Aitor Albaina

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 October 2009–30 November 2009

This article documents the addition of 411 microsatellite marker loci and 15 pairs of Single Nucleotide Polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Acanthopagrus schlegeli, Anopheles lesteri, Aspergillus clavatus, Aspergillus flavus, Aspergillus fumigatus, Aspergillus oryzae, Aspergillus terreus, Branchiostoma japonicum, Branchiostoma belcheri, Colias behrii, Coryphopterus personatus, Cynogolssus semilaevis, Cynoglossus semilaevis, Dendrobium officinale, Dendrobium officinale, Dysoxylum malabaricum, Metrioptera roeselii, Myrmeciza exsul, Ochotona thibetana, Neosartorya fischeri, Nothofagus pumilio, Onychodactylus fischeri, Phoenicopterus roseus, Salvia officinalis L., Scylla paramamosain, Silene latifo, Sula sula, and Vulpes vulpes. These loci were cross‐tested on the following species: Aspergillus giganteus, Colias pelidne, Colias interior, Colias meadii, Colias eurytheme, Coryphopterus lipernes, Coryphopterus glaucofrenum, Coryphopterus eidolon, Gnatholepis thompsoni, Elacatinus evelynae, Dendrobium loddigesii Dendrobium devonianum, Dysoxylum binectariferum, Nothofagus antarctica, Nothofagus dombeyii, Nothofagus nervosa, Nothofagus obliqua, Sula nebouxii, and Sula variegata. This article also documents the addition of 39 sequencing primer pairs and 15 allele specific primers or probes for Paralithodes camtschaticus.

Multiple SNP Markers Reveal Fine-Scale Population and Deep Phylogeographic Structure in European Anchovy (Engraulis encrasicolus L.)

Geographic surveys of allozymes, microsatellites, nuclear DNA (nDNA) and mitochondrial DNA (mtDNA) have detected several genetic subdivisions among European anchovy populations. However, these studies have been limited in their power to detect some aspects of population structure by the use of a single or a few molecular markers, or by limited geographic sampling. We use a multi-marker approach, 47 nDNA and 15 mtDNA single nucleotide polymorphisms (SNPs), to analyze 626 European anchovies from the whole range of the species to resolve shallow and deep levels of population structure. Nuclear SNPs define 10 genetic entities within two larger genetically distinctive groups associated with oceanic variables and different life-history traits. MtDNA SNPs define two deep phylogroups that reflect ancient dispersals and colonizations. These markers define two ecological groups. One major group of Iberian-Atlantic populations is associated with upwelling areas on narrow continental shelves and includes populations spawning and overwintering in coastal areas. A second major group includes northern populations in the North East (NE) Atlantic (including the Bay of Biscay) and the Mediterranean and is associated with wide continental shelves with local larval retention currents. This group tends to spawn and overwinter in oceanic areas. These two groups encompass ten populations that differ from previously defined management stocks in the Alboran Sea, Iberian-Atlantic and Bay of Biscay regions. In addition, a new North Sea-English Channel stock is defined. SNPs indicate that some populations in the Bay of Biscay are genetically closer to North Western (NW) Mediterranean populations than to other populations in the NE Atlantic, likely due to colonizations of the Bay of Biscay and NW Mediterranean by migrants from a common ancestral population. Northern NE Atlantic populations were subsequently established by migrants from the Bay of Biscay. Populations along the Iberian-Atlantic coast appear to have been founded by secondary waves of migrants from a southern refuge.

SNP Discovery in European Anchovy (Engraulis encrasicolus, L) by High-Throughput Transcriptome and Genome Sequencing

Increased throughput in sequencing technologies has facilitated the acquisition of detailed genomic information in non-model species. The focus of this research was to discover and validate SNPs derived from the European anchovy (Engraulis encrasicolus) transcriptome, a species with no available reference genome, using next generation sequencing technologies. A cDNA library was constructed from four tissues of ten fish individuals corresponding to three populations of E. encrasicolus, and Roche 454 GS FLX Titanium sequencing yielded 19,367 contigs. Additionally, the European anchovy genome was sequenced for the same ten individuals using an Illumina HiSeq2000. Using a computational pipeline for combining transcriptome and genome information, a total of 18,994 SNPs met the necessary minor allele frequency and depth filters. A series of further stringent filters were applied to identify those SNPs likely to succeed in genotyping assays, and for filtering of those in potential duplicated genome regions. A novel method for detecting potential intron-exon boundaries in areas of putative SNPs has also been applied in silico to improve genotyping success. In all, 2,317 filtered putative transcriptome SNPs suitable for genotyping primer design were identified. From those, a subset of 530 were selected, with the genotyping results showing the highest reported conversion and validation rates (91.3% and 83.2%, respectively) reported to date for a non-model species. This study represents a promising strategy to discover genotypable SNPs in the exome of non-model organisms. The genomic resource generated for E. encrasicolus, both in terms of sequences and novel markers, will be informative for research into this species with applications including traceability studies, population genetic analyses and aquaculture.

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 October 2011 - 30 November 2011: PERMANENT GENETIC RESOURCES NOTEM

This article documents the addition of 139 microsatellite marker loci and 90 pairs of single‐nucleotide polymorphism sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Aglaoctenus lagotis, Costus pulverulentus, Costus scaber, Culex pipiens, Dascyllus marginatus, Lupinus nanus Benth, Phloeomyzus passerini, Podarcis muralis, Rhododendron rubropilosum Hayata var. taiwanalpinum and Zoarces viviparus. These loci were cross‐tested on the following species: Culex quinquefasciatus, Rhododendron pseudochrysanthum Hay. ssp. morii (Hay.) Yamazaki and R. pseudochrysanthum Hayata. This article also documents the addition of 48 sequencing primer pairs and 90 allele‐specific primers for Engraulis encrasicolus.

18S rRNA V9 metabarcoding for diet characterization: a critical evaluation with two sympatric zooplanktivorous fish species

Abstract The potential of the 18S rRNA V9 metabarcoding approach for diet assessment was explored using MiSeq paired‐end (PE; 2 × 150 bp) technology. To critically evaluate the method′s performance with degraded/digested DNA, the diets of two zooplanktivorous fish species from the Bay of Biscay, European sardine (Sardina pilchardus) and European sprat (Sprattus sprattus), were analysed. The taxonomic resolution and quantitative potential of the 18S V9 metabarcoding was first assessed both in silico and with mock and field plankton samples. Our method was capable of discriminating species within the reference database in a reliable way providing there was at least one variable position in the 18S V9 region. Furthermore, it successfully discriminated diet between both fish species, including habitat and diel differences among sardines, overcoming some of the limitations of traditional visual‐based diet analysis methods. The high sensitivity and semi‐quantitative nature of the 18S V9 metabarcoding approach was supported by both visual microscopy and qPCR‐based results. This molecular approach provides an alternative cost and time effective tool for food‐web analysis.

Insights on the origin of invasive copepods colonizing Basque estuaries; a DNA barcoding approach

BackgroundThe introduction of NIS to estuaries and coastal embayment is of great concern. Commercial ships’ ballast water discharge and the northwards progression of species due to the ongoing climate change arise as the main factors explaining the rising occurrence of NIS species in Northern Atlantic waters. In this regard, regular monitoring of the plankton communities is paramount as to be able to respond properly to this potential issue.ResultsWhile monitoring the invasive copepod Acartia tonsa populations in the estuaries of Bilbao and Urdaibai (Basque country, Spain), we report here the Asian copepod Pseudodiaptomus marinus for the first time in the Iberian Peninsula waters. Individuals from both species were collected from July to October, 2013 for DNA sequencing of mitochondrial cytochrome c oxidase subunit I gene (MT-CO1). Phylogenetic analysis of MT-CO1 confirmed P. marinus identity.ConclusionsPhylogeographic distribution of A. tonsa haplotypes in Europe along with the Bilbao port traffic patterns suggested a secondary invasion from an European source to Basque estuaries. The successful establishment of the A. tonsa population and the appearance of Pseudodiaptomus marinus confirm the need for regular plankton monitoring of estuarine and port waters. This applies also to nearby systems as these populations could represent a source of future dispersal.

Unraveling the environmental and anthropogenic drivers of bacterial community changes in the Estuary of Bilbao and its tributaries

In this study, 16S rRNA gene sequencing was used to characterize the changes in taxonomic composition and environmental factors significantly influencing bacterial community structure across an annual cycle in the Estuary of Bilbao as well as its tributaries. In spite of this estuary being small and characterized by a short residence time, the environmental factors most highly correlated with the bacterial community mirrored those reported to govern larger estuaries, specifically salinity and temperature. Additionally, bacterial community changes in the estuary appeared to vary with precipitation. For example, an increase in freshwater bacteria (Comamonadaceae and Sphingobacteriaceae) was observed in high precipitation periods compared to the predominately marine-like bacteria (Rhodobacterales and Oceanospirillales) that were found in low precipitation periods. Notably, we observed a significantly higher relative abundance of Comamonadaceae than previously described in other estuaries. Furthermore, anthropic factors could have an impact on this particular estuary’s bacterial community structure. For example, ecosystem changes related to the channelization of the estuary likely induced a low dissolved oxygen (DO) concentration, high temperature, and high chlorophyll concentration period in the inner euhaline water in summer (samples with salinity >30 ppt). Those samples were characterized by a high abundance of facultative anaerobes. For instance, OTUs classified as Cryomorphaceae and Candidatus Aquiluna rubra were negatively associated with DO concentration, while Oleiphilaceae was positively associated with DO concentration. Additionally, microorganisms related to biological treatment of wastewater (e.g Bdellovibrio and Zoogloea) were detected in the samples immediately downstream of the Bilbao Wastewater Treatment Plant (WWTP). There are several human activities planned in the region surrounding the Estuary of Bilbao (e.g. sediment draining, architectural changes, etc.) which will likely affect this ecosystem. Therefore, the addition of bacterial community profiling and diversity analysis into the estuary’s ongoing monitoring program would provide a more comprehensive view of the ecological status of the Estuary of Bilbao.

Spatial dynamics and mixing of bluefin tuna in the Atlantic Ocean and Mediterranean Sea revealed using next-generation sequencing

The Atlantic bluefin tuna is a highly migratory species emblematic of the challenges associated with shared fisheries management. In an effort to resolve the species’ stock dynamics, a genomewide search for spatially informative single nucleotide polymorphisms (SNPs) was undertaken, by way of sequencing reduced representation libraries. An allele frequency approach to SNP discovery was used, combining the data of 555 larvae and young‐of‐the‐year (LYOY) into pools representing major geographical areas and mapping against a newly assembled genomic reference. From a set of 184,895 candidate loci, 384 were selected for validation using 167 LYOY. A highly discriminatory genotyping panel of 95 SNPs was ultimately developed by selecting loci with the most pronounced differences between western Atlantic and Mediterranean Sea LYOY. The panel was evaluated by genotyping a different set of LYOY (n = 326), and from these, 77.8% and 82.1% were correctly assigned to western Atlantic and Mediterranean Sea origins, respectively. The panel revealed temporally persistent differentiation among LYOY from the western Atlantic and Mediterranean Sea (FST = 0.008, p = .034). The composition of six mixed feeding aggregations in the Atlantic Ocean and Mediterranean Sea was characterized using genotypes from medium (n = 184) and large (n = 48) adults, applying population assignment and mixture analyses. The results provide evidence of persistent population structuring across broad geographic areas and extensive mixing in the Atlantic Ocean, particularly in the mid‐Atlantic Bight and Gulf of St. Lawrence. The genomic reference and genotyping tools presented here constitute novel resources useful for future research and conservation efforts.