Early replication banding in <Emphasis Type="Italic">Leporinus</Emphasis> species (Osteichthyes,Characiformes) bearing differentiated sex chromosomes (ZW)

There are few examples of differentiated sex chromosomes in fishes. In the genus Leporinus, seven species present a highly differentiated ZW system, derived from heterochromatinization process. Cytogenetic analyses carried out in three of these fish species, Leporinus obtusidens, L. elongatus and L. reinhardti, through RBG-banding, showed late replication bands, coincident with heterochromatic regions in both Z and W chromosomes. A similar interstitial early replication segment was observed in the complex heterochromatic region along the Wq arms in the three species, which might correspond to a pseudoautosomal segment (SD, sex determining locus). Asynchrony related to the replication pattern among different Z chromosomes was not observed. When the identification of nuclear organizer regions by silver nitrate was performed over chromosomal preparations previously exposed to 5-bromo-2′-deoxyuridine (BrdU), remarkable positive signals at interstitial and telomeric position were observed on the q arms of W chromosomes in the species L. elongatus and L. reinhardti. The absence of 18S ribosomal RNA gene loci in this region, formerly demonstrated by FISH, indicates that this argentophilic behavior is putatively due to heterochromatin decondensation caused by BrdU incorporation, favoring such Ag+ reaction. Early and late replication bands were also observed in the heterochromatic portions of Z and W chromosomes, indicating that euchromatic and heterochromatic regions are interspersed. The present data suggest a significant level of heterochromatic complexity in the sex chromosomes of each species. On the other hand, the replication pattern shared by them supports a monophyletic origin.     

Karyomorphological studies onWoodwardia sensu lato of Japan

Karyomorphological comparisons were made of five species of JapaneseWoodwardia. There were no marked differences at interphase and prophase among the five species.Woodwardia japonica, W. prolifera, andW. unigemmata were diploid with 2n=68 and the formulas of their metaphase karyotypes uniformly 4m(median centromeric chromosomes)+12sm(submedian)+52(st+t)(subterminal and terminal).Woodwardia orientalis was tetraploid with 2n=136 and 8m+24sm+104(st+t), and the ratio of each chromosomal type to total complement was identical to that of three diploid species. These four species had several characteristics in common:x=34, the longest chromosome of sm, and a mean chromosome length over 3.0 μm. AlthoughWoodwardia orientalis showed some similarity toW. prolifera, it seems to be an allotetraploid which originated by chromosome doubling of a hybrid ofW. prolifera and a diploid species as yet karyomorphologically unknown.Woodwardia kempii was tetraploid with 2n=124 and 8m+24sm+92(st+t), and differed from the others in havingx=31, the longest chromosomes of t, and a mean chromosome length under 3.0 μm. This species has been classified as an independent genus,Chieniopteris, and our karyomorphological study supports this treatment.     

Karyomorphology ofCrossostylis (Rhizophoraceae)

We present the first report on somatic chromosome numbers and morphology in eight of 13 recorded species ofCrossostylis, one of inland genera of Rhizophoraceae. The chromosome number ofCrossostylis is 2n=28 in all species examined; therefore, the genus hasx=14, a number which is the smallest and unknown elsewhere in the family. Based onCrossostylis raiateensis, we further present that 24 of 28 chromosomes at metaphase have centromeres at median position, and the remaining four at submedian or subterminal position. The chromosome morphology seems to imply thatCrossostylis might be a tetraploid with the original base numberx=7, but an extensive study in the other inland genera is needed to find such a small chromosome number.     

SUPERNUMERARY CHROMOSOMES IN SAXIFRAGA VIRGINIENSIS (SAXIFRAGACEAE)

Acetocarmine squashes of root tips have demonstrated that 2n = 20 and 38 in Saxifraga virginiensis. These contrast with the earlier reported count of 2n = 28 for this species. In several populations supernumerary chromosomes were detected. Both intrapopulational and interpopulational variation in supernumerary chromosome number were detected, with the largest number of supernumerary chromosomes observed being six. Because these supernumerary chromosomes are equal in size to many of the smaller A chromosomes during mitotic metaphase, the presence of supernumerary chromosomes in this species could not be ascertained by analysis of mitotic metaphase preparations alone. During mitotic prophase, however, the supernumerary chromosomes of S. virginiensis are highly heterochromatic, appearing more densely coiled and darkly stained than the A chromosomes. This characteristic facilitated the recognition of supernumerary chromosomes in this species. The similarity in size of A and supernumerary chromosomes during mitotic metaphase and the observation of six supernumerary chromosomes in one population suggest that the count of 2n = 28 reported earlier for S. virginiensis may actually be a misinterpretation of 2n = 20 plus 8 supernumerary chromosomes. Furthermore, these findings and the observation of this same supernumerary chromosome phenomenon in other species of Saxifraga raise the possibility that some of the many disparate chromosome counts attributed to aneuploidy in the large genus Saxifraga may also be the result of misinterpretations of supernumerary chromosomes as A chromosomes.     

TheTy1-copia group retrotransposons inVicia species: copy number, sequence heterogeneity and chromosomal localisation

We present an in-depth study of theTy1-copia group of retrotransposons within the plant genusVicia, which contains species with widely differing genome sizes. We have compared the numbers and sequence heterogeneities of these genetic elements in three diploidVicia species chosen to represent large (V. faba, 1C=13.3 pg), medium (V. melanops, 1C=11.5 pg) and small (V. sativa, 1C=2.3 pg) genomes within the genus. The copy numbers of the retrotransposons are all high but vary greatly, withV. faba containing approximately 10⁶ copies,V. melanops about 1000 copies andV. sativa 5000 copies. The degree of sequence heterogeneity ofTy1-copia group elements correlates with their copy number within each genome, but neither heterogeneity nor copy number are related to the genome size of the host. In situ hybridization to metaphase chromosomes shows that the retrotransposons inV. faba are distributed throughout all chromosomes but are much less abundant in certain heterochromatic regions. These results are discussed in the context of plant retrotransposon evolution.     

Cytological studies on meiotic configurations of intraspecific aneuploids ofCarex blepharicarpa (Cyperaceae) in Japan

Chromosome configurations at meiotic metaphase I ofCarex blepharicarpa were determined for 245 individuals collected from 11 localities in the Chugoku District of Japan. Nine intraspecific aneuploids, 2n=26–33 and 41, were found. The most common diploid number was 29, and found in 73 individuals. No clear geographical pattern was suggested by a distribution of these aneuploids. A consecutive series of chromosome numbers from 2n=26 to 32 was found in two populations from Okayama Prefecture. At meiotic metaphase I, univalents and multivalents were found, and one to three trivalents were observed in each aneuploid. Heteromorphic chain trivalents comprising large, medium and small chromosomes were prevalent. Homomorphic trivalents, which are thought to be originated from chromosome duplications caused by unreduced gametes, were found in only one individual with 2n=41. The high frequency of heteromorphic trivalents in this species indicates that most aneuploidy probably results from fission and/or fusion of chromosomes.     

国产毛茛科驴蹄草属两种植物的细胞学研究

为探讨国产毛茛科(Ranunculaceae)驴蹄草属(Caltha L.)植物的细胞学特征,对驴蹄草(C.palustris L.)3个居群和花葶驴蹄草(C.scaposa Hook.f.&Thoms.)5个居群进行了细胞学研究。驴蹄草贵州纳雍居群的染色体数目为2n=32(四倍体),两个云南中甸居群的染色体数目均为2n=64(八倍体)。花葶驴蹄草四川红原、康定、石渠居群的染色体数目均为2n=32(四倍体),该数目为首次报道;西藏林芝和云南德钦居群的染色体数目均为2n=64(八倍体)。驴蹄草的染色体比花葶驴蹄草大。这两种植物的32或64条染色体分别以4条或8条为单位大致能够排列为8组同源染色体,但同一组内的染色体经常具有明显的异形性(heteromorphy),不同居群的核型组成多少具有差异。同时,还分析了驴蹄草和花葶驴蹄草的不同倍性细胞型在我国的地理分布式样。     

Karyomorphology and evolution in some Hamamelidaceae and Platanaceae (Hamamelididae; Hamamelidales)

Karyomorphology in 14 species of 12 genera representing a variation of Hamamelidaceae and in one species of Platanaceae (Platanus only) is investigated in an effort to contribute to an understanding of chromosome evolution and inter- and intrafamilial relationships. All genera investigated show similar chromosome features at resting stage and prophase, excepting that at resting stageRhodoleia shows the simple, rather than the simple-complex, chromocenter type as in other genera. At metaphase all the genera investigated of Hamamelidaceae, like other ‘lower’ Hamamelididae, have chromosomes with median centromeres (m-chromosomes), those with submedian centromeres (sm-chromosomes) and those with subterminal (or terminal) centromeres (st-t-chromosomes) at different frequencies, although frequencies ofst-t-chromosomes are always less than 33%. InPlatanus,m-chromosomes are lacking and insteadst-t-chromosomes are predominant (86%), a feature seemingly very specialized. We confirmedx=7 in Platanaceae,x=12 in Hamamelidoideae and Rhodoleioideae, andx=8 in Exbucklandioideae and Altingioideae (Hamamelidaceae). An analysis of chromosome morphology supports the hypothesis thatx=12 in the former two subfamilies is of tetraploid origin fromx=6, rather than of triploid origin fromx=8. We further give brief comments on the suprageneric classification of Hamamelidaceae that was recently proposed by Endress.     

Die intraspezifische Variabilität des heterochromatischen Armes eines Chromosoms bei der GattungCarabus L. (Coleoptera)

The chromosome complement ofC. auronitens Fabr. is 2n _♂=26+XY. One autosomal pair—called A-chromosomes—is relatively long.A-chromosomes consist of a euchromatic and a heterochromatic arm. Labelling of mitotic chromosomes with³H-thymidine shows that replication of the heterochromatic arm continues when it has ended in the euchromatic arm. In males and females the length of the heterochromatic arm varies intraindividually. In 47 of 99 males the heterochromatic arms were heteromorphic. Calculations of the quotient length of the euchromatic/length of the heterochromatic arm have shown that at least 6 different types of the A-chromosome exist. These types differ from each other in the number of heterochromatic sections separated by constrictions. The longest heterochromatic arm observed consisted of 8 such sections. The genetic significance of the heterochromatin in the genus ofCarabus is at present unknown (Zusammenfassung see p.305).      

Diversity of centromeric repeats in two closely related wild rice species, Oryza officinalis and Oryza rhizomatis

Oryza officinalis (CC, 2n=24) and Oryza rhizomatis (CC, 2n=24) belong to the Oryza genus, which contains more than 20 identified wild rice species. Although much has been known about the molecular composition and organization of centromeres in Oryza sativa, relatively little is known of its wild relatives. In the present study, we isolated and characterized a 126-bp centromeric satellite (CentO-C) from three bacterial artificial chromosomes of O. officinalis. In addition to CentO-C, low abundance of CentO satellites is also present in O. officinalis. In order to determine the chromosomal locations and distributions of CentO-C (126-bp), CentO (155 bp) and TrsC (366 bp) satellite within O. officinalis, fluorescence in situ hybridization examination was done on pachytene or metaphase I chromosomes. We found that only ten centromeres (excluding centromere 7 and 2) contain CentO-C arrays in O. officinalis, while centromere 7 comprises CentO satellites, and centromere 2 is devoid of any detectable satellites. For TrsC satellites, it was detected at multiple subtelomeric regions in O. officinalis, however, in O. rhizomatis, TrsC sequences were detected both in the four centromeric regions (CEN 3, 4, 10, 11) and the multiple subtelomeric regions. Therefore, these data reveal the evolutionary diversification pattern of centromere DNA within/or between close related species, and could provide an insight into the dynamic evolutionary processes of rice centromere.     

Cytogenetic studies on Apareiodon affinis (Pisces, Characiformes) from Paraná river basin: sex chromosomes and polymorphism

Comparative cytogenetic studies were carried out on Apareiodon affinis from an important hydrographic system at South America, the Paraná river basin. Two distant regions were chosen, which were separated by Guaíra Falls (formerly Sete Quedas); the region in the upper part of the hydrographic basin is called Upper Paraná (Brazil), whereas and the other in the lower part is called Lower Paraná (Argentina). Individuals from Upper Paraná have diploid numbers of 2n = 54 (NF = 108) for males and 2n = 55 (NF = 110) for females, showing female heterogamety with a ZZ/ZW₁W₂ multiple sex chromosomes system that is endemic for the region. In different localities at Lower Paraná, the specimens presented diploid number of 2n = 54 for both sexes, without any sex chromosomes heteromorphism. However, they have an accentuated polymorphism characterized by variation in number of acrocentric chromosomes, constituting something new for family Parodontidae. The most likely hypothesis to explain the origin of such polymorphism is based on successive pericentric inversions giving rise to acrocentric chromosomes. Thus, it was possible to detect 10 cytotypes along the Lower Paraná basin. Such chromosomal variations possibly are the consequence of an adaptative process. Our data probably indicate the occurrence of distinct species in each region that share the same denomination. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytogeographical study of four aneuploids ofCarex oxyandra Kudo in Japan

Chromosome numbers were determined for 342 clones ofCarex oxyandra collected from 35 localities in Hokkaido, Honshu, Shikoku and Kyushu, Japan. Four intraspecific aneuploids, 2n=18, 20, 24 and 26, were found. In meiotic division, only bivalent chromosomes were observed in all clones at metaphases I and II, suggesting that the aneuploids are established gamodemes. In the mitotic metaphase chromosomes, trimodal variation in chromosome length was observed. The 2n=26 clones found on Mt. Hiko had two particularly small chromosomes. The cytodemes with higher number of chromosomes are distributed in more southern areas of Japan.Carex oxyandra, therefore, accompanied with chromosome fragmentations, might spread the geographical distribution to the southern parts. The morphological characters of leaves, spikes, scales, perigynia and nuts were similar among the four cytodemes, except for the small leaves on plants from Yaku Island.     

Chromosome analysis of five Brazilian species of poison frogs (Anura: Dendrobatidae)

Dendrobatid frogs have undergone an extensive systematic reorganization based on recent molecular findings. The present work describes karyotypes of the Brazilian species Adelphobates castaneoticus, A. quinquevittatus, Ameerega picta, A. galactonotus and Dendrobates tinctorius which were compared to each other and with previously described related species. All karyotypes consisted of 2n = 18 chromosomes, except for A. picta which had 2n = 24. The karyotypes of the Adelphobates and D. tinctorius species were highly similar to each other and to the other 2n = 18 previously studied species, revealing conserved karyotypic characteristics in both genera. In recent phylogenetic studies, all Adelphobates species were grouped in a clade separated from the Dendrobates species. Thus, we hypothesized that their common karyotypic traits may have a distinct origin by chromosome rearrangements and mutations. In A. picta, with 2n = 24, chromosome features of pairs from 1 to 8 are shared with other previously karyotyped species within this genus. Hence, the A. picta data reinforced that the C-banding pattern and the NOR location are species-specific traits in the genus Ameerega. Moreover, the Ameerega monophyletism proposed by previous phylogenetic studies indicates that the karyotypic differences among species in this genus result from a long divergence time.     

Karyotype variation in Drosophila birchii

Drosophila birchii, a member of the melanogaster species group of the subgenus Sophophora, is common in the tropical rain forests of the Australia-New Guinea areas. Chromosome squashes are easily prepared from the larval ganglion cells and the sex chromosomes are readily recognizable. The species exhibits a remarkable karyotype variation. The metaphase plate figures, in general, show two pairs of V's, one pair of dots and one pair of sex chromosomes. Variations in metaphase chromosome morphology are found in the X (with four types), the Y (with three types) and chromosome IV (with two types). Chromosomal interchanges between X- and Y-chromosomes Type I are postulated to be involved in the differentiation of sex chromosome morphology while the modification of chromosome IV seems likely to be a result of the acquisition of extra heterochromatin. These chromosome types form seven distinct metaphase plate figures, all encountered in wild populations, thus giving D. birchii the most variable karyotype in the genus Drosophila.     

Chromosome plasticity in Ctenomys (Rodentia Octodontidae): chromosome 1 evolution and heterochromatin variation

A chromosome 1 (Cr1) pericentric inversion is described in six of seven species in the genus Ctenomys (tuco-tucos) from Uruguay. The inversion was inferred from G-band analyses of subtelocentric Cr1 hypothesised to be derived from the ancestral metacentric condition. Cr1 varies across species in heterochromatin amount and localisation including a metacentric chromosome without positive C-bands in C. torquatus, a subtelocentric chromosome with heterochromatic short arms in C. rionegrensis, and a subtelocentric chromosome negative after C-banding in five of the species analysed here. Pachytene chromosomes from C. rionegrensis, a species with the highest heterochromatin content, and C. torquatus, one of the species with the lowest heterochromatin content, were analysed in order to assess possible mechanisms of heterochromatin evolution. This analysis revealed the presence of three heterochromatic chromocenters in C. rionegrensis where bivalents converge, while in C. torquatus only one chromocenter was observed. In both species, highly repetitive DNA was observed, localised in chromocenters after “in situ” hybridisation. Heterochromatin associated protein M31 was localised in chromocenters of both species after immuno-detection. The spread of heterochromatin in Ctenomys chromosomes could be produced by chromatin exchanges at the chromocenter level. We propose the exchange of this DNA associated proteins between non-homologous chromosomes in pachytene to be the responsible for the spread of heterochromatin through the karyotypes of species like C. rionegrensis     

Karyotype with 210 chromosomes in guaraná (<Emphasis Type="Italic">Paullinia cupana</Emphasis> ‘Sorbilis’)

The genus Paullinia includes the economically important P. cupana, known as guaraná in Brazil and more recently in the world market. Native Americans of the Maué and Andirá tribes cultivated P. cupana ‘Sorbilis’ in central Amazon, and the Barés cultivated the ‘Typica’ variety in the upper Negro River (Brazil). Cytological studies in the Sapindaceae family have concentrated on the diversity in number (from 2n = 14 to 96) and size of the chromosomes. In Paullinia, seven species have been karyotyped and all show 2n = 24. Meristem maceration, cellular dissociation and air-drying techniques were used for cytogenetic preparations and DNA content was determined by flow cytometry. Chromosome characterization and DNA content of Paullinia cupana Kunth ‘Sorbilis’ (Mart.) Ducke (Sapindaceae) were studied. The high chromosome number (2n = 210) fall into two cytomorphological groups: (a) a metacentric and submetacentric group showing 25 sets of three pairs of chromosomes (2–76); (b) a group containing only acrocentric showing 12 sets of two pairs of chromosomes (82–105), a homologous submetacentric pair (1) and an acrocentric pair (81). Mean nuclear DNA content of guaraná was 2C = 22.8 pg. A karyogram was set up showing a high chromosome number complement.     

Chromosomal diversification of reef fishes from genus <Emphasis Type="Italic">Centropyge</Emphasis>(Perciformes,Pomacanthidae)

The genus Centropyge is remarkable for species richness, composing a highly specialized fish group amongst members from family Pomacanthidae. However, cytogenetical reports are nearly absent in these animals. New data are provided from karyotypical studies carried out on Centropyge aurantonotus from the Brazilian coast of the Atlantic Ocean and C. ferrugatus from the Philippines Sea of the Indo-Pacific Ocean. Both species present 2n=48 but karyotypes are differentiated by fundamental number. C. aurantonotus has a great number of biarmed chromosomes (4 m + 14 sm+16 st+4 a), while C. ferrugatus presents only acrocentric chromosomes. Single nucleolar organizer regions (NORs) are located at interstitial position of an acrocentric pair in C. ferrugatus and on short arms of a subtelocentric pair in C. aurantonotus, as confirmed by fluorescent in situ hybridization (FISH) with 18S rDNA probes. Heterochromatin is distributed over NOR and centromeric regions in both species, but additional GC-rich heterochromatic blocks on short arms of up to eight chromosomal pairs can be detected in C. aurantonotus. 5S rDNA segments were located interstitially on two chromosomal pairs in C. ferrugatus and on nine pairs in C. aurantonotus, mostly equivalent to heterochromatic blocks on short arms of biarmed chromosomes. C. ferrugatus can be considered a species in which basal chromosomal features proposed for modern Teleosteans were conserved. The derived karyotype pattern of C. aurantonotus seems to be determined by pericentric inversions and heterochromatin addition which probably determined the notorious dispersion of 5S rRNA (pseudo)genes. It is demonstrated that, even within a group generally characterized by cytogenetical homogeneity as the family Pomacanthidae, diversified karyotypes can be found.     

Differentielle Darstellung der Metaphasechromosomen von Cypripedium debile mit Chinacrin- und Giemsa-Färbung

The somatic metaphase chromosomes of C. debile stain differentially with quinacrine and Giemsa respectively. After quinacrine staining the heterochromatic regions show fainter fluorescence than euchromatic regions. With Giemsa the heterochromatic regions are more deeply stained than the euchromatic regions. This is true also for slides which were pre-treated with trypsin or trichloroacetic acid. From these findings it may be supposed, that the heterochromatic regions are G-C rich. Furthermore, differences in protein composition are expected between hetero- and euchromatic regions. The results of differential staining of metaphase chromosomes of C. debile, including those which were published in previous papers, are summarized here.     

Chromosomal polymorphism and small karyotypic differentiation in a group of Ctenomys species from Central Argentina (Rodentia: Octodontidae)

Cytogenetic analysis was performed in six nominal taxa of the genus Ctenomys with a diploid number of 2n=47/48. The studied species were: C. australis (2n=48, FN=76); C. mendocinus (2n=47/48, FN=68/75/76); C. porteousi (2n=47/48, FN=71/72/73); C. azarae (2n=47, FN=71); C. sp. (chasiquensis) (2n=47/48); and C. talarum (2n=48, FN=80). The first three species shared the whole complement, C. talarum shares with them 19 arms from a total of 43 (44%). In all species analyzed constitutive heterochromatin was detected in most short arms, and in several centromeres. Polymorphisms for several pairs involving the heterochromatic short arms together with a complex polymorphism of pair A1 were found in C. azarae, C. sp., C. mendocinus and C. porteousi. Intraindividual variation found in one specimen of C. porteousi involving heterochromatic arms is discussed.These results lead us to propose the inclusion of all species except C. talarum, within a complex called the mendocinus-group. A new case of conservatism for chromosomal number in the genus Ctenomys is found in this group.     

Molecular cytogenetic characterization of <Emphasis Type="Italic">Rumex papillaris</Emphasis>, a dioecious plant with an XX/XY<Subscript>1</Subscript>Y<Subscript>2</Subscript> sex chromosome system

Rumex papillaris Boiss, & Reut., an Iberian endemic, belongs to the section Acetosa of the genus Rumex whose main representative is R. acetosa L., a species intensively studied in relation to sex-chromosome evolution. Here, we characterize cytogenetically the chromosomal complement of R. papillaris in an effort to enhance future comparative genomic approaches and to better our understanding of sex chromosome structure in plants. Rumex papillaris, as is common in this group, is a dioecious species characterized by the presence of a multiple sex chromosome system (with females 2n = 12 + XX and males 2n = 12 + XY₁Y₂). Except for the X chromosome both Y chromosomes are the longest in the karyotype and appear heterochromatic due to the accumulation of at least two satellite DNA families, RAE180 and RAYSI. Each chromosome of pair VI has an additional major heterochromatin block at the distal region of the short arm. These supernumerary heterochromatic blocks are occupied by RAE730 satellite DNA family. The Y-related RAE180 family is also present in an additional minor autosomal locus. Our comparative study of the chromosomal organization of the different satellite-DNA sequences in XX/XY and XX/XY₁Y₂ Rumex species demonstrates that of active mechanisms of heterochromatin amplification occurred and were accompanied by chromosomal rearrangements giving rise to the multiple XX/XY₁Y₂ chromosome systems observed in Rumex. Additionally, Y₁ and Y₂ chromosomes have undergone further rearrangements leading to differential patterns of Y-heterochromatin distribution between Rumex species with multiple sex chromosome systems.