Museum samples could help to reconstruct the original distribution of Salmo trutta complex in Italy.

Partial D-loop sequences of museum specimens of brown trout and marble trout (Salmo trutta species complex) collected from Mediterranean rivers in the late 19th century were analysed to help to describe the native distribution of these species. All the individuals studied carried native haplotypes, the geographic distribution of which is consistent with published data. These results indicate that museum specimens from the 19th century could represent an opportunity to get a picture of the original genetic diversity distribution of this species complex.

New insights into sex chromosome evolution in anole lizards (Reptilia, Dactyloidae).

Anoles are a clade of iguanian lizards that underwent an extensive radiation between 125 and 65 million years ago. Their karyotypes show wide variation in diploid number spanning from 26 (Anolis evermanni) to 44 (A. insolitus). This chromosomal variation involves their sex chromosomes, ranging from simple systems (XX/XY), with heterochromosomes represented by either micro- or macrochromosomes, to multiple systems (X1X1X2X2/X1X2Y). Here, for the first time, the homology relationships of sex chromosomes have been investigated in nine anole lizards at the whole chromosome level. Cross-species chromosome painting using sex chromosome paints from A. carolinensis, Ctenonotus pogus and Norops sagrei and gene mapping of X-linked genes demonstrated that the anole ancestral sex chromosome system constituted by microchromosomes is retained in all the species with the ancestral karyotype (2n = 36, 12 macro- and 24 microchromosomes). On the contrary, species with a derived karyotype, namely those belonging to genera Ctenonotus and Norops, show a series of rearrangements (fusions/fissions) involving autosomes/microchromosomes that led to the formation of their current sex chromosome systems. These results demonstrate that different autosomes were involved in translocations with sex chromosomes in closely related lineages of anole lizards and that several sequential microautosome/sex chromosome fusions lead to a remarkable increase in size of Norops sagrei sex chromosomes.

Karyological characterization of the endemic Iberian rock lizard, Iberolacerta monticola (Squamata, Lacertidae): insights into sex chromosome evolution.

Rock lizards of the genus Iberolacerta constitute a promising model to examine the process of sex chromosome evolution, as these closely related taxa exhibit remarkable diversity in the degree of sex chromosome differentiation with no clear phylogenetic segregation, ranging from cryptic to highly heteromorphic ZW chromosomes and even multiple chromosome systems (Z1Z1Z2Z2/Z1Z2W). To gain a deeper insight into the patterns of karyotype and sex chromosome evolution, we performed a cytogenetic analysis based on conventional staining, banding techniques and fluorescence in situ hybridization in the species I. monticola, for which previous cytogenetic investigations did not detect differentiated sex chromosomes. The karyotype is composed of 2n = 36 acrocentric chromosomes. NORs and the major ribosomal genes were located in the subtelomeric region of chromosome pair 6. Hybridization signals of the telomeric sequences (TTAGGG)n were visualized at the telomeres of all chromosomes and interstitially in 5 chromosome pairs. C-banding showed constitutive heterochromatin at the centromeres of all chromosomes, as well as clear pericentromeric and light telomeric C-bands in several chromosome pairs. These results highlight some chromosomal markers which can be useful to identify species-specific diagnostic characters, although they may not accurately reflect the phylogenetic relationships among the taxa. In addition, C-banding revealed the presence of a heteromorphic ZW sex chromosome pair, where W is smaller than Z and almost completely heterochromatic. This finding sheds light on sex chromosome evolution in the genus Iberolacerta and suggests that further comparative cytogenetic analyses are needed to understand the processes underlying the origin, differentiation and plasticity of sex chromosome systems in lacertid lizards.

Genome duplication in early vertebrates: insights from agnathan cytogenetics.

Agnathans represent a remnant of a primitive offshoot of the vertebrates, and the long evolutionary separation between their 2 living groups, namely hagfishes and lampreys, could explain profound biological differences, also in karyotypes and genome sizes. Here, cytogenetic studies available on these vertebrates were summarized and data discussed with reference to the recently demonstrated monophyly of this group and to the 2 events of whole genome duplication (1R and 2R) characterizing the evolution of vertebrates. The comparison of cytogenetic data and phylogenetic relationships among agnathans and gnathostomes seems to support the hypothesis that 1R and 2R occurred before the evolutionary divergence between jawless and jawed vertebrates.

Isolation of novel microsatellite loci in the black goby Gobius niger and cross-amplification in other gobiid species (Perciformes, Gobiidae).

Twelve microsatellite loci were isolated from and characterized for the black goby Gobius niger. These loci were tested on a total of 48 individuals from two geographically distant locations (Orbetello and Cattolica) and the number of alleles ranged from two to 18, with expected (H(e)) and observed (H(o)) heterozygosities ranging from 0.042 to 0.941 and from 0.042 to 0.917, respectively. The loci described were used to cross-amplify other gobiid species belonging to Gobius, Zosterisessor, Lesueurigobius and Aphia.

Characterisation of a GC-rich telomeric satellite DNA in Eumeces schneideri Daudin (Reptilia, Scincidae).

A hitherto undescribed satellite DNA family (AvaII satDNA) has been isolated and characterised in Eumeces schneideri, a squamate reptile belonging to the family Scincidae. AvaII satDNA is characterised by a monomer length of 208 bp, a GC content of 59% and exhibits a certain degree of CpG methylation. FISH experiments with AvaII satDNA probe produced bright signals (i) at the end of the short arms of all subtelocentric chromosomes except for pair 14, in which the signal was at the end of the long arms, (ii) at the ends of both arms of the small metacentric chromosomes 12, and (iii) in a terminal position on the acrocentric chromosomes 11 and 13. AvaII satDNA repeats were not found in the metacentric pair 3, whereas only a weak interstitial signal occurred in the metacentric pairs 1 and 2. C-banding showed that this satellite represents most of the constitutive heterochromatin in the genome of this skink, and chromomycin A(3) staining produced a clear signal overlapping with the satellite, except for NOR-associated heterochromatin. In addition, quantitative dot blot analysis showed that these repetitive sequences constitute about 3% of the genomic DNA of this lizard. AvaII satDNA sequence analysis revealed the occurrence of short guanine residue stretches for which a function in structural stability of these sequences and a role in recombination with telomeric sequences can be hypothesised. Fibre FISH experiments showed that on some chromatin fibres telomeric sequences and AvaII satellite DNA repeats are intermingled or overlapping.

Chromosomal study of native and hatchery trouts from Italy (Salmo trutta complex, Salmonidae): conventional and FISH analysis.

A cytogenetic analysis was carried out using conventional staining, banding techniques and fluorescence in situ hybridization (FISH) in Italian populations of brown trout (Salmo truttacomplex). All individuals analysed, belonging to the Atlantic (At), Marmoratus (Ma), Adriatic (Ad) and Mediterranean (Me) lineages, showed remarkable karyotype uniformity, with diploid complement of 2n = 80 chromosomes, arm number (NF) of 102 and invariable karyotype composition. Such uniformity was also observed with respect to the location of 5S rDNA and the active, i.e. silver-positive NOR sites. On the contrary, FISH with 28S ribosomal probe and fluorescent staining with CMA3 revealed that inactive NOR sites are more numerous in Ad and Me than in At and Ma lineages. A centromeric sequence was successfully isolated from Salmo trutta individuals by polymerase chain reaction (PCR)-based cloning, using primers designed from published Atlantic salmon (Salmo salar) satellite DNA sequences. This sequence had high AT content (65.3%) and short consensus motif (A/T)(G/C)AAA(T/C) similar to other centromeric satellite repeats. The isolated satellite DNA clones were localized with FISH in the centromeric regions of the brown trout chromosomes, showing lineage-specific patterns. Because it is well known that AT-rich sequences can induce a pronounced DNA curvature, which in turn would promote faster and higher chromatin spiralization, it may be hypothesised that the wide distribution of this satellite in the S. trutta genome may have played a role in its karyotype stability. The presence of this sequence in other salmonid species was also tested by Southern blot hybridization and used to analyze its evolution within salmonids.

Skinks (Reptilia: Scincidae) have highly conserved karyotypes as revealed by chromosome painting.

Skinks represent the most diversified squamate reptiles with a great variation in body size and form, and are found worldwide in a variety of habitats. Their remarkable diversification has been accompanied by only a few chromosome rearrangements, resulting in highly-conservative chromosomal complements of these lizards. In this study cross-species chromosome painting using Scincus scincus (2n = 32) as the source genome, was used to detect the chromosomal rearrangements and homologies between the following skinks: Chalcides chalcides (2n = 28), C. ocellatus (2n = 28), Eumeces schneideri (2n = 32), Lepidothyris fernandi (2n = 30), Mabuya quinquetaeniata (2n = 32). The results of this study confirmed a high degree of chromosome conservation between these species. The main rearrangements in the studied skinks involve chromosomes 3, 5, 6 and 7 of S. scincus. These subtelocentric chromosomes are homologous to the p and q arms of metacentric pair 3 and 4 in C. chalcides, C. ocellatus, L. fernandi, and M. quinquetaeniata, while they are entirely conserved in E. schneideri. Other rearrangements involve S. scincus 11 in L. fernandi and M. quinquetaeniata, supporting the monophyly of Lygosominae, and one of the chromosomes S. scincus 12-16, in M. quinquetaeniata. In conclusion, our data support the monophyly of Scincidae and confirm that Scincus-Eumeces plus Chalcides do not form a monophyletic clade, suggesting that the Scincus-Eumeces clade is basal to other members of this family. This study represents the first time the whole genome of any reptile species has been used for cross-species chromosome painting to assess chromosomal evolution in this group of vertebrates.

Mitochondrial DNA variation of an isolated population of the Adriatic brook lamprey Lampetra zanandreai (Agnatha: Petromyzontidae): phylogeographic and phylogenetic inferences.

Two mitochondrial genes were examined to compare an isolated population of the Adriatic brook lamprey Lampetra zanandreai in central Italy with other populations in the species range (Po plain) and with parasitic and freshwater lampreys. A single haplotype, identical to one in a Venetian sample, was found in 10 individuals from the isolated population. The reduced variability is consistent with a history of dispersal after the Pleistocene expansion of the Po basin. The results support the hypothesis of an origin of L. zanandreai and L. fluviatilis-L. planeri from a common anadromous ancestor.

Chromosome banding and molecular cytogenetic study of two Mediterranean trachinoid fish species (Teleostei: Trachinidae, Uranoscopidae).

The chromosomes of Echiichthys vipera (Trachinidae) and Uranoscopus scaber (Uranoscopidae) were analyzed by means of various banding methods and fluorescence in situ hybridization (FISH) with telomeric and major rDNA probes, respectively. The karyotype of E. vipera was composed of 48 acrocentric chromosomes and NOR sites, as revealed by all detection methods, were situated pericentromerically on a single pair of middle-sized chromosomes. Blocks of constitutive heterochromatin were present in the pericentromeric regions of all pairs of chromosomes. The karyotype of U. scaber showed three karyomorphs: 2n = 30 (18 m + 12 a/st [m = metacentric, a = acrocentric and st = subtelocentric]), 2n = 28 (20 m + 8 a/st), 2n = 27 (21 m + 6 a/st). NORs, as revealed by FISH, were situated pericentromerically on a single pair of middle-sized chromosomes in spite of Ag-positive signals in the centromeres of all pairs of chromosomes. Robertsonian fusions were hypothesized for observed variation due to invariable number of chromosome arms FN = 48.