Accumulation of Y-specific satellite DNAs during the evolution of <Emphasis Type="Italic">Rumex acetosa</Emphasis> sex chromosomes

The study of the molecular structure of young heteromorphic sex chromosomes of plants has shed light on the evolutionary forces that control the differentiation of the X and Y during the earlier stages of their evolution. We have used the model plant Rumex acetosa, a dioecious species with multiple sex chromosomes, 2n = 12 + XX female and 2n = 12 + XY₁Y₂ male, to analyse the significance of repetitive DNA accumulation during the differentiation of the Y. A bulk segregant analysis (BSA) approach allowed us to identify and isolate random amplified polymorphic DNA (RAPD) markers linked to the sex chromosomes. From a total of 86 RAPD markers in the parents, 6 markers were found to be linked to the Ys and 1 to the X. Two of the Y-linked markers represent two AT-rich satellite DNAs (satDNAs), named RAYSII and RAYSIII, that share about 80% homology, as well as with RAYSI, another satDNA of R. acetosa. Fluorescent in situ hybridisation demonstrated that RAYSII is specific for Y₁, whilst RAYSIII is located in different clusters along Y₁ and Y₂. The two satDNAs were only detected in the genome of the dioecious species with XX/XY₁Y₂ multiple sex chromosome systems in the subgenus Acetosa, but were absent from other dioecious species with an XX/XY system of the subgenera Acetosa or Acetosella, as well as in gynodioecious or hermaphrodite species of the subgenera Acetosa, Rumex and Platypodium. Phylogenetic analysis with different cloned monomers of RAYSII and RAYSIII from both R. acetosa and R. papillaris indicate that these two satDNAs are completely separated from each other, and from RAYSI, in both species. The three Y-specific satDNAs, however, evolved from an ancestral satDNA with repeating units of 120 bp, through intermediate satDNAs of 360 bp. The data therefore support the idea that Y-chromosome differentiation and heterochromatinisation in the Rumex species having a multiple sex chromosome system have occurred by different amplification events from a common ancestral satDNA. Since dioecious species with multiple XX/XY₁Y₂ sex chromosome systems of the section Acetosa appear to have evolved from dioecious species with an XX/XY system, the amplification of tandemly repetitive elements in the Ys of the section Acetosa is a recent evolutionary process that has contributed to an increase in the size and differentiation of the already non-recombining Y chromosomes.     

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.     

Independent Origins of New Sex-Linked Chromosomes in the <Emphasis Type="Italic">melanica</Emphasis> and <Emphasis Type="Italic">robusta</Emphasis> Species Groups of <Emphasis Type="Italic">Drosophila</Emphasis>

Background  

Recent translocations of autosomal regions to the sex chromosomes represent important systems for identifying the evolutionary forces affecting convergent patterns of sex-chromosome heteromorphism. Additions to the sex chromosomes have been reported in the melanica and robusta species groups, two sister clades of Drosophila. The close relationship between these two species groups and the similarity of their rearranged karyotypes motivates this test of alternative hypotheses; the rearranged sex chromosomes in both groups are derived through a common origin, or the rearrangements are derived through at least two independent origins. Here we examine chromosomal arrangement in representatives of the melanica and the robusta species groups and test these alternative hypotheses using a phylogenetic approach.     

New insights on the origin of B chromosomes in <Emphasis Type="Italic">Astyanax scabripinnis</Emphasis> obtained by chromosome painting and FISH

Chromosome painting (CP) with a probe of B chromosome obtained by microdissection and fluorescence in situ hybridization (FISH) with probes of As51 satellite DNA, C _o t−1 DNA, and 18S and 5S rDNA confirmed sharing of some repetitive DNA but not rDNA between A and B chromosomes in the fish Astyanax scabripinnis. Meiotic analysis revealed a pachytene B chromosome bivalent nearly half the size of its mitotic configuration, suggesting a self-pairing of B chromosome arms. Such an isochromosome nature of somatic B chromosome was further evidenced by CP and FISH. All the findings obtained suggest (i) intraspecific origin of B chromosome, and (ii) evolutionary enrichment of repetitive DNA classes, especially those contained in the C _o t−1 and the As51 probes, in B chromosome. However, the precise origin of B chromosome in the present species remains to be elucidated by further molecular cytogenetic analysis because of painting of some A chromosome regions with the B chromosome-derived probe.     

Two new chromodomain-containing proteins that associate with heterochromatin in <Emphasis Type="Italic">Sciara coprophila</Emphasis> chromosomes

We report here the molecular and cytological characterization of two proteins, ScoHET1 and ScoHET2 (for Sciara coprophila heterochromatin), which associate to constitutive heterochromatin in the dipteran S. coprophila. Both proteins, ScoHET1 of 37 kDa and ScoHET2 of 44 kDa, display two chromodomain motifs that contain the conserved residues essential for the recognition of methylated histone H3 at lysine 9. We raised antibodies to analyze the chromosomal location of ScoHET1 and ScoHET2 in somatic and germline cells. In S. coprophila polytene chromosomes, both proteins associate to the pericentromeric regions and to the heterochromatic subterminal bands of the chromosomes. In germinal nuclei, ScoHET1 and ScoHET2 proteins distribute to the heterochromatic regions of the regular chromosome complement and are abundantly present along the heterochromatic germline-limited “L” chromosomes. We investigated histone methylation modifications and found that all heterochromatic regions enriched in ScoHET1/ScoHET2 proteins exhibit high levels of di- and tri-methylated histone H3 at lysine 9. Taken together, our results support that the association of ScoHET1/ScoHET2 to heterochromatin is mediated by histone H3K9 methylation. Using 5-methylcytosine antibodies, we proved the cytological detection of DNA methylation in S. coprophila. From our observations in L germline chromosomes, heterochromatin in S. coprophila is highly enriched in DNA 5-methylcytosine residues. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

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.     

Localization of BAC clones on mitotic chromosomes of <Emphasis Type="Italic">Musa acuminata</Emphasis> using fluorescence <Emphasis Type="Italic">in situ</Emphasis> hybridization

A bacterial artificial chromosome (BAC) library of banana (Musa acuminata) was used to select BAC clones that carry low amounts of repetitive DNA sequences and could be suitable as probes for fluorescence in situ hybridization (FISH) on mitotic metaphase chromosomes. Out of eighty randomly selected BAC clones, only one clone gave a single-locus signal on chromosomes of M. acuminata cv. Calcutta 4. The clone localized on a chromosome pair that carries a cluster of 5S rRNA genes. The remaining BAC clones gave dispersed FISH signals throughout the genome and/or failed to produce any signal. In order to avoid the excessive hybridization of repetitive DNA sequences, we subcloned nineteen BAC clones and selected their ‘low-copy’ subclones. Out of them, one subclone gave specific signal in secondary constriction on one chromosome pair; three subclones were localized into centromeric and peri-centromeric regions of all chromosomes. Other subclones were either localized throughout the banana genome or their use did not result in visible FISH signals. The nucleotide sequence analysis revealed that subclones, which localized on different regions of all chromosomes, contained short fragments of various repetitive DNA sequences. The chromosome-specific BAC clone identified in this work increases the number of useful cytogenetic markers for Musa.     

Giant sex chromosomes retained within the Portuguese lineage of the field vole (<Emphasis Type="Italic">Microtus agrestis</Emphasis>)

The field vole (Microtus agrestis) is characterised by extremely large blocks of heterochromatin on both the X and Y chromosome. Some other Microtus also have blocks of heterochromatin on their sex chromosomes but not as extensive and always of independent origin from the heterochromatic expansion found in M. agrestis. Coupled with evidence of geographic variation in large heterochromatic blocks within other species (e.g. in the western hedgehog Erinaceus europaeus), it might be expected that field voles would show substantial variation in size and disposition of the sex chromosome heterochromatin. In fact, only minor variation has been described up to now. Those studies conducted previously were largely on field voles from central and northern Europe. Here, we describe the karyotype of field voles from Portugal, of interest because recent molecular studies have shown field voles from western Iberia to be a separate evolutionary unit that might be considered a cryptic species, distinct from populations further to the east. The two Portuguese field voles (one female, one male) that we examined also had essentially the same karyotype as seen in other field voles, including the giant sex chromosomes, but with small differences in the structure of the Y chromosome from that described previously. The finding that field voles throughout Europe show relatively little variation in their giant sex chromosomes is consistent with molecular data which suggest a recent origin for this complex of species/near-species.     

Evolutionary dynamics of heterochromatin in the genome of <Emphasis Type="Italic">Dichotomius</Emphasis> beetles based on chromosomal analysis

We comparatively analyzed six Dichotomius species (Coleoptera: Scarabainae) through cytogenetic methods and mitochondrial genes sequencing in the aim to identify patterns of chromosomal evolution and heterochromatin differentiation in the group. The chromosomal data were accessed through the classical analysis of heterochromatin and mapping of high and moderately repeated DNAs (C ₀ t-1 DNA fraction). Mitochondrial data were obtained from nucleotide sequences of the cytochrome oxidase I (COI) and 16S rRNA genes. The heterochromatin distribution was conserved but revealed variability in the base pair richness and repetitive DNA content, and an intense turnover of heterochromatic associated sequences seems to have occurred during Dichotomius speciation. Specifically for D. bos, an interesting pattern was observed, indicating apparently the presence of heterochromatic sequences composed of low copy-number sequences. Moreover, highly conserved terminal/sub-terminal sequences that could act as a telomeric or telomere-associated DNA were observed. The heterochromatin diversification patterns observed in Dichotomius were not accomplished by the diversification of the species studied, which may be a consequence of the intense dynamics that drive the evolution of repeated DNA clusters in the genome. Finally our findings also suggest that the use of C ₀ t-1 DNA fraction represents a powerful, inexpensive and not time consuming tool to be applied in understanding heterochromatin and repetitive DNA organization.     

XY<Subscript>1</Subscript>Y<Subscript>2</Subscript> chromosome system in <Emphasis Type="Italic">Salinomys delicatus</Emphasis> (Rodentia,Cricetidae)

Salinomys delicatus is considered a rare species due to its restricted and patchy distribution, poor records and low abundances. It is also the phyllotine with the lowest known diploid chromosome number (2n = 18), however its sex chromosome system has never been described. Here, we studied the chromosomes of six females and three males with bands G, C, DAPI/CMA₃ and meiosis. In males, the chromosome number was 2n = 19, with one large metacentric X-chromosome and two medium-sized acrocentrics absent in females. The karyotype of females was the same as previously described (2n = 18, FN = 32), with X-chromosomes being metacentric and the largest elements of the complement. In males, the two acrocentrics and the large metacentric form a trivalent in meiotic prophase. This indicates that S. delicatus has XY₁Y₂ sex chromosomes, which is confirmed by G and DAPI bands. Constitutive heterochromatin (CH) is restricted to small pericentromeric blocks in all chromosomes. The X-chromosome shows the largest block of centromeric CH, which could favor the establishment of this X-autosome translocation. This sex chromosome system is rare in mammals and, compared with other phyllotine rodents, S. delicatus seems to have undergone a major chromosome restructuring during its karyotypic evolution.     

Tracking the evolutionary pathway of sex chromosomes among fishes: characterizing the unique XX/XY<Subscript>1</Subscript>Y<Subscript>2</Subscript> system in <Emphasis Type="Italic">Hoplias malabaricus</Emphasis> (Teleostei,Characiformes)

The Neotropical fish, Hoplias malabaricus, is one of the most cytogenetically studied fish taxon with seven distinct karyomorphs (A–G) comprising varying degrees of sex chromosome differentiation, ranging from homomorphic to highly differentiated simple and multiple sex chromosomes. Therefore, this fish offers a unique opportunity to track evolutionary mechanisms standing behind the sex chromosome evolution and differentiation. Here, we focused on a high-resolution cytogenetic characterization of the unique XX/XY₁Y₂ multiple sex chromosome system found in one of its karyomorphs (G). For this, we applied a suite of conventional (Giemsa-staining, C-banding) and molecular cytogenetic approaches, including fluorescence in situ hybridization FISH (with 5S and 18S rDNAs, 10 microsatellite motifs and telomeric (TTAGGG) _n sequences as probes), comparative genomic hybridization (CGH), and whole chromosome painting (WCP). In addition, we performed comparative analyses with other Erythrinidae species to discover the evolutionary origin of this unique karyomorph G-specific XY₁Y₂ multiple sex chromosome system. WCP experiments confirmed the homology between these multiple sex chromosomes and the nascent XX/XY sex system found in the karyomorph F, but disproved a homology with those of karyomorphs A–D and other closely related species. Besides, the putative origin of such XY₁Y₂ system by rearrangements of several chromosome pairs from an ancestral karyotype was also highlighted. In addition, clear identification of a male-specific region on the Y₁ chromosome suggested a differential pattern of repetitive sequences accumulation. The present data suggested the origin of this unique XY₁Y₂ sex system, revealing evidences for the high level of plasticity of sex chromosome differentiation within the Erythrinidae.     

Repetitive DNA in the automictic fungus <Emphasis Type="Italic">Microbotryum violaceum</Emphasis>

The small genomes of fungi are expected to have little repetitive content other than rDNA genes. Moreover, among asexual or highly selfing lineages, the diversity of repetitive elements is also expected to be very low. However, in the automictic fungus Microbotryum violaceum, a very large proportion of random DNA fragments from the autosomes and the fungal sex chromosomes are repetitive in nature, either as retrotransposon or helicase sequences. Among the retrotransposon sequences, examples were found from each major kind of elements, including copia, gypsy, and non-LTR sequences. The most numerous were copia-like elements, which are believed to be rare in fungi, particularly among basidiomycetes. The many helicase sequences appear to belong to the recently discovered Helitron type of transposable elements. Also, sequences that could not be identified as a known type of gene were also very repetitive within the database of random fragments from M. violaceum. The differentiated pair of fungal sex chromosomes and suppression of recombination may be the major forces determining the highly repetitive content in the small genome of M. violaceum.     

Development of a sex-specific molecular marker for Japanese hop <Emphasis Type="Italic">Humulus Japonicus</Emphasis> Siebold &; Zucc

Japanese hop (Humulus japonicus Siebold & Zucc.) is a dioecious plant and a suitable model for studying the XX/XY₁Y₂ system of sex chromosomes. To develop a sex-specific marker, 12 RAPD and 36 ISSR markers were analyzed on the basis of pools of male and female plants identified after flowering. We were the first to identify ISSR marker K-16, which manifested stable amplification of an approximately 300-bp fragment in male plants and the absence of amplification in female plants in the populations examined. Marker effectiveness was confirmed in several Japanese hop populations of different origin.     

Karyotypic and molecular polymorphisms in <Emphasis Type="Italic">Ctenomys</Emphasis><Emphasis Type="Italic">torquatus</Emphasis> (Rodentia: Ctenomyidae): taxonomic considerations

The rodent genus Ctenomys (tuco-tucos) comprises more than 60 described species, and shows extraordinary inter- and intraspecific karyotypic variation. The most widely distributed species of Ctenomys in Brazil is C. torquatus. Although several cytogenetic studies have been done, the karyotypic variability of this species is still poorly known. In this paper we report two new diploid numbers for C. torquatus: 2n = 40 and 2n = 42, both showing AN = 72. The new distribution limits of C. torquatus here reported include localities in the southern, central and western parts of Rio Grande do Sul (RS) State in southern Brazil. The phylogenetic relationship between C. torquatus from Alegrete, RS, and Ctenomys sp. from Corrientes, Argentina, is described by means of mtDNA cytochrome b analysis. Although both entities share similar karyotypes and sperm morphology, these two species are not phylogenetically close.     

Chromosome banding and essential oils composition of Brazilian accessions of <Emphasis Type="Italic">Lippia alba</Emphasis> (Verbenaceae)

The essential oil components and a karyotypic analysis of five Lippia alba (Verbenaceae) accessions from Brazil were performed with the objective of investigating the variation among different populations. The chemistry analysis allowed the grouping of the accessions in two main chemotypes: neral chemotype (LaCat, LaJF and LaRJ) and linalool chemotype (LaGua and LaVC). However, large karyotypic differences, verified by different chromosome banding techniques, were not detected. The results presented the same chromosome number for all accessions (2n = 30) with 10 metacentric chromosomes and 5 submetacentric. The chromosome banding showed great blocks of constitutive heterochromatin (C-bands) around the centromeric region, which was rich in AT bases (DAPI₊), while the CMA bands were observed only in terminal regions of six chromosomes. Through Ag-NOR techniques, only two active pairs of NORs were detected on the three pairs of secondary constrictions (the NOR activity is discussed). This work relates the pattern of heterochromatin for Lippia alba for the first time.     

Male gametophyte development and two different DNA classes of pollen grains in <Emphasis Type="Italic">Rumex acetosa</Emphasis> L., a plant with an XX/XY<Subscript>1</Subscript>Y<Subscript>2</Subscript> sex chromosome system and a female-biased sex ratio

Female-biased sex ratio is an interesting phenomenon observed in Rumex acetosa, a dioecious plant with an XX/XY₁Y₂ sex chromosome system. Previous authors have suggested that the biased sex ratio in this species is conditioned not only postzygotically (sex-differential sporophytic mortality) but also prezygotically, because the sex ratio of seeds is also female-biased, although to a lesser extent than the sex ratio of flowering plants. The mechanisms underlying female bias in Rumex seeds are only poorly understood. To gain more knowledge of them, we analysed male gametophyte development and used flow cytometry to determine the frequency of female-determining (n = 7, A + X) and male-determining (n = 8, A + Y₁Y₂) pollen grains in anthers. Embryological studies showed a regular course of male gametophyte development in R. acetosa. There were no signs of degeneration of microspores or disturbances in pollen divisions (irregular nuclei, micronuclei, delayed chromosomes and anaphase bridges). The Alexander test revealed only 1.6% nonviable pollen grains within anthers. All mature pollen grains were uniformly equipped with starch granules. The two sexes were shown to substantially differ in nuclear 2C DNA amount in somatic tissues (7.00 pg in 2A + XX females and 7.50 pg in 2A + XY₁Y₂ males), and two clearly different DNA classes of mature pollen grains, with lower and with higher DNA amounts (16.8% difference) were found. Most probably the grains with the lower DNA amount possess seven chromosomes, and grains with the higher DNA amount eight of them. The quantitative ratio of these grains in anthers at anthesis was 1:1.2, very close to the sex ratio of seeds observed by the majority of previous authors. All these observations support the opinion that the sex-ratio bias in Rumex is determined prezygotically to some extent.     

Multicolor FISH mapping of the dioecious model plant,<Emphasis Type="Italic"> Silene latifolia</Emphasis>

Silene latifolia is a key plant model in the study of sex determination and sex chromosome evolution. Current studies have been based on genetic mapping of the sequences linked to sex chromosomes with analysis of their characters and relative positions on the X and Y chromosomes. Until recently, very few DNA sequences have been physically mapped to the sex chromosomes of S. latifolia. We have carried out multicolor fluorescent in situ hybridization (FISH) analysis of S. latifolia chromosomes based on the presence and intensity of FISH signals on individual chromosomes. We have generated new markers by constructing and screening a sample bacterial artificial chromosome (BAC) library for appropriate FISH probes. Five newly isolated BAC clones yielded discrete signals on the chromosomes: two were specific for one autosome pair and three hybridized preferentially to the sex chromosomes. We present the FISH hybridization patterns of these five BAC inserts together with previously described repetitive sequences (X-43.1, 25S rDNA and 5S rDNA) and use them to analyze the S. latifolia karyotype. The autosomes of S. latifolia are difficult to distinguish based on their relative arm lengths. Using one BAC insert and the three repetitive sequences, we have constructed a standard FISH karyotype that can be used to distinguish all autosome pairs. We also analyze the hybridization patterns of these sequences on the sex chromosomes and discuss the utility of the karyotype mapping strategy presented to study sex chromosome evolution and Y chromosome degeneration.Communicated by J.S. Heslop-Harrison     

Comparative molecular cytogenetic analysis of three <Emphasis Type="Italic">Leuciscus</Emphasis> species (Pisces,Cyprinidae) using chromosome banding and FISH with rDNA

A comparative molecular cytogenetic analysis was performed on three species of the genus Leuciscus viz. ide L. idus, chub L. cephalus and dace L. leuciscus distributed in Poland, using C-, Ag- and chromomycin A₃ (CMA₃)-stainings and fluorescence in situ hybridization (FISH) with 5.8S + 28S rDNA as a probe. Although the three species examined shared 2n = 50 chromosomes and the largest acrocentric chromosome pair in the complement, they were characterized with karyotypic differences in terms of the number of uni- and biarmed chromosomes and the localization of nucleolar organizer regions (NORs) revealed by Ag-staining and FISH. L. idus and L. cephalus showed the rDNA sites on the long arms of one submetacentric (SM) chromosome pair and on the short arms of one subtelocentric (ST) chromosome pair, respectively. These NORs were CMA₃-positive, GC-rich and C-positive heterochromatic sites in both species. Such chromosome banding features were also true for four NORs localizing on one of each SM and ST pair in L. leuciscus, but considerable numerical NOR polymorphism became apparent with Ag-staining and FISH due to a different combination of these NOR-bearing SMs and STs in this dace. The present results indicate that the molecular cytogenetic analysis applied herein may become useful to elucidate the karyotype evolution and phylogenetic relationships among the species in the genus Leuciscus and other related groups.     

X-linked heterochromatin distribution in the holocentric chromosomes of the green apple aphid <Emphasis Type="Italic">Aphis pomi</Emphasis>

Chromatin organization in the holocentric chromosomes of the green apple aphid Aphis pomi has been investigated at a cytological level after C-banding, NOR, Giemsa, fluorochrome staining and fluorescent in situ hybridization (FISH). C-banding technique showed that heterochromatic bands are exclusively located on X chromosomes. This data represents a peculiar feature that clearly contradicts the equilocal distribution of heterochromatin typical of monocentric chromosomes. Moreover, silver staining and FISH carried out with a 28S rDNA probe localized rDNA genes on one telomere of each X chromosome; CMA₃ staining reveals that these silver positive telomeres are the only GC-rich regions among A. pomi heterochromatin, whereas all other C-positive bands are DAPI positive thus containing AT-rich DNA.     

Genetic variation in populations of <Emphasis Type="Italic">Allothrombium pulvinum</Emphasis> (Acari: Trombidiidae) from Northern Iran revealed by mitochondrial <Emphasis Type="Italic">cox</Emphasis>I and nuclear rDNA <Emphasis Type="Italic">ITS</Emphasis>2 sequences

Allothrombium pulvinum Ewing is a common natural enemy of aphids and some other arthropods. So far, there are no studies that have addressed genetic variation of this predatory mite. We investigated genetic variation of A. pulvinum across its whole known range in Iran. A 410 bp portion of the mitochondrial cytochrome c oxidase subunit I gene (coxI) and 797–802 bp portion of the internal transcribed spacer 2 of rDNA (ITS2) were sequenced for 55 individuals from 11 populations, resulting in 12 and 26 haplotypes, respectively. In the coxI region, haplotype and nucleotide diversities varied among populations from 0.00 to 0.90 and from 0.0000 to 0.0110, respectively. In the ITS2 region they varied from 0.20 to 0.91 and from 0.0006 to 0.0023, respectively. For both gene regions the highest haplotype and nucleotide diversities were detected in population Mahmoud Abad from northern Iran. Statistically significant population differentiation (F _ST) was detected in most pair-wise population comparisons. The results of population differentiation for both gene regions were generally congruent indicating that A. pulvinum from Iran consists of genetically different populations. This suggests that A. pulvinum comprises at least two geographically distinct populations or even more than one species. This study is an initial step towards understanding genetic variation of A. pulvinum, a taxon for which little molecular information is available. More intensive sampling and analysis of additional DNA regions are necessary for more detailed classification of this taxon.