Chloroplast DNA study in sweet cherry cultivars (Prunus avium L.) using PCR-RFLP method

The chloroplast DNA of 96 sweet cherry cultivars (Prunus avium L.) and five cultivars of sour cherry (P. cerasus L.) were analysed to reveal their haplotypes using PCR-RFLP (Polymerase Chain Reaction - Restriction Fragment Length Polymorphism) method. The main advantages of the PCR-RFLP technique are: resolutive, time and cost effective and reproducible. Approximately 9.6% of the chloroplast genome was analysed, using six universal primer pairs and two restriction enzymes. All the mutations detected were insertion-deletions, ranging between 5–30 bp. The combination of all the mutations resulted in three haplotypes (H1, H2 and H3) in the 96 sweet cherry cultivars and a single haplotype (H4) in the five cultivars of sour cherry. The cpDNA polymorphism determined by PCR-RFLP markers helped to understand the maternal inheritance of chloroplast genome in sweet cherry, clearly distinguished the sour from the sweet cherry, and supported that P. avium is not the maternal species of P. cerasus.     

Glacial refugia: hotspots but not melting pots of genetic diversity

Glacial refuge areas are expected to harbor a large fraction of the intraspecific biodiversity of the temperate biota. To test this hypothesis, we studied chloroplast DNA variation in 22 widespread European trees and shrubs sampled in the same forests. Most species had genetically divergent populations in Mediterranean regions, especially those with low seed dispersal abilities. However, the genetically most diverse populations were not located in the south but at intermediate latitudes, a likely consequence of the admixture of divergent lineages colonizing the continent from separate refugia.     

Mitochondrial D‐loop analysis reveals low diversity in Mangalica pigs and their relationship to historical specimens

The genetic relationship between 195 Mangalica and 79 non‐Mangalica pigs was studied using mitochondrial D‐loop SNP genotyping. Altogether, 35 polymorphic sites and 27 haplotypes were identified. Of the haplotypes, eight and 16 are Mangalica and non‐Mangalica specific, respectively, while three contain both Mangalica and non‐Mangalica individuals. Genetic distance values and phylogenetic analysis indicate that Mangalica individuals are very closely related, and five haplotypes represent approximately 92% of the Mangalica pigs involved in the study, thus determining the major maternal lineages. In contrast to previous microsatellite studies, individuals of Mangalica could not be distinguished as three separate breeds using mtDNA genotyping. Comparing modern and archaeological mtDNA sequences revealed that present day Mangalica is related to pigs that lived in the Carpathian basin where postulated ancestors of Mangalica also lived. This is the first DNA‐based genetic evidence to support the described breeding history of Mangalica.     

A comprehensive survey on selective breeding programs and seed market in the European aquaculture fish industry

The use of selective breeding is still relatively limited in aquaculture species. Information on such activities is sparse, hindering an overall evaluation of their success. Here, we report on the results of an online survey of the major aquaculture breeding companies operating in Europe. Six main reared fish species were targeted. A total of 31 respondents contributed to the survey, representing 75 % of European breeding organizations. Family-based breeding schemes were predominant, but individual selection was more frequently applied in marine species. Artificial fertilization is the preferred means of reproduction; however, mass spawning is often used as a fallback method. The most frequently selected trait is growth performance, but the number of selected traits has been increasing over the years through the addition of traits such as disease resistance or product quality. The use of molecular tools is now common in all programs, mainly for pedigree traceability. An increasing number of programs use either genomic or marker-assisted selection. Results related to the seed production market confirmed that for Atlantic salmon there are a few dominant players at the European level, with 30–50 % market share. Only part of the European fish aquaculture industry today fully exploits selective breeding to the best advantage. A larger impact assessment still needs to be made by the remainder, particularly on the market share of fish seed (eggs, larvae or juveniles) and its consequences for hatchery stability.     

Detection of Neospora caninum in semen of bulls

In cattle, transplacental infection is the main route of Neospora caninum transmission, but postnatal transmission by the oral uptake of sporozoite-containing oocysts shed by dogs may also be possible. Other routes of horizontal transmission, such as the venereal route, have not been investigated. In this study, we evaluated the presence of N. caninum DNA by a nested-PCR in fresh non-extended semen and frozen extended semen straws of five Holstein-Friesian bulls with naturally-acquired neosporosis. The infection status was assessed by an immunofluorescent antibody test (IFAT) and confirmed by immunoblotting (IB). Because of inhibitory components of semen, a protocol was developed to purify N. caninum DNA from bovine semen. Sporadically, N. caninum DNA was detected in non-extended fresh semen samples and frozen extended semen straws of the five seropositive bulls. In all positive samples, specific DNA was consistently found in the cell fraction of semen and not in seminal plasma. The parasite mean load in positive fresh semen samples determined by a real-time PCR was low oscillating between 1 and 2.8 parasites/ml of semen (maximum parasite load detected in one sample was 7.5 parasites/ml of semen). In parallel, another three similar but uninfected bulls acted as controls and no N. caninum DNA was amplified in any of their fresh and straw semen samples assayed. Whether venereal transmission plays a role in the spread of bovine neosporosis needs to be determined.     

Fingerprinting and genetic variability in cork oak (Quercus suber L.) elite trees using ISSR and SSR markers

Quercus suber L., is a socially, economically and ecologically important forest species in rural areas of the Mediterranean basin. Fifty three elite-trees from nine stands of four provenance regions in the Community of Extremadura (Spain) were analysed with the aim to establish their DNA-fingerprinting and the genetic relationships among them. Two types of molecular markers, microsatellites and intermicrosatellites, were used for tree genotyping. Microsatellite markers could distinguish 94% of the trees. Observed and expected heterozygosities, and effective number of alleles were correlated with the polymorphism information content (PIC). Intermicrosatellite patterns were used to construct a dendrogram. They showed high levels of genetic diversity in these elite trees, without any clear relationship with provenance regions. Both types of markers detected a high level of intra-stand variability, which is common in Quercus species, probably due to its mating system, the low distance among stands and the small size of stands. Others factors that could affect this result, such as introgression between Q. suber L. and Q. ilex L., are also commented. The high level of genetic variability detected in these elite trees can be useful for improvement programs. Also the usefulness of SSR and ISSR markers to establish the DNA-fingerprinting of each tree could be focused to select clonal lines of commercial interest.     

An exploration of wild <Emphasis Type="Italic">Brassica oleracea</Emphasis> L. germplasm in Northern Spain

An intense exploration of the Spanish Cantabrian coast for the presence or absence of wild Brassica oleracea L., yielded 24 new localities to be added to the 21 previously known. Of the resulting 45 localities, 22 correspond to Asturias, 11 to Cantabria and 12 to the Basque Country. Data on the habitat requirements of this plant have been annotated, the conservation status of each population has been estimated and seed samples have been collected for long-term preservation. As a whole, wild B. oleracea is not threatened in Northern Spain, but some populations and/or sub-populations are at risk either because of their small size or some detectable human impact.     

Water temperature,time of exposure and population density are key parameters in Enteromyxum leei fish-to-fish experimental transmission

Enteromyxum leei is a myxozoan histozoic parasite that infects the intestine of several teleost fish species. In gilthead sea bream (Sparus aurata), it provokes a chronic disease, entailing anorexia, delayed growth, reduced marketability and mortality. Direct fish-to-fish transmission, relevant in aquaculture conditions, has been demonstrated for E. leei via effluent, cohabitation, and oral and anal routes. However, the minimum time of exposure for infection has not been established, nor the possible effect on the fish immune response. Two effluent trials were performed at different temperatures (high: average of 25.6°C; and low: constant at 18°C), different times of exposure to the effluent (1, 3, 5 and 7 weeks) and different population densities. The results showed that 1 week was enough to infect 100% of fish at high temperature and 58.3% at low temperature. High temperature not only increased the prevalence of infection in posterior intestine, but also induced a higher production of specific antibodies, limiting the progression of the infection along the intestine. Longer time of exposure to the parasite and higher fish densities facilitated E. leei infection. These results show that effective diagnosis, lowering animal density and removal of infected fish are key aspects to manage this disease in aquaculture facilities.     

Seasonal changes in otolith increment width trajectories and the effect of temperature on the daily growth rate of young sardines

We studied the otolith microstructure and growth of sardine, Sardina pilchardus, in the North Aegean Sea (eastern Mediterranean Sea), using samples of larvae and juveniles that had hatched in winter (November–January) and winter–spring (February–May), respectively. The juveniles had developed during an extended period coinciding with marked pelagic ecosystem changes (from winter, mixed conditions to summer, stratified waters). To examine the relationship between environmental changes and the observed variability in their otolith increment–width trajectories (width‐at‐age), we summarized the shape of trajectories with a four‐parameter set estimated from a growth model fit to each width trajectory. The individual parameter sets were then related to the potential oceanographic conditions that fish experienced during their development, derived from a hydrodynamic–biogeochemical model (POM‐ERSEM), implemented in the sampling area. Substantial seasonal effects were demonstrated on the otolith microstructure (platykurtic versus leptokurtic trajectories in winter‐mixed versus summer‐stratified conditions), which were related to the progressive sea surface warming. In a subsequent step, in order to study the effect of oceanographic conditions on larval and juvenile daily growth rates, a GAM (Generalized Additive Model) analysis of otolith increment widths was carried out, using model‐derived oceanographic parameters and taking into account the ‘inherent otolith growth’, expressed by the explanatory variables ‘previous increment width’ and ‘Age’. Results showed a strong and positive, linear effect of temperature on the growth rate of winter‐caught larvae, whereas in juveniles, which had developed within a wide range of temperatures, an optimum temperature for growth was observed at around 24°C.     

A new approach to prioritizing marine fish and shellfish populations for conservation

There has been increasing awareness of the vulnerability of marine organisms to population extirpation and species extinction. While very few documented cases of species extinction exist in the marine environment, it is anticipated that managers will face the dilemma of prioritizing populations of marine fish and shellfish for protection in the near future. Current prioritization procedures have been developed from salmonid models with the intent of applying them to all marine organisms, and in some cases to freshwater and terrestrial taxa. In this review we provide evidence for the relevance of such a process for marine species and further suggest five broad categories of marine organisms that have distinctive traits influencing their genetic structure. The current prioritization models have been adapted to account for each of these species groups. Emphasis is placed on ‘Classical Marine Species’ which represent the opposite end of the continuum from the salmon model, displaying high within‐population genetic variance. From this category, three cod (Gadus morhua) stocks were selected to evaluate a revised scheme developed specifically for ‘Classical Marine Species’ that includes performance measures such as (i) reduction in number of spawning populations; (ii) reduction of Ne : Nc (ratio of effective to census population size); (iii) changes in life‐history traits; (iv) critical density for spawning success; and (v) patchy vs. continuous distribution pattern. When the salmonid scheme was applied, the cod examples were allocated low values, indicating that they were not under threat. However, when the revised scheme was applied, all three cod stocks were allocated high values, indicating that the revised scheme was more reflective of the particular life‐history traits of this category of organisms.