Squalius Alburnoides Research Articles

Lack of correspondence between CMA3-, Ag-positive signals and 28S rDNA loci in two Iberian minnows (Teleostei, Cyprinidae) evidenced by sequential banding

Despite the growing outcome of results that put doubt upon the reliability of silver (Ag) staining and chromomycin A₃ (CMA₃) fluorescent banding in the detection of major ribosomal gene sites (NORs), these methods have been widely used, especially in fishes. In order to clarify the previous patterns obtained with those techniques, we performed fluorescence in situ hybridisation (FISH) with 28S rDNA probe followed by sequential CMA₃ and Ag staining in diploid non-hybrid males of the Squalius alburnoides complex and in Squalius pyrenaicus. The results from all the studied specimens revealed a lack of correlation between classical and molecular techniques. Not just some other regions besides NORs were stained with CMA₃ and Ag, but also the majority of the 28S rDNA sites were not detected. Care should then be taken in considering CMA₃- and Ag-stained sites as NORs since their accuracy for that purpose may not always correspond to the expectations.

Contrasting views over a hybrid complex: Between speciation and evolutionary “dead-end”

The Squalius alburnoides complex, of hybrid origin, comprises diploid, triploid and tetraploid forms and has a widespread distribution in the Iberian waters. The southern populations of this complex, sympatric with S. pyrenaicus, show high genetic variability, diversity of forms and reproductive modes which create pathways that may allow for the establishment of a new species in the future. Here we report a contrasting view over the S. alburnoides complex: in the Mondego River basin (northern Portugal), nuclear “non-hybrid” and tetraploid forms are absent and a clearly impoverished genetic diversity is observed, contributing to a general scarcity of possibilities of generating novel genetic material. Moreover, the bisexual species involved in the maintenance of the complex in this basin ( S. carolitertii) exhibits a considerably lower genetic variability, when compared with S. pyrenaicus. The observed differences suggest that, despite being originated by similar hybridization events and maintained by analogous reproductive mechanisms, different populations of the complex were exposed to distinct evolutionary constrains, which in some cases resulted in diversification and speciation while in others led to a compromising situation in terms of evolutionary potential. Additionally, and for the first time all forms were used in the calculation of genetic distances and diversity indices, widening the possibilities of analysis of the complex by allowing the inclusion of a large part of the available data, irrespective of ploidy level.

Variability in diet and foraging behaviour between sexes and ploidy forms of the hybridogeneticSqualius alburnoidescomplex (Cyprinidae) in the Guadiana River basin, Portugal

Samples of theSqualius alburnoidescomplex, a hybridogenetic Iberian minnow composed of diploid, triploid and, less commonly, tetraploid forms, were collected from a tributary of the Ardila River (Guadiana River basin) between February 1999 and January 2001. Seasonal variation in diet was evident, and was probably linked to prey availability. Distinct foraging behaviours between ploidy forms were found towards several prey items, suggesting that diploid adult males fed mostly near the surface, whereas diploid adult females tended to feed near the bottom and submerged vegetation. Triploid females exhibited an intermediate foraging behaviour, although there was greater affinity towards diploid male feeding behaviour. Diploid males which, in contrast to diploid and triploid females, have non‐hybrid genomes in the Guadiana drainage, exhibited a higher specialization for food. Despite considerable dietary overlap, there appeared to be spatial segregation of feeding niches between the three forms, especially during dry periods when prey availability was lower, which may be a strategy for diminishing competition for food. Therefore, considering asexual generalist and specialist hypotheses, it appears that the different ploidy levels are generalist, opportunistic feeders that partition the resources when limited.

Heterozygous indels as useful tools in the reconstruction of DNA sequences and in the assessment of ploidy level and genomic constitution of hybrid organisms

In this paper we describe a simple approach using double peaks in chromatograms generated as artefacts in the vicinity of heterozygous indels, to identify the specific sequences present in individual strands of a given DNA fragment. This method is useful to assign bases in individuals that are heterozygous at multiple sites. In addition, the relative sizes of the double peaks help to determine the ploidy level and the relative contribution of the parental genomes in hybrids. Our interpretation was confirmed with the analysis of artificial mixtures of DNA of two different species. Results were robust with varying PCR and sequencing conditions. The applicability of this method was demonstrated in hybrids of the Squalius alburnoides complex and in heterozygotes of Chondrostoma oligolepis. Far from being limited to these fish models and the gene where it was tested (beta-actin), this sequence reconstruction methodology is expected to have a broader application.

Ecología trófica de la nutria paleártica Lutra lutra en el río Francia (Cuenca del Tajo, Salamanca)

Trophic ecology of the otter Lutra lutra in the River Francia (Salamanca, Central Spain). The trophic ecology of the paleartic otter (Lutra lutra) is analyzed on the basis of its spraints collected along the river Francia (Salamanca), belonging to the hydrographic basin of the river Tajo. The study analyzes the changes in diet, both seasonally during a complete year and altitudinally along the river Francia on a stretch 24.5 Km long and approximately 1000 m of difference in height. Fish are the fundamental prey while the remainder of groups are accidental. Nine species of fish were found during the study, the principal one being Squalius pyrenaicus, followed by Barbus sp, Chondrostoma lemmingii and Salmo trutta. Chondrostoma arcasii is consumed in a secondary way, whereas Micropterus salmoides, Cobitis vettonica, Squalius alburnoides and Cyprinus carpio are accidental in the diet. Among the fish, the Cyprinids are the most consumed in the middle and lower zones, whereas in the high stretches it is the trout. In the zones where trout and small Cyprinids coexist, the otter preys more on the Cyprinids. Lutra lutra does not seem to be selective with regard to the fish species on which it preys, but rather on the size of the prey and prefers, on occasions, to capture fish of small or medium size to large ones, thus exploiting in an intensive manner the zones of deep water more than open water. After fish, the more important prey is the American red crayfish (Procambarus clarkii), followed by amphibians and reptiles, mainly Rana iberica and Rana perezi during the summer, and the viperine snake (Natrix maura) in the autumn. Birds and mammals comprise a small contribution to the diet, though the occasional captures of water voles (Arvicola sapidus) and the Iberian desman (Galemys pyrenaicus) are qualitatively noteworthy. With regard to diversity in the diet, it is greater on the high stretches of the river than along the lower zones, as in the summer period in contrast to the winter, in accordance with the availability in the environment.

Polymorphism of major ribosomal gene chromosomal sites (NOR-phenotypes) in the hybridogenetic fish Squalius alburnoides complex (Cyprinidae) assessed through crossing experiments

Chromosomal locations of major ribosomal sites, i.e. NOR-phenotypes, were assigned in Squalius alburnoides complex using sequential chromomycin A3 (CMA3)- and silver (Ag)-staining. This hybridogenetic Iberian minnow comprises diploid, triploid and tetraploid forms that arose by interspecific hybridisation between S. pyrenaicus and an unknown species. Inheritance of NOR patterns was studied by means of crossing experiments involving most diploid-polyploid forms of the S. alburnoides complex with identified specific genotype constitution. In all the specimens studied, the NORs were localised in the short arms of submetacentric chromosomes. Although S. pyrenaicus presented only one pair of NOR-bearing chromosomes, the data from experimental crosses evidenced that S. alburnoides complex was characterised by a multiple NOR phenotype composed of one chromosome pair with stable NORs and two chromosome pairs with NOR site polymorphism of presence/absence type. These data suggest that the karyotype of the unknown parental species of the S. alburnoides complex should have a multiple NOR pattern and emphasised the role of the all-male diploid linage in the dynamics and evolutionary potential of the S. alburnoides complex allowing the preservation of the missing ancestor genome. Cross-analyses evidenced that in spite of the high polymorphic nature of NORs in this fish complex, we have no reason to reject the hypothesis that their inheritance patterns were in accordance with Mendelian segregation.

Growth and reproductive traits of diploid and triploid forms of the Squalius alburnoides cyprinid complex in a tributary of the Guadiana River, Portugal

The Squalius alburnoides hybrid complex has diploid, triploid and some rare tetraploid male and female forms, with triploid females predominating in most populations. The reproductive traits of diploids and triploids from a population in a tri- butary of the Guadiana River, Portugal, were compared, during a two-years period. Few differences were found in the growth and reproductive traits of diploid and trip- loid females. Marginal differences were found in the longevity, with a few triploid females living up to six years compared with a maximum of five years for diploid females and a maximum of four years for diploid males. Both diploid and triploid females exhibited rapid growth in the first two years of life followed by rapid drop off linked to attaining sexual maturity. Reproductive effort increased with age but was mainly linked to number of eggs produced and not to egg size, since oocyte size did not vary with age or length of fish in either ploidy forms. Both forms exhibited a pro- tracted spawning period between March and June, suggesting multiple-spawning be- haviour. Triploid females had a slightly smaller oocyte diameter that may be due to the production of reduced (haploid) oocytes while diploid females produced diploid (unre- duced) oocytes The potentially higher production of diploid oocytes (which become triploid eggs after fertilisation) by diploid females could at least partially account for the higher percentage of triploids in natural populations. In addition, the production of bigger (diploid) oocytes, with possibly higher energetic content could lead to bigger fish larvae, which tend to have a higher survival rate. The time of spawning of both diploids and triploids appears to be synchronised, although differences were found in

The genetics of maintenance of an all‐male lineage in the Squalius albuvnoides complex

Squalius alburnoides is a complex of minnows common to the Iberian Peninsula, consisting of two distinct forms. The most common form is comprised of diploid and polyploid asexual hybrids heterozygous for several diagnostic allozyme loci contributed by Squalius pyrenaicus or Squalius carolitertii and a missing ancestor. The other form is diploid and homozygous for the allele contributed by the missing ancestor at these same loci. Present results from analyses of sex ratio and cytochrome b sequences are not consistent with the evolutionary distinctiveness of this non‐hybrid form and suggest that it represents an all‐male lineage imbedded within an almost all‐female complex. This all‐male lineage allowed preservation of the ancestral paternal nuclear genome after the paternal ancestor became extinct in all or most drainages, withimportant evolutionary implications.

The genetics of maintenance of an all-male lineage in the Squalius alburnoides complex

The genetics of maintenance of an all-male lineage in the Squalius alburnoides complex