C-heterochromatin and extra (B) chromosome distribution in six species of the Nabis (Heteroptera, Nabidae) with the modal male karyotype 2n = 16 + XY

The basic male karyotype of the six Nabis species (Heteroptera, Nabidae) is confirmed as being 2n=16+XY. The chromosomes are holokinetic while male meiosis is achiasmatic. The sex chromosomes undergo postreduction and in second metaphase show distance pairing, registered in all nabid species examined so far. Using C-banding technique for the first time in the family Nabidae, the heterochromatin was revealed on chromosomes of six species. The species showed different amount and distribution of C-heterochromatin. Only in Nabis (Dolichonabis) limbatus did the C-bands distribution make possible the identification of every chromosome pair in the karyotype. In other species, C-bands were found in some of the autosomes and the X, localized either interstitially or at telomeres. Only the Y usually showed relative stability ofthe C-banding pattern. In four of six species, extra (B) chromosomes were observed and their behaviour in meiosis described.     

Comparative Mitogenomic Analysis of Damsel Bugs Representing Three Tribes in the Family Nabidae (Insecta: Hemiptera)

Background

Nabidae, a family of predatory heteropterans, includes two subfamilies and five tribes. We previously reported the complete mitogenome of Alloeorhynchus bakeri, a representative of the tribe Prostemmatini in the subfamily Prostemmatinae. To gain a better understanding of architecture and evolution of mitogenome in Nabidae, mitogenomes of five species representing two tribes (Gorpini and Nabini) in the subfamily Nabinae were sequenced, and a comparative mitogenomic analysis of three nabid tribes in two subfamilies was carried out.

Methodology/Principal Findings

Nabid mitogenomes share a similar nucleotide composition and base bias, except for the control region, where differences are observed at the subfamily level. In addition, the pattern of codon usage is influenced by the GC content and consistent with the standard invertebrate mitochondrial genetic code and the preference for A+T-rich codons. The comparison among orthologous protein-coding genes shows that different genes have been subject to different rates of molecular evolution correlated with the GC content. The stems and anticodon loops of tRNAs are extremely conserved, and the nucleotide substitutions are largely restricted to TψC and DHU loops and extra arms, with insertion-deletion polymorphisms. Comparative analysis shows similar rates of substitution between the two rRNAs. Long non-coding regions are observed in most Gorpini and Nabini mtDNAs in-between trnI-trnQ and/or trnS2-nad1. The lone exception, Nabis apicalis, however, has lost three tRNAs. Overall, phylogenetic analysis using mitogenomic data is consistent with phylogenies constructed mainly form morphological traits.

Conclusions/Significance

This comparative mitogenomic analysis sheds light on the architecture and evolution of mitogenomes in the family Nabidae. Nucleotide diversity and mitogenomic traits are phylogenetically informative at subfamily level. Furthermore, inclusion of a broader range of samples representing various taxonomic levels is critical for the understanding of mitogenomic evolution in damsel bugs.     

Parallel occurrence of asynaptic sex chromosomes in gray voles (Microtus Schrank, 1798)

In many eutherian species, pairing and recombination of X and Y chromosomes are indispensable for normal meiotic progression and correct segregation of sex chromosomes. The rodent subfamily Arvicolinae provides an interesting exception. The majority of arvicoline species with asynaptic sex chromosomes belong to the genus Microtus sensu lato. However, some vole species of the genus Microtus and other genera display normal X-Y pairing in meiosis. These observations indicate that synaptic condition was typical for the common ancestor of all voles, but the gaps in taxonomic sampling makes impossible to identify a lineage or lineages, in which the asynapsis occurred. The methods of electron and fluorescent microscopy were used to study the synapsis of sex chromosomes in males of some additional species of the subfamily Arvicolinae. This extended taxonomic list allowed us to identify asynaptic species in every large lineage of the tribe Microtini. Apparently, the ability of sex chromosomes to pair and recombine in male meiosis was lost in arvicoline evolution for at least three times independently. Our results indirectly suggest the unnecessity of sex chromosome pairing in male meiosis of arvicoline rodents, and presence of alternate molecular mechanism of sex chromosome segregation in this large mammalian tribe.     

Karyotype of Brazilian Anopheles albitarsis sensu lato (Diptera:Culicidae)

Anopheles (Nyssorhynchus) albitarsis sensu lato is an important malaria vector in Brazil, especially in the Brazilian Amazon region. Chromosome preparations of fourth-instar larvae of A. albitarsis from Iranduba and Coari (AM) and Ilha Comprida (SP) were analyzed for karyotype determination and to improve cytogenetic identification of this species. Anopheles albitarsis possesses 2n = 6 chromosomes, with two pairs (submetacentric and metacentric) of autosomes and one pair of sex chromosomes, with X-Y dimorphism. The sex pair is homomorphic and acrocentric in females and heteromorphic in males, with a punctiform Y chromosome. Somatic pairing was detected in the prometaphase and metaphase chromosomes of the three A. albitarsis populations. Apparently, sex chromosome evolution in the Culicidae does not function as does evolution in the Culicidae, since it occurs in the subfamily Anophelinae, which possesses heteromorphic sex chromosomes and is regarded as primitive, based on several criteria. These karyotype data on the albitarsis complex reinforce the hypothesis that sex chromosome evolution in the subfamily Anophelinae is conserved, and the variation revealed in the mean size of chromosomes in three populations indicates that selective pressure in these populations is occurring only at a genetic level.     

Karyotypic evolution and phylogeny of Mexican Passalidae (Coleoptera: Polyphaga: Scarabaeoidea)

The chromosomes of 26 taxa from Mexico of the tribes Passalini (three species) and Proculini (23 species) have been studied, increasing the karyotypically known species of the family Passalidae to 56. Karyotypic dynamism is high since the diploid number varies from 18 to 44 in the tribe Proculini. and from 25 to 31 in the tribe Passalini. In addition, supernumerary chromosomes, chromosome heteromorphism, translocations and possible sex multivalents have been found. These results contrast with the numerical conservatism found in related families of the superfamily Scarabaeoidea. However, both tribes are conservative with regard to sex determination, as all species of Proculini have male XY chromosomes whereas species of the Passalini have male XO chromosomes. It is postulated that differences in patterns shown by these two tribes are mainly due to population structure, because many species of Proculini are endemic to restricted areas of Meso and South America, favouring the settlement of karyotypic changes, whereas species of Passalini are distributed over large areas in the lowlands. It is also postulated that the ancestral karyotypic formula of the family is close to 12–14 pairs of autosomes although the ancestral male sex determination may be either XY or XO. At present only a weak relationship between morphological and karyotypic evolution has been found, which together with the marked numerical variability found within and between genera make it difficult to obtain phylogenetic conclusions from karyotypic results.     

The species and chromosomal distribution of the centromeric alpha-satellite I sequence from sheep in the tribe Caprini and other Bovidae

The evolution of chromosomes in species in the family Bovidae includes fusion and fission of chromosome arms (giving different numbers of acrocentric and metacentric chromosomes with a relatively conserved total number of arms) and evolution in both DNA sequence and copy number of the pericentromeric alpha-satellite I repetitive DNA sequence. Here, a probe representing the sheep alpha-satellite I sequence was isolated and hybridized to genomic DNA digests and metaphase chromosomes from various Bovidae species. The probe was highly homologous to the centromeric sequence in all species in the tribe Caprini, including sheep (Ovis aries), goat (Capra hircus) and the aoudad or Barbary sheep (Amnotragus lervia), but showed no detectable hybridization to the alpha-satellite I sequence present in the tribe Bovini and at most very weak to species in the tribes Hippotragini, Alcelaphini or Aepycerotini. The sex chromosomes of sheep, goat and aoudad did not contain detectable alpha-satellite I sequence; in sheep, one of the three metacentric autosomal chromosomes does not carry the sequence, while in aoudad, it is essentially absent in three large autosomal pairs as well as the large metacentric chromosome pair. The satellite probes can be used as robust chromosome and karyotype markers of evolution among tribes and increase the resolution of the evolutionary tree at the base of the Artiodactyla.     

Distribution and Evolution of Repeated Sequences in Genomes of Triatominae (Hemiptera-Reduviidae) Inferred from Genomic In Situ Hybridization

The subfamily Triatominae, vectors of Chagas disease, comprises 140 species characterized by a highly homogeneous chromosome number. We analyzed the chromosomal distribution and evolution of repeated sequences in Triatominae genomes by Genomic in situ Hybridization using Triatoma delpontei and Triatoma infestans genomic DNAs as probes. Hybridizations were performed on their own chromosomes and on nine species included in six genera from the two main tribes: Triatomini and Rhodniini. Genomic probes clearly generate two different hybridization patterns, dispersed or accumulated in specific regions or chromosomes. The three used probes generate the same hybridization pattern in each species. However, these patterns are species-specific. In closely related species, the probes strongly hybridized in the autosomal heterochromatic regions, resembling C-banding and DAPI patterns. However, in more distant species these co-localizations are not observed. The heterochromatic Y chromosome is constituted by highly repeated sequences, which is conserved among 10 species of Triatomini tribe suggesting be an ancestral character for this group. However, the Y chromosome in Rhodniini tribe is markedly different, supporting the early evolutionary dichotomy between both tribes. In some species, sex chromosomes and autosomes shared repeated sequences, suggesting meiotic chromatin exchanges among these heterologous chromosomes. Our GISH analyses enabled us to acquire not only reliable information about autosomal repeated sequences distribution but also an insight into sex chromosome evolution in Triatominae. Furthermore, the differentiation obtained by GISH might be a valuable marker to establish phylogenetic relationships and to test the controversial origin of the Triatominae subfamily.     

Cytogenetics of the true bug infraorder Cimicomorpha (Hemiptera, Heteroptera): a review

The Cimicomorpha is one of the largest and highly diversified infraorders of the Heteroptera. This group is also highly diversified cytogenetically and demonstrates a number of unusual cytogenetic characters such as holokinetic chromosomes; m-chromosomes; multiple sex chromosome systems; post-reduction of sex chromosomes in meiosis; variation in the presence/absence of chiasmata in spermatogenesis; different types of achiasmate meiosis. We present here a review of essential cytogenetic characters of the Cimicomorpha and outline the chief objectives and goals of future investigations in the field.     

Karyotype and male pre-reductional meiosis of the sharpshooter Tapajosa rubromarginata (Hemiptera: Cicadellidae)

Cicadellidae in one of the best represented families in the Neotropical Region, and the tribe Proconiini comprises most of the xylem-feeding insects, including the majority of the known vectors of xylem-born phytopathogenic organisms. The cytogenetics of the Proconiini remains largely unexplored. We studied males of Tapajosa rubromarginata (Signoret) collected at El Manantial (Tucumán, Argentina) on native spontaneous vegetation where Sorghum halepense predominates. Conventional cytogenetic techniques were used in order to describe the karyotype and male meiosis of this sharpshooter. T. rubromarginata has a male karyological formula of 2n = 21 and a sex chromosome system XO:XX (male:female). The chromosomes do not have a primary constriction, being holokinetic and the meiosis is pre-reductional, showing similar behavior both for autosomes and sex chromosomes during anaphase I. For this stage, chromosomes are parallel to the acromatic spindle with kinetic activities in the telomeres. They segregate reductionally in the anaphase I, and towards the equator during the second division of the meiosis. This is the first contribution to cytogenetic aspects on proconines sharpshooters, particularly on this economic relevant Auchenorrhyncha species.     

Cytogenetic and molecular evaluation of centromere-associated DNA sequences from a marsupial (Macropodidae: Macropus rufogriseus) X chromosome

The constitution of the centromeric portions of the sex chromosomes of the red-necked wallaby, Macropus rufogriseus (family Macropodidae, subfamily Macropodinae), was investigated to develop an overview of the sequence composition of centromeres in a marsupial genome that harbors large amounts of centric and pericentric heterochromatin. The large, C-band-positive centromeric region of the X chromosome was microdissected and the isolated DNA was microcloned. Further sequence and cytogenetic analyses of three representative clones show that all chromosomes in this species carry a 178-bp satellite sequence containing a CENP-B DNA binding domain (CENP-B box) shown herein to selectively bind marsupial CENP-B protein. Two other repeats isolated in this study localize specifically to the sex chromosomes yet differ in copy number and intrachromosomal distribution. Immunocytohistochemistry assays with anti-CENP-E, anti-CREST, anti-CENP-B, and anti-trimethyl-H3K9 antibodies defined a restricted point localization of the outer kinetochore at the functional centromere within an enlarged pericentric and heterochromatic region. The distribution of these repeated sequences within the karyotype of this species, coupled with the apparent high copy number of these sequences, indicates a capacity for retention of large amounts of centromere-associated DNA in the genome of M. rufogriseus.     

Meiotic chromosomes of the tree frog Smilisca baudinii (Anura: Hylidae)

The Mexican tree frog Smilisca baudinii, is a very common frog in Central America. In spite their importance to keep the ecological equilibrium of the rainforest, its biology and genetics are poorly known. In order to contribute with its biological knowledge, we described the typical meiotic karyotype based in standard cytogenetic protocols to specimens collected in Tabasco, Mexico. The study was centered in the analysis of 131 chromosome spreads at meiotic stage from two adults of the species (one female and one male). The metaphase analysis allowed the establishment of the modal haploid number of 1n = 12 bivalent chromosomes. The chromosomic formulae from the haploid bivalent karyotype was integrated by 12 biarmed chromosomes characterized by twelve pairs of metacentric-submetacentric (msm) chromosomes. The meiotic counting gives the idea that diploid chromosome number is integrated by a complement of 2n = 24 biarmed chromosomes. The presence of sex chromosomes from female and male meiotic spreads was not observed. Current results suggest that S. baudinii chromosome structure is well shared among Hylidae family and "B" chromosomes are particular structures that have very important evolutionary consequences in species diversification.     

A cytogenetic view of sex chromosome evolution in plants

The recent origin of sex chromosomes in plant species provides an opportunity to study the early stages of sex chromosome evolution. This review focuses on the cytogenetic aspects of the analysis of sex chromosome evolution in plants and in particular, on the best-studied case, the sex chromosomes in Silene latifolia. We discuss the emerging picture of sex chromosome evolution in plants and the further work that is required to gain better understanding of the similarities and differences between the trends in animal and plant sex chromosome evolution. Similar to mammals, suppression of recombination between the X and Y in S. latifolia species has occurred in several steps, however there is little evidence that inversions on the S. latifolia Y chromosome have played a role in cessation of X/Y recombination. Secondly, in S. latifolia there is a lack of evidence for genetic degeneration of the Y chromosome, unlike the events documented in mammalian sex chromosomes. The insufficient number of genes isolated from this and other plant sex chromosomes does not allow us to generalize whether the trends revealed on S. latifolia Y chromosome are general for other dioecious plants. Isolation of more plant sex-linked genes and their cytogenetic mapping with fluorescent in situ hybridisation (FISH) will ultimately lead to a much better understanding of the processes driving sex chromosome evolution in plants.     

Karyotypes of nine species of Cicindelini and cytotaxonomic notes on Cicindelinae (Coleoptera,Carabidae)

The male meioformula of seven species of Cicindelini from the Iberian Peninsula and one species from Canada is 9+XXXY. In addition, female data of Cylindera paludosa corroborate previous reports of males. The Palearctic species show a generalized karyotype which presents specific modifications, but it is not possible to establish if it is also present in species of other faunas.Present data indicate that multiple sex chromosomes is an ancestral condition for the subfamily Cicindelinae. It is postulated that the meioformula of the tribe Cicindelini is 9 or 10 plus XXY and that trends in karyotypic evolution are partially related to the geographic distribution of lineages. Numerical deviations observed in the tribe can be explained by changes occurring independently in autosomes and heterosomes, so that autosome-heterosome fusions do not seem to be a generalized event. A number of criteria may be developed for determining the polarity of change of karyotypic characters, thus making cytotaxonomic studies a valuable tool for understanding the systematics of Cicindelini.     

Cytogenetics characterization of <Emphasis Type="Italic">Crenuchus spilurus</Emphasis> (Günther, 1863): a remarkable low diploid value within family Crenuchidae (Characiformes)

Conventional and molecular cytogenetic analyses were performed in specimens of the Neotropical Crenuchus spilurus freshwater fish species from a single location (Caeté River, Brazil). All specimens presented diploid values of 2n?=?38 chromosomes (12 m?+?4sm?+?2st?+?20a), the lowest reported for family Crenuchidae up to now. A single pair of nucleolar organizing regions (NORs) was detected in the subtelocentric chromosome pair no. 9 by silver-staining and fluorescence in situ hybridization (FISH) with 18S rDNA sequence-specific probe. Two pairs of 5S rRNA gene clusters were found either interstitial or terminally located in the long arms of the acrocentric chromosome pairs nos. 10 and 13. Heterochromatic regions were clearly observed in the short arms of the NOR-bearing chromosome pair and weakly-positive to the pericentromeric regions of most acrocentric chromosomes. Additionally, no sex chromosomes were identified in the surveyed specimens. Crenuchidae have signals of several mechanisms involved in karyotype diversification within this family: differential location of heterochromatin-rich regions, multiplication, and translocation of rDNA clusters, presence/absence of sex chromosomes, macrostructural changes in morphology and number of chromosomes. This variety of karyotype patterns reveals the importance of widening cytogenetic studies to more taxa for better know the chromosomal evolution occurred in this group.     

Phylogenetic Reconstruction by Cross-Species Chromosome Painting and G-Banding in Four Species of Phyllostomini Tribe (Chiroptera,Phyllostomidae) in the Brazilian Amazon: An Independent Evidence for Monophyly

The subfamily Phyllostominae comprises taxa with a variety of feeding strategies. From the cytogenetic point of view, Phyllostominae shows different rates of chromosomal evolution between genera, with Phyllostomus hastatus probably retaining the ancestral karyotype for the subfamily. Since chromosomal rearrangements occur rarely in the genome and have great value as phylogenetic markers and in taxonomic characterization, we analyzed three species: Lophostoma silvicola (LSI), Phyllostomus discolor (PDI) and Tonatia saurophila (TSA), representing the tribe Phyllostomini, collected in the Amazon region, by classic and molecular cytogenetic techniques in order to reconstruct the phylogenetic relationships within this tribe. LSA has a karyotype of 2n=34 and FN=60, PDI has 2n=32 and FN=60 and TSA has 2n=16 and FN=20. Comparative analysis using G-banding and chromosome painting show that the karyotypic complement of TSA is highly rearranged relative to LSI and PHA, while LSI, PHA and PDI have similar karyotypes, differing by only three chromosome pairs. Nearly all chromosomes of PDI and PHA were conserved in toto, except for chromosome 15 that was changed by a pericentric inversion. A strongly supported phylogeny (bootstrap=100 and Bremer=10 steps), confirms the monophyly of Phyllostomini. In agreement with molecular topologies, TSA was in the basal position, while PHA and LSI formed sister taxa. A few ancestral syntenies are conserved without rearrangements and most associations are autapomorphic traits for Tonatia or plesiomorphic for the three genera analyzed here. The karyotype of TSA is highly derived in relation to that of other phyllostomid bats, differing from the supposed ancestral karyotype of Phyllostomidae by multiple rearrangements. Phylogenies based on chromosomal data are independent evidence for the monophyly of tribe Phyllostomini as determined by molecular topologies and provide additional support for the paraphyly of the genus Tonatia by the exclusion of the genus Lophostoma.     

Numerical and structural variations of the X chromosomes and no. 2 autosomes in mandarin vole, Microtus mandarinus (Rodentia)

Here we describe our studies on Microtus mandarinus faeceus of Jiangyan in Jiangsu province of China. By karyotype and G-banding analysis we have found variation in chromosome number and polymorphisms of the X chromosome and the second pair of autosomes of the subspecies. Chromosome number of the subspecies is 2n=47-50. The subspecies has three kinds of chromosomal sex: XX, XO and XY, among which one of the X chromosomes is subtelocentric (X(ST)) and the other is metacentric (X(M)). After comparing karyotypes of different subspecies, we found the specific cytogenetic characteristics of Microtus mandarinus, that is they have three kinds of chromosomal sex: XX, XO and XY; X chromosomes are heteromorphic; the chromosome number of female individuals are one less than male individuals; chromosome number of XX individuals are equal to that of XO ones. We hypothesize that Robertsonian translocation is the main reason of the polymorphism of the second pair of autosomes and variety of chromosome number, and it also causes the chromosome number evolution in different subspecies of Microtus mandarinus.     

Sex chromosome differentiation in Belostoma (Insecta: Heteroptera: Belostomatidae)

Belostoma, a genus of the family Belostomatidae, includes species of great ecological importance as biocontrol agents. Few species of these species have been the subject of cytogenetic analyses. Karyotypic evolution in this genus involves agmatoploidy and simploidy; there are also different sex chromosome systems. We examined two Belostoma species (B. dilatatum and B. candidulum) collected from the Paranapanema River Basin (Brazil). Mitotic and meiotic analysis revealed 2n(♂) = 26 + X(1)X(2)X(3)Y for B. dilatatum and 2n(♂) = 14 + XY for B. candidulum; both karyotypes have holokinetic chromosomes. Differences in heterochromatin distribution were also observed between the species, besides variation in the localization of CMA(3)(+)/DAPI(-) blocks. The existence of different types of sex chromosome systems in these species was confirmed based on arrangements of the chromosomes in different meiotic stages. We identified a new sex system in B. dilatatum, and make the first cytogenetic report on B. candidulum.     

Chromosome numbers, sex determining systems, and patterns of the C-heterochromatin distribution in 13 species of lace bugs (Heteroptera, Tingidae).

The karyotypes, sex chromosome systems, and male meiotic patterns in 13 species belonging to 10 genera of the family Tingidae were studied. Data on eleven species, one subgenus, and 5 genera are presented for the first time, and the chromosome formula of Acalypta parvula is revised. Karyotypes of all species included six pairs of autosomes. Most of the species displayed an XY sex chromosome system, in four species, belonging to genera of Acalypta and Kalama, the X0 system was found. Male meiosis is chiasmatic for autosomes. Sex chromosomes are achiasmatic and undergo pre-reductional meiosis. Using C-banding technique, for the first time constitutive heterochromatin was localized on chromosomes in all the species studied. The heterochromatin was found either in telomeres or in some species in interstitial locations, evidencing that a quite substantial redistribution of chromosome material within chromosomes might occur without fragmentations or fusions. In two species, a supernumerary (B) chromosome was found. In addition, the male reproductive system of four species was examined and the number of testicular follicles was determined as two per testis.     

High-density sex-specific linkage maps of a European tree frog (Hyla arborea) identify the sex chromosome without information on offspring sex

Identifying homology between sex chromosomes of different species is essential to understanding the evolution of sex determination. Here, we show that the identity of a homomorphic sex chromosome pair can be established using a linkage map, without information on offspring sex. By comparing sex-specific maps of the European tree frog Hyla arborea, we find that the sex chromosome (linkage group 1) shows a threefold difference in marker number between the male and female maps. In contrast, the number of markers on each autosome is similar between the two maps. We also find strongly conserved synteny between H. arborea and Xenopus tropicalis across 200 million years of evolution, suggesting that the rate of chromosomal rearrangement in anurans is low. Finally, we show that recombination in males is greatly reduced at the centers of large chromosomes, consistent with previous cytogenetic findings. Our research shows the importance of high-density linkage maps for studies of recombination, chromosomal rearrangement and the genetic architecture of ecologically or economically important traits.