concentrations are a useful indicator in fish of reduced resistance to bacterial infection. Taking advantage of the hydrophobic properties of silicone rubber, we developed an optical enzyme sensor system for measuring plasma cholesterol concentrations in fish. The system was constructed using an immobilized enzyme membrane, optical oxygen probe, flow system, and personal computer. The enzyme membrane was prepared from cholesterol esterase, cholesterol oxidase, and silicone rubber. The calibration curves were linear in the range of 55–85 mg dl⁻¹ for yellowtail and 20–90 mg dl⁻¹ for rainbow trout. Assays could be completed within 3 min, and the sensor response was stabilized by the hydrophobic properties of silicone rubber. Storage of immobilized enzyme membranes at 5°C permitted stable measurements for 58 days. The sensor system was applied to determine plasma total cholesterol concentrations in fish. Good correlations were obtained between results obtained using the sensor and results obtained using conventional methods (correlation coefficients: yellowtail 0.9751, rainbow trout 0.9947). Our method required much less time than conventional cholesterol assays and can be used economically for continuous determination of plasma total cholesterol in fish.     

Development of biosensor for measuring oxidative stress of fish

Fisheries Science - To elucidate the dynamics of oxidative stress in fish, it is necessary to know the concentration of superoxide anions as a precursor to various reactive oxygen species in the...     

计算机视觉技术在鱼类应激状态监测中应用研究

从行为参数的量化方法、应激状态下的行为变化两方面介绍了计算机视觉技术在鱼类应激状态监测方面的研究进展,并提出了进一步研究的方向是实际应用过程中量化鱼行为的新方法研究,以及根据鱼在不同条件下行为参数的变化特点,结合统计方法和人工智能算法建立自动判断应激的方法。     

Determination of DNA content of whole fish

ABSTRACT:   The DNA content of whole fish from 31 different North Atlantic fish species was determined. Sample preparation was performed on board the fishery research vessel 'Walter Herwig III' directly after catch and sorting. Up to three homogenates were prepared of each 10 specimens per species by mincing the whole fish. The homogenates were heated to destroy nuclease activity, and then stored at frozen temperature until being analyzed in the laboratory. Measurement of DNA content was carried out using the DNA specific fluorescent dye Hoechst 33258 (Serva Biochemica, Heidelberg, Germany). Matrix effects as light scattering and quenching could be estimated by use of standard addition of calf thymus DNA. DNA contents were referred to dry weight of sample material, and ranged from 570 µg to 3500 µg/g dry weight of homogenate. The coefficient of variation did not exceed 25% of mean for one species; coefficient of variation for all investigated species did not exceed 30% of mean, which was 1020 µg/g dry weight.     

Mediator-type biosensor for real-time wireless monitoring of blood glucose concentrations in fish

We developed a wireless mediator-type biosensor system to rapidly monitor blood glucose concentrations in free-swimming fish. Glucose oxidase was used to determine the glucose concentration, but fluctuations in the oxygen concentrations in the samples affected the measurements. In this work, we used a mediator, a synthetic electron acceptor, as a substitute in the oxidation–reduction reaction, thus ensuring that changes in the concentration of oxygen do not affect the measurements of glucose concentration performed by the sensor. We especially focused on the use of ferrocene as a mediator. The resulting enzyme sensor utilized a Pt–Ir wire (diameter 0.178 mm) as the working electrode and Ag/AgCl paste as the reference electrode. Chitosan was used to fix ferrocene and glucose oxidase to the working electrode. The characteristics of the sensor were tested, and it was found to be capable of measuring glucose independent of the oxygen concentration of the sample. Its reproducibility was also tested, and the results showed that the sensor was suitable for performing measurements in vivo. The sensor was then inserted into to eyeball interstitial sclera fluid in order to wirelessly monitor the glucose concentration in free-swimming fish. Measurements were taken for a total of 48 h. During these measurements, artificial stress was applied to the fish. The glucose concentrations of fish rise with their stress levels. The output current from the sensor gradually increased during the application of stress, which hinted that the stress was monitored by this system.     

次黄嘌呤生物传感器检测鱼类新鲜度之研究

检测鱼类新鲜度的常规方法是测定其总挥发性盐基氮（ＴＶＢ－Ｎ）或细菌菌落总数，手续繁琐且代价较高，本文提出的方法是一种新颖的快速检测法。实验表明，在鱼的腐败过程中，总挥发性盐基氮（ＴＶＢ－Ｎ，国标）、细菌落落总数和次黄嘌呤（Ｈｘ）的量之间皆存在着良好的相关性。因此，鱼腐烂而累积于组织的次黄嘌呤量可以指示鱼类的新鲜度，生物传感器快速检测法将取代现有的常规手段。     

鱼类铁调素（Hepcidin）的研究进展

铁调素是一类具有调节铁代谢功能的抗菌肽,在鱼类中广泛存在。文章介绍了Hepcidin的基本结构、生理功能,并总结了鱼类Hepcidin基因的克隆和表达、抗菌活性、铁代谢活性的研究进展,以期为鱼类Hepcidin的开发和利用奠定一定的基础。     

Quantification of cholesterol oxidation products in commercial fish meals and their formation during storage

Twenty‐two samples of commercial fish meals from Norway, Chile and Peru were analysed for cholesterol and oxysterols using gas chromatography. Cholesterol content ranged from 25.2 to 64.8 g kg^?1 total lipids. Detectable levels of the oxysterols 7β‐hydroxycholesterol and 7‐ketocholesterol were found and their identity was confirmed by gas chromatography‐mass spectrometry. Samples of fish meal exhibited wide variability in oxysterols content, 7β‐hydroxycholesterol ranging from 3.9 to 105.6 mg kg^?1 total lipids (0.4–9.4 mg kg^?1 dry matter) and 7‐ketocholesterol from 2.0 to 56.2 mg kg^?1 total lipids (0.2–5.0 mg kg^?1 dry matter). The formation of 7β‐hydroxycholesterol and 7‐ketocholesterol in one sample of Norse LT94 fish meal stored at room temperature was also studied. Oxysterol content increased during the first 42 days of storage by about 350% and then decreased with further storage. The low amount of oxysterols measured indicates a limited degree of cholesterol oxidation in commercially available fish meals.     

The use of anglers' catches to monitor fish populations in large water bodies, with particular reference to the River Derwent, Derbyshire, England

Abstract. Nearly 2000 marked coarse fish of four species were introduced into a stretch in the middle reaches of the River Derwent. Derbyshire, England in an attempt to estimate the fish population size from catch returns. It was estimated that 800 dace, 1975 roach, 1481 chub and 366 bream were present in the reach and the density of the fish population (0·35 fish m^?2) was low. The validity of employing anglers as samplers in a mark-recapture exercise to estimate fish population size in large fisheries is discussed. Angler catch data were collected from angling clubs holding competitions on the Hoveringham Fishery on the lower reaches of the River Derwent over the period 1973/1974 to 1983/1984 to assess the impact of achange in management strategy on the fishery in 1979. Catch rates improved gradually until 1979/1980 when a dramatic increase occurred. This elevated catch rate, in excess of 400 g man-h^?1, was maintained and, in recent years, further enhanced (>500 g man-h^?1). The improvement was the result of a strong year-class of chub entering the catchable cohort and dominating anglers' catches and not the change in management policy.     

Pyridinoline concentrations in muscular and skin collagen of fish and relationship between collagen solubility and pyridinoline concentration in fish muscular collagen

Pyridinoline (Pyr), one of the mature crosslinks of collagen, was determined in muscular collagen of three species of fish. The amounts of muscular Pyr in red sea bream, yellowtail, and tiger puffer were 3.4, 8.8, and 50.3 mmol/mol collagen, respectively, indicating that the Pyr concentration in muscular collagen differs greatly among fish species. The Pyr concentration in tiger puffer muscular collagen was the greatest, but it was only one-fourth that in rabbit muscle. As in mammalian skin collagen, Pyr was not detected in skin collagen of red sea bream and yellowtail. However, tiger puffer skin contained Pyr (3.75 mmol/mol collagen). The presence of Pyr would have a relationship to some features of tiger puffer skin, such as mechanical strength and thickness. Pyr concentrations in acid-soluble collagen (ASC), pepsin-solubilized collagen (PSC), and insoluble collagen (ISC) in muscles of three species of fish were determined. Pyr was found in ISC > PSC > ASC, from the highest to the lowest concentration, and the concentration in ISC was 45–200 times that in ASC. Therefore, Pyr crosslinks that are formed between collagen molecules would have a close relationship to collagen solubility.     

Recovery of fish from functional and haemolytic anaemia after brief exposure to a lethal concentration of nitrite

Total haemoglobin and methaemoglobin concentration in blood was analyzed in sea bass (Dicentrarchus labrax) exposed for 18 h to 150 mg/l NO₂-N in sea water. Observations of recovery mechanisms carried out for 24 days following the nitrite exposure showed that, after a rapid disappearance of methaemoglobinaemia, a strong anaemia occurred, due to a drop in the total haemoglobin concentration. Complete recovery from this anaemia occurred only after 24 days. The mechanism of recovery from nitrite intoxication is discussed with a view to devising treatment that might help to minimize losses at fish-farming facilities.     

The whole amino acid profile as indicator of the nutritional condition in cultured marine fish larvae

The effect of variations in the amount or quality of food provided on the amino acid profile of larvae, was tested in two marine fish species, the Senegal sole (Solea senegalensis) and the sea bass (Dicentrarchus labrax). The objective was to assess whether such nutritionally suboptimum dietary patterns, which simulate situations that may occur in hatcheries, result in variations in the concentration of one or more amino acids that could be used as indicators of nutritional condition of larvae. Restriction in the normal food ration by 60% had no clear effect on the amino acid profile of sea bass, while the concentration of some amino acids (Arg, Ala and Phe) showed significant variations in sole. Feeding on artificial diets, which have shown their inadequacy a priori, resulted also in no significant effect on the profile of sea bass, but in a dramatic modification of the amino acid profile in sole. In addition, changes in the amino acid profiles considered as a whole were evaluated using the cluster analysis instead of paired comparisons between amino acid concentrations in each treatment. The analysis clearly separated profiles of larvae fed restrictedly or inadequately from their controls, irrespective of the species. This demonstrated the potential value of evaluating whole changes in amino acid profile as nutritional indicator. Besides, it was demonstrated that sensitivity of larvae to nutritional changes that may be reflected in the amino acid profile is greatly conditioned by the species and developmental stage.     

灯笼鱼资源的开发与利用

灯笼鱼属于中深海鱼类,广泛分布于全球各大洋中,资源量巨大。灯笼鱼通常被用于加工饲料鱼粉和鱼油,市场价值较低,因此被称为低值鱼类,长期不被人们重视。近年来,随着海洋渔业资源日趋紧张,人们开始重新关注全球的灯笼鱼资源,在灯笼鱼的资源调查、探捕和产品加工方面进行了大量的研究和实践,开发出了可供人类消费的灯笼鱼食品。尽管在灯笼鱼的研究和开发利用方面还有许多问题尚待解决,但是越来越多的渔业专家对灯笼鱼资源开发利用的前景抱有信心。     

Comparing geometric morphometrics and outline fitting methods to monitor fish shape variability of Diplodus puntazzo (Teleostea: Sparidae)

The definition of a remote system in the monitoring of fin fish shape variability relies on the appropriate shape analysis tools. In this paper, different methods are proposed and compared. Growth trajectories, computed using geometric morphometrics, allow the pattern of size-related shape change and visualize growth allometry to be analyse. Geometric morphometrics is used to describe shape change in three samples (507 5100 days after hatching) of 119 individuals of the Sparid Diplodus puntazzo. The results are potentially interesting but the method still relies on the manual detection of homologous landmarks (coordinates of points located unambiguously on each specimen’s profile or structure) which are basic for such analysis. In an attempt to proceed towards the remote estimation and the automation of data collection, two alternative approaches are tested. In particular, Elliptic Fourier analysis and Bezier function were applied to the same samples, and results compared with geometric morphometrics. Fourier analysis’ growth curves are very similar to geometric morphometrics’ ones and allows for an appropriate visualisation of global shape changes. At the same time, Fourier methods can be applied to remotely and automatically detected profiles. This method is thus considered as the best candidate in the context of automated fish shape monitoring. The Bezier method performed poorly, both in terms of statistical output and shape visualization.     

Fish welfare assurance system: initial steps to set up an effective tool to safeguard and monitor farmed fish welfare at a company level

The objective was to take a first step in the development of a process-oriented quality assurance (QA) system for monitoring and safeguarding of fish welfare at a company level. A process-oriented approach is focused on preventing hazards and involves establishment of critical steps in a process that requires careful control. The seven principles of the Hazard Analysis Critical Control Points (HACCP) concept were used as a framework to establish the QA system. HACCP is an internationally agreed approach for management of food safety, which was adapted for the purpose of safeguarding and monitoring the welfare of farmed fish. As the main focus of this QA system is farmed fish welfare assurance at a company level, it was named Fish Welfare Assurance System (FWAS). In this paper we present the initial steps of setting up FWAS for on growing of sea bass (Dicentrarchus labrax), carp (Cyprinus carpio) and European eel (Anguilla anguilla). Four major hazards were selected, which were fish species dependent. Critical Control Points (CCPs) that need to be controlled to minimize or avoid the four hazards are presented. For FWAS, monitoring of CCPs at a farm level is essential. For monitoring purposes, Operational Welfare Indicators (OWIs) are needed to establish whether critical biotic, abiotic, managerial and environmental factors are controlled. For the OWIs we present critical limits/target values. A critical limit is the maximum or minimum value to which a factor must be controlled at a critical control point to prevent, eliminate or reduce a hazard to an acceptable level. For managerial factors target levels are more appropriate than critical limits. Regarding the international trade of farmed fish products, we propose that FWAS needs to be standardized in aquaculture chains. For this standardization a consensus on the concept of fish welfare, methods to assess welfare objectively and knowledge on the needs of farmed fish are required.     

A method for the histochemical localization of digestive enzymes in whole freeze-substituted larval fish embedded in glycol methacrylate

In order to study the localization of digestive enzymes in larval walleye ( Sander vitreus vitreus), a novel method of low-temperature processing of whole, unfixed larvae and subsequent embedding in glycol methacrylate (GMA) was developed. Larval walleye aged 2–10 days post hatch were flash-frozen in liquid nitrogen and transferred into pre-chilled acetone for 12 h at a temperature of −25 °C. Acetone was gradually replaced with increasing concentrations of GMA resin monomer over a 6-h period. The polymer (100%) was further allowed to infiltrate the larvae for 36 h. Resin-embedded larvae were transferred to embedding moulds and polymerized overnight at −25 °C. Four micrometre sections were stained to identify either alkaline phosphatase, non-specific esterase, aminopeptidase M or dipeptidyl peptidase IV. All enzymes studied were readily detected and accurately localized, and no enzyme diffusion was observed. Therefore, freeze substitution combined with low-temperature GMA embedding allows the maintenance of excellent tissue morphology and accurate enzyme localization in whole larval walleye specimens from 2 to 10 days post hatch. It is recommended, however, that samples be frozen in pre-cooled fluorocarbon-based liquid coolants in order to assure optimal tissue preservation.     

鱼病防治的给药方法及效果评价

为了充分发挥药物在水产养殖鱼类疾病上的预防和治病作用，选择正确的给药方法至关重要。渔药的使用方法不同，不仅会影响发病鱼体对药物吸收的量和速度，而且还与药物对病原体的作用力的强弱密切相关，甚至会引起药物作用性质的改变。在水产养殖鱼类的疾病防治中，体外用药主要是发挥局部作