Understanding behaviour to improve the welfare of an ornamental fish

Some common practices in aquaculture, ornamental trade and fish facilities may disturb the behavioural repertoire of fish and its natural adaptive value, reducing welfare and impairing fish production. Hence, it is necessary to understand fish behaviour, as well as the factors affecting it, to improve the quality of fish's life under artificial environment. Here, we reviewed the behaviour of the angelfish Pterophyllum scalare, an Amazonian cichlid used worldwide both as an ornamental fish and as a fish model in scientific research. We characterized social, reproductive and feeding behaviour, as well as the amazing cognitive ability of the angelfish. In addition, we reviewed the effects of environmental enrichment and suggested some important variables that need to be considered for rearing P. scalare. In this review, we show for the first time a synthesis on behaviour and a best practice overview to improve the welfare of angelfish as a target species. Nonetheless, most topics reviewed fit a broader set of fish species, particularly ornamental ones. This synthesis can therefore open a path for further behavioural research applied to the welfare of angelfish and bring insights to other fish species.     

Ethical considerations in fish research

Fishes are used in a wide range of scientific studies, from conservation research with potential benefits to the species used to biomedical research with potential human benefits. Fish research can take place in both laboratories and field environments and methods used represent a continuum from non-invasive observations, handling, through to experimental manipulation. While some countries have legislation or guidance regarding the use of fish in research, many do not and there exists a diversity of scientific opinions on the sentience of fish and how we determine welfare. Nevertheless, there is a growing pressure on the scientific community to take more responsibility for the animals they work with through maximising the benefits of their research to humans or animals while minimising welfare or survival costs to their study animals. In this review, we focus primarily on the refinement of common methods used in fish research based on emerging knowledge with the aim of improving the welfare of fish used in scientific studies. We consider the use of anaesthetics and analgesics and how we mark individuals for identification purposes. We highlight the main ethical concerns facing researchers in both laboratory and field environments and identify areas that need urgent future research. We hope that this review will help inform those who wish to refine their ethical practices and stimulate thought among fish researchers for further avenues of refinement. Improved ethics and welfare of fishes will inevitably lead to increased scientific rigour and is in the best interests of both fishes and scientists.     

Gamete interactions in teleost fish: the egg envelope. Basic studies and perspectives as environmental biomonitor

The current knowledge about teleost fish egg envelope is summarized. The paper analyzes the organization and deposition process of the protein composition and genes involved in the synthesis of teleost fish egg envelopes and their role in gamete interaction during fertilization. Pelagic and demersal species that our research group is working with are especially considered. The vertebrate ZP family of proteins, the evolution and relationship among the different genes and their expression are taken into account. We consider fish envelope as a possible biomonitor for ecological contaminants. The biotechnological applications for aquaculture and genomic and post-genomic approaches are auspicious.     

Iridoviruses associated with epizootic haematopoietic necrosis (EHN) in aquaculture

Systemic infections of teleost fishes caused by iridoviruses have recently been recognized in Australia, Asia, Europe and the USA. These iridoviruses are different from those of the established genera Lymphocystivirus and Goldfish Virus 1-like Viruses of the family Iridoviridae. The agents exhibit similar physicochemical properties, are antigenically related and prove to be of high virulence to different teleost fishes in aquaculture. The first iridovirus, epizootic haematopoietic necrosis virus, responsible for an epizootic outbreak of haematopoietic necrosis in redfin perch, was reported in Australia. Some years later, similar iridovirus epizootics occurred in sheatfish and catfish in Europe. The Australian and the European isolates proved to be antigenically related and showed properties in common with frog virus 3, the type species of the genus Ranavirus of the Iridoviridae. Further iridovirus isolates from fish, amphibians and reptiles exhibited a close relationship with each other and with frog virus 3. It is important to note that the Australian amphibian iridovirus, Bohle iridovirus, was experimentally transmitted to teleost fish inducing high mortalities. The occurrence of similar viruses in different host species in the aquatic environment and their inter-species transmission emphasize the importance of health control in aquaculture.     

Current issues in fish welfare

Human beings may affect the welfare of fish through fisheries, aquaculture and a number of other activities. There is no agreement on just how to weigh the concern for welfare of fish against the human interests involved, but ethical frameworks exist that suggest how this might be approached. Different definitions of animal welfare focus on an animal's condition, on its subjective experience of that condition and/or on whether it can lead a natural life. These provide different, legitimate, perspectives, but the approach taken in this paper is to focus on welfare as the absence of suffering. An unresolved and controversial issue in discussions about animal welfare is whether non‐human animals exposed to adverse experiences such as physical injury or confinement experience what humans would call suffering. The neocortex, which in humans is an important part of the neural mechanism that generates the subjective experience of suffering, is lacking in fish and non‐mammalian animals, and it has been argued that its absence in fish indicates that fish cannot suffer. A strong alternative view, however, is that complex animals with sophisticated behaviour, such as fish, probably have the capacity for suffering, though this may be different in degree and kind from the human experience of this state. Recent empirical studies support this view and show that painful stimuli are, at least, strongly aversive to fish. Consequently, injury or experience of other harmful conditions is a cause for concern in terms of welfare of individual fish. There is also growing evidence that fish can experience fear‐like states and that they avoid situations in which they have experienced adverse conditions. Human activities that potentially compromise fish welfare include anthropogenic changes to the environment, commercial fisheries, recreational angling, aquaculture, ornamental fish keeping and scientific research. The resulting harm to fish welfare is a cost that must be minimized and weighed against the benefits of the activity concerned. Wild fish naturally experience a variety of adverse conditions, from attack by predators or conspecifics to starvation or exposure to poor environmental conditions. This does not make it acceptable for humans to impose such conditions on fish, but it does suggest that fish will have mechanisms to cope with these conditions and reminds us that pain responses are in some cases adaptive (for example, suppressing feeding when injured). In common with all vertebrates, fish respond to environmental challenges with a series of adaptive neuro‐endocrine adjustments that are collectively termed the stress response. These in turn induce reversible metabolic and behavioural changes that make the fish better able to overcome or avoid the challenge and are undoubtedly beneficial, in the short‐term at least. In contrast, prolonged activation of the stress response is damaging and leads to immuno‐suppression, reduced growth and reproductive dysfunction. Indicators associated with the response to chronic stress (physiological endpoints, disease status and behaviour) provide a potential source of information on the welfare status of a fish. The most reliable assessment of well‐being will be obtained by examining a range of informative measures and statistical techniques are available that enable several such measures to be combined objectively. A growing body of evidence tells us that many human activities can harm fish welfare, but that the effects depend on the species and life‐history stage concerned and are also context‐dependent. For example, in aquaculture, adverse effects related to stocking density may be eliminated if good water quality is maintained. At low densities, bad water quality may be less likely to arise whereas social interactions may cause greater welfare problems. A number of key differences between fish and birds and mammals have important implications for their welfare. Fish do not need to fuel a high body temperature, so the effects of food deprivation on welfare are not so marked. For species that live naturally in large shoals, low rather than high densities may be harmful. On the other hand, fish are in intimate contact with their environment through the huge surface area of their gills, so they are vulnerable to poor water quality and water borne pollutants. Extrapolation between taxa is dangerous and general frameworks for ensuring welfare in other vertebrate animals need to be modified before they can be usefully applied to fish. The scientific study of fish welfare is at an early stage compared with work on other vertebrates and a great deal of what we need to know is yet to be discovered. It is clearly the case that fish, though different from birds and mammals, however, are sophisticated animals, far removed from unfeeling creatures with a 15 s memory of popular misconception. A heightened appreciation of these points in those who exploit fish and in those who seek to protect them would go a long way towards improving fish welfare.     

The control of chemicals used in aquaculture in Europe

A range of chemicals are used in European marine aquaculture and these may be categorized as disinfectants, antifoulants and medicines (includes vaccines). This article provides a review of chemicals used in aquaculture in Europe, their regulatory status, and a checklist of points considered best practice in the use (and avoidance of use) of medicines in marine aquaculture. The release of antifoulants and disinfectants into the marine environment is controlled by local and/or national waste discharge regulations that may in turn be guided by wider environmental quality objectives. The authorization of veterinary medicines, biologicals (vaccines) and pharmaceuticals (chemicals), in Europe is the subject of several EC Directives. Registration dossiers address the issues of product quality, safety and efficacy and include environmental and consumer safety where the product is destined for use in a food‐producing animal. Fish farmers, like all livestock producers, must have access to a range of properly authorized medicines to safeguard animal health and welfare. The distribution and supply of medicines must be appropriately controlled and their authorization appropriately includes environmental risk assessment to a common European Union (EU) or international standard. There is progress towards the harmonization of the authorization process within the EU and this will help to ensure the continued availability of medicines for fish. Consumer safety is addressed by the setting of maximum residue limits (MRLs) derived through toxicological risk assessment and by surveillance of food for residues of veterinary medicines. The system for the environmental risk assessment of chemicals used in aquaculture is being developed and is outlined in the present article. It is recommended that the supply and use of fish medicines is uniformly regulated in the EU and supported by appropriate codes of best practice. A number of codes of practice that include reference to the use of medicines have been produced both at a European level and in member states. It is recommended that all European marine aquaculture producers adopt a code of best practice for the use of medicinal and other chemicals their industry. Medicines are one part of an integrated package in dealing with animal health. This includes environmental conditions, nutrition and hygiene. The best practice guidelines presented here are based on the outcome of three European workshops as part of the EU MARAQUA project that involved industry, government and research scientists. They cover the avoidance and minimizing of the need to use medicines and other chemicals, to recording and monitoring their use and effectiveness (in case of resistance development), exchange of experiences within the industry, and staff training. Recommendations are also included for manufacturers of medicines and other chemicals, and for regulatory authorities. Minimizing the need to use medicines and other chemicals requires attention to a healthy source of fish stock. Staff must be appropriately trained in fish husbandry (to minimize stress), hygiene and disease recognition and treatment, including management of the farm site to keep it disease free. The latter may require single generations of fish per site to allow a fallow period during which a disease or parasite cycle is broken. These recommendations and guidelines are in accordance with the current codes of practice being developed by different sectors of the aquaculture industry in different countries. They do not necessarily involve significantly higher production costs and indeed are more likely to save costs as medicines and disease impacts are very costly to industry.     

Stress and welfare in ornamental fishes: what can be learned from aquaculture?

The ornamental fish trade is estimated to handle up to 1·5 billion fishes. Transportation and handling of fishes imposes a range of stressors that can result in mortality at rates of up to 73%. These rates vary hugely, however, and can be as low as 2%, because they are generally estimated rather than based on experimental work. Given the numbers of ornamental fishes traded, any of the estimated mortality rates potentially incur significant financial losses and serious welfare issues. Industry bodies, such as the Ornamental Aquatic Trade Association (OATA), have established standards and codes of best practice for handling fishes, but little scientific research has been conducted to understand the links between stress, health and welfare in ornamental species. In aquaculture, many of the same stressors occur as those in the ornamental trade, including poor water quality, handling, transportation, confinement, poor social and physical environment and disease and in this sector directed research and some resulting interventions have resulted in improved welfare standards. This review considers the concept of welfare in fishes and evaluates reported rates of mortality in the ornamental trade. It assesses how the stress response can be quantified and used as a welfare indicator in fishes. It then analyses whether lessons from aquaculture can be usefully applied to the ornamental fish industry to improve welfare. Finally, this analysis is used to suggest how future research might be directed to help improve welfare in the ornamental trade.     

Dietary self-selection in fish: a new approach to studying fish nutrition and feeding behavior

The principles of modern aquaculture encourage the development of fish feeds containing low fish meal content and several types of plant ingredients plus nutrients to avoid depleting global fish stocks and to reduce costs. However, food constituents can affect animal nutrition and feeding behavior, so the effect of different diets on fish behavior and growth needs to be understood to optimize the use of nutrients and to improve fish welfare. The development of multiple-choice self-feeding systems led to a new perspective for investigating these issues in aquaculture species. Our purpose with this review is to summarize the information that has been published to date on this topic and to identify gaps in knowledge where research is needed. Key subjects are assessed under the following major headings: How do we study dietary selection in fish? What food signals do fish use to choose the right diet? and How do fish respond to food challenges? The present review will provide a picture of the main results obtained to date in these studies in aquaculture fish species, as well as perspectives for future research in the field.     

Taking account of fish welfare: lessons from aquaculture

This paper explores the possibility that lessons learned from aquaculture might contribute to current debate on welfare and fisheries. After looking briefly at the history of research interest in the welfare of farmed fishes, some implications of using different definitions of and approaches to the concept of welfare are discussed. Consideration is given to the way in which the aquaculture industry has responded to public concern about fish welfare and, for cases where these responses have been effective, why this might be the case. Finally, possible cross‐over points between aquaculture and fisheries in the context of fish welfare, as well as experience and expertise that might be shared between these two areas, are identified.     

Analysis of heterologous and homologous promoters and enhancers in vitro and in vivo by gene transfer into Japanese medaka (Oryzias latipes) Xiphophorus.

Efficient expression systems are required for analysis of gene regulation and function in teleost fish. To develop such systems, a number of inducible or constitutive promoter and enhancer sequences of fish or higher vertebrate origin were tested for activity in a variety of fish cell lines and in embryos of the Japanese medaka fish (Oryzias latipes) and Xiphophorus. The activity of the different promoter-enhancer combinations were quantitated. Considerable differences were found for some constructs if tested in vitro or in vivo. Fro the data obtained, a set of expression vectors for basic research as well as for aquaculture purposes were established.     

Viral infections of aquatic animals with special reference to Asian aquaculture

Worldwide, the number of communicable diseases of animals raised in aquaculture continue to increase. Viral infections of cultivated shellfish, crustacea, and finfish have been frequently recognized in the past few years. In the Asian regions, penaeid shrimp and several teleost fish underwent epizootics associated with heavy losses in aquaculture. Baculoviruses are particularly harmful to shrimp and prawns. Herpes-, irido-, reo-, or rhabdovirus-like agents can cause outbreaks in fish farms. Viral diseases are important limiting factors in the expansion of aquaculture. However, studies on viral infections of aquatic animals have been focused primarily on economically important farmed fish. Therfore, certain viral diseases of teleost fish are relatively well understood. In contrast, our knowledge of viral infections of farmed aquatic invertebrates is still very spare. Although a great number of viruses have been detected in farmed molluscs and crustaceans, the pathogenicity and epizootiology of most of the agents is not known.     

Comparison of hypoxia-inducible factor-1 alpha in hypoxia-sensitive and hypoxia-tolerant fish species

Levels of oxygen can vary dramatically in aquatic environments. Aquatic organisms, including fishes, have adapted accordingly to survive. As there are both phylogenetically closely related fish species with differing oxygen requirements and distantly related species with similar oxygen requirements, fishes are good candidates for examining oxygen-related functions in vertebrates. We set out to investigate if sequence variation in the hypoxia-inducible factor-1 alpha (HIF-1α) gene is associated with variations in oxygen requirements. Since the teleost HIF-1α sequences available in databases represent a very limited dataset both phylogenetically and with regard to oxygen requirements, we have sequenced the protein coding sequence for HIF-1α from an additional 9 fish species. Our results indicate that the deduced HIF-1α proteins of teleost fishes are somewhat shorter than those of tetrapods. Additionally, the results suggest that tetrapod sequences more closely resemble the ancestral form of the protein than do teleost sequences. No clear signatures which could be associated with the oxygen requirements of the species were found. This study suggests that if species-specific differences in HIF-1α function with regards to oxygen dependence have evolved, they do not occur in the protein coding sequence but at other levels of the HIF-1α pathway.     

Altering fish embryos with aquaporin-3: an essential step toward successful cryopreservation

Fish populations are globally threatened by overharvesting and habitat degradation. The ability to bank fish embryos by cryopreservation could be crucial for preserving species diversity, for aquaculture (allowing circannual fish farming), and for managing fish models used in human biomedical research. However, no nonmammalian embryo has ever been successfully cryopreserved. For fish, low membrane permeability prevents cryoprotectants from entering the yolk to prevent cryodamage. Here, we present evidence of a membrane mechanism hindering cryopreservation of fish and propose a novel solution to this obstacle. Zebrafish (Danio rerio) embryos have rectifying membranes that allow water to leave but not to reenter readily. This feature may be an evolutionary trait that allows freshwater embryos to grow in hypoosmotic environments without osmoregulatory organs. However, this trait may also prevent successful fish embryo cryopreservation because both water and cryoprotectants must move into and out of cells. As a solution, we injected zebrafish embryos with mRNA for the aquaporin-3 water channel protein and demonstrated increased membrane permeability to water and to a cryoprotectant. Modeling indicates that sufficient cryoprotectant enters aquaporin-3-expressing zebrafish embryos to allow cryopreservation.     

A comparative view on mechanisms and functions of skeletal remodelling in teleost fish, with special emphasis on osteoclasts and their function

Resorption and remodelling of skeletal tissues is required for development and growth, mechanical adaptation, repair, and mineral homeostasis of the vertebrate skeleton. Here we review for the first time the current knowledge about resorption and remodelling of the skeleton in teleost fish, the largest and most diverse group of extant vertebrates. Teleost species are increasingly used in aquaculture and as models in biomedical skeletal research. Thus, detailed knowledge is required to establish the differences and similarities between mammalian and teleost skeletal remodelling, and between distantly related species such as zebrafish (Danio rerio) and medaka (Oryzias latipes). The cellular mechanisms of differentiation and activation of osteoclasts and the functions of teleost skeletal remodelling are described. Several characteristics, related to skeletal remodelling, distinguish teleosts from mammals. These characteristics include (a) the absence of osteocytes in most species; (b) the absence of haematopoietic bone marrow tissue; (c) the abundance of small mononucleated osteoclasts performing non‐lacunar (smooth) bone resorption, in addition to or instead of multinucleated osteoclasts; and (d) a phosphorus‐ rather than calcium‐driven mineral homeostasis (mainly affecting the postcranial dermal skeleton). Furthermore, (e) skeletal resorption is often absent from particular sites, due to sparse or lacking endochondral ossification. Based on the mode of skeletal remodelling in early ontogeny of all teleosts and in later stages of development of teleosts with acellular bone we suggest a link between acellular bone and the predominance of mononucleated osteoclasts, on the one hand, and cellular bone and multinucleated osteoclasts on the other. The evolutionary origin of skeletal remodelling is discussed and whether mononucleated osteoclasts represent an ancestral type of resorbing cells. Revealing the differentiation and activation of teleost skeletal resorbing cells, in the absence of several factors that trigger mammalian osteoclast differentiation, is a current challenge. Understanding which characters of teleost bone remodelling are derived and which characters are conserved should enhance our understanding of the process in fish and may provide insights into alternative pathways of bone remodelling in mammals.     

Using model selection to choose a size-based condition index that is consistent with operational welfare indicators

Quantitative and qualitative measures of fish health and welfare are essential for management of both wild capture and aquaculture species. These measures include morphometric body condition indices, energetic condition and aquaculture operational welfare indicators (OWIs). Measures vary in ease of measurement (and may require destructive sampling), and it is critical to know how well they correlate with fish health and welfare so appropriate management decisions can be based on them. Lumpfish (Cyclopterus lumpus) is a new farming species that needs nondestructive OWIs to be developed and validated. In this study, we developed a C. lumpus fin damage score. Four different body condition indexes based on individual weight relative to either length–weight relationships or relative to other fish in its local environment were tested (using model selection) as predictors of individual fin damage. Results showed severity of fin damage was predicted by small size relative to the other individuals in the tank or cage. Body condition based on length–weight relationship was not found to predict fin damage, indicating that using established indices from fisheries or from other species would not predict welfare risks from fin damage. Implications are that especially in hatchery conditions grading will improve the condition index, and is expected to mitigate fin damage, but that low weight at length was not of use in predicting fin damage. Model selection to choose between a suite of possible indices proved powerful and should be considered in other applications where an easily measured index is needed to correlate with other health measures.     

Genetic interactions between marine finfish species in European aquaculture and wild conspecifics

The principal species of marine aquaculture in Europe are Atlantic salmon (Salmo salar), sea bass (Dicentrarchus labrax) and sea bream (Sparus auratus). For Atlantic salmon and sea bass, a substantial part of total genetic variation is partitioned at the geographical population level. In the case of sea bream, gene flow across the Azores/Mediterranean scale appears to be extensive and population structuring is not detected. For Atlantic salmon and sea bass, natural population structure is at risk from genetic interaction with escaped aquaculture conspecifics. The locally adaptive features of populations are at risk from interbreeding with non‐local aquaculture fish. Wild populations, generally, are at risk from interactions with aquaculture fish that have been subject to artificial selection or domestication. Atlantic salmon is the main European aquaculture species and its population genetics and ecology have been well‐studied. A general case regarding genetic interactions can be based on the information available for salmon and extended to cover other species, in the appropriate context. A generalized flow chart for interactions is presented. Salmon escape from aquaculture at all life stages, and some survive to breed among wild salmon. Reproductive fitness in the escaped fish is lower than in native, wild fish because of behavioural deficiencies at spawning. However, as the number of salmon in aquaculture greatly exceeds the number of wild fish, even small fractional rates of escape may result in the local presence of large numbers, and high frequencies, of escaped fish. At present, policy and legislation in relation to minimizing genetic interactions between wild and aquaculture fish is best developed for Atlantic salmon, through the recommendations of the Oslo Agreement developed by the North Atlantic Salmon Conservation Organization and subsequent agreements on their implementation. In future, the potential use of genetically modified fish in aquaculture will make additional policy development necessary. Improved containment is recommended as the key to minimizing the numbers and therefore the effects of escaped fish. Emergency recovery procedures are recommended as a back‐up measure in the case of containment failure. Reproductive sterility is recommended as a future key to eliminating the genetic potential of escaped fish. The maintenance of robust populations of wild fish is recommended as a key to minimizing the effects of escaped fish on wild populations.     

Fish host range of seven isolates of Cryptocaryon irritans (Ciliophora)

Host specificity of the marine ciliate Cryptocaryon irritans was investigated using seven parasite isolates and six experimental fish species. The low degree of host specificity of C. irritans to teleost fish has been confirmed. Furthermore, five fish species occurring outside the geographical range of this parasite were also susceptible to infection; three of these could be at risk to Cryptocaryon through aquaculture practices.     

Generation time of zebrafish (Danio rerio) and medakas (Oryzias latipes) housed in the same aquaculture facility

The zebrafish and the medaka are both important model organisms in biomedical research. Both species are frequently characterized as having a generation time of approximately 2-4 months, but the precise onset of sexual maturity and the variability of reproductive success with age have not been previously examined. The authors studied reproduction in replicate groups of wild-type zebrafish (strain AB) and medakas (strain Cab) that were maintained together in the same aquaculture system. Length, weight and survival of the fish were measured and recorded once per week. Reproductive success and viability of offspring were also evaluated. Both zebrafish and medakas began producing viable embryos within 60 d post-fertilization. These findings show that it is possible to successfully maintain populations of both species within the same research infrastructure without compromising reproductive success or embryo viability.     

In search of determinants: gene expression during gonadal sex differentiation

The diversity of inputs that guide sexual fate during development is both intriguing and daunting. In the field of fish biology, the study of sex determination is of great importance. For example, in aquaculture, sexually dimorphic growth rates and overall size leads to one sex being more marketable than the other. Moreover, for breeding purposes it is important to maintain balanced sex ratios. Furthermore, sex determination is sensitive to environmental factors, such as temperature and contaminants, which can lead to skewed sex ratios, intersexes and sterility in wild or farmed fish. The gonad is typically the first organ to exhibit morphological signs of sexual dimorphism and therefore is likely to be the primary organ system whose fate is controlled by the sex determination cues in many fish species. Additionally, the sexual fate of the gonad has been shown to fully or partially control organismal sex differentiation. Thus, understanding the genetic regulation of gonadal sex differentiation is critical in studies of fish sex determination. This review summarizes recent knowledge of genes expressed during gonadal sex differentiation in gonochoristic teleost fish. Three species are discussed, which serve as excellent model systems for probing teleost sex differentiation: the Oreochromis niloticus, Oryzias latipes and Danio rerio. The similarities and differences between gonadal gene expression in these three species and in comparison to mammals suggest conserved roles during vertebrate gonadal sex differentiation. In the future, it will be essential to develop tools to assay the function of genes expressed during gonadal sex differentiation in fish.