Development and application of biological technologies in fish genetic breeding

Fish genetic breeding is a process that remolds heritable traits to obtain neotype and improved varieties.For the purpose of genetic improvement,researchers can select for desirable genetic traits,integrate a suite of traits from different donors,or alter the innate genetic traits of a species.These improved varieties have,in many cases,facilitated the development of the aquaculture industry by lowering costs and increasing both quality and yield.In this review,we present the pertinent literatures and summarize the biological bases and application of selection breeding technologies(containing traditional selective breeding,molecular marker-assisted breeding,genome-wide selective breeding and breeding by controlling single-sex groups),integration breeding technologies(containing cross breeding,nuclear transplantation,germline stem cells and germ cells transplantation,artificial gynogenesis,artificial androgenesis and polyploid breeding)and modification breeding technologies(represented by transgenic breeding)in fish genetic breeding.Additionally,we discuss the progress our laboratory has made in the field of chromosomal ploidy breeding of fish,including distant hybridization,gynogenesis,and androgenesis.Finally,we systematically summarize the research status and known problems associated with each technology.     

Genomics to accelerate genetic improvement in tilapia

Selective breeding of tilapia populations started in the early 1990s and over the past three decades tilapia has become one of the most important farmed freshwater species, being produced in more than 125 countries around the globe. Although genome assemblies have been available since 2011, most of the tilapia industry still depends on classical selection techniques using mass spawning or pedigree information to select for growth traits with reported genetic gains of up to 20% per generation. The involvement of international breeding companies and research institutions has resulted in the rapid development and application of genomic resources in the last few years. GWAS and genomic selection are expected to contribute to uncovering the genetic variants involved in economically relevant traits and increasing the genetic gain in selective breeding programs, respectively. Developments over the next few years will probably focus on achieving a deep understanding of genetic architecture of complex traits, as well as accelerating genetic progress in the selection for growth-, quality- and robustness-related traits. Novel phenotyping technologies (i.e. phenomics), lower-cost whole-genome sequencing approaches, functional genomics and gene editing tools will be crucial in future developments for the improvement of tilapia aquaculture.     

Biotechnology and fish culture

Biotechnology can currently be considered of importance in aquaculture. The increase in the production of aquatic organisms over the last two decades through the use of biotechnology indicates that in a few generations biotechnology may overtake conventional techniques, at least for the commercially more valuable species. In the last few years, genetics has contributed greatly to fish culture through the application of the more recent techniques developed in biotechnology and in genetic engineering. At present, the most commonly used methods in fish biotechnology are chromosome manipulation and hormonal treatments, which can be used to produce triploid, tetraploid, haploid, gynogenetic and androgenetic fish. These result in the production of individuals and lineages of sterile, monosex or highly endogamic fish. The use of such strategies in fish culture has as a practical objective the control of precocious sexual maturation in certain species; other uses are the production of larger specimens by control of the reproductive process and the attainment of monosex lines containing only those individuals of greater commercial value. The use of new technologies, such as those involved in gene transfer in many species, can result in modified individuals of great interest to aquaculturists and play important roles in specific programmes of fish production in the near future.     

Measuring farmers' attitude towards breeding tools: the Livestock Breeding Attitude Scale

Under-use of genetic improvement tools and low participation in breeding programmes are key drivers of breeding programmes under-performance. Both aspects are heavily influenced by farmers attitudes which, to date, have not been analysed in an objective and systematic manner. A key factor constraining the implementation of attitudinal studies towards livestock breeding tools is the lack of a reference scale for measuring attitudes. In this research, we provide the livestock breeding sector with such a reference measure. We developed the scale following the standardized psychometric methodologies and statistical tools. Then, as a case study, we used the scale to explore the attitudes of beef and dairy sheep farmers in Australia, New Zealand and Spain and analysed farmer and farming system factors related to those attitudes. Fourteen sheep and beef breed associations facilitated the implementation of a survey of 547 farmers, generating data that was used for the scale evaluation. The relationship between attitudinal factors and farmer and farming system factors was analysed using generalized linear models across and within breeds. The results suggest that the 8-item definitive scale we have developed is appropriate to measure farmer attitudes. We found that attitudes towards genetic improvement tools have two components; i) traditional selection and ii) genetic and genomic selection combined. This means that positive attitudes towards traditional phenotypic selection do not necessarily imply a negative attitude towards genetic and genomic selection tools. Farmer attitudes varied greatly not only across the studied breeds, species and countries, but also within them. High-educated farmers of business-oriented farms tend to have the most negative attitude towards traditional selection. However, attitudes towards genetic and genomic selection tools could not be linked to these factors. Finally, we found that the breed raised had a large effect on farmer attitude. These findings may help in the evolution of breeding programmes by identifying both the farmers most inclined to uptake breeding innovations in the early stages of its establishment and the farmers who would be more reluctant to participate in such programmes, thus informing where to focus extension efforts.     

The evolving technology of DNA fingerprinting and its application to fisheries and aquaculture

In 1985, Alec Jeffreys reported the development of multilocus DNA fingerprinting by Southern blot-detection of hypervariable minisatellites or variable number of tandem repeat (VNTR) loci. This technology found immediate application to various forensic and scientific problems, including fisheries and aquaculture. By 1989, however, it was recognized by many researchers that inherent problems exist in the application of multilocus fingerprinting to large sample sizes as might occur in fisheries and aquaculture genetic studies. As such, individual VNTRs were cloned for single-locus DNA fingerprinting. Although single-locus fingerprinting ameliorates many of the problems associated with multilocus DNA fingerprinting, it suffers from the problem that electrophorectic anomalies of band migration within and between gels necessitates binning of alleles, thus underestimating genetic variability in a given population. Amplification of microsatellite loci by the polymerase chain reaction, however, solved many of the problems of Southern blot-based DNA fingerprinting. Moreover, microsatellites exhibit attributes that make them particularly suitable as genetic markers for numerous applications in aquaculture and fisheries research: (1) they are abundant in the genome; (2) they display varying levels of polymorphism; (3) alleles exhibit codominant Mendelian inheritance; (4) minute amounts of tissue are required for assay (e.g., dried scales or otoliths); (5) loci are conserved in related species; (6) potential for automated assay. Recent innovations in DNA fingerprinting technology developed over the past 5 years are discussed with special emphasis on microsatellites and their application to fisheries and aquaculture, e.g., behavioural and population genetics of wild species, and selection and breeding programmes for aquaculture broodstock.     

In silico whole-genome EST analysis reveals 2322 novel microsatellites for the silver-lipped pearl oyster, Pinctada maxima

Molecular stock improvement techniques such as marker assisted selection have great potential in accelerating selective breeding programmes for animal production industries. However, the discovery and application of trait/marker associations usually requires a large number of genome-wide polymorphic loci. Here, we present 2322 unique microsatellites for the silver-lipped pearl oyster, Pinctada maxima, a species of aquaculture importance throughout the Indo-Australian Archipelago for production of the highly valued South Sea pearl. More than 1.2 million Roche 454 expressed sequence tag (EST) reads were screened for microsatellite repeat motifs. A total of 12,604 sequences contained either a di, tri, tetra, penta or hexa microsatellite repeat motif (n ≥ 6), with 6435 of these sequences having sufficient flanking regions for primer development. All identified microsatellites with designed primers were condensed into 2322 unique clusters (i.e., unique loci) of which 360 were shown to be polymorphic based on multiple sequence reads with different repeat motifs. Genotyping of five microsatellite loci demonstrated that in silico evaluation of polymorphism levels was a very useful method for identification of polymorphic loci, with the variation uncovered being a lower bound. Gene Ontology annotations of sequences containing microsatellites suggest that most are derived from a diverse array of unique genes. This EST derived microsatellite database will be a valuable resource for future studies in genetic map construction, diversity analysis, quantitative trait loci analysis, association mapping and marker assisted selection, not only for P. maxima, but also closely related species within the genus Pinctada.     

Genetic Manipulation of Sex Ratio for the Large-Scale Breeding of YY Super-Male and XY All-Male Yellow Catfish (Pelteobagrus fulvidraco (Richardson))

Yellow catfish has become one of the most important freshwater aquaculture species in China. The mono-sex male yellow catfish has important application value in aquaculture because the male grows generally faster than the sibling females under the same conditions. This study has screened YY super-male and YY physiological female yellow catfish by sex reversal, gynogenesis, and progeny testing, which can help to achieve the large-scale production of YY super-male and XY all-male. From 2008 to 2010, about 123,000 YY super-male were produced, and about 81 million XY all-male fry were produced with 100 % male rate by random sampling. Therefore, these results indicate that YY super-male and YY physiological female yellow catfish can be viable and fertile. We conclude that the mono-sex breeding technique by YY super-male yellow catfish is stable and reliable, which has great potential for application in yellow catfish aquaculture.     

Genetic manipulations in aquaculture: a review of stock improvement by classical and modern technologies

The aim of this review was to highlight the extent to which the genetic technologies are implemented by the aquaculture industry. The review shows that some of the modern genetic technologies are already extensively applied by the diverse aquaculture industries, though not to the same extent for all important aquacultured species (according to FAO 1998 figures). Some species (common carp, Atlantic salmon, rainbow trout, channel catfish, Nile tilapia, and the Pacific oyster) received concentrated breeding efforts, while other major cultured species (Chinese and Indian carps and the giant tiger shrimp) received, so far, relatively limited attention, and a few species (Yesso scallop, blue mussel, white Amur bream, and milkfish) have, apparently, not been genetically improved at all. Most of the genetically improved strains reaching the aquaculture industry were developed through traditional selective breeding (selection, crossbreeding, and hybridization). Emerging, more modern technologies for genetic manipulation seem to take 10-20 years from being established experimentally until applications affect the industry. Thus, chromosome-set and sex manipulations started to affect the industry during the 1980's and 1990's. DNA marker technology and gene manipulations have yet hardly affected the industry. The former have not matured yet, but hold much promise. The latter could have affected the industry already had it not been restricted by public concern.     

Application of sperm sorting and associated reproductive technology for wildlife management and conservation

Efforts toward the conservation and captive breeding of wildlife can be enhanced by sperm sorting and associated reproductive technologies such as sperm cryopreservation and artificial insemination (AI). Sex ratio management is of particular significance to species which naturally exist in female-dominated social groups. A bias of the sex ratio towards females of these species will greatly assist in maintaining socially cohesive groups and minimizing male-male aggression. Another application of this technology potentially exists for endangered species, as the preferential production of females can enable propagation of those species at a faster rate. The particular assisted reproductive technology (ART) used in conjunction with sperm sorting for the production of offspring is largely determined by the quality and quantity of spermatozoa following sorting and preservation processes. Regardless of the ART selected, breeding decisions involving sex-sorted spermatozoa should be made in conjunction with appropriate genetic management. Zoological-based research on reproductive physiology and assisted reproduction, including sperm sorting, is being conducted on numerous terrestrial and marine mammals. The wildlife species for which the technology has undergone the most advance is the bottlenose dolphin. AI using sex-sorted fresh or frozen-thawed spermatozoa has become a valuable tool for the genetic and reproductive management of captive bottlenose dolphins with six pre-sexed calves, all of the predetermined sex born to date.     

DNA分子标记、基因组作图及其在植物遗传育种上的应用

本文总结了现代分子生物学所发展的各类分子标记及其在遗传图谱构建中的作用,还综述了遗传图谱与物理图谱的构建、数量与质量性状定位及分子标记在作物育种中的应用。     

Reintroduction of the European Capercaillie from the Capercaillie Breeding Centre in Wis?a Forest District: Genetic Assessments of Captive and Reintroduced Populations

The Western capercaillie (Tetrao urogallus) is a specific bird species, which, despite its very broad distribution and large global population size, is highly endangered in many Western and Central European countries. According to the species situation, in many countries (including Poland), breeding and reintroduction programmes have been started. One of the most complex and large-scale reintroduction programmes was started in Bory Dolnośląskie Forest, and the Capercaillie Breeding Centre in Wisła Forest District was used as one of the sources of individuals for reintroduction. As genetic tools provide essential knowledge about species biodiversity, which is crucially important during the breeding process and reintroduction, both captive and reintroduced grouse populations were genetically analysed. We were particularly interested in genetic diversity of the individuals in both populations and the genetic relationship between them, as well as between them and other capercaillie representatives from their current range. To fulfil these goals we determined nine microsatellite loci along with a fragment of the mitochondrial control region. Genetic diversity parameters were moderate to high compared to populations from other Central and Western European countries. Both populations were clustered into three distinct genetic clades based on microsatellites. Phylogenetic analysis placed all mitochondrial haplotypes we revealed in the Eurasian clade. The present results will play an important role as they will help to preserve and maximize genetic diversity in captive populations, and will provide a basis for future monitoring of the reintroduction process.     

Determination of n-3 HUFA content in Atlantic salmon flesh based on the lipid content, morphometric measurements and blood fatty acid composition: A modeling approach

Fish are the most important source of n-3 highly unsaturated fatty acids (HUFA) in the human diet and, with declining wild stocks, an increasing proportion is being provided by aquaculture. Paradoxically, aquaculture fish diets have traditionally incorporated fish oil and meal derived from wild fisheries. Continued aquaculture development requires fish oil to be replaced with vegetable oils, the only sustainable alternative. However, vegetable oils lack n-3 HUFA and so flesh from fish reared on these diets can also have reduced n-3 HUFA and thus reduced nutritional quality. This accepted, the flesh n-3 HUFA content should be an economically important trait, however to be included in the breeding goal the trait must be measurable. In the present study, we investigated whether the flesh n-3 HUFA content of salmon can be estimated in a non-fatal way. We showed that a general regression model based on flesh lipid content, morphometric and blood fatty acid measurements could estimate and predict flesh n-3 HUFA content. This would allow a choice from a range of selection methods, including mass selection or within-family selection, if this important flesh quality trait is to be included in future salmon breeding programmes.     

Genetic monitoring of the Amazonian fish matrinchã (Brycon cephalus) using RAPD markers: insights into supportive breeding and conservation programmes

The importance of genetic evaluations in aquaculture programmes has been increased significantly not only to improve effectiveness of hatchery production but also to maintain genetic diversity. In the present study, wild and captive populations of a commercially important neotropical freshwater fish, Brycon cephalus (Amazonian matrinchã), were analyzed in order to evaluate the levels of genetic diversity in a breeding programme at a Brazilian research institute of tropical fish. Random Amplified Polymorphic DNA fingerprinting was used to access the genetic variability of a wild stock from the Amazon River and of three captive stocks that correspond to consecutive generations from the fishery culture. Although farmed stocks showed considerably lower genetic variation than the wild population, a significantly higher level of polymorphism was detected in the third hatchery generation. The results seem to reflect a common breeding practice on several hatchery fish programmes that use a small number of parents as broodstocks, obtaining reproductive success with few non‐identified mating couples. The obtained data were useful for discussing suitable strategies for the genetic management and biodiversity conservation of this species.     

Genetic resource banks in wildlife conservation

Recent advances in reproductive technologies for animal breeding, together with improvements in techniques for storage of gametes and embryos, have encouraged the view that the time is now appropriate for developing systematic policies of germplasm banking. Such activities would aim to support more conventional breeding programmes for threatened species, by providing the opportunity to store valuable genetic material for use on some future occasion. A number of pertinent issues should be addressed, however, before embarking upon the large scale implementation of genetic bank programmes. This review raises and discusses some of the issues involved.     

Fish Growth in Marine Culture Systems: A Challenge for Biotechnology

Aquaculture production is constrained largely by the growth efficiency of the species being produced. Nutritional approaches have played an important part in improving this situation, but, it is argued, the room for further improvement using such established techniques is limited. Alternative ways of improving fish production by utilizing recent biotechnological advances are explored and assessed as to their potential for commercialization in the near future. Transgenic technologies promise a revolution in aquaculture, but it is considered that consumer resistance may delay the use of transgenic fish for food production. An alternative approach could be the breeding of transgenic fodder plants without the amino acid deficiencies of existing alternatives to fish meal in aquaculture diets. The use of probiotics could reduce antibiotic use on fish farms while they might also provide the basis for ``smart' diets, tailored to specific purposes by the inclusion of microorganisms. The selection and genetic engineering of nitrifying and denitrifying bacteria could also pave the way for fully enclosed, recirculating marine culture systems, which would allow control of the environmental variables that currently restrain marine fish culture. Received August 10, 1998; accepted October 8, 1998.     

Inheritance of RAPD markers in channel catfish (Ictalurus punctatus), blue catfish (I. furcatus), and their F1, F2 and backcross hybrids

The channel catfish ( Ictalurus punctatus ) has become the most important aquaculture species in the USA. A genetic linkage map in catfish is needed to improve efficiency of breeding by marker-assisted selection (MAS) and for identification of economically important genes such as disease resistance genes. To identify DNA-based genetic polymorphism, the present authors tested 42 randomly amplified polymorphic DNA (RAPD) primers for their utility in identifying genetic polymorphism in catfish. Out of these primers, 22 generated 171 highly reproducible RAPD markers, producing almost eight polymorphic bands per primer. The remaining 20 primers produced an additional 20 polymorphic bands. The RAPD markers were highly reproducible, transmitted to F1 hybrids, and segregated in F2 or backcross progeny in ratios that did not differ from Mendelian expectations. Because the interspecific hybrids of channel catfish and blue catfish are fertile, RAPD markers using the interspecific hybrid system will be useful for rapid construction of genetic linkage maps of catfish and for analysis of important quantitative trait loci.     

Development and testing of a combined species SNP array for the European seabass (Dicentrarchus labrax) and gilthead seabream (Sparus aurata)

SNP arrays are powerful tools for high-resolution studies of the genetic basis of complex traits, facilitating both selective breeding and population genomic research. The European seabass (Dicentrarchus labrax) and the gilthead seabream (Sparus aurata) are the two most important fish species for Mediterranean aquaculture. While selective breeding programmes increasingly underpin stock supply for this industry, genomic selection is not yet widespread. Genomic selection has major potential to expedite genetic gain, particularly for traits practically impossible to measure on selection candidates, such as disease resistance and fillet characteristics. The aim of our study was to design a combined-species 60 K SNP array for European seabass and gilthead seabream, and to test its performance on farmed and wild populations from numerous locations throughout the species range. To achieve this, high coverage Illumina whole-genome sequencing of pooled samples was performed for 24 populations of European seabass and 27 populations of gilthead seabream. This resulted in a database of ~20 million SNPs per species, which were then filtered to identify high-quality variants and create the final set for the development of the ‘MedFish’ SNP array. The array was then tested by genotyping a subset of the discovery populations, highlighting a high conversion rate to functioning polymorphic assays on the array (92% in seabass; 89% in seabream) and repeatability (99.4–99.7%). The platform interrogates ~30 K markers in each species, includes features such as SNPs previously shown to be associated with performance traits, and is enriched for SNPs predicted to have high functional effects on proteins. The array was demonstrated to be effective at detecting population structure across a wide range of fish populations from diverse geographical origins, and to examine the extent of haplotype sharing among Mediterranean farmed fish populations. In conclusion, the new MedFish array enables efficient and accurate high-throughput genotyping for genome-wide distributed SNPs for each fish species, and will facilitate stock management, population genomics approaches, and acceleration of selective breeding through genomic selection.     

Review: Towards the agroecological management of ruminants,pigs and poultry through the development of sustainable breeding programmes. II. Breeding strategies

Agroecology uses ecological processes and local resources rather than chemical inputs to develop productive and resilient livestock and crop production systems. In this context, breeding innovations are necessary to obtain animals that are both productive and adapted to a broad range of local contexts and diversity of systems. Breeding strategies to promote agroecological systems are similar for different animal species. However, current practices differ regarding the breeding of ruminants, pigs and poultry. Ruminant breeding is still an open system where farmers continue to choose their own breeds and strategies. Conversely, pig and poultry breeding is more or less the exclusive domain of international breeding companies which supply farmers with hybrid animals. Innovations in breeding strategies must therefore be adapted to the different species. In developed countries, reorienting current breeding programmes seems to be more effective than developing programmes dedicated to agroecological systems that will struggle to be really effective because of the small size of the populations currently concerned by such systems. Particular attention needs to be paid to determining the respective usefulness of cross-breeding v. straight breeding strategies of well-adapted local breeds. While cross-breeding may offer some immediate benefits in terms of improving certain traits that enable the animals to adapt well to local environmental conditions, it may be difficult to sustain these benefits in the longer term and could also induce an important loss of genetic diversity if the initial pure-bred populations are no longer produced. As well as supporting the value of within-breed diversity, we must preserve between-breed diversity in order to maintain numerous options for adaptation to a variety of production environments and contexts. This may involve specific public policies to maintain and characterize local breeds (in terms of both phenotypes and genotypes), which could be used more effectively if they benefited from the scientific and technical resources currently available for more common breeds. Last but not least, public policies need to enable improved information concerning the genetic resources and breeding tools available for the agroecological management of livestock production systems, and facilitate its assimilation by farmers and farm technicians.     

Harvesting the Promising Fruits of Genomics: Applying Genome Sequencing Technologies to Crop Breeding

Next generation sequencing (NGS) technologies are being used to generate whole genome sequences for a wide range of crop species. When combined with precise phenotyping methods, these technologies provide a powerful and rapid tool for identifying the genetic basis of agriculturally important traits and for predicting the breeding value of individuals in a plant breeding population. Here we summarize current trends and future prospects for utilizing NGS-based technologies to develop crops with improved trait performance and increase the efficiency of modern plant breeding. It is our hope that the application of NGS technologies to plant breeding will help us to meet the challenge of feeding a growing world population.

This article is part of the PLOS Biology Collection “The Promise of Plant Translational Research.”