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We report the first genetic linkage map of the catfish Lophiosilurus alexandri, a South American threatened and promising aquaculture catfish species. Using the progeny of three full-sib families, with 141, 74, and 49 offspring, respectively, we could genotype 2351 SNP markers using the ddRAD technology, shared by the male and the female maps constructed. The averaged, female and male maps spanned a total length of 2201.3 cM, 2481.9 cM, and 1872.8 cM, respectively, and comprehended the expected 27 linkage groups according to the karyotype information of the species (2n = 54). The recombination rate was nearly twice higher in the female than in the male map. The average map was used to estimate the historical effective population size (Ne) of the species from linkage disequilibrium between pairs of SNPs using parental individuals and revealed a remarkable drop in Ne about 20–25 generations in the past. The construction of the biggest artificial reservoir in Latin America and associated nutrient retention is pointed as a possible reason for such a reduction and suggests a reevaluation of the conservation status of the species. The current map lays the groundwork for understanding the genetic basis of economically important traits in breeding programs and will be useful for the genome assembly of this important commercial species

The common cockle (Cerastoderma edule) plays an important role in marine ecosystems and represents a valuable socioeconomic resource for coastal communities. In 2012, the cockle beds from Rı́a de Arousa (Galicia, NW Spain) were seriously decimated by the protozoan Marteilia cochillia responsible for marteiliosis. We aimed to identify single nucleotide polymorphisms (SNP) markers potentially associated with resilience to marteiliosis to be used in marker-assisted selection programs for restoring affected cockle beds and recovering their production. For this, we carried out a population genomics approach using 2b-RADseq, where 38 naive samples (before the first detection of M. cochillia in 2012) from two beds of Rı́a de Arousa were compared with 39 affected samples collected in 2018/2019 (after several years of marteiliosis occurring in the area), collected either before (15 non-exposed samples) or during (24 exposed samples) the marteiliosis outbreak. Additionally, 767 differentially expressed genes (DEG) from a previous transcriptomic study addressed during the aforementioned 2018/19 marteiliosis outbreak, were evaluated to identify SNPs showing signals of selection. Using 2b-RADseq, 9,154 SNPs were genotyped and among them, 110 consistent outliers for divergent selection were identified. This set of SNPs was able to discriminate the samples according to their marteiliosis status (naive vs affected; exposed vs non-exposed), while another 123 SNPs were identified linked to DEGs associated with the level of infection across a temporal series. Finally, combining the population genomics and transcriptomics information, we selected the 60 most reliable SNPs associated with marteiliosis resilience. These SNPs were close to or within DEGs, and many of them were related to immune response (phagocytosis and cell adhesion), defence, such as apoptosis, stress, and cellular cycle, among other functions. This set of SNPs will eventually be validated to develop a cost-effective g

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis

European flat oyster (Ostrea edulis) is an ecologically and economically important marine bivalve, that has been severely affected by the intracellular parasite Bonamia ostreae. In this study, a flat oyster SNP array (~14,000 SNPs) was used to validate previously reported outlier loci for divergent selection associated with B. ostreae exposure in the Northeast Atlantic Area. A total of 134 wild and hatchery individuals from the North Sea, collected in naïve (NV) and long-term affected (LTA) areas, were analysed. Genetic diversity and differentiation were related to the sampling origin (wild vs. hatchery) when using neutral markers, and to bonamiosis status (NV vs. LTA) when using outlier loci for divergent selection. Two genetic clusters appeared intermingled in all sampling locations when using outlier loci, and their frequency was associated with their bonamiosis status. When both clusters were compared, outlier data sets showed high genetic divergence (FST > 0.25) unlike neutral loci (FST not ≠ 0). Moreover, the cluster associated with LTA samples showed much higher genetic diversity and significant heterozygote excess with outlier loci, but not with neutral data. Most outliers mapped on chromosome 8 (OE-C8) of the flat oyster genome, supporting a main genomic region underlying resilience to bonamiosis. Furthermore, differentially expressed genes previously reported between NV and LTA strains showed higher mapping density on OE-C8. A range of relevant immune functions were specifically enriched among genes annotated on OE-C8, providing hypotheses for resilience mechanisms to an intracellular parasite. The results suggest that marker-assisted selection could be applied to breed resilient strains of O. edulis to bonamiosis, if lower parasite load and/or higher viability of the LTA genetic cluster following B. ostreae infection is demonstrated

Background: The irruption of Next-generation sequencing (NGS) and restriction site-associated DNA sequencing (RAD-seq) in the last decade has led to the identification of thousands of molecular markers and their genotyping for refined genomic screening. This approach has been especially useful for non-model organisms with limited genomic resources. Many building-loci pipelines have been developed to obtain robust single nucleotide polymorphism (SNPs) genotyping datasets using a de novo RAD-seq approach, i.e. without reference genomes. Here, the performances of two building-loci pipelines, STACKS 2 and Meyer’s 2b-RAD v2.1 pipeline, were compared using a diverse set of aquatic species representing different genomic and/or population structure scenarios. Two bivalve species (Manila clam and common edible cockle) and three fish species (brown trout, silver catfish and small-spotted catshark) were studied. Four SNP panels were evaluated in each species to test both different building-loci pipelines and criteria for SNP selection. Furthermore, for Manila clam and brown trout, a reference genome approach was used as control. Results: Despite different outcomes were observed between pipelines and species with the diverse SNP calling and filtering steps tested, no remarkable differences were found on genetic diversity and differentiation within species with the SNP panels obtained with a de novo approach. The main differences were found in brown trout between the de novo and reference genome approaches. Genotyped vs missing data mismatches were the main genotyping difference detected between the two building-loci pipelines or between the de novo and reference genome comparisons. Conclusions: Tested building-loci pipelines for selection of SNP panels seem to have low influence on population genetics inference across the diverse case-study scenarios here studied. However, preliminary trials with different bioinformatic pipelines are suggested to evaluate their influence on pop

Background: Understanding sex determination (SD) across taxa is a major challenge for evolutionary biology. The new genomic tools are paving the way to identify genomic features underlying SD in fish, a group frequently showing limited sex chromosome differentiation and high SD evolutionary turnover. Turbot (Scophthalmus maximus) is a commercially important flatfish with an undifferentiated ZW/ZZ SD system and remarkable sexual dimorphism. Here we describe a new long-read turbot genome assembly used to disentangle the genetic architecture of turbot SD by combining genomics and classical genetics approaches. Results: The new turbot genome assembly consists of 145 contigs (N50 = 22.9 Mb), 27 of them representing >95% of its estimated genome size. A genome wide association study (GWAS) identified a ~ 6.8 Mb region on chromosome 12 associated with sex in 69.4% of the 36 families analyzed. The highest associated markers flanked sox2, the only gene in the region showing differential expression between sexes before gonad differentiation. A single SNP showed consistent differences between Z and W chromosomes. The analysis of a broad sample of families suggested the presence of additional genetic and/or environmental factors on turbot SD. Conclusions: The new chromosome-level turbot genome assembly, one of the most contiguous fish assemblies to date, facilitated the identification of sox2 as a consistent candidate gene putatively driving SD in this species. This chromosome SD system barely showed any signs of differentiation, and other factors beyond the main QTL seem to control SD in a certain proportion of families

The European flat oyster (Ostrea edulis) is a highly appreciated mollusk with an important aquaculture production throughout the 20th century, in addition to playing an important role on coastal ecosystems. Overexploitation of natural beds, habitat degradation, introduction of non‐native species, and epidemic outbreaks have severely affected this important resource, particularly, the protozoan parasite Bonamia ostreae, which is the main concern affecting its production and conservation. In order to identify genomic regions and markers potentially associated with bonamiosis resistance, six oyster beds distributed throughout the European Atlantic coast were sampled. Three of them have been exposed to this parasite since the early 1980s and showed some degree of innate resistance (long‐term affected group, LTA), while the other three were free of B. ostreae at least until sampling date (naïve group, NV). A total of 14,065 SNPs were analyzed, including 37 markers from candidate genes and 14,028 from a medium‐density SNP array. Gene diversity was similar between LTA and NV groups suggesting no genetic erosion due to long‐term exposure to the parasite, and three population clusters were detected using the whole dataset. Tests for divergent selection between NV and LTA groups detected the presence of a very consistent set of 22 markers, located within a putative single genomic region, which suggests the presence of a major quantitative trait locus associated with B. ostreae resistance. Moreover, 324 outlier loci associated with factors other than bonamiosis were identified allowing fully discrimination of all the oyster beds. A practical tool which included the 84 highest discriminative markers for tracing O. edulis populations was developed and tested with empirical data. Results reported herein could assist the production of stocks with improved resistance to bonamiosis and facilitate the management of oyster beds for recovery production and ecosystem services provided b

Unraveling adaptive genetic variation represents, in addition to the estimate of population demographic parameters, a cornerstone for the management of aquatic natural living resources, which, in turn, represent the raw material for breeding programs. The turbot (Scophthalmus maximus) is a marine flatfish of high commercial value living on the European continental shelf. While wild populations are declining, aquaculture is flourishing in southern Europe. We evaluated the genetic structure of turbot throughout its natural distribution range (672 individuals; 20 populations) by analyzing allele frequency data from 755 single nucleotide polymorphism discovered and genotyped by double‐digest RAD sequencing. The species was structured into four main regions: Baltic Sea, Atlantic Ocean, Adriatic Sea, and Black Sea, with subtle differentiation apparent at the distribution margins of the Atlantic region. Genetic diversity and effective population size estimates were highest in the Atlantic populations, the area of greatest occurrence, while turbot from other regions showed lower levels, reflecting geographical isolation and reduced abundance. Divergent selection was detected within and between the Atlantic Ocean and Baltic Sea regions, and also when comparing these two regions with the Black Sea. Evidence of parallel evolution was detected between the two low salinity regions, the Baltic and Black seas. Correlation between genetic and environmental variation indicated that temperature and salinity were probably the main environmental drivers of selection. Mining around the four genomic regions consistently inferred to be under selection identified candidate genes related to osmoregulation, growth, and resistance to diseases. The new insights are useful for the management of turbot fisheries and aquaculture by providing the baseline for evaluating the consequences of turbot releases from restocking and farming.

Highly dense linkage maps enable positioning thousands of landmarks useful for anchoring the whole genome and for analysing genome properties. Turbot is the most important cultured flatfish worldwide and breeding programs in the fifth generation of selection are targeted to improve growth rate, obtain disease resistant broodstock and understand sex determination to control sex ratio. Using a Restriction-site Associated DNA approach, we genotyped 18,214 single nucleotide polymorphism in 1,268 turbot individuals from 31 full-sibling families. Individual linkage maps were combined to obtain a male, female and species consensus maps. The turbot consensus map contained 11,845 markers distributed across 22 linkage groups representing a total normalised length of 3,753.9 cM. The turbot genome was anchored to this map, and scaffolds representing 96% of the assembly were ordered and oriented to obtain the expected 22 megascaffolds according to its karyotype. Recombination rate was lower in males, especially around centromeres, and pairwise comparison of 44 individual maps suggested chromosome polymorphism at specific genomic regions. Genome comparison across flatfish provided new evidence on karyotype reorganisations occurring across the evolution of this fish group

The impact of escapees from aquaculture is of general concern for the sustainability of natural resources. Turbot Scophthalmus maximus is a marine flatfish of great commercial value whose land-based aquaculture started approx. 40 yr ago; hence, a low impact of escapees is expected on wild populations. However, enhancement of wild stocks using farmed turbot has been carried out along the Northeast Atlantic coasts in the last decades. Recently, a broad panel of single nucleotide polymorphism (SNP) markers (755 SNPs; 1 SNP Mb−1) has been used to evaluate the genetic structure of turbot throughout its distribution range, constituting the baseline to evaluate the impact of farmed fish in the wild. Two distinct origins were identified for farmed turbot (F_ORI1 and F_ORI2; FST = 0.049), which differentiated from wild populations after 5 generations of selection (average FST = 0.059), and consistent evidence of adaptation to domestication was de - tected. A notable proportion of fish of farmed ancestry was detected in the wild (15.5%), mainly in the North Sea, where restocking activities have taken place, determining genetic introgression in wild populations. Conversely, effects of land-based aquaculture appear negligible. A simulation exercise supported panels of 40 and 80 SNPs to identify fishes of F_ORI1 and F_ORI2 ancestry in the wild, respectively. Application to empirical data showed an assignment success (wild/farmed ancestry) of approx. 95% in comparison with the full SNP dataset. The SNP tools will be useful to monitor turbot of farmed ancestry in the wild, which might represent a risk, considering the lower fitness of farmed individuals