Nature and distribution of constitutive heterochromatin and NOR location in the grasshopper Phaeoparia megacephala (Romaleidae: Orthoptera)

The constitutive heterochromatin (CH) of Phaeoparia megacephala was studied using C-banding and fluorochrome staining (CMA3, DAPI and acridine orange). The nucleolar organizer regions (NOR) were identified with silver staining. The chromosome complement of this species was 2n = 23, XO in males, and 2n = 24, XX in females. The CH was pericentromeric in all chromosomes. L1, L2, L3 and X chromosomes showed large blocks of CH, while the medium and small chromosomes had small blocks. The staining procedure with acridine orange revealed the same pattern. All the pericentromeric regions showed small blocks of CMA3-positive constitutive heterochromatin (GC-rich regions), while only part of the large C-band positive chromosome segments (L1, L2, L3 and X) were CMA3 positive. This character demonstrates an uncommon heterogeneity of constitutive heterochromatin in P. megacephala. The fluorochrome DAPI did not reveal DAPI-positive regions (AT-rich regions). Silver staining revealed only one pair of medium chromosomes with NOR.     

Polymorphic karyotypes and sex chromosomes in the tufted deer (Elaphodus cephalophus): cytogenetic studies and analyses of sex chromosome-linked genes

Different diploid chromosome numbers have been reported for the tufted deer Elaphodus cephalophus (female, 2n = 46/47; male, 2n = 47/48) in earlier reports. In the present study, chromosomal analysis of seven tufted deer (5 male symbol, 2 female symbol) revealed that the karyotype of these animals contains 48 chromosomes, including a pair of large heteromorphic chromosomes in the male. C-banding revealed these chromosomes to be very rich in constitutive heterochromatin. Chromosome banding and PCR of sex chromosome-linked genes (SRY, ZFX, ZFY) performed on DOP-PCR products of single microdissected X and Y chromosomes confirmed that the large telocentric chromosome without secondary constriction is the X chromosome whereas the subtelocentric chromosome is the Y. The increased size of both, the X and Y chromosome, appears to be at least partially attributable to the presence of substantial amounts of heterochromatin.     

First report on C-banding and fluorescent banding in species of Dieuches (Rhyparochrominae: Lygaeidae: Heteroptera)

Although the karyotypes of twelve species of Dieuches Dohrn, 1860 belonging to Rhyparochrominae have been described so far, there is no information about heterochromatin and its characterization in terms of base composition for any of the species. In the present paper, C-banding and fluorescent banding have been applied for the first time to three species of Dieuches : D. uniguttatus, D. insignis (2n = 12 = 8A + 2m + XY) and D. coloratus (2n = 14 = 10A + 2m + XY). Dieuches uniguttatus and D. insignis show distinct terminal C-bands along with a few interstitial bands in all the autosomal bivalents, whereas in D. coloratus , one autosomal pair is almost completely heterochromatic, three show C-positive bands while one is totally euchromatic. The sex chromosomes too show heterogeneity in distribution of C-heterochromatin among three Dieuches species. Characterization of heterochromatin in D. uniguttatus and D. insignis using DAPI/CMA₃ staining reveals that in D. uniguttatus , C- heterochromatin blocks of all the autosomal bivalents, which are predominantly A–T rich, whereas in D. insignis , these are rich both in A–T and G–C. In D. uniguttatus, sex chromosomes X and Y have localized G–C rich regions whereas in D. insignis , these are scattered in X and absent in Y. As variations in the heterochromatin represent the main source of karyological differentiation among and within species, it seems that there occurred extensive redistribution of heterochromatin within the complement as the three species evolved. There is need for cytological details of more species to understand evolutionary aspects in the genus Dieuches .     

Karyology of the Savi pine vole, Microtus savii (De Sélys-Longchamps, 1838) (Rodentia, Arvicolidae): G-, C-, DA/DAPI-, and AluI-bands.

Specimens of the Savi pine vole (Microtus savii) were collected from three localities in central (Pisa and Viterbo) and southern Italy (Rosarno, Calabria) and were karyotyped using G-, C-, DA/DAPI-, and AluI-banding. All karyotypes had 2n = 54 chromosomes and seemingly identical autosomal banding. The sex chromosomes of the southern Italian specimens, M. savii brachycercus, showed additional large blocks of heterochromatin. In the northern specimens, M. savii savii, the X chromosome is metacentric, whereas in the southern specimens of M. savii brachycercus the X chromosome is a much larger submetacentric chromosome, and the Y chromosome is more than twice the size of the Y in the northern specimens. DA/DAPI staining reveals three levels of fluorescent intensity in the sex chromosomes of the Calabrian specimens. The sex chromosomes of M. savii brachycercus also have the only AluI bands seen in either chromosome set. These data suggest a heterogeneous origin and composition of the C-band regions of these chromosomes. Preliminary data suggest that fertility is reduced in crosses between the two karyomorphs.     

Variation of constitutive heterochromatin in the sex chromosomes of the rodent Bandicota bengalensis bengalensis (Gray)

Bandicota bengalensis bengalensis (Gray) trapped from different localities of India and Nepal exhibited a marked variation in the size and morphology of sex chromosomes. Three types of X's were found; A) simple acrocentric, B) composite subtelocentric and C) composite submetacentric X with their relative sizes 5.9%, 7.5% and 9.6% of the genome respectively. The autosomes remained unaltered. It was shown that this variation in the size of sex chromosomes was caused by deletion of constitutive heterochromatin. The Y chromosome was also found to be variable. Usually a large X was combined with a large Y. The preponderance of homozygotes for each type of X chromosome in populations, suggested the probable role of sex chromosomes heterochromatin in speciation.     

白眉长臂猿(Hylobates hoolock leuconedys)的染色体研究

本文对两只雄性白眉长臂猿的染色体的C带、G带及Ag-NORs分布进行了较详细的分析,证实染色体数2n=38,并对该种的分类地位提出了一些新看法。     

Cytogenetic analysis of the neotropical spider Nephilengys cruentata (Araneomorphae, Tetragnathidae): standard staining nors, C-bands and base-specific fluorochromes.

The aim of this work is to characterize Nephilengys cruentata in relation to the diploid number, chromosome morphology, type of sex determination chromosome system, chromosomes bearing the Nucleolar Organizer Regions (NORs), C-banding pattern, and AT or GC repetitive sequences. The chromosome preparations were submitted to standard staining (Giemsa), NOR silver impregnation, C-banding technique, and base-specific fluorochrome staining. The analysis of the cells showed 2n = 24 and 2n = 26 chromosomes in the embryos, and 2n = 26 in the ovarian cells, being all the chromosomes acrocentric. The long arm of the pairs 1, 2 and 3 showed an extensive negative heteropycnotic area when the mitotic metaphases were stained with Giemsa. The sexual chromosomes did not show differential characteristics that allowed to distinguish them from the other chromosomes of the complement. Considering the diploid numbers found in N. cruentata and the prevalence of X1X2 sex determination chromosome system in Tetragnathidae, N. cruentata seems to possess 2n = 24 = 22 + X1X2 in the males, and 2n = 26 = 22 + X1X1X2X2 in the females. The pairs 1, 2 and 3 showed NORs which are coincident with the negative heteropycnotic patterns. Using the C-banding technique, the pericentromeric region of the chromosomes revealed small quantity or even absence of constitutive heterochromatin, differing of the C-banding pattern described in other species of spiders. In N. cruentata the fluorochromes DAPI/DA, DAPI/MM and CMA3/DA revealed that the constitutive heterochromatin is rich in AT bases and the NORs possess repetitive sequences of GC bases.     

Heterochromatin variation and sex chromosome polymorphism in Nesokia indica: a population study

Nesokia indica, the Indian mole rat, exhibits extensive variability (polymorphism) for the constitutive heterochromatin of the X and Y chromosomes. These polymorphic X and Y types range from a large metacentric chromosome to a small acrocentric one and occur in different frequencies in the population. On the assumption that there is random mating among individuals carrying these various X and Y chromosomes, the population shows Hardy-Weinberg proportions for the genotypes. However, notwithstanding the partial or total loss of constitutive heterochromatin of the X and Y chromosomes in a few individuals, its retention in most of the animals seems obligatory to the population at large. Hence, we suggest that the C-heterochromatin plays a "regulatory" role in the population dynamics of this species.     

5-Azacytidine- and Hoechst-induced aneuploidy in Indian muntjac

Hoechst 33258 (bis-benzimidazole) and 5-azacytidine (5-AC) cause decondensation of the pericentric heterochromatin in mouse and aberrations in the sequence of centromere separation apparently by different mechanisms. We treated the male Indian muntjac cells (2n=7), which do not undergo decondensation of the pericentric heterochromatin, to study if these chemicals would result in induction of aneuploidy limited to the Y(2) chromosome. This paper reports that both agents result in aneuploidy primarily limited to one chromosome, the Y(2). It is likely that other chromosomes are not tolerated in aneuploid condition because every chromosome carries some household genes including those essential for mitotic progression. The loss/gain of the Y(2) chromosome is tolerated because it is the smallest chromosome and is almost entirely composed of constitutive heterochromatin. Since Indian muntjac has only three pairs of large chromosomes comprising its basic genome, which can be clearly viewed under high dry objective, these cells are very suitable for the preliminary analysis of aneuploidy-inducing ability of various chemicals.     

The karyotype of Adenomera diptyx (Boettger 1885) (Anura, Leptodactylidae) from northeastern Argentina

In this work we analyzed the karyotype of five populations of Adenomera diptyx from Argentina after conventional staining, Ag-NOR and C-banding. All specimens presented 2n = 26 and FN = 34. The karyotype was formed by three submetacentric, one metacentric and nine telocentric pairs. Silver staining revealed that the NOR was located on a secondary constriction in pair 7. C- banding evidenced constitutive heterochromatin at the pericentromeric region of all chromosomes. The karyotype of A. diptyx was similar to that of A. hylaedactyla (2n = 26, FN = 34) and different from that of A. andreae (2n = 26, FN = 40) in the fundamental number and secondary constriction position. It also differed from the karyotypes of A. marmorata (2n = 24, FN = 34 and 36) and of A. aff. bokermanni (2n = 23, FN = 34) in diploid number. Until a comprehensive cytogenetic analysis of all the species of the genus is performed, their chromosome evolution will remain poorly understood.     

Cytogenetic analysis of Astylus antis (Perty, 1830) (Coleoptera, Melyridae): Karyotype, heterochromatin and location of ribosomal genes

Cytogenetic analysis of Astylus antis using mitotic and meiotic cells was performed to characterize the haploid and diploid numbers, sex determination system, chromosome morphology, constitutive heterochromatin distribution pattern and chromosomes carrying nucleolus organizer regions (NORs). Analysis of spermatogonial metaphase cells revealed the diploid number 2n = 18, with mostly metacentric chromosomes. Metaphase I cells exhibited 2n = 8II+Xyp and a parachute configuration of the sex chromosomes. Spermatogonial metaphase cells submitted to C-banding showed the presence of small dots of constitutive heterochromatin in the centromeric regions of nearly all the autosomes and on the short arm of the X chromosome (Xp), as well as an additional band on one of the arms of pair 1. Mitotic cells submitted to double staining with base-specific fluorochromes (DAPI-CMA(3) ) revealed no regions rich in A+T or G+C sequences. Analysis of spermatogonial mitotic cells after sequential Giemsa/AgNO (3) staining did not reveal any specific mark on the chromosomes. Meiotic metaphase I cells stained with silver nitrate revealed a strong impregnation associated to the sex chromosomes, and in situ hybridization with an 18S rDNA probe showed ribosomal cistrons in an autosomal bivalent.     

Autosomal and sex chromosomal polymorphisms with multiple rearrangements and a new karyotype in the genus Rhipidomys (Sigmodontinae, Rodentia)

Two diploid numbers and five karyomorphs were found in ten specimens of Rhipidomys (Sigmodontinae, Rodentia) from three states in Brazil: 2n = 50 from Amazonas, and 2n = 44 from Mato Grosso and Bahia. CBG, GTG, and RBG-banding and Ag-NOR analyses were performed, as well as fluorescence in situ: hybridization with (T2AG3)7 probes. The new diploid number of 2n = 50 was associated with two different fundamental numbers (FN = 71 and 72) as a result of pericentric inversions and addition/deletion of constitutive heterochromatin. The samples from two localities (Aripuan? and Vila Rica) in the state of Mato Grosso shared 2n = 44 and FN = 52, but their karyotypes differed because of pericentric inversions. Although the single specimen from Bahia had the same diploid number as the samples from Mato Grosso, its karyotype and FN were completely distinctive. Karyological comparison of GTG-banding patterns revealed total homology between the karyotypes of the specimens from Bahia and Mato Grosso, implying the occurrence of 14 autosomal pericentric inversions. Homologies between ten of the autosomes in the karyotypes with 2n = 50 (FN = 72) and 2n = 44 (from Vila Rica, MT) were demonstrated. The differentiation between 2n = 44 and 2n = 50 involved five pericentric inversions, addition/deletion of constitutive heterochromatin in both autosomes and sex chromosomes, at least one Robertsonian rearrangement and other not detected rearrangements. Despite the remarkable number of rearrangements, interstitial telomeric sites (ITS) were not detected. Sex chromosomes also exhibited polymorphism in size and morphology.     

Chromosomes and DNA of Mus

Using C-banded preparations of Mus dunni it is possible to study the behavior of constitutive heterochromatin in early stages of meiotic prophase. The X and the Y chromosomes, both of which contain a large amount of heterochromatin, lie apart in leptotene but move toward each other during zygotene. They then form the sex vesicle at late zygotene. In autosomes zygotene pairing appears to start from the telomeric ends. The centromere of the Y chromosome associates end-to-end with the terminal end of the long arm of the X chromosome. The autosomal heterochromatic short arms show forked morphology in certain bivalents at pachytene, suggesting probable incomplete synapsis.     

Synaptonemal complexes in Gerbillidae: probable role of intercalated heterochromatin in gonosome-autosome translocations

A study of sex chromosomes and synaptonemal complexes in male specimens of Gerbillus chiesmani, G. nigeriae, G. hoogstrali, and Taterillus pygargus is reported. In each of these Gerbillidae species there are two or three translocations of autosomes with X and Y chromosomes. Analysis of mitotic chromosomes consistently shows the presence of constitutive heterochromatin on the der t(X;autosome) at the X-autosome junction and on the der t(Y;autosome). Analysis of the synaptonemal complexes shows the existence of an unusual structure, lightly stained, at the X-autosome junction and at the Y-autosome junction, which is probably heterochromatic in nature, thus corresponding to the mitotic patterns. This heterochromatin separates the autosomal and gonosomal segments, which behave independently and normally. By analogy with findings from humans and other mammals, a general hypothesis is proposed on the role of intercalated heterochromatin between translocated gonosomes and autosomes. This hypothesis explains why the pathological consequences of these translocations may be very different in males and females. The role of intercalated heterochromatin would be to avoid the pathological consequences of gonosome-autosome translocations resulting from inactivation of the sex chromosomes in female somatic cells and male germinal cells.     

Constitutive heterochromatin variation in two species of rattus with apparently similar karyotypes

Rattus blanfordi and R. cutchicus medius both have a chromosome complement of 2n=36 and all chromosomes except the submetacentric Y of R. blanfordi are acrocentric. The apparently similar karyotypes of the two species, however, show variations in the nature and quantity of C-band-positive constitutive heterochromatin (C-heterochromatin) as revealed by C- and G-banding and Hoechst 33258 fluorescence. R. blanfordi with large-sized X and Y chromosomes and conspicuously larger centromeric heterochromatin in all the autosomes as compared to that of R. cutchicus medius has much more C-heterochromatin in its genome than the latter. The variation in the quantity of C-heterochromatin has been accomplished without altering the morphology of the acrocentric chromosomes unlike other mammals in which variations have been reported to result generally in the addition or deletion of a totally heterochromatic second arm.     

白颊长臂猿染色体的研究

本文对我国云南南部的白须长臂猿（Ｈ．ｌｅｕｃｏｇｅｎｙｓ）染色体的Ｇ带、Ｃ带、晚复制带及Ａｇ－ＮＯＲｓ进行了较为详细的研究。它的２ｎ＝５２,核型公式为４４（Ｍ或ＳＭ）＋６（Ａ）,ＸＹ（Ｍ,Ａ）。Ｃ带表明一些染色体着丝点Ｃ带弱化;有的染色体出现插入的和端位的Ｃ带;Ｘ染色体两臂有端位Ｃ带,Ｙ染色体是Ｃ带阳性和晚复制的。Ａｇ－ＮＯＲｓ的数目,雌体有４个,雄体有５个,Ｙ染色体上具ＮＯＲ。本文对白颊长臂猿与其它长臂猿间的亲缘关系、核型进化的可能途径进行了讨论。     

Silver staining of the centromeric area of acrocentric and Robertsonian metacentric mouse chromosomes

A silver stain (Kt) technique was used to analyze the centromeric area in metacentric chromosomes originating from Robertsonian rearrangements in the mouse. The 2n=40 all-acrocentric mouse karyotype and two Robertsonian-rearranged karyotypes (2n=24 and 2n=26 from Upper Valtellina) were used. The existence was demonstrated of a single centromeric pattern common to metacentric and to acrocentric chromosomes except for the Y, and consisting of two deeply stained dots, one per chromatid. In many cells this technique stains the nucleolar organizers and resolves the paracentromeric constitutive heterochromatin in chromomeres.     

The founder effect theory: quantitative variation and mdg-1 mobile element polymorphism in experimental populations of Drosophila melanogaster

The chromosomes of 14 specimens of the genus Reithrodon from three different localities of Argentina and two localities of Uruguay were studied using G-and C-banding techniques. Specimens of Uruguay showed a karyotype of 2n=28 chromosomes having a large metacentric X, and a telocentric Y chromosome. This karyotype is very similar to that recently described in a sample from southern Brazil, differing only in the nature of the Y chromosome, which is metacentric in the Brazilian form. All specimens from Argentina showed a 2n=34 karyotype, differing from the Brazilian karyotype by two centric fusions, an acquisition of chromosome material, and at least one pericentric inversion, and by the telocentric nature of both the X and the Y chromosomes. G-and C-banding suggest that the metacentric gonosomes in the Brazilian form resulted from a double autosomal-X-Y Robertsonian translocation. The Uruguayan cytotype is interpreted as derived from a hypothetical neo-X/Y₁Y₂ ancestral form by the secondary loss of the Y₁ chromosome. The karyotypic differences between the Brazilian-Uruguayan and the Argentinian forms afford evidence of species differentiation. It is proposed to assign the former to Reithrodon typicus, and the later to R. auritus.     

A new allopatric lineage of the rodent Deltamys (Rodentia: Sigmodontinae) and the chromosomal evolution in Deltamys kempi and Deltamys sp

Deltamys Thomas 1917 is a poorly studied and rarely collected taxon of Akodontini (Sigmodontinae). The single described species, Deltamys kempi (DKE), has a basic karyotype with a diploid number of 2n = 37 in males and 2n = 38 in females, a fundamental number FN = 38 for both sexes, and an X(1)X(1)X(2)X(2)/X(1)X(2)Y sex determination system. Herein, a new allopatric form, Deltamys sp. (DSP), is reported, based on specimens from southern Brazil, with 2n = 40, FN = 40 and XX/XY sex chromosomes. We describe the karyotype and mechanism of chromosomal differentiation between both Deltamys complements. Phylogenetic analyses, based on the complete sequence (1,140 bp) of the mitochondrial cytochrome b gene, grouped Deltamys sp. as sister species to D. kempi, with up to 12% genetic divergence between them. The GTG-banding patterns show complete autosomal correspondence between D. kempi and Deltamys sp. and identify a tandem rearrangement involving DSP7, DSP19 and DKE4 that is responsible for the differences in 2n and FN. Chromosome painting with Akodon paranaensis chromosome 21 (a small metacentric akodont marker) paint revealed total homology with the smallest acrocentric Deltamys sp. chromosome, DSP19. This suggests the occurrence of a pericentric inversion or centromeric shift when compared to other akodontines, with a posterior tandem rearrangement giving rise to DKE4. In DKE, large blocks of pericentromeric constitutive heterochromatin are present on the autosomes and the X, and the Y/autosome has an entirely heterochromatic short arm. In DSP, small heterochromatic blocks are observed on autosomes and X, and the Y is a very small, mostly heterochromatic acrocentric. The cytogenetic analyses suggest that the Deltamys sp. karyotype is ancestral, with the derived condition resulting from a tandem fusion (DSP7 + DSP19) and the Y/autosome translocation giving rise to the multiple sex chromosome system. The autosomal rearrangements, the differences in CBG-banding patterns and Ag-NOR localization, as well as the presence of X(1)X(1)X(2)X(2)/X(1)X(2)Y and XX/XY sex determination mechanisms, possibly acting as a reproductive barrier, and the phylogenetic position within the Deltamys genus, with high genetic divergence, call for a taxonomic review of the genus.     

The creeping vole, Microtus oregoni: karyotype and sex-chromosome differences between two geographical populations

The G-banded karyotype of the creeping vole, Microtus oregoni, prepared from animals trapped in Oregon and Washington, is presented. The two populations had similar autosomal banding patterns but exhibited striking differences in their sex chromosomes. The X chromosome of voles captured in Oregon was 39% longer than that of voles trapped in Washington. The length difference was primarily due to an increase in size of light G-bands, which, in both populations, comprised large segments of the X chromosome. On C-banding, the X chromosome exhibited major blocks of constitutive heterochromatin corresponding to the light G-bands. In contrast, the Y chromosome of the Oregon voles was 24% shorter than that of the Washington voles and lacked the short arm and some terminal bands present in the Washington voles.