Species identification of two closely exploited flatfish, turbot (Scophthalmus maximus) and brill (Scophthalmus rhombus), using a ddRADseq genomic approach

Abstract

1. Application of genomics tools for conservation purposes (i.e. conservation genomics) allows looking deep into the genetic structure and adaptations of populations and species in order to define management units, trace exploited stocks, and identification of species.2. In this work, a double-digestion restriction-site-associated DNA sequencing (ddRADseq) approach was performed to identify individuals belonging to two important commercial flatfish, the turbot (Scophthalmus maximus) and the brill (Scophthalmus rhombus), and their potential hybrids. A totalof 83 diagnostic single nucleotide polymorphisms (SNPs) between brill and turbot were detected in the genomic RAD-tag database created with 122 turbot and 20 brill individuals. Markers consisted of an SNP with alternative alleles fixed in each species. From these diagnostic markers, a molecular tool comprising 10 SNPs located far apart in the turbot genome (>100 kb) was successfully designed and validated using the SNaPshot® method from a new sample set of 154 brill and turbot samples distributed throughout the natural range of both species. Additionally, 10 individuals (two turbot and eight brill) previously genotyped with ddRADseq were genotyped with SNaPshot to check for genotyping accuracy. 4. ddRADseq and SNaPshot genotyping of these 10 markers were identical in the 10 individuals used as controls. Species assignment based on the SNP tool corresponded with phenotypic identification for all samples. Using the SNP tool in a set of 29 unknown samples, all individuals were identified as S. maximus except one identified as a hybrid (heterozygous for all diagnostic SNPs).5. The methodology (i.e. ddRADseq) has enabled the development of a helpful traceability tool for sustainable management and conservation of turbot and brill resources. Similar approaches could also be applied to other non-model organisms despite lacking an assembled genome. * These authors contributed equally to this work and are first co-authors.

Scophthalmus maximus (turbot)

“…Some subpopulations also differ in their adaptation to local temperature and oxygen availability (Andersen et al 2020). This species is similar to that of Brill, Scophthalmus rhombus, and hybridization has been documented between the two species (Maroso et al 2018).…”

A Useful SNP Panel to Distinguish Two Cockle Species, Cerastoderma edule and C. glaucum, Co-Occurring in Some European Beds, and Their Putative Hybrids

“…E-96 mollusk DNA kit (OMEGA Bio-tech, Norcross, GA, USA), following the manufacturer recommendations. SNP identification and selection, as well as genotyping and validation protocols, were similar to those described by Maroso et al [21]. Briefly, genomic DNA was cut using the AlfI IIb restriction enzyme (RE), and then 2b–RAD libraries were constructed by joining adaptors to both fragment ends, followed by PCR amplification using primers targeting specific regions within adaptors.…”

“…The development of cutting-edge genomic technologies and protocols in the last decade has opened the possibility to develop a wide range of genotyping by sequencing methods that enable simultaneously identifying and genotyping thousands of single nucleotide polymorphisms (SNP) at low cost [20]. This approach has also been applied to compare large portions of the genomes of individuals from closely related species to look for diagnostic markers that make possible suitable discrimination between taxa [21]. Furthermore, these SNP-based tools enable distinguishing F1, F2, and backcrosses with high confidence, thus allowing a refined evaluation of the hybridization degree of individuals and populations.…”

Genomic Hatchery Introgression in Brown Trout (Salmo trutta L.): Development of a Diagnostic SNP Panel for Monitoring the Impacted Mediterranean Rivers

“…In our laboratories, without taking into account DNA extraction, the current price of RE digestion for LDH-C * genotyping using the methodology developed by McMeel et al (2001) [ 43 ] would be around EUR 7/individual to test just one SNP, while using the developed tool would be around EUR 5/individual for genotyping 20 SNPs, reducing drastically the cost per SNP. Other SNP tools were successfully applied for the identification of hybrids in aquatic organisms [ 104 , 105 , 106 , 107 ]. In brown trout, the detection of diagnostic native alleles may support the successful management practices addressed to identify black spots from past releases leading to almost naturalized populations (such as RQS18 in the present study).…”