Jose L. Martinez

Sex chromosome linkage of 5S rDNA in rainbow trout (Oncorhynchus mykiss)

The karyotype of the rainbow trout is characterized by a primitive XX/XY sex-determining chromosomal system. (Thorgaard et al., 1977). In the present study using FISH we have physically linked the 5S rRNA genes to the partially undifferentiated X chromosome pair. PCR amplified 5S rDNA was used for FISH and hybridization signals indicated that the genes were duplicated, present in one acrocentric and one metacentric pair of chromosomes. After analyzing several individuals, the female metaphases showed four fluorescent signals whereas males presented only three signals. Two of the three signals obtained in males corresponded to the metacentric pair whereas the single signal was mapped to the heterochromatin that cytologically differentiates the X chromosome from the Y chromosome. Double FISH experiments demonstrated that the 5S rDNA which is not sex linked is located at the NOR bearing arm close to the major ribosomal RNA genes (5.8S, 18S and 28S), similar to the situation observed in Atlantic salmon (Pendas et al., 1994a).

Applications of 5S rDNA in Atlantic salmon, brown trout, and in Atlantic salmon brown trout hybrid identification

The members of the salmonid family form perhaps the most economically important group of the world’s fish species. In Europe the most important species employed in fish farming are the introduced rainbow trout (Oncorhynchus mykiss) and the Atlantic salmon (Salmo salnr), whereas interest in the brown trout (Salmo trufta) is based on its ecological diversity and sport fishing. These two Salmo species are very difficult to distinguish using morphological characters, not only during their first months of life (eggs and alevins) but also at the returning adult stage. Isoenzyme genetic analysis, which has usually been used for their identification, has often demonstrated that anglers have confused adults of brown trout with Atlantic salmon, or with salmon x trout hybrids (Leaniz & Verspoor 1989). Caution must therefore be taken in enhancement programmes since the artificial spawning of adults caught in the river could lead to restocking with salmon x trout hybrids. Another area of interest is the identification of manufactured products of these species (i.e. smoked). Genetic analysis potentially provides the only reliable method to unambiguously determine their species identity. In addition, the genetic marker employed should be a nuclear marker if interspe cific hybrids among these species have to be recognized. From a practical point of view, the procedure chosen must be fast, extremely robust, and be able to utilize tiny quantities of any tissue. This last requirement excludes most isoenzyme techniques. At present, PCR-based methodologies constitute the most reliable techniques available for this purpose. Most of these approaches are, however, based on the use of conserved mitochondria1 DNA primers (for the cytochrome b gene) and sequencing of the amplified fragment (Paabo et al. 1989; Bartlett h Davidson 1992). Instead, we have focused on the 5s

Multiple paternity increases effective size of southern Atlantic salmon populations

Genetic analyses were performed on the progeny of Atlantic salmon (Salmo salar L.) sampled in natural redds of three rivers flowing into the Bay of Biscay, the Nivelle, the Mandeo and the Sella. These rivers are at the southern limit of the European distribution of the species and their populations are small and endangered by human activities. Nine variable number of tandem repeat (VNTR) loci (five minisatellites and four microsatellites) were used for parentage analysis. Multiple male participation was recognized in the fertilization of eggs. A large proportion was fertilized by precociously mature parr. We demonstrate that multiple paternity derived from mature parr is crucial for the conservation of genetic variability in small populations of Atlantic salmon.

Chromosomal localization of the major and 5S rRNA genes in the European eel (Anguilla anguilla)

The major (18S, 5.8S, and 28S) and 5S rRNA genes have been mapped by double fluorescent in situ hybridization to European eel metaphase chromosomes. The major rRNA genes were localized to a submetacentric pair of chromosomes that showed a consistent size polymorphism among the individuals studied. The 5S rRNA genes were clustered in a single locus that mapped to the centromeric region of an acrocentric pair. In contrast to the major rRNA genes, no detectable polymorphism, in either size or intensity of the fluorescent signal, was observed. The chromosomal organization of both families of rRNA genes are discussed in terms of genomic organization and chromosomal evolution.

High Level of Mislabeling in Spanish and Greek Hake Markets Suggests the Fraudulent Introduction of African Species

DNA analysis of hake products commercialized in southern European (Spanish and Greek) market chains have demonstrated more than 30% mislabeling, on the basis of species substitution. Tails and fillets were more mislabeled than other products, such as slices and whole pieces. African species were substitute species for products labeled as American and European species, and we suggest it is a case of deliberate economically profitable mislabeling because real market prices of European and American hake products are higher than those of African in Spanish market chains. The presented results suggest fraud detection that disadvantages African producers. Government-mandated genetic surveys of commercial hakes and the use of subsequent statements of fair trade on labels of seafood products could help to reduce fraud levels in a global market of increasingly conscious consumers sensitive to ethical issues.

Detection of Mislabeling in Hake Seafood Employing mtSNPs-Based Methodology with Identification of Eleven Hake Species of the Genus Merluccius

Species-specific DNA-based tags are valuable tools for the management of both fisheries and commercial fish products. Eleven hake species of the genus Merluccius have been identified employing mtSNPs-based methodology. The method is highly reproducible, fast, and technically easy. It is a reliable tool, allowing for routine analysis of commercial seafood. It can be applied by nonexperts in genetics because both laboratory handling and interpretation of results are easy and direct. The convenience of routine surveys in fish markets has been clearly established with a survey of commercial hake batches imported in south Europe. A total of 40 commercial processed hake were analyzed in this study. More than 20% of mislabeling has been detected.

Dealcoholisation of Cider by Supercritical Extraction with Carbon Dioxide

A study of cider dealcoholisation by supercritical extraction with carbon dioxide using pilot-plant-scale equipment has been carried out. The effects of pressure, temperature and solvent to feed ratio have been examined. Supercritical and liquid carbon dioxide have been used to extract ethanol and aromas from cider at pressures of 80-250 bar and temperatures of 20 and 40°C.

Interspecific barriers between salmonids when hybridisation is due to sneak mating

Male sneaking behaviour can lead to interspecific hybridisation if sneakers attempt to fertilise ova in heterospecific mating, contributing to break down of interspecific barriers. In south European rivers, sneaking Atlantic salmon males fertilise an important proportion of ova from adult females in heterospecific crosses, up to 65%. In a south French flow-controlled stream, we found that they were able to naturally fertilise brown trout ova in absence of brown trout males. Aggressiveness of brown trout males towards sneaking salmon males and low survival of hybrids issued from salmon sneakers are found to be interspecific barriers.

Fine Spatial Structure of Atlantic Hake (Merluccius merluccius) Stocks Revealed by Variation at Microsatellite Loci

Genetic variation at 5 microsatellite loci was analyzed for European hake Merluccius merluccius sampled from 9 different regions in the Atlantic Ocean and the Mediterranean Sea. Significant genetic differentiation was found between samples, suggesting a fine subdivision of Atlantic and Mediterranean hake stocks. These results are discussed in the context of the decline of demersal fish species, probably due to overfishing.

Detecting nuisance species using NGST: Methodology shortcomings and possible application in ballast water monitoring

Detecting the presence of potential invasive species in ballast water is a priority for preventing their spread into new environments. Next generation sequencing technologies are being increasingly used for exploring and assessing biodiversity from environmental samples. Here we apply high throughput sequencing from DNA extracted from ballast water (BW) samples employing two different platforms, Ion Torrent and 454, and compare the putative species catalogues from the resulting Operational Taxonomic Units (OTU). Water samples were taken from the RV Polastern ballast tank in five different days between the second and the twentieth navigation day. Pronounced decrease of oxygen concentration and increase of temperature occurred in the BW during this time, coincident with a progressively higher proportion of unassigned OTU and short reads indicating DNA degradation. Discrepancy between platforms for species catalogues was consistent with previously published bias in AT-rich sequences for Ion Torrent platform. Some putative species detected from the two platforms increased in frequency during the Polarstern travel, which suggests they were alive and therefore tolerant to adverse conditions. OTU assigned to the highly invasive red alga Polysiphonia have been detected at low but increasing frequency from the two platforms. Although in this moment NGST could not replace current methods of sampling, sorting and individual taxonomic identification of BW biota, it has potential as an exploratory methodology especially for detecting scarce species.