Location Of Nucleolus Organizer Regions Research Articles

Citogenética comparativa em felídeos (Carnivora: Felidae): Leopardus wiedii, Panthera onca, Puma concolor e Felis catus

ABSTRACT This study aims to conduct a cytogenetic analysis on four South American feline species: the jaguar (Panthera onca), the puma (Puma concolor), the ocelot (Leopardus wiedii), and the domestic cat (Felis catus). We discovered that the jaguar and the puma mainly differ in their fundamental number of chromosomes. Furthermore, we observed a morphological difference in a pair of chromosomes between the jaguar and the domestic cat. This suggests that, over the course of evolution, there was a pericentric inversion in a pair of B4 group chromosomes, leading them to become part of the A group in the jaguar. When analyzing the puma, we noticed the presence of a fourth pair of chromosomes in the E group, along with a deletion in the F group. Additionally, we observed the presence of an additional pair of chromosomes in the C group in the ocelot. Another interesting aspect of the study was the location of nucleolar organizer regions (NORs) on the chromosomes. To elucidate the origin of other chromosome pairs, we employed chromosomal banding techniques, allowing us to better understand the evolution of the C3 pair in the Leopardus genus and the E4 pair in the Puma genus.

Silver Nitrate Staining of Nucleolar Organizer Regions (Ag-NORs) in Plant Chromosomes.

Silver nitrate staining to evidence the location of nucleolar organizer regions (Ag-NORs) in chromosomes is widely used as a classical method in plant cytogenetics. Here, we present the most used procedures and highlight some aspects in terms of their replicability by plant cytogeneticists. Some technical features described are materials and methods used, procedures, protocol modifications, and precautions in order to obtain positive signals. The methods to obtain Ag-NOR signals have different degrees of replicability, but do not require any sophisticated technology or equipment for their application.

Rebuilding the Cytogenetics of Atelognathus (Anura: Batrachylidae): Half a Century of Confusion

The genus Atelognathus is composed of five species that occur in the southernmost regions of Argentina and Chile. It belongs to the Batrachylidae, one of the few endemic anuran families from Patagonia. More than 50 yr ago, it was concluded that A. reverberii had a diploid chromosomal complement of 2N = 26, a feature later reported for other congenerics. In this work, we describe the cytogenetics of all known species of Atelognathus (A. nitoi, A. patagonicus, A. praebasalticus, A. reverberii, and A. solitarius) and two additional undetermined populations. Unexpectedly, our analyses showed that all have a diploid chromosomal complement 2N = 24, which contrasts with conclusions of previous studies. We conclude that a karyotype of 2N = 24 is shared in Atelognathus as a derived condition within Batrachylidae. Additionally, we describe and discuss other cytogenetics characters (i.e., nucleolar organizer region location and C-banding patterns) of Atelognathus and related genera of Batrachylidae in an evolutionary framework.

Genomic Organization of the Repetitive Sequences in Centropomus undecimalis (Perciformes, Centropomidae): Implications for Hybridization and Aquaculture Programs.

The family Centropomidae includes a number of fish species of high commercial value. One of these species, Centropomus undecimalis, is a target of artisanal, industrial, and sports fisheries and has also considerable potential for captive breeding, which has led to its inclusion in several aquaculture programs. While the biology and ecology of C. undecimalis are relatively well documented, few karyological data are available on this species, and they are still scarce for other centropomids. The few chromosomal data available on this family indicate a conserved karyotype 2n = 48, but it is unclear whether the chromosome microstructure is also conserved. In this study, new cytogenetic data are presented on C. undecimalis from the Amazon coastal zone, including C-banding, Ag-NOR, in situ hybridization with repetitive DNA probes (5S and 18S ribosomal genes), and telomeric (TTAGGG)n sequences. The diploid number of the species was 2n = 48, with heterochromatic blocks in the centromeric and pericentromeric regions, as well as distal signals; the nucleolus organizer regions (NORs) were associated with the heterochromatic region. The 18S and 5S recombinant DNA (rDNA) clusters were located in the distal region of chromosome pairs 1 and 11, respectively. The similarities of the karyotype macrostructure found among the centropomid species reinforce their exceptional chromosomal stability. However, the presence of heterochromatic blocks and location of NORs suggest the occurrence of structural rearrangements, which indicates that evolutionary dynamics at the microstructural level in this group may be relatively complex and should be evaluated carefully in any study that targets the production of hybrids for aquaculture.

Molecular Clock as Insight to Estimate the Evolutionary History and Times of Divergence for 10 Nominal Astyanax Species (Characiformes, Characidae): An Evolutionary Approach in Species with 2n = 36, 46, 48, and 50 Chromosomes

Astyanax is a genus with a wide distribution ranging from the south United States to north of Patagonia (Argentina). The available cytogenetic data on Astyanax indicate a high karyotypic diversity, with diploid number of 36-52 chromosomes, presence of B chromosomes, heterochromatin polymorphism, and variations with respect to the number and localization of nucleolar organizer regions (NORs) and 18S and 5S ribosomal DNA sites. In the present study, we estimated the evolutionary history and times of divergence for 10 nominal Astyanax species from the South and Central/North American (Cna) continents, which present distinct chromosomal characteristics, based on molecular clocks inferred from mitochondrial DNA sequence. The molecular clock results indicate the origin of three distinct clades (Humeral dark spot [Hds]; Diffuse humeral spot [Dhs]; Cna group) during the late Miocene about 11.2 million years ago (Mya). Thus, Astyanax mexicanus (Cna) represent a species that diverged a long time ago (∼8.6 Mya) from the Hds group, and Astyanax schubarti is the oldest species (∼6.5 Mya) among the Dhs species.

The karyotype of the great sculpin, Myoxocephalus polyacanthocephalus (Pallas, 1814) (Pisces: Cottidae), from the Russian part of the species range

The karyotype of the great sculpin, Myoxocephalus polyacanthocephalus (Pallas, 1814) (Pisces: Cottidae) from the Sea of Okhotsk and the Sea of Japan has been studied for the first time. The karyotype is stable; it consists of 40 chromosomes (4 metacentric, 2 submeta-subtelocentric, 20 subtelocentric, and 14 acrocentric chromosomes); the number of chromosomal arms is 44 + 2. Nucleolar organizer regions (NOR) are found in the telomeric region of the arm in one homologue of a pair of small metacentric chromosomes, using the Ag-NOR banding technique. A comparative analysis of the karyotype of M. polyacanthocephalus and the karyotypes of other Myoxocephalus species (M. stelleri, M. brandtii, M. jaok, M. ochotensis, and M. scorpius) has been carried out based on the main karyotype characters, as well as on the number and localization of NORs. The identified differences make it possible to differentiate the studied species, whereas the general traits indicate their taxonomic proximity.

Chromosomal similarities between Nephilidae and Tetragnathidae indicate unique evolutionary traits among Araneoidea

Nephilid systematics has been subject to several changes in the last years, and the use of non-classical characters could be useful for evolutionary considerations. In this study, we analyzed the mitotic chromosomes of two nephilid spiders, Nephila clavipes and Nephila sexpunctata, using standard staining, silver nitrate impregnation and C-banding techniques, aiming to discuss the chromosomal similarities of Nephilidae and Tetragnathidae, and chromosome evolution within Nephila and Nephilingis. The basic karyotype characteristics observed in these two species (2n♂ = 22 + X1X20 and monoarmed chromosomes) were similar to those registered for most araneoid families, i.e., Araneidae, Linyphiidae, Nephilidae, Nesticidae and Tetragnathidae. However, the occurrence of both prominent secondary constrictions and nucleolar organizer regions (NORs) is a shared characteristic between Nephilidae and Tetragnathidae, considering that these regions were not observed in any other Araneoidea species cytogenetically examined. Furthermore, in the present study we showed that within Nephila and Nephilingis species, change in the number and location of NORs as well as in the quantity and distribution of constitutive heterochromatin were the main events responsible for chromosome evolution, and that these differences can be useful in the cytotaxonomy of this group.

The morphological, histological and cytogenetic characteristics of goldfishCarassius auratus(L.) × common carpCyprinus carpio(L.) hybrids

Morphology of hybrids between the goldfish (Carassius auratus) and common carp (Cyprinus carpio) are studied by biometric and meristic methods. Additionally, histological analyses concerning gonads as well as genetic characteristics of chromosomes are performed. Studied hybrids were morphologically very similar to a goldfish but possessed one pair of reduced posterior barbels. Most meristic and biometric indices had intermediate values when compared to parental species. Histological study showed that investigated hybrids produce gonads with germ cells. All studied males had gonads at the same stage of maturity (III stage). By contrast, female gonads were in various stages of development – from sexually immature to post-spawning stage. Karyotype characteristics were assessed in terms of chromosomes number, fundamental arm number and location of nucleolus organizer regions (NOR). The data obtained were compared with baseline karyotypes of species used for hybridization, based on information from the literat...

Heterochromatin characterization and ribosomal gene location in two monotypic genera of bloodsucker bugs (Cimicidae, Heteroptera) with holokinetic chromosomes and achiasmatic male meiosis.

Members of the family Cimicidae (Heteroptera: Cimicomorpha) are temporary bloodsuckers on birds and bats as primary hosts and humans as secondary hosts. Acanthocrios furnarii (2n=12=10+XY, male) and Psitticimex uritui (2n=31=28+X1X2Y, male) are two monotypic genera of the subfamily Haematosiphoninae, which have achiasmatic male meiosis of collochore type. Here, we examined chromatin organization and constitution of cimicid holokinetic chromosomes by determining the amount, composition and distribution of constitutive heterochromatin, and number and location of nucleolus organizer regions (NORs) in both species. Results showed that these two bloodsucker bugs possess high heterochromatin content and have an achiasmatic male meiosis, in which three regions can be differentiated in each autosomal bivalent: (i) terminal heterochromatic regions in repulsion; (ii) a central region, where the homologous chromosomes are located parallel but without contact between them; and (iii) small areas within the central region, where collochores are detected. Acanthocrios furnarii presented a single NOR on an autosomal pair, whereas P. uritui presented two NORs, one on an autosomal pair and the other on a sex chromosome. All NORs were found to be associated with CMA3 bright bands, indicating that the whole rDNA repeating unit is rich in G+C base pairs. Based on the variations in the diploid autosomal number, the presence of simple and multiple sex chromosome systems, and the number and location of 18S rDNA loci in the two Cimicidae species studied, we might infer that rDNA clusters and genome are highly dynamic among the representatives of this family.

Activity patterns of nucleolar organizer region during spermatogenesis of different curimatid species (Characiformes: Curimatidae)

The nucleolus is an important nuclear structure where transcription of ribosomal DNA (rDNA) takes place. During mitotic division, the nucleolus passes through different processes that inactivate rDNA transcription; in meiosis, its reassembly takes place during telophase II. The objective of this study was to identify the activity patterns and localization of nucleolar organizer regions (NORs) during meiotic division in fish species of the family Curimatidae. For this analysis, the meiotic division in five curimatid species was studied using silver nitrate impregnation, fluorescent in situ hybridization (FISH), and base-specific fluorochrome staining. Silver nitrate staining indicated the presence of a nucleolus in interphase nuclei, one chromosome pair in the spermatogonial metaphases, and one bivalent at the pachytene stage. No Ag-NORs were identified for cells at the diplotene, diakinesis, metaphase I, or metaphase II stages; however, FISH confirmed the presence of Ag-NORs in the nuclei, in spermatogonia, and at the pachytene phase. FISH identified this region during the other stages of meiosis, as did fluorochrome CMA3 staining, which revealed fluorescent marks corresponding to NORs during all stages of meiosis analyzed. The gene activity and localization of this ribosomal sequence during the different stages involved will also be discussed.

Description of Karyotype of Kerodon acrobata, an endemic rodent in Brazilian Cerrado.

Kerodon acrobata is a caviidae rodent endemic from Brazilian Cerrado. It was described only in 1997 and the data about it is very scarce. The aim of this work was to characterize the karyotype of K. acrobata. Giemsa staining, nucleolar organizer region (NOR) banding, C-positive heterochromatin banding and DAPI fluorescence were used in N metaphases of a specimen collected in Asa Branca Farm, in Aurora do Tocantins municipality, Tocantins state, Brazil. K. acrobata showed the same diploid number, fundamental number and chromosome morphology as Kerodon rupestris. But its NOR location and heterochromatin distribution patterns indicated a unique cytogenetic profile when compared to its sister species, emphasizing the evolutionary uniqueness of this relatively new and unknown species. This record also extends the distribution of this species northward.

Dynamics of chromosomal evolution in the genus Hypsiboas (Anura: Hylidae).

Hylidae is one of the most species-rich families of anurans, and 40% of representatives in this group occur in Brazil. In spite of such remarkable diversity, little is known about this family and its taxonomical and systematic features. Most hylids have 2n = 24, even though most of the cytogenetic data are mainly obtained based on the conventional chromosomal staining and are available for only 16% of Hypsiboas species, a genus accounting for about 10% of the hylid diversity. In this study, cytogenetic data of distinct species and populations of Hypsiboas were analyzed, and the evolutionary dynamics of chromosomal macro- and microstructure of these amphibians were discussed. Contrary to the conservativeness of 2n = 24, this genus is characterized by a high variation of chromosomal morphology with as much as 8 karyotype patterns. Differences in the number and location of nucleolus organizer regions and C-bands allowed the identification of geographical variants within nominal species and cytotaxonomical chromosomal markers. Comparative analyses revealed a strong phylogeographic relationship between chromosomal patterns in this group.

Cytogenetics and sperm ultrastructure of Atelopus spumarius (Anura, Bufonidae) from the Brazilian Amazon.

The current taxonomy of most Atelopus species is based on morphological and color data only. Recent studies suggest that A. spumarius may represent a species complex assigned under the same name. Karyotypic data and description of sperm ultrastructure for 13 specimens of A. spumarius are presented here for the first time. A chromosomal analysis revealed 2n = 22 chromosomes, with centromeric heterochromatin in all pairs and a nucleolar organizer region (NOR) on the telomere of pair 7. The sperm was of the bufonoid type, presenting a filiform nucleus covered by an acrosomal complex and a mitochondrial collar in the neck region. The tail was composed of an axoneme, an undulating membrane and an axial rod. A karyotype analysis of A. spumarius showed the same chromosome number and similar chromosomal morphology as described for congeneric species, with slight differences probably resulting from pericentric inversions. The NOR location (on pair 7) was the same as that observed for species belonging to the genus Rhinella. The spermatological findings indicate a close relationship between Atelopus and the bufonoid lineage. The present data are useful for reference in future studies to determine whether more than one species are assigned to A. spumarius.

Mapping five repetitive DNA classes in sympatric species of Hypostomus (Teleostei: Siluriformes: Loricariidae): analysis of chromosomal variability

Fish belonging to the genus Hypostomus are known for exhibiting a striking diversity in its karyotype structure, however the knowledge concerning the distribution patterns of heterochromatin and location of repetitive DNA sequences in the karyotypes is still limited. Aiming a better understanding of the chromosomal organization in this group, we analyzed three sympatric species of Hypostomus collected in the Hortela stream, a component of the Paranapanema River basin, Botucatu/SP/Brazil. The analyses involved the cytogenetic characterization and chromosomal mapping of repetitive sequences and intra/interspecific comparisons using sequences of the cytochrome C oxidase subunit I. The results revealed that H. ancistroides presents a karyotype with 2n = 68 chromosomes, H. strigaticeps 2n = 72 chromosomes, and H. nigromaculatus 2n = 76 chromosomes. In addition to differences found in the diploid number, it was also observed variations in karyotypic formulae, amount of constitutive heterochromatin, and location of nucleolus organizer regions. The cytogenetic mapping of 5S and 18S rDNA, as well as of the H3 histone gene, disclosed a differential dispersion process among the three species. In some cases the Rex1 transposable element showed to be co-located with 5S rDNA sites. The molecular analyses support the cytogenetic data and represent an additional tool for the characterization of the analyzed species. The results evidenced that chromosomal variations are not restricted to differences in diploid number or karyotypic macrostructure in the genus Hypostomus, indicating that events such as transposition of heterochromatin and rDNA segments may participate in the differentiation process occurred in these species.

Cytogenetic and molecular analyses in troglobitic and epigean species of Pimelodella (Siluriformes: Heptapteridae) from Brazil

Samples from seven different locations of the genus Pimelodella were genetically examined, two caves (exclusively subterranean, upper Tocantins River and São Francisco River) and five epigean (from upper Paraná River basin). Cytogenetic analyses revealed the same diploid number (2n=46) for all species besides similarities in both number and location of nucleolar organizer regions and C bands. FISH with 5S rDNA probes and CMA3 staining indicated significant differences among the studied species. Application of PCR-RFLP in ATPase 6 and 8 mitochondrial genes allowed building a minimum evolution phenogram identifying the close evolutionary relationship among groups. Both chromosomal and molecular data were useful to infer the relationships among studied Pimelodella species.

Molecular Insights into X;BTA5 Chromosome Rearrangements in the Tribe Antilopini (Bovidae)

For a clade that includes Antilope, Gazella,Nanger and Eudorcas (Antilopinae), X;BTA5 translocation is a synapomorphy. Using a combination of fluorescence in situ hybridization (FISH) probes and polymerase chain reaction techniques, we provide (i) the first insight into the X;BTA5 architecture which differs in the species under study: Antilope cervicapra (genus Antilope), Gazella leptoceros (genus Gazella) and Nanger dama ruficollis (genus Nanger), (ii) determination of interstitial satellite DNA at the X;BTA5 junctions, and (iii) determination of repetitive sequences occupying constitutive heterochromatin of Xp arms in the studied species. The distribution of 2 repetitive DNA families in the centromeric regions of all chromosomes has been investigated by FISH with probes representing satellite I and satellite II DNA in all studied species. In this context, we discuss a markedly smaller centromere in the BTA5 (Y2) unfused chromosomes in males in the XY1Y2 determining system in comparison with other acrocentrics. An analysis of karyotypic data described in current published studies revealed a disparity with the data determined by FISH. In this report, we document chromosomal fusions in the 3 species mentioned resulting from FISH with painting probes prepared from cattle (Bos taurus). The number and chromosomal location of nucleolus organizer regions were determined by FISH. In the present study, we emphasize the importance of chromosomal rearrangement verification, particularly, if they are used for phylogenetic analysis.

Localization of Nucleolar Organizer Regions (NORs) of 4 Gibbon Species in Thailand by Ag-NOR Banding Technique

Summary Localization of nucleolar organizer regions (NORs) of four gibbon species in Thailand was studied. Blood samples were taken from 1 male each of the following 3 species: white-handed gibbon (Hylobates lar), pileated gibbon (H. pileatus), and white-cheeked gibbon (Nomascus leucogenys): and 1 female of the dark-handed gibbon (H. agilis). These were subjected to standard whole blood T-lymphocyte culture. The samples were harvested by colchicine-hypotonic-fixation-air-drying technique and were followed by the Ag-NOR banding technique. The results showed that the H. lar had NORs on chromosome pair 13 (submetacentric chromosomes) while the H. agilis had NORs on chromosome pair 10 (submetacentric chromosomes) and H. pileatus had NORs on chromosome pair 15 (metacentric chromosomes). In addition, the N. leucogenys showed 5 positions with NORs on chromosomes pairs 23, 24 (acrocentric chromosomes) and the Y-chromosome (submetacentric chromosome). These results are useful to confirm marker chromosomes of gibbons and also comprise basic genetic information for these animals. Kew words Gibbon, Karyotype, Chromosome, Idiogram, Ag-NOR banding. Thailand is tropical country thus its weather and surrounding are suitable for organism that causes high diversity. Wild animals are one element that show the diversity and also important in ecology and food chain. The destruction of forest causes wild animal extinction or risk of extinction includes animal in family Hylobatidae. There were 13 families, 60 genera and 232 species of animals in order Primate (Wilson and Cole 2000); however, only 3 families, 5 genera, and 13 species were found in Thailand. Thai gibbon species consists of 4 species, white-handed gibbon (Hylobates lar Linnaeus 1771), agile gibbon or dark-handed gibbon (Hylobates agilis Cuvier 1821), pileated gibbon (Hylobates pileatus Gray 1842), and white-cheeked gibbon (Nomascus leucogenys Ogilby 1840) (Lekagul and McNeely 1988, Giessmann 1995) (Fig. 1). These species are listed as protected species in Wild Animal Reservation and Protection Act (1992), and in the International Union for the Conservation of Nature (IUCN) as vulnerable species. The Convention on International Trade in Endangered Species (CITES) of wild fauna and flora also listed these species in Appendix I (Brokelman 1981).

Chromosomal dynamics of nucleolar organizer regions (NORs) in the house mouse: micro-evolutionary insights

Variation in the number and chromosomal location of nucleolar organizer regions (NORs) was studied in the house mouse, Mus musculus (2n=40). From an origin in Western Asia, this species colonized the Middle East, Europe and Asia. This expansion was accompanied by diversification into five subspecies. NOR diversity was revealed by fluorescence in situ hybridization using 18S and 28S probes on specimens spanning Asia to Western Europe. The results showed that the house mouse genome possessed a large number of NOR-bearing autosomes and a surprisingly high rate of polymorphism for the presence/absence of rRNA genes on all these chromosomes. All NOR sites were adjacent to the centromere except for two that were telomeric. Subspecific differentiation established from the NOR frequency data was concordant with the overall pattern of radiation proposed from molecular studies, but highlighted several discrepancies that need to be further addressed. NOR diversity in M. musculus consisted of a large number of polymorphic NORs that were common to at least two subspecies, and a smaller number of NORs that were unique to one subspecies. The most parsimonious scenario argues in favor of a subspecific differentiation by lineage sorting of ancestral NOR polymorphisms; only the unique NORs would have appeared by inter-chromosomal transposition, except for the two telomeric ones that may have originated by hybridization with another species. Such a scenario provides an alternative view from the one prevailing in most systematic and phylogenetic analyses that NORs have a high transposition rate due to concerted evolution of rRNA genes.

Chromosome analyses in species of Pareiorhina (Siluriformes, Loricariidae) and cytotaxonomic considerations on the group

The genus Pareiorhina, formerly considered monotypic, was recently described to comprise two new species P. carrancas and P. brachyrhyncha. Karyotype analysis allowed to infer cytotaxonomical diagnostic characters for P. rudolphi and P. brachyrhyncha in the present study. The species showed the same diploid number (2n = 54) and karyotypes with 18 m, 32sm, 4st and 18 m, 30sm, 6st chromosomes, respectively. In addition, we found that the nucleolus organizer regions (NORs) are located on different chromosomes. Localization of NORs has been ascertained by fluorescent in situ hybridization (FISH) and colloidal silver nitrate stain (Ag-NORs). P. rudolphi shows NORs in interstitial position of the long arm of chromosome pair number 11, while P. brachyrhyncha shows subterminal NORs only in the short arm of chromosome pair 25. Occurrence of diploid chromosome number and presence of only one pair of NOR bearing chromosomes makes these species to comprised of most plesiomorphic karyotype of Loricariidae and supports its clustering with the genus Neoplecostomus. The chromosome types and diversification in the location of NORs corroborate the description of the species P. brachyrhyncha.

Chromosomal painting and ZW sex chromosomes differentiation in Characidium (Characiformes, Crenuchidae)

BackgroundThe Characidium (a Neotropical fish group) have a conserved diploid number (2n = 50), but show remarkable differences among species and populations in relation to sex chromosome systems and location of nucleolus organizer regions (NOR). In this study, we isolated a W-specific probe for the Characidium and characterized six Characidium species/populations using cytogenetic procedures. We analyzed the origin and differentiation of sex and NOR-bearing chromosomes by chromosome painting in populations of Characidium to reveal their evolution, phylogeny, and biogeography.ResultsA W-specific probe for efficient chromosome painting was isolated by microdissection and degenerate oligonucleotide primed-polymerase chain reaction (DOP-PCR) amplification of W chromosomes from C. gomesi. The W probe generated weak signals dispersed on the proto sex chromosomes in C. zebra, dispersed signals in both W and Z chromosomes in C. lauroi and, in C. gomesi populations revealed a proximal site on the long arms of the Z chromosome and the entire W chromosome. All populations showed small terminal W probe sites in some autosomes. The 18S rDNA revealed distinctive patterns for each analyzed species/population with regard to proto sex chromosome, sex chromosome pair, and autosome location.ConclusionsThe results from dual-color fluorescence in situ hybridization (dual-color FISH) using W and 18S rDNA probes allowed us to infer the putative evolutionary pathways for the differentiation of sex chromosomes and NORs, from structural rearrangements in a sex proto-chromosome, followed by gene erosion and heterochromatin amplification, morphological differentiation of the sex chromosomal pair, and NOR transposition, giving rise to the distinctive patterns observed among species/populations of Characidium. Biogeographic isolation and differentiation of sex chromosomes seem to have played a major role in the speciation process in this group of fish.