Mitochondrial phylogeny and biogeographic affinities of sea breams of the genus Diplodus (Sparidae)

Phylogenetic analyses, using 482 bp of the mitochondrial 16S rDNA and 461 bp of the control region of 16 Diplodus species and Oblada melanura, Pagellus bogaraveo and Pagellus acarne , all close relatives of Diplodus , identified the two representatives of Pagellus as the sister group of Diplodus. Oblada melanura was confirmed as the sister taxon of D. puntazzo , despite its different dental morphology and ecology. Within the genus Diplodus , three clades were identified, the first containing D. annularis and D. bellottii , the second D. vulgaris and D. prayensis , and the third comprising three subclades. These were formed by O. melanura clustering with D. puntazzo, D. fasciatus with D. cervinus , and by the Diplodus sargus sub-species assemblage which also included the West Atlantic taxa D. argenteus, D. bermudensis, D. holbrooki , and the Red Sea endemic D. noct. All members of the D. sargus assemblage were genetically closely related. Among them, D. sargus lineatus from the Cape Verde islands was resolved as most ancestral branch, pointing to the possibility that the diversification and spread of the D. sargus assemblage originated in this region. The hypothesis of stepwise speciation following colonization events within the D. sargus complex is fully supported by phylogenetic reconstruction.     

Relationship between shape and trophic ecology of selected species of Sparids of the Caprolace coastal lagoon (Central Tyrrhenian sea)

Synopsis The shapes of juveniles of nine species of the family Sparidae (Diplodus annularis, Diplodus puntazzo, Diplodus sargus, Diplodus vulgaris, Lithognathus mormyrus, Pagellus acarne, Sarpa salpa, Sparus aurata, Spondyliosoma cantharus), collected in the tide channel of the Caprolace Lagoon (Central Tyrrhenian Sea—Italy) were studied using geometric morphometry. These species have different ecologies. The trophic ecology of each species, reported as TROPH values, are related to shape. For the first time the relationship between shape and trophic ecology in sparids was studied in a quantitative way giving an ecomorphological meaning to the shape differences. Mean shapes of carnivorous, omnivorous and herbivorous fish were extracted, analyzed and found to be unique. Strict herbivores such as Sarpa salpa have a small mouth gap; omnivores such as the four species of the genus Diplodus have a higher body (discoidal) and a caudal peduncle shorter and higher; carnivorous species such as Lithognathus mormyrus, possess a relatively larger head region, a larger mouth gap, a longer body and a caudal peduncle longer and narrower.     

Conventional karyotype, nucleolar organizer regions and genome size in five Mediterranean species of Syngnathidae (Pisces, Syngnathiformes)

Conventional karyotypes, NOR-bearing chromosomes by means of silver impregnation and genome size were investigated in five Mediterranean species in three genera of the Syngnathidae. A karyotype of 48 subtelocentric-acrocentric chromosomes was found in the seahorse Hippocampus hippocampus (FN=48) while a diploid value of 44 occurred in H. guttulatus (2 sm-m+42 a; FN=46) and the pipefish Syngnathus abaster (44 a; FN=44) and S. typhle (44 a; FN=44). The pipefish Nerophis ophidion , possessing a diploid chromosomal set of 58 made up of 50 meta-submetacentric and eight subteloacrocentric elements (FN=108) and a genome size three to four times larger than those known to date, differs cytogenetically from all other Syngnathids studied so far. A single pair of active NOR-bearing chromosomes was found in both species of the genus Hippocampus while in Syngnathus and Nerophis species more than two silver positive chromosomes were found to be involved in nucleolus organization giving rise to NOR-bearing chromosome polymorphism. The possible evolutionary routes of quantitative and qualitative changes in chromosome and DNA are discussed. The resulting phylogenetic scheme is shown to coincide with that constructed from morphological characters.     

Nucleolus organizer activity and the origin of Robertsonian translocations.

Chromosomes with active nucleolus organizer regions (NOR's) were identified by combined Q-banding (in some cases), and silver staining in mouse cell lines. NOR-bearing chromosomes were overrepresented among the chromosomes involved in Robertsonian translocations in LM(TK-), A9, and RAG cell lines. Usually only one NOR-bearing chromosome was seen in any biarmed chromosome; relatively few contained two NOR-bearing chromosomes. Thus the nucleolus plays an important role, but nucleolar fusion is relatively unimportant, in the origin of Robertsonian translocations in the mouse.     

Shape changes and growth trajectories in the early stages of three species of the genus Diplodus (Perciformes, Sparidae)

The larvae of three species of the genus Diplodus (Diplodus vulgaris, D. sargus, and D. puntazzo) colonize shallow waters along the Mediterranean coasts and, after a short period spent in the water column, they settle. For all three species this habitat transition is characterized by important shape changes mostly related to swimming capacity and feeding behavior. In this study, geometric morphometrics are used to characterize shape changes during the early juvenile life of specimens collected in a single locality in order to compare growth curves and allometric relationships. Size-related shape changes proved to be similar for all three species and are consistent with the ecological transition. A nonparametric smoothing technique (Loess) was used to fit the scatter of shape on size. The graphical representation (of most size-related shape variability) of this fitting technique shows how major shape changes are rapid for small sizes and slow down successively. The approach allows for the visualization of allometry and the fitting technique might help in defining the allometric growth pattern, thus contributing to the study of the autoecology of the species and in establishing terms for comparison with other ecologically or phylogenetically related species.     

Description of three species of Holorchis Stossich, 1901 (Digenea: Lepocreadiidae) from marine fishes off Corsica

Three species of Holorchis are redescribed from marine fishes from off Corsica. They are: Holorchis pycnoporus from Diplodus vulgaris, D. sargus, Lithognathus mormyrus, Pagellus erythrinus, Sparus pagrus and Symphodus roissali; H. micracanthum from Sparus pagrus and Pagellus erythrinus; and H. legendrei from Mullus surmuletus. The characteristics differentiating these species are discussed, in particular those separating H. pycnoporus and H. legendrei, species which previously have been considered synonyms. The internal morphology of the cirrus-sac differs distinctly between these forms, which also have microhabitat and host differences.     

Chromosome banding studies in an Indian mullet: evidence of structural rearrangements from NOR locations

C-, G- and NOR bands have been studied in the female sex of Rhinomugil corsula. (Mugilidae, Pisces) by deploying the conventional methodologies with suitable modifications of minor nature. The diploid metaphase complements contained 48 acrocentric chromosomes. The localization of C-band heterochromatin was found to be mostly at or near the centromeric regions of the acrocentric chromosomes. The G-type bands were not so well defined, but some of the G-banded chromosomes also contained C-bands. Interestingly, silver-positive NORs were found at the telomeric ends of five acrocentric chromosomes, including one homologous pair having NORs in both chromatids, while one chromosome showed NORs in both of its chromatids and the other two had only one NOR localized at one of its chromatids. This would suggest that one homologue of the second pair of NOR-bearing chromosomes possibly underwent a chromatid exchange with a non-NOR bearing chromosome. This is quite a unique situation not reported earlier in any species of fish., though some other form of NOR-polymorphism/heteromorphism has rarely been reported. Therefore, further exploration in natural populations of this species to examine the other sex and to verify if there also exists other chromosomally polymorphic races (in respect of NOR-polymorphism) of this species, would be rewarding.     

Karyotypes of 5 species of Soricidae-Crocidura in the fauna of the USSR

The chromosomes of five Crocidura species have been studied by means of routine, G-, C- and Ag NOR-banding techniques. The karyotypes of the species with 2n = 40, i.e. C. gueldenstaedtii, C. suaveolens and C. sibirica, are identical. Short arms of the Y-chromosome of C. sibirica are larger. The karyotypes of C. leucodon (2n = 28) and C. zarudnyi (2n = 22) could be obtained as a result of tandem and centric fusion and pericentric inversions. Small amounts of heterochromatin were detected in autosomes of the species investigated. The Y-chromosomes are completely heterochromatic. Four NOR-bearing pairs are detected in the species investigated.     

Constitutive heterochromatin,G-bands and nucleolus-organizer regions in four species of Didelphidae (Marsupialia)

Chromosomes of Didelphis albiventris, D. marsupialis, Philander opossum and Lutreolina crassicaudata, four species of marsupials with very similar karyotypes and 2n=22 were studied. All the chromosomes were acrocentrics except the X in L. crassicaudata, which is a metacentric.The G-band patterns of these species are similar but the distribution of constitutive heterochromatin differs among them as shown by C-banding. The hypothesis that the X in L. crassicaudata might be an isochromosome derived from the acrocentric X in the other species is discarded since G-and C-banding patterns differ in the two arms.In D. marsupialis the Ag-NORs are terminal and located in both arms of one pair and in the long arms of two pairs of medium-sized autosomes. In P. opossum the NOR-bearing chromosomes could be precisely identified through simultaneous silver staining and G-banding. The Ag-NORs are terminal and located at the short arm of pair 5 and the long arm of pair 7.     

Comparative chromosomal studies on two minnow fish, Phoxinus phoxinus (Linnaeus, 1758) and Eupallasella perenurus (Pallas, 1814); an associated cytogenetic-taxonomic considerations

The present work provides new data on the banding pattern of two cyprinid fish species Phoxinus phoxinus and Eupallasella perenurus from Poland. C-banding, silver-staining (Ag), and fluorescent staining with chromomycin A₃ techniques were used to describe the karyotypes. Both of the species karyotypes of 2n=50 were characterised by one pair of acrocentric chromosomes, the largest in the set, and by two pairs of NOR-bearing chromosomes. In the chromosome set of Ph. phoxinus Ag-stained NORs were located on telomeres of two metacentric and two submetacentric chromosomes, but in most metaphases only one of the two homologous was observed. The karyotype of E. perenurus was characterised by Ag-NOR regions at a telomeric position on the shorter arm of two submetacentric chromosome pairs. In most metaphases only three NOR-bearing chromosomes were observed. In both investigated species the location of the A₃ positive signals corresponded with the location of Ag-stained NORs and these sites were associated with heterochromatin shown as C-bands. The results of cytogenetical studies on other related, mainly the North American phoxinins, species are compared and discussed.     

Studies on the chromosomes of genusNycticebus

The karyotypes of three species (N. coucang, N. intermedius, andN. pygmaeus) of genusNycticebus, collected from the southern Yunnan of China, have been studied. All individuals from three species possess 2n=50 chromosomes, and all chromosomes in their complement are biarm chromosomes. The karyotype of slow loris (N. coucang) is characterized by having a secondary constriction and Ag-NORs in the short arms of pair No. 1. The G-banding patterns of three species are very similar. Three species are found to have multiple Ag-NORs. InN. coucang, NORs were observed on five pairs (Nos. 1, 6, 9, 15, and 23) and inN. intermedius andN. pygmaeus, NORs were found on four pairs (Nos. 6, 9, 15, and 20). This finding indicates that slow lorises, as primitive primates, also have multiple NOR-bearing chromosomes. Finally, the classification of genusNycticebus by karyotype analysis is discussed, and our results suggest that there are at least two valid species, namely:N. coucang andN. pygmaeus.     

The chromosomes of Nycticebus coucang (Boddaert, 1785) (Primates: Prosimii)

The high-quality karyotype of a specimen of Nycticebus coucang is described and illustrated. The X chromosome is found to be indistinguishable from that of the greater galagos, and may represent a synapomorphic trait. The Y chromosome is a medium to small submetacentric (3.2% TCL) and constitutes one of the larger Y chromosomes known in primates. N. coucang is found to have multiple NOR-bearing chromosomes in contrast to the single pair found in galagine and catarrhine monkeys. Since a single NOR-bearing pair is often considered ancestral for primates, this new finding may have important implications for the evolution of these cistrons. One of the chromosomal polymorphisms in this specimen is a pericentric inversion, involving a NOR-bearing autosomal pair (no. 6), that alters the position of the active site. Further, homologues 2p differ by aparacentric inversion. These results confirm that lorisiforms are characterized by considerable chromosomal polymorphism.     

Description de deux nouvelles espèces de Lamellodiscus Johnston & Tiegs, 1922 (Monogenea: Diplectanidae) du groupe morphologique ‘ignoratus’, parasites de Diplodus sargus et D. vulgaris (Teleostei: Sparidae)

The study of the diplectanid gill parasites of Diplodus Rafinesque (Teleostei: Sparidae) from off the Algerian coast revealed the presence of several species of Lamellodiscus Johnston & Tiegs, 1922 on D. sargus (Linnaeus) and D. vulgaris (Geoffroy Saint-Hilaire). Some of these species differ from the others by having a copulatory organ "en lyre" and lamellodiscs of the morphological group 'ignoratus' of Oliver (1987). In Lamellodiscus, the subgroup 'ignoratus', proposed by Amine & Euzet (2005), characterised by a simple lateral dorsal bar, includes L. ignoratus Palombi, 1943 and L. fraternus Bychowsky, 1957. Two new species, L. falcus n. sp. and L. neifari n. sp. form part of the 'ignoratus' subgroup. These species are distinguished by the morphology and the size of the sclerotised parts of the haptor and copulatory organ.     

Cytogenetic analyses of two Curimatidae species (Pisces; Characiformes) from the Paranapanema and Tietê Rivers.

Cytogenetic analyses were performed in two Curimatidae species (Steindachnerina insculpta and Cyphocharax modesta) from the Paranapanema and Tietê Rivers (S?o Paulo State, Brazil), showing a karyotype composed of 54 meta-submetacentric chromosomes in both species. Silver- and chromomycyn-staining and fluorescent in situ hybridization (FISH) using a 18S rDNA probe indicated that the nucleolar organizer regions (NORs) of both species are localized in the terminal region of the long arm of two metacentric chromosomes. Although a single NOR system was evidenced in both analyzed species, S. insculpta and C. modesta presented the nucleolar organizer regions in distinct chromosome pairs, indicating that these cistrons can be considered cytogenetic markers. Variation on the amount and distribution of the constitutive heterochromatin (C-bands) could also be detected between the two species - while S. insculpta presented few heterochromatic blocks, intensely stained C-bands were evidenced in C. modesta specially in the terminal region of the long arm of the NOR-bearing chromosomes. Although most Curimatidae species have been characterized by homogeneous karyotypes, isolated populations could be established under different environmental conditions leading to karyotype micro-structure variations specially related to the NORs localization and C-banding distribution. The obtained data were useful for the cytogenetic characterization and differentiation of S. insculpta and C. modesta and could be used in evolutionary inferences in the Curimatidae group.     

Identification of Mediterranean Diplodus spp. and Dentex dentex (Sparidae) by means of DNA Inter-Simple Sequence Repeat (ISSR) markers

The aim of this paper was to test the effectiveness of Inter-Simple Sequence Repeat (ISSR) technique in detecting specific molecular markers to genetically discriminate some species of teleost fish. As model species, we used four Mediterranean sea bream species (i.e. Diplodus annularis, D. puntazzo, D. sargus sargus, and D. vulgaris), and the common Mediterranean dentex Dentex dentex. Furthermore, the occurrence in our samples of two partially albino sparids (suspected, on a morphological basis, to probably belong to D. sargus sargus and to D. dentex, respectively) allowed ISSRs to be tested on doubtful specimens. Eight ISSR primers produced 95 out of 97 scorable polymorphic loci with a large number (63) of species-private bands. Maximum parsimony analysis, ΔK statistics, and assignment test were consistent in separating the five species and solving the taxonomic position of specimens of dubious attribution. The results obtained evidence the usefulness of ISSR markers in specimen and species identification, particularly when the morphological traits of fish make the classification problematic.     

H-Y antigen as a tool for the determination of the heterogametic sex in Amphibia

H-Y antigen was investigated in three amphibian species with different degrees of sex-chromosome differentiation: Bufo bufo, Triturus vulgaris, and Pyxicephalus adspersus. No heteromorphic sex chromosomes were found in B. bufo, but an examination of the progeny of hermaphrodites (Ponse, 1942) indicated that the female of this species was heterogametic (ZW). Sex chromosomes differing only by a very small heterochromatic region at their telomeres were found in the male of T. vulgaris (XY). Pyxicephalus adspersus revealed high differentiated ZW sex chromosomes. The results of the H-Y antigen studies on these three species indicate that H-Y antigen is expressed only in the heterogametic sex, irrespective of differences in morphological differentiation of the sex chromosomes. Therefore, H-Y antigen could be a valuable tool in determining the heterogametic sex, not only in Amphibia but possibly also in other vertebrate species that have either evolved no heteromorphic sex chromosomes or where sex-reversal experiments are not possible.     

NORs inheritance analysis in crossings including individuals from two stocks of rainbow trout (Oncorhynchus mykiss)

Silver nitrate staining of rainbow trouts (Oncorhynchus mykiss) chromosomes, for the identification of the nucleolar organizing regions (NORs), revealed that in individuals from Núcleo Experimental de Salmonicultura de Campos do Jord?o (Brazil) NORs were located in the long arms of submetacentric pair while in specimens from Mount Shasta (USA) NORs were located in the short arms of a submetacentric pair. Cytogenetic analysis of the offspring, obtained through artificial crosses including individuals from both stocks, allowed the identification of NORs in two submetacentric chromosomes, one in the short arms and the other in the long arms, confirming the effectiveness of the hybridization process. Complementary results obtained using the FISH technique with 18S and 5S rDNA probes showed that NOR-bearing chromosomes exhibited a cluster of 5S genes located in tandem with the 18S gene cluster in both stocks. The results allow us to suggest that the difference in NOR-bearing chromosomes found between the two stocks is likely to be due to pericentric inversion involving the chromosome segment where 18S and 5S rDNA genes are located. The presence of ribosomal genes in the long arms of a submetacentric chromosome is apparently a particular characteristic of the rainbow trout stock of Campos do Jord?o and might be used as a chromosome marker in studies of controlled crosses in this species.     

Microhabitat requirements for settlement of juvenile sparid fishes on Mediterranean rocky shores

Microhabitat attributes were characterized for recently settled juvenile fishes in six species of Sparidae (Diplodus annularis, D. puntazzo, D. sargus, D. vulgaris, Oblada melanura, and Sarpa salpa) on the rocky shore near Marseille, French Mediterranean coast. Depth, slope, type of substratum, biotic cover and hydrodynamic conditions were recorded wherever sparid recruits occurred. Juvenile Diplodus annularis settled in Posidonia oceanica seagrass beds at 5–8 m deep. The five other sparid species recruited in very shallow water (<2 m). D. puntazzo, D. sargus and D. vulgaris were observed in sites presenting a gentle slope with coarse sand, gravel, pebbles or boulders. Sarpa salpa was less dependent on slope for recruitment, but was always associated with microalgae. Oblada melanura settlement occurred on rocky areas with various slopes, and was favoured by the presence of overhangs. When growing, juvenile sparids extended their home range vertically into deeper zones, and laterally in more exposed areas. Time partitioning in the use of suitable microhabitats for recruitment was observed for some species. Juveniles of Diplodus puntazzo and D. vulgaris shared the same sites from March to May, both exhibiting relatively low abundances. D. sargus recruited in far higher abundance in the same sites from May to September. Specific improvements in coastal man-made structures (harbours, artificial beaches) are suggested in order to increase the recruitment of some littoral fish species.

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Résumé Les caractéristiques des microhabitats des jeunes recrues ont été déterminées pour six espèces de Sparidae (Diplodus annularis, D. puntazzo, D. sargus, D. vulgaris, Oblada melanura, et Sarpa salpa) sur les côtes rocheuses prés de Marseille, en Méditerranée. La profondeur, la pente, le type de substrat, la couverture biotique et les conditions hydrodynamiques ont été relevés dans tous les sites où des juvéniles de Sparidae ont été observés. Les juvéniles de Diplodus annularis recrutent dans les herbiers à Posidonia oceanica vers 5–8 m de profondeur. Les cinq autres espéces de Sparidae recrutent dans les petits fonds inférieurs à 2 m de profondeur. D. puntazzo, D. sargus and D. vulgaris ont été observés dans des zones calmes en pente douce avec des fonds de sable grossier, de gravier, de galets ou de petits blocs. Les jeunes de Sarpa salpa dépendent moins de la pente du substrat pour leur recrutement, mais sont toujours associés à des peuplements de macroalgues. Les juvéniles d'Oblada melanura recrutent dans des zones rocheuses de diverses pentes et sont favorisés par la présence de surplombs. Au cours de leur croissance, les jeunes Sparidae élargissent leur domaine vital verticalement en profondeur et latéralement vers des zones plus agitées. Une partition temporelle de l'utilisation des microhabitats favorables au recrutement a été observée chez certaines espèces. Les juvéniles de Diplodus puntazzo et D. vulgaris partagent les mêmes sites de mars à mai, les deux espèces présentant des densités de populations relativement faibles. D. sargus recrute en beaucoup plus grand nombre dans les mêmes sites entre mai et septembre. Cette étude a permis de suggérer des aménagements spécifiques des zones littorales artificielles pour accroître le recrutement des juvéniles de certaines espèces de poissons.

     

Validation of daily increment formation in otoliths for three Diplodus species in the Mediterranean sea

The efficacy of two fluorochromes, chlorhydrate of tetracycline (CHTC) and alizarin complexone (ALC), to induce a label on the otoliths of juvenile sparid fishes by immersion techniques was tested in different experimental conditions. CHTC did not mark otoliths in natural sea water and was toxic in divalent cation-free sea water. Immersing fish in a 50-mg 1⁻¹ ALC natural seawater solution for 24 h induced a well-defined mark on the otoliths and had no effect on survival. A daily periodicity of increment formation on otoliths was observed in captivity for Diplodus vulgaris and D. puntazzo , and was conserved in natural environment for D. sargus . The frequency of increment deposition did not vary with the age of juvenile fishes. Thus, otolith microstructure analysis will be a reliable method to give age estimates in these three sparid species during their juvenile life.