Two dispersed highly repeated DNA families of Triturus vulgaris meridionalis (Amphibia,Urodela) are widely conserved among Salamandridae

Two BamHI families of repeated sequences were characterized from the genome of the Italian smooth newt, Triturus vulgaris meridionalis (Amphibia, Urodela). The first family, which is divided into subfamilies, consists of tandemly arranged arrays whose basic repeat is around 398 bp long; these arrays are dispersed throughout the entire chromosome sets of the various species of Triturus tested. Moreover the family is widely conserved among Salamandridae, being detected by genomic DNA blotting of Notophthalmus viridescens, Taricha granulosa, Salamandrina terdigitata and Euproctus platycephalus. The second BamHI family is represented by a cloned sequence of 419 bp, which is dispersed in the chromosome set of several species of Triturus. The sequence is also conserved in S. terdigitata and in E. platycephalus but is not detectable in N. viridescens or T. granulosa. The cloned sequence is most probably only part of a longer unit interspersed within the Triturus genome.by H.C. Macgregor     

Priority effects of the early breeding fire salamander on the late breeding banded newt

Early breeding intraguild predators may have advantages over late breeding predators via priority effects; early breeding predators may reduce shared prey resources before late breeders appear and may also prey upon the late breeders. Here we show that predatory larvae of the late-breeding predatory banded newt, Triturus vittatus vittatus, occupy the same temporary pond toward the end of the developmental period of the early-breeding predatory fire salamander, Salamandra salamandra, resulting in a large size disparity between larvae of these two species while they co-occur. We conducted outdoor artificial pool experiments to assess priority effects of large larval Salamandra at the end of their larval development period, on recently hatched larval Triturus. We also assessed how artificial vegetation may influence larval Triturus performance in the presence or absence of Salamandra Salamandra, introduced into the experimental pools two weeks prior to the newt larvae, strongly reduced invertebrate prey abundance shared by these two predatory urodeles and with only a one week period of overlap, strongly reduced abundance of Triturus larvae. The artificial vegetation had only a small ameliorating effect on Triturus survival when Salamandra was present. Triturus size at metamorphosis (snout-tail length) was significantly larger in the Salamandra pools, presumably due to a combination of a strong “thinning effect” and greater vulnerability of smaller Triturus individuals to predation by Salamandra. Time to metamorphosis was not significantly affected by Salamandra. These results have conservation implications as T. v. vittatus is listed as highly endangered and may also explain the largely negative spatial association of the two species. Handling editor: K. Martens     

The maps of the lampbrush chromosomes of Triturus (Amphibia urodela)

The lampbrush chromosomes of Triturus vulgaris meridionalis were isolated from the germinal vesicle of medium and large-sized oocytes and studied with phase-contrast microscope. The maps were constructed on the basis of the lengths and major morphological features of the chromosomes. The length of each map is equal to the mean of the relative lengths of the corresponding chromosome from different oocytes (the relative length of each chromosome is represented by the ratio between its absolute length and that of chromosome XII from the same complement, conventionally considered as 100 units long). The maps arranged in decreasing length order, were oriented according to the most frequent position of chiasmata, as centromeres were not always evident. — Chromosomes VI and XI bear a sphere in subterminal position. Landmarks typical for T. vulgaris meridionalis are the loops inserted on chromosomes VIII (47 units), X (23 units), XI (34 units) and XII (34 units) frequently presenting themselves under the form of double loop bridges of considerable extension. On chromosomes I (4 units), VI (13 units), X (4 units) and XI (36 units) giant bodies were found that are sometimes comparable to dense-matrix loops. Chromosome XI includes a nucleolus-organizing region, sometimes identifiable by the presence of an inserted nucleolus. Normal and granular loops (much extended at times), axial granules, globules, and loopless bars supplement the morphology of the lampbrush chromosomes of this species.     

Evolution of highly repeated DNA within the genusTriturus (Amphibia,Urodela)

Highly repeated DNA is a main feature of urodele amphibian genomes. InTriturus this class of DNA consists of several sequence families differently arranged at both the molecular and the chromosomal level, showing varying degrees of conservation across species. Present data on highly repeated DNA inTriturus are here summarized and discussed with regard to the evolution and possible functional role of these sequences.     

Expression of amino acid transporter genes in developmental stages and adult tissues of Antarctic echinoderms

Epithelial cells in the body wall of adult and developmental stages of marine invertebrates absorb dissolved organic material directly from seawater. Despite over a century of study, little is known about the molecular biological mechanisms responsible for this transport process. Previous studies on embryonic and larval Antarctic echinoderms show that amino acid uptake could provide an important supplement of metabolic substrates. In the present study, partial cDNA sequences of 11 putative amino acid transporter genes were isolated from six species of Antarctic echinoderms including the Antarctic sea stars Acodontaster hodgsoni, Diplasterias brucei, Odontaster meridionalis, Odontaster validus, and Perknaster fuscus, and the Antarctic sea urchin Sterechinus neumayeri. Conserved domains of cDNA-deduced amino acid sequences characterized these genes as being members of a family of amino acid transporters (solute carrier family 6). Expression of these genes was detected throughout embryonic and larval development of two species that have contrasting developmental modes (A. hodgsoni: lecithotrophic; O. meridionalis: planktotrophic). In all six species studied, the expression of amino acid transporter genes was detected in tube feet and digestive organs of adult animals, demonstrating that members of a single amino acid transporter gene family are expressed during the entire life history of a marine invertebrate. The identification of these genes is an important step toward developing a mechanistic understanding of amino acid transport capacities in Antarctic marine invertebrates.     

Chromosome polymorphism in the Italian newt,Triturus italicus

A chromosomal variation, changing shape and C-banding pattern of chromosome XII of Triturus italicus was detected among the offspring of two F₁ hybrid families of T. italicus × T. vulgaris meridionalis . In both families a number of individuals appeared to have a metacentric instead of the expected subtelocentric chromosome XII of T. italicus. — Investigations in three well separated localities in the range of the species showed the polymorphism to have a wide distribution and to be part of a complex pattern involving at least two inversions and (presumably) deficiencies of large C-bands. At meiosis, the shape of bivalent XII, and the location and frequency of chiasmata in the bivalent varied with the karyomorph involved. It is suggested that large rearrangements may still play an important role in the karyological evolution of Triturus.     

Molecular and chromosomal organization of two repetitive DNA sequences with intercalary locations in sugar beet and other Beta species

 In a search for repetitive DNA sequences in the sugar beet genome, two sequences with repeat unit lengths of 143 and 434 bp were isolated and characterized. The pSV family showed an unusual conservation of restriction sites reflecting homogenization of the analyzed repeats. Members of the family are organized as tandem repeats as revealed by PCR and sequencing of dimeric units. The pSV satellite occurs in large intercalary arrays which are present on all chromosome arms of sugar beet. The pSV sequence family is present in different abundance in the sections Beta, Corollinae and Nanae but is not detectable by Southern hybridization in the section Procumbentes. The pDRV family is characterized by an interspersed genomic organization. The sequence is detectable in all sections of the genus and is amplified in species of the section Beta but was also detected, although at lower abundance, in the remaining three sections. Fluorescent in situ hybridization has shown that the pDRV sequence family is dispersed over all chromosomes of the sugar beet complement with some regions of clustering and centromeric depletion. Received: 18 March 1998 / Accepted: 31 March 1998     

Characterisation of a short,highly repeated and centromerically localised DNA sequence in crested and marbled newts of the genus Triturus

A 32–33 bp highly repeated DNA sequence, TkS1, has been isolated from genomic DNA of the newt Triturus karelini digested with the restriction endonucleases HaeIII or AluI. TkS1 is known to be localised in the centromeric heterochromatin of all the chromosomes in T. karelini and the related species T. cristatus TkS1 has been shown to be present in varying amounts in the genomic DNA of a range of species of Triturus, including representatives of the two main subgenera Triturus and Palaeotriton. A programme of sequencing of monomers, dimers and trimers of TkS1 was carried out in order to determine the level of conservation of the sequence within and between species of Triturus. Altogether 204 monomer (32/33 bp) clones were made of TkS1 from three individuals of T. karelini, and one individual each of T. cristatus, T. carnifex, T. dobrogicus and T. marmoratus, all members of the subgenus Triturus and the cristatus species group. A number of dimer (64 bp) and trimer (96 bp) clones were also made from DNA of a single specimen of T. karelini digested with HaeIII or AluI. Three distinct types of TkS1 were identified in all species examined, except for T. marmoratus where only two of the types were found. The types were distinguished on the basis of certain recurring divergent patterns in monomers sequenced from T. karelini. Type 1 is mainly characterised by the presence of an AluI site at positions 24–27 and type 3 mainly by the presence of an additional base (C) at position 14. Type 2 normally lacks the AluI site and the C at position 14, as well as having a number of other distinguishing features. TkS1 and its three types have remained remarkably constant in sequence since before the divergence of T. marmoratus from other species in the cristatus species group, about 10 million years ago. Examination of all 204 monomer clones and comparison with consensus sequences for the three types shows less than 5% divergence at any one position in the sequence. There is good evidence from examination of dimer and trimer clones of TkS1 that the different types are intermingled with each other, and all three types are likely to be present on all chromosomes. Dimeric (64 bp) TkS1 clones constructed from AluI fragments of T. karelini DNA show evidence of a trimeric (96 bp) supertype with the pattern type 1-type 3-type 1 that is much more common than would be expected on a random basis. The properties of TkS1 are discussed in relation to models for the involvement of highly repeated satellite DNA in chromosomal growth and evolution.W. Hennig     

Cloning and characterization of a centromere-specific repetitive DNA element from Sorghum bicolor

 A 823-bp Sau3AI fragment (pSau3A10) was subcloned from a sorghum bacterial artificial chromosome (BAC) clone, 13I16, that contains DNA sequences specific to the centromeres of grass species. Sequence analysis showed that pSau3A10 consists of six copies of an approximately 137-bp monomer. The six monomers were organized into three dimers. The monomers within the dimers shared 62–72% homology and the dimers were 79–82% homologous with each other. Fluorescence in situ hybridization (FISH) analysis indicated that the Sau3A10 family is present only in the centromeres of sorghum chromosomes. Sequencing, Southern hybridization, and Fiber-FISH analyses indicated that the Sau3A10 family is tandemly arranged and is present in uninterrupted stretches of up to at least 81 kb of DNA. Slot-blot analysis estimated that the Sau3A10 family constitutes 1.6–1.9% of the sorghum genome. The long stretches of Sau3A10 sequences were interrupted by other centromeric DNA elements. Southern analysis indicated that the Sau3A10 sequence is one of the most abundant DNA families located in sorghum centromeres and is conserved only in closely related sorghum species. Methylation experiments indicated that the cytosine of the CG sites in sorghum centromeric regions is generally methylated. The structure and organization of the Sau3A10 family shared similarities with centromeric DNA repeats in other eukaryotic species. It is suggested that the Sau3A10 family is probably an important part of sorghum centromeres. Received: 11 November 1997 / Accepted: 17 November 1997     

Genomic organization of ribosomal DNA and related sequences inTriturus

The ribosomal RNA genes ofTriturus vulgaris meridionalis are clustered at variable and often multiple chromosomal loci. The rDNA repeats exhibit only a limited and discrete length heterogeneity which is accounted for by the non-transcribed spacer (NTS). Interestingly, sequences homologous to the NTS are clustered outside the ribosomal loci. Clones containing such non ribosomal sequences have been isolated from a genomic library ofT. v. meridionalis and analyzed by restriction mapping. These sequences appear to consist mostly of repetitive Bam HI fragments ranging from 500 bp to 1000 bp. The Bam HI fragments are internally repetitious and highly homologous to each other.     

Banding patterns in newt chromosomes by the giemsa stain

Specific banding patterns can be produced on the mitotic chromosomes of the newt species Triturus vulgaris meridionalis and T. italicus by using the Giemsa stain technique. These bands are most useful cytogenetic markers in karyotyping, since they facilitate identification of the individual elements of the complements. Evaluation of the shape of chromosomes as well as of the banding patterns produced by the Giemsa stain indicates that the karyotypes of T. vulgaris meridionalis and T. italicus are differentiated: hence the specific distinction of the two Salamandrids, still debated by taxonomists, appears supported by chromosome evidence. — Most of the bands seem to correspond to the heterochromatic tracts observable on mitotic chromosomes from embryos and larvae either untreated or submitted to cold treatment. Besides, the comparison of mitotic karyotypes and lampbrush maps shows that the bands located near the centromeric regions of mitotic chromosomes probably correspond to the so-called bars visible on either side of centromeres of lampbrush chromosomes, while some of the subterminal bands may correspond to the sphere.This work was financially supported by C. N. R., Roma.     

DNA content in nine species of Nematocera with special reference to the sibling species of the Anopheles maculipennis group and the Culex pipiens group

DNA values were determined for the nine species: Telmatoscopus meridionalis (family Psychodidae), Dixa obscura (family Dixidae), and Culiseta litorea, Culex pipiens, Aedes caspius, Anopheles labranchiae, Anopheles atroparvus, Anopheles stephensi, and Anopheles freeborni (family Culicidae). The DNA content indicated that the species could be divided into three categories: The Culex group with Culex pipiens, Culiseta litorea and Aedes caspius containing 1.02, 0.92 and 0.99 pg DNA in the haploid set, the Anopheles group including the four Anopheles species A. labranchiae, A. atroparvus, A. stephensi, A. freeborni and Telmatoscopus meridionalis with 0.23, 0.24, 0.24, 0.29 and 0.24 pg DNA and the third group with Dixa obscura containing 0.16 pg DNA. Within the sibling species of the Anopheles maculipennis group DNA differences were found to be present between the palaearctic and the nearctic populations. No deviations could be found between two Culex pipiens populations which were reproductively isolated.This work was supported by a grant of the Volkswagenstiftung (Germany) and by grant No. 70.1146.04 from C.N.R. (Italian Centre of Researches).     

Characterization of Charr Chromosomes Using Fluorescence in Situ Hybridization

The chromosomal locations of several families of tandem repetitive DNA sequences and the 5S rDNA were determined using fluorescence in situ hybridization (FISH) in the five North American charr species: Salvelinus namaycush, S. fontinalis, S. alpinus, S. malma, and S. confluentus. The pattern of hybridization of three centromeric repetitive sequences previously isolated from S. namaycush and S. alpinus was unique in each species. Dual-color FISH experiments showed that in several species many of the centromeres had the EcoRI-DraI family in addition to either the AluI-RsaI type A or type B families. The EcoRI-DraI family which was found only at the centromeres of acrocentric chromosomes in S. namaycush, S. fontinalis and S. malma was also found at centromeres of selected metacentrics in S. alpinus (one pair) and S. confluentus (four pairs) whose chromosomes have undergone additional centric fusions compared to the other species. The locations of 5S rDNA sequences were different in each species except for the two most closely related (S. alpinus and S. malma). Two whole-arm chromosome paint probes, one specific for the short and the other for the long arm of the lake charr sex chromosomes, hybridize to the same chromosome pair in all species. Results with other paint probes suggest that independent centric fusions have occurred in S. alpinus and S. confluentus which is consistent with the phylogenetic tree obtained previously for Salvelinus with cytogenetic and DNA data.     

A cereal centromeric sequence

We report the identification of a family of sequences located by in situ hybridisation to the centromeres of all the Triticeae chromosomes studied, including the supernumerary and midget chromosomes, the centromeres of all maize chromosomes and the heterochromatic regions of rice chromosomes. This family of sequences (CCS1), together with the cereal genome alignments, will allow the evolution of the cereal centromeres and their sites to be studied. The family of sequences also shows homology to the CENP-B box. The centromeres of the cereal species and the proteins that interact with them can now be characterised. Received: 11 July 1996; in revised form: 19 September 1996 / Accepted: 24 September 1996     

Boom-Bust Turnovers of Megabase-Sized Centromeric DNA in Solanum Species: Rapid Evolution of DNA Sequences Associated with Centromeres

Centromeres are composed of long arrays of satellite repeats in most multicellular eukaryotes investigated to date. The satellite repeat–based centromeres are believed to have evolved from “neocentromeres” that originally contained only single- or low-copy sequences. However, the emergence and evolution of the satellite repeats in centromeres has been elusive. Potato (Solanum tuberosum) provides a model system for studying centromere evolution because each of its 12 centromeres contains distinct DNA sequences, allowing comparative analysis of homoeologous centromeres from related species. We conducted genome-wide analysis of the centromeric sequences in Solanum verrucosum, a wild species closely related to potato. Unambiguous homoeologous centromeric sequences were detected in only a single centromere (Cen9) between the two species. Four centromeres (Cen2, Cen4, Cen7, and Cen10) in S. verrucosum contained distinct satellite repeats that were amplified from retrotransposon-related sequences. Strikingly, the same four centromeres in potato contain either different satellite repeats (Cen2 and Cen7) or exclusively single- and low-copy sequences (Cen4 and Cen10). Our sequence comparison of five homoeologous centromeres in two Solanum species reveals rapid divergence of centromeric sequences among closely related species. We propose that centromeric satellite repeats undergo boom-bust cycles before a favorable repeat is fixed in the population.     

Ontogeny of skull size and shape changes within a framework of biphasic lifestyle: a case study in six <Emphasis Type="Italic">Triturus</Emphasis> species (Amphibia,Salamandridae)

As with many other amphibians, Triturus species are characterized by a biphasic life cycle with abrupt changes in the cranial skeleton during metamorphosis. The post-metamorphic shape changes of the cranial skeleton were investigated using geometric morphometric techniques in six species: Triturus alpestris, T. vulgaris, T. dobrogicus, T. cristatus, T. carnifex, and T. karelinii. The comparative analysis of ontogenetic trajectories revealed that these species have a conserved developmental rate with divergent ontogenetic trajectories of the ventral skull shape that mainly reflect phylogenetic relatedness. A striking exception in the ontogenetic pattern was possibly found in T. dobrogicus, characterized by a marked increase in the developmental rate compared to the other newt species. The size-related shape changes explained a large proportion of shape change during post-metamorphic growth within each species, with marked positive allometric growth of skull elements related to foraging.     

Repeatless and Repeat-Based Centromeres in Potato: Implications for Centromere Evolution

Centromeres in most higher eukaryotes are composed of long arrays of satellite repeats. By contrast, most newly formed centromeres (neocentromeres) do not contain satellite repeats and instead include DNA sequences representative of the genome. An unknown question in centromere evolution is how satellite repeat-based centromeres evolve from neocentromeres. We conducted a genome-wide characterization of sequences associated with CENH3 nucleosomes in potato (Solanum tuberosum). Five potato centromeres (Cen4, Cen6, Cen10, Cen11, and Cen12) consisted primarily of single- or low-copy DNA sequences. No satellite repeats were identified in these five centromeres. At least one transcribed gene was associated with CENH3 nucleosomes. Thus, these five centromeres structurally resemble neocentromeres. By contrast, six potato centromeres (Cen1, Cen2, Cen3, Cen5, Cen7, and Cen8) contained megabase-sized satellite repeat arrays that are unique to individual centromeres. The satellite repeat arrays likely span the entire functional cores of these six centromeres. At least four of the centromeric repeats were amplified from retrotransposon-related sequences and were not detected in Solanum species closely related to potato. The presence of two distinct types of centromeres, coupled with the boom-and-bust cycles of centromeric satellite repeats in Solanum species, suggests that repeat-based centromeres can rapidly evolve from neocentromeres by de novo amplification and insertion of satellite repeats in the CENH3 domains.     

Sexual dimorphism in the skull geometry of newt species of <Emphasis Type="Italic">Ichthyosaura,Triturus</Emphasis> and <Emphasis Type="Italic">Lissotriton</Emphasis> (Salamandridae,Caudata, Amphibia)

In this study, we applied geometric morphometrics to explore variations in the level and pattern of sexual size dimorphism (SSD) and sexual shape dimorphism (SShD) of the ventral cranium in three different Modern Eurasian newt taxa (Ichthyosaura alpestris, Triturus species group and Lissotriton vulgaris). The ventral cranium is the part of the skull that is more directly related to foraging and feeding. Our results indicate that the level and pattern of sexual dimorphism in the ventral cranium differ among Modern Eurasian newt taxa. Regarding sexual dimorphism in skull size, Ichthyosaura alpestris and Triturus species show female-biased patterns (females are larger than males), whereas Lissotriton vulgaris appears to be non-dimorphic in skull size. In I. alpestris and Triturus species, SShD is mostly absent, whereas in L. vulgaris, SShD is more pronounced. A high level of variation between populations in both SSD and SShD indicates that local conditions may have a profound effect on the magnitude and direction of sexual dimorphism. The significant sexual differences in ventral cranium size and shape indicate possible subtle intersexual differences in ecological demands due to diet specialisation, in spite of similar general ecological settings.     

HYBRID ZONES AND HOST-PARASITE RELATIONSHIPS: EFFECT ON THE EVOLUTION OF PARASITIC SPECIFICITY

In southern France, Diplozoon gracile (Monogenea, Polyopisthocotylea), parasitizes four sympatric cyprinids. One of these host species, Barbus meridionalis, naturally hybridizes with another species of barbel, Barbus barbus, which is never parasitized by D. gracile under natural conditions. This hybridization has previously been studied and described as an introgression of B. barbus by B. meridionalis. The hybrids are parasitized by D. gracile, and parasite prevalence increases in proportion to the introgression rate, i.e., the percentage of B. meridionalis genes. The causes for this preferential distribution of the parasite in the hybrid population are analysed on the basis of ecological and ethological differences between the two parent species.     

NTRS, a new family of highly repetitive DNAs specific for the T1 chromosome of tobacco

Species-specific repeated DNAs are important for identifying genomic components of hybrid organisms in plant breeding and in taxonomic studies, and we have previously described the HRS60 and GRS families of highly repetitive DNA sequences in tobacco. Here we describe a new family of highly repetitive DNA sequences termed NTRS (SspI family) that we have isolated from Nicotiana tomentosiformis (Goodspeed) and characterized and that is specific for the genomes of several species of the subgenus Tabacum. In situ hybridization showed that NTRS sequences are present in three pairs of chromosomes of N. tomentosiformis, six pairs of chromosomes of N. kawakamii, and only one pair of chromosomes of N. tabacum at an intercalary site. The NTRS family is not present in the N. otophora genome. The majority of NTRS sequences appeared to be organized in tandem arrays in which local DNA structures sensitive to single strand-specific chemical probes, potassium permanganate, and osmium tetroxide complexed with pyridine revealed a periodicity of 220 bp, equal to the length of the repeat unit. The inner cytosine in CCGG and CC(A/T)GG sequences of the NTRS family is frequently methylated. Cloned and sequenced NTRS monomeric units are 212–219 bp in length and show 83.5%–95% mutual homology. They exhibit properties characteristic for molecules that possess stable intrinsic curvature, but there are differences among individual monomers in the degree of curvature. NTRS sequences like HRS60 and GRS sequences, were found to specify nucleosome positions. Received: 12 November 1996 / in revised form: 12 May 1997 / Accepted: 12 May 1997