progress
November 2014
The principal objective in WP4 is to evaluate the genetic variability of captive pikeperch (Sander lucioperca) broodstocks in commercial RAS farms in Europe and then compare this variability to those found in wild populations in order to define how future genetic breeding program should be established for sustainable optimal performances. We have so far received approximately 700 fin-clips (non-invasive DNA sampling method in fish) from 12 broodstocks samples belonging to 7 commercial companies in central and northern Europe; following standard protocols, DNA was extracted from all fish and amplified by PCR (Polymerase Chain Reaction) using eleven pairs of microsatellite primers described in phylogenetically close species (e.g. the yellow perch Perca flavescens, the walleye Stizostedion vitreum and the Rhone streber Zingel asper). Preliminary analysis was based on population genetics parameters (allelic richness, heterozygosity indices, inbreeding coefficients) calculated for all domesticated stocks and a wild/natural one used as a control to see whether there is loss of genetic variability due to breeding practices which theoretically will limit the potential for future genetic improvement from selection practices.
The principal objective in WP4 is to evaluate the genetic variability of captive pikeperch (Sander lucioperca) broodstocks in commercial RAS farms in Europe and then compare this variability to those found in wild populations in order to define how future genetic breeding program should be established for sustainable optimal performances. We have so far received approximately 700 fin-clips (non-invasive DNA sampling method in fish) from 12 broodstocks samples belonging to 7 commercial companies in central and northern Europe; following standard protocols, DNA was extracted from all fish and amplified by PCR (Polymerase Chain Reaction) using eleven pairs of microsatellite primers described in phylogenetically close species (e.g. the yellow perch Perca flavescens, the walleye Stizostedion vitreum and the Rhone streber Zingel asper). Preliminary analysis was based on population genetics parameters (allelic richness, heterozygosity indices, inbreeding coefficients) calculated for all domesticated stocks and a wild/natural one used as a control to see whether there is loss of genetic variability due to breeding practices which theoretically will limit the potential for future genetic improvement from selection practices.
Participating organizations (WP leader in bold): P1. HCMR and P9. UL
Task 4.1 Evaluation of the genetic variation in available domesticated broodstocks of pikeperch (led by UL). Up to now, there has been no evaluation of the current genetic diversity of captive pikeperch. Yet, because there are only a few commercial hatcheries that produce pikeperch (around 10 farms), it is likely that the genetic diversity might be very low compared to the genetic variability of natural populations. Each pikeperch farm uses its own isolated stock captured either from the wild or supplied by another farmer. Therefore, pikeperch populations differ from one farm to another depending upon the geographical origin of the wild populations from which the captive stocks derived.
Several studies have been published on the variability of wild pikeperch. For instance, Saisa et al. (2010) have studied the genetic variability of three coastal and five freshwater populations in the northern part of the Baltic sea by using 12 microsatellite loci previously developed for the walleye Sander vitreum and the yellow perch Perca flavescens. They found that the coastal populations differ genetically from the lake populations, which present a higher genetic diversity. Saliminen et al., (2012) have further studied the genetic consequences and gene flow of pikeperch stocking in three lakes by comparing the pre- and post-release patterns using a subset of 9 microsatellites loci from Saisa et al. (2010). The genetic structures of populations were disrupted by the releases of fish. Other studies have analysed the genetic structure and dynamics of pikeperch at a regional scale using some of the above cited microsatellites loci.
Our first objective is to develop for the species a highly informative and efficient microsatellite multiplex consisted of more than 10 markers, which will allow the adequate genotyping of the fish sampled. Microsatellite loci will be first ordered by increasing size in base pairs (bp) and size range, and in each range one of the primers for each microsatellite locus, e.g. the reverse, will be fluorescently labelled with the dyes conformed to the ABI technology systems (FAM, NED, VIC and PET) and using the Qiagen multiplex PCR kit. The PCR conditions will be optimized in order to finally have a powerful molecular tool for genotyping. The above-mentioned commercial kit gives the advantage of maximal transferability of molecular protocols between labs.
Few microsatellites have already been developed in pikeperch , but numerous have proved to cross-amplify from phylogenetically close Percid species (i.e., walleye Sander vitreum, Eurasian perch Perca fluviatilis, Yellow perch Perca flavescens). Among these genetic markers, microsatellite loci showing a high level of polymorphism will be used to characterize the genetic diversity of the available captive pikeperch populations in parallel to wild stocks from which these stocks were founded and across European geographical regions (from North to South and from fresh and brackish water). Sampled populations will be from commercial farms, and at least eight populations with more than 50 fish each will be sampled and analyzed through genotyping with microsatellite markers. Basic population genetics parameters (allelic richness, heterozygosity indices, inbreeding coefficients) are going to be estimated with open access software, and also whether or not there is substantial genetic structure will be investigated since it is of particular importance not only for the management but also for the traceability of the species products.
Task 4.2 Evaluation of the genetic variation in non-domesticated broodstocks of pikeperch (led by HCMR). Since differences in biological characteristics may be related to genetic background, it can be expected that captive populations founded from wild populations of different geographical regions may display different zootechnical performances, as already shown in a preliminary study on Eurasian perch. Moreover, during the first period of habituation to captivity conditions, the rearing process is often conducted empirically (i.e., selecting fish on the basis of reproductive and growth performances and using a founding population of small size), without any real management of the genetic variability as for Eurasian perch culture. In other fish species it has been shown that such rearing practices result rapidly in a decrease in genetic variability and/or in a genetic drift of captive stocks. In the Eurasian perch, a 2-3fold decrease in allelic diversity is already observed after 4-5 generations reared under captive conditions. Ensuring sufficient genetic variation within populations is fundamental, because it determines the potential of adaptation to hostile changes in environmental/rearing conditions. Moreover, loss of genetic variability within the first generations of breeding practices will limit the potential for future genetic improvement from artificial selection in that cultured stock.
For this purpose, more than five wild pikeperch populations (at least 50 fish from each population) will be sampled from natural fisheries that have been identified by the consortium participants. Wild fish will be caught, anesthetised, a fin clip taken and released. The fin clips will be analyzed through genotyping with the same set of microsatellite markers as above. Genetic differentiation between wild samples and between wild and domesticated will be finally estimated following standard methodologies. Basic population genetics parameters (allelic richness, heterozygosity and inbreeding coefficients in each stock and phylogeographic relationships between populations) will be estimated in order to describe the genetic status of wild broodstock and for comparison with captive broodstocks. The genetic characterisation of all these stocks will be used to propose strategies to establish founding broodstocks that provide the genetic basis for the domestication of pikeperch through the selection for sustainable optimal culture performance.
Task 4.1 Evaluation of the genetic variation in available domesticated broodstocks of pikeperch (led by UL). Up to now, there has been no evaluation of the current genetic diversity of captive pikeperch. Yet, because there are only a few commercial hatcheries that produce pikeperch (around 10 farms), it is likely that the genetic diversity might be very low compared to the genetic variability of natural populations. Each pikeperch farm uses its own isolated stock captured either from the wild or supplied by another farmer. Therefore, pikeperch populations differ from one farm to another depending upon the geographical origin of the wild populations from which the captive stocks derived.
Several studies have been published on the variability of wild pikeperch. For instance, Saisa et al. (2010) have studied the genetic variability of three coastal and five freshwater populations in the northern part of the Baltic sea by using 12 microsatellite loci previously developed for the walleye Sander vitreum and the yellow perch Perca flavescens. They found that the coastal populations differ genetically from the lake populations, which present a higher genetic diversity. Saliminen et al., (2012) have further studied the genetic consequences and gene flow of pikeperch stocking in three lakes by comparing the pre- and post-release patterns using a subset of 9 microsatellites loci from Saisa et al. (2010). The genetic structures of populations were disrupted by the releases of fish. Other studies have analysed the genetic structure and dynamics of pikeperch at a regional scale using some of the above cited microsatellites loci.
Our first objective is to develop for the species a highly informative and efficient microsatellite multiplex consisted of more than 10 markers, which will allow the adequate genotyping of the fish sampled. Microsatellite loci will be first ordered by increasing size in base pairs (bp) and size range, and in each range one of the primers for each microsatellite locus, e.g. the reverse, will be fluorescently labelled with the dyes conformed to the ABI technology systems (FAM, NED, VIC and PET) and using the Qiagen multiplex PCR kit. The PCR conditions will be optimized in order to finally have a powerful molecular tool for genotyping. The above-mentioned commercial kit gives the advantage of maximal transferability of molecular protocols between labs.
Few microsatellites have already been developed in pikeperch , but numerous have proved to cross-amplify from phylogenetically close Percid species (i.e., walleye Sander vitreum, Eurasian perch Perca fluviatilis, Yellow perch Perca flavescens). Among these genetic markers, microsatellite loci showing a high level of polymorphism will be used to characterize the genetic diversity of the available captive pikeperch populations in parallel to wild stocks from which these stocks were founded and across European geographical regions (from North to South and from fresh and brackish water). Sampled populations will be from commercial farms, and at least eight populations with more than 50 fish each will be sampled and analyzed through genotyping with microsatellite markers. Basic population genetics parameters (allelic richness, heterozygosity indices, inbreeding coefficients) are going to be estimated with open access software, and also whether or not there is substantial genetic structure will be investigated since it is of particular importance not only for the management but also for the traceability of the species products.
Task 4.2 Evaluation of the genetic variation in non-domesticated broodstocks of pikeperch (led by HCMR). Since differences in biological characteristics may be related to genetic background, it can be expected that captive populations founded from wild populations of different geographical regions may display different zootechnical performances, as already shown in a preliminary study on Eurasian perch. Moreover, during the first period of habituation to captivity conditions, the rearing process is often conducted empirically (i.e., selecting fish on the basis of reproductive and growth performances and using a founding population of small size), without any real management of the genetic variability as for Eurasian perch culture. In other fish species it has been shown that such rearing practices result rapidly in a decrease in genetic variability and/or in a genetic drift of captive stocks. In the Eurasian perch, a 2-3fold decrease in allelic diversity is already observed after 4-5 generations reared under captive conditions. Ensuring sufficient genetic variation within populations is fundamental, because it determines the potential of adaptation to hostile changes in environmental/rearing conditions. Moreover, loss of genetic variability within the first generations of breeding practices will limit the potential for future genetic improvement from artificial selection in that cultured stock.
For this purpose, more than five wild pikeperch populations (at least 50 fish from each population) will be sampled from natural fisheries that have been identified by the consortium participants. Wild fish will be caught, anesthetised, a fin clip taken and released. The fin clips will be analyzed through genotyping with the same set of microsatellite markers as above. Genetic differentiation between wild samples and between wild and domesticated will be finally estimated following standard methodologies. Basic population genetics parameters (allelic richness, heterozygosity and inbreeding coefficients in each stock and phylogeographic relationships between populations) will be estimated in order to describe the genetic status of wild broodstock and for comparison with captive broodstocks. The genetic characterisation of all these stocks will be used to propose strategies to establish founding broodstocks that provide the genetic basis for the domestication of pikeperch through the selection for sustainable optimal culture performance.