Diploid Number 2n Research Articles

Morphological variation in house mice from the Robertsonian polymorphism area of Barcelona

Morphometric variation in the Robertsonian polymorphism zone of Barcelona of Mus musculus domesticus was studied by geometric morphometrics. This system is characterized by populations of reduced diploid number (2n = 27–39) surrounded by standard populations (2n = 40). We investigated the morphological variation in mice from this area, as well as the effect of geographical distance and karyotype on this variation. We also investigated the degree of co-variation between the two functional units of the mandible to explore the origin of this system (primary intergradation or secondary contact). The size and shape of the cranium, mandible and scapula were analysed for 226 specimens grouped by population, chromosome number and structural heterozygosity. Size was estimated as the centroid size, and shape was estimated after Procrustes superimposition. No significant differences in size between populations or chromosomal groups were detected. Diploid number, structural heterozygosity and local geographical isolation contributed to the differentiation in shape. Morphological differentiation between standard mice and Robertsonian specimens was observed, suggesting genetic isolation between these groups. Co-variation between the ascending ramus and alveolar region of the mandible was quantified by the trace correlation between landmark subsets of these modules. The trace values showed an ascending trend, correlated with the distance from the centre of the polymorphism area, a pattern consistent with a primary intergradation scenario. © 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 97, 555–570.

Banded karyotypes of mole rats (Spalax, Spalacidae, Rodentia) from Turkey: a comparative analysis

Chromosome banding (G-, C- and Ag-NOR) analysis was carried out on 27 specimens of Sphalax ehrenbergi from seven localities and two specimens of S. leucodon from one locality, all from Turkey. No chromosomal variation was detected in S. ehrenbergi populations from Elazig, Siverek, Diyarbakir and Birecik having the same diploid numbers (2n = 52) and morphology of chromosomes (NFa = 72). The karyotypes of mole rats from Tarsus and Gaziantep possessed the identical diploid number (2n = 56) but different numbers of autosomal arms: NFa = 68 in the Tarsus and NFa = 78 in the Gaziantep populations. Chromosomes of S. leucodon from Malaty (2n = 60, NFa = 74) differed distinctly in the C-banding pattern from all S. ehrenbergi cytotypes by the almost entire absence of heterochromatin in acrocentric autosomes and the presence of heterochromatin arms iin subtelocentric autosomes. Nucleolar organizing regions were found mainly on three pairs of chromosomes, but some differences in their localization were revealed. Comparison of G-banded chromosomes showed, that most chromosomes have a similar pattern. The types of chromosomal rearrangemetns were revealed due to the banding methods.

Intriguing evidence of translocations in Discus fish (Symphysodon, Cichlidae) and a report of the largest meiotic chromosomal chain observed in vertebrates

As part of a program to understand the genetics of Amazonian ornamental fish, classical cytogenetics was used to analyze Symphysodon aequifasciatus, S. discus and S. haraldi, popular and expensive aquarium fishes that are endemic to the Amazon basin. Mitotic analyses in Symphysodon have shown some odd patterns compared with other Neotropical cichlids. We have confirmed that Symphysodon species are characterized by chromosomal diversity and meiotic complexity despite the fact that species share the same diploid number 2n=60. An intriguing meiotic chromosomal chain, with up to 20 elements during diplotene/diakinesis, was observed in S. aequifasciatus and S. haraldi, whereas S. discus only contains typical bivalent chromosomes. Such chromosomal chains with a high number of elements have not been observed in any other vertebrates. We showed that the meiotic chromosomal chain was not sex related. This observation is unusual and we propose that the origin of meiotic multiples in males and females is based on a series of translocations that involved heterochromatic regions after hybridization of ancestor wild Discus species.

Frequency of aneuploids in progenies of autotriploid barley,Hordeum vulgare L.

Chromosome counts of 863 progeny plants originating from 68 autotriploid barley plants revealed a considerable variation in chromosome numbers ranging from the diploid number (2n= 14) to 2n= 39. The most frequent groups were plants with 15 and 16 chromosomes each constituting about 27% of all progeny plants. Diploid plants and plants with 1 to 5 extra chromosomes constituted about 85% of the progeny plants. The remaining 15% had chromosome numbers from 20 to 39. Plants with the euploid numbers of chromosomes. 21, 28, and 35, were not more frequent than plants with aneuploid numbers between these groups. The results show that barley, although a diploid species, has a high tolerance for extra chromosomes.

Cytogenetic analysis of Astyanax laticeps (Cope, 1894) (Ostariophysi: Characidae) from the laguna dos Patos system

The genus Astyanax is prominent among Characiformes, due to the large number of species found and its wide geographic distribution. In this work, Astyanax laticeps specimens from the laguna dos Patos system were cytogenetically analyzed. A diploid number of 2n = 50 chromosomes distributed into 6m+16sm+16st+12a (FN = 88) was found, without differences between males and females. A few small heterochromatin blocks were observed, besides three more conspicuous C-bands, corresponding to NORs, as confirmed by silver nitrate and CMA3 staining, FISH, and DAPI negative staining. These regions were located in a medium-sized subtelocentric and in a large subtelocentric chromosomal pair, probably because of a deletion of this region in one homologous chromosome, or due to a transposition event between them.

Biogeography and comparative cytogenetics between two populations of Hoplias malabaricus (Bloch, 1794) (Ostariophysi: Erythrinidae) from coastal basins in the State of Bahia, Brazil

The species Hoplias malabaricus is a predator fish found in nearly all cis-Andean basins. From a cytogenetic point of view, this species comprises, at least, seven differentiated karyomorphs. Several localities have been formerly analyzed in Brazil, however, some regions, such as Bahia State, remain underrepresented. Recently, the Brazilian Environment Ministry classified both Itapicuru and Contas river basins (entirely located within Bahia territory) as priority conservation areas, whose biodiversity status lacks enough information. Therefore, the goal of the present work was to characterize, cytogenetically, populations of H. malabaricus from both basins, by using conventional staining, Ag-NOR and C-banding techniques. All specimens presented a diploid number of 2n = 40 with metacentric/submetacentric chromosomes, without differences between sexes, thereby representing the so-called "karyomorph F". The first metacentric pair presented a remarkably larger size in relation to the other pairs. The NORs were multiple, comprising the terminal region on long arms of two chromosomal pairs in both populations. However, the C-banding pattern was somewhat distinguishable between samples. Although sharing heterochromatic blocks at centromeric region of all chromosomes, the population from Itapicuru River basin appeared to have some more conspicuous blocks than those observed in the population from Contas River basin. The similar karyotype observed in both populations suggests a common geological history between them. The present results represent an advance in the knowledge about the cytogenetic pattern of H. malabaricus populations from poorly studied basins.

Chromosome evolution in the subtribe Bovina (Mammalia, Bovidae): The karyotype of the Cambodian banteng (Bos javanicus birmanicus) suggests that Robertsonian translocations are related to interspecific hybridization

Three subspecies of banteng (Bos javanicus) have been described: B. j. javanicus in Java, B. j. lowi in Borneo, and B. j. birmanicus in Cambodia, Lao PDR, Myanmar, Thailand and Vietnam. In this paper we provide the first description of the karyotype of the Cambodian banteng. The chromosomal complement of B. j. birmanicus differs from that of B. j. javanicus, which was previously found to be similar to that of cattle, Bos taurus (2n = 60). The Cambodian banteng karyotype has a diploid number of 2n = 56 (FN = 62) and the karyotype consists of 26 pairs of acrocentric chromosomes and two pairs of submetacentric chromosomes. Comparisons with other species of the subtribe Bovina show that the two pairs of bi-armed chromosomes resulted from two centric fusions involving the equivalent of cattle chromosomes 1 and 29, and 2 and 28, respectively. Cross-species fluorescence in-situ hybridization (FISH) with B. taurus whole chromosome paints and satellite DNA I probes was used to identify the chromosomes involved in the translocations, and their orientation. We suggest that Robertsonian translocations (1;29) and (2;28) have been fixed in the common ancestor of Cambodian banteng as a consequence of hybridization with the kouprey (Bos sauveli) during the Pleistocene epoch.

Cariotipo del pejelagarto tropical Atractosteus tropicus (Lepisosteiformes: Lepisosteidae) y variación cromosómica en sus larvas y adultos

The karyotype of the tropical gar Atractosteus tropicus is described from conventional Giemsa-staining of 295 mitotic chromosome slides from 120 larvae and 15 adults (five females and ten males) from Tabasco, southern of Mexico. The diploid number 2n = 56 chromosomes was calculated (73 spreads from 206 larval and 208 adult metaphases). Variation on chromosome number was from 46 (4.4%) to 64 (3.9%) chromosome elements on larval samples, whereas 58 (11.7%) chromosomes were the second most abundant after the diploid number of 2n = 56 (35%). Such variation was related with the presence of mobile microchromosomes. The karyotype was determined from six clear chromosome spreads photographed from three females and three males. The averaged karyotype was integrated by eight pairs of metacentric (m) chromosomes, four submetacentric (sm) pairs, eight telocentric (t) pairs and eight pairs of telocentric microchromosomes (*t). The fundamental number was FN = 80 chromosome arms. We saw no sexual differences on chromosome structure.

Chromosome studies of European cyprinid fishes: interspecific homology of leuciscine cytotaxonomic marker—the largest subtelocentric chromosome pair as revealed by cross-species painting

Leuciscine cyprinids possess a nearly invariant diploid number (2n = 50) with an extremely uniform karyotype comprising of 8 pairs of metacentric, 13-15 pairs of submetacentric and 2-4 pairs of subtelocentric (st) to acrocentric (a) chromosomes. The largest pair is characteristically an st/a element-the 'leuciscine' cytotaxonomic marker. Previously, the interspecific homology of this chromosome pair could not be assessed owing to the inability to produce euchromatic or serial banding patterns. In the present study, we used laser-microdissection (15-20 copies of the marker chromosome) to construct a whole chromosome probe (WCP) from the marker chromosome of the roach Rutilus rutilus to ascertain the interspecific homology of marker chromosomes by cross-species in-situ hybridization. WCP was hybridized to chromosomes of widely distributed (Abramis brama, Alburnoides bipunctatus, Alburnus alburnus, Aspius aspius, Ballerus ballerus, B. sapa, Blicca bjoerkna, Chondrostoma nasus, Leucaspius delineatus, Leuciscus leuciscus, L. idus, R. rutilus, Scardinius erythrophthalmus, Squalius cephalus, and Vimba vimba) and Iberian endemic species (Achondrostoma oligolepis, Iberochondrostoma almacai, I. lusitanicum, Pseudochondrostoma duriense, S. alburnoides and S. pyrenaicus). Cross-species in-situ hybridization to chromosomes of Phoxinus phoxinus, a representative of leuciscine sister lineage, showed the same pattern as in all of the leuciscins. The probe consistently hybridized to the distal part of the short arm of the marker chromosome, indicating sequence homology.

Mesabolivar brasiliensis (Moenkhaus 1898) and Mesabolivar cyaneotaeniatus (Keyserling 1891) (Araneomorphae, Pholcidae): close relationship reinforced by cytogenetic analyses

Pholcidae is the most diverse family among haplogyne spiders but only 15 species have been analyzed cytogenetically. These studies revealed that the diploid number varies from 2n 5 15 to 2n 5 32, that there are three types of sex chromosome systems in males (X, X1X2 and X1X2Y), and that the chromosomes are predominantly biarmed. Within the genus Mesabolivar, only Mesabolivar luteus (Keyserling 1891) has been karyotyped, and it showed 2n 5 15 5 14 + X, with all chromosomes being metacentric. In the present work, we characterize the mitotic and meiotic chromosomes of Mesabolivar brasiliensis (Moenkhaus 1898) and Mesabolivar cyaneotaeniatus (Keyserling 1891). Male mitotic metaphases of the two species showed the diploid number 2n 5 17 5 16 + X; oogonial cells of M. brasiliensis showed 2n 5 18 5 16 + XX. In both species, the chromosomes were exclusively biarmed, and the X chromosome was the largest element of the karyotype. Diplotene spermatocytes of the two species exhibited 8II + X and the occurrence of only one terminal or interstitial chiasma per bivalent. In M. cyaneotaeniaus, metaphases II with n 5 9 5 8 + X and n 5 8 were found, indicating the regular segregation of all chromosomes during meiosis I. Mitotic metaphases of M. brasiliensis stained with CMA3/DA/ DAPI revealed GC-rich chromatin in the terminal region of almost all autosomes, especially in pair 2. An earlier revision of the New World pholcids grouped M. brasiliensis and M. cyaneotaeniatus in a ''southern group'' and placed M. luteus in a ''miscellaneous group.'' A molecular study showed a closer relationship between M. brasiliensis and M. cyaneotaeniatus than between M. luteus and either of these two species. The 2n 5 17 found in M. brasiliensis and M. cyaneotaeniatus corroborates this hypothesis, given that M. luteus has a diploid number of 2n 5 15. According to Platnick (2008), the family Pholcidae Koch 1851 includes 999 extant species, and thus constitutes the most diverse family among haplogyne spiders. A phylogenetic analysis based on morphological characters separated pholcids into four clades: ''ninetines,'' ''pholcines,'' ''holocnemines,'' and the ''New World clade'' (Huber 2000). The genus Mesabolivar Gonzalez-Sponga 1998, which ranges from northern South America to northern Argentina, is included in the ''New World clade'' and possesses 45 species (Huber 2000, 2008). Moreover, the genus is divided into four operational groups based on morphological characters: a ''northern group with spines on male metatarsi,'' a ''northern group without spines on male metatarsi,'' a ''southern group,'' and a ''miscellaneous group.'' The species Mesabolivar brasiliensis (Moenkhaus 1898) and Mesabolivar cyaneotaenia- tus (Keyserling 1891), both part of the ''southern group,'' are distributed in southern and eastern Brazil and northern

Comparative karyological analysis of three members of Allocreadiidae (Digenea): taxonomic and phylogenetic implications

Chromosome number and morphology were studied in three allocreadiid species from mitotic metaphases of intramolluscan stages using karyometric analysis. Bunodera luciopercae has a diploid number 2n=14, with two large pairs of metacentrics, one pair of submeta-subtelocentrics, and four pairs of acrocentrics. Karyotype of Allocreadium isoporum, 2n=14, with two large pairs of metacentrics and five pairs of acrocentrics was recognized to be closely related to that of B. lucioperca. The most conspicuous interspecific difference occurs in the centromere position of the chromosomes of the pair 3. The possible reasons of the discrepancies between data reported therein and the karyological information previously published on these two species are discussed. Crepidostomum sp. has 2n=12, with one pair of large metacentrics and five pairs of acrocentrics and is more distantly related karyologically to the other two members of the family. Evolutionary considerations based on comparison of the chromosomes of Allocreadiidae and other studied families of the suborder Xiphidiata suggest that allocreadiids are karyotypically distinct. The placement of Allocreadiidae and Opecoelidae in the superfamily Allocreadioidea is questioned.

A new species of the genus Alsodes (Anura: Neobatrachia) from the Nothofagus forest, Coastal Range, Southern Chile, identified by its karyotype

Based on morphology, distribution, and on the presence of a novel diploid number (2n = 30 chromosomes), a new species of frog Alsodes norae from the Coastal Range, Valdivia Province temperate Nothofagus forest, Southern Chile, is described. Hypothetical chromosomal rearrangements and the origin of its unusual karyotype are discussed. Among the South American frogs, the genus Alsodes is recognized by its complex relationships, and based on karyological and geographical distribution information its species are grouped in two larger groups, herein: the Andean group which is conservative, and the Coastal group which is far more variable in gross chromosome morphology. Additionally, the inclusion of Alsodes in the new family Cycloramphidae is commented.

High levels of chromosomal differentiation in Euchroma gigantea L. 1735 (Coleoptera, Buprestidae)

Euchroma gigantea was karyotypically studied using conventional staining, C-banding, silver nitrate staining and ribosomal fluorescent in situ hybridization (rDNA FISH). Broad wide autosomal polymorphism and a complex sex determination system were found in this beetle. Karyotype complements ranging from 2n = 32, X1X2X3Y1Y2Y3 to 2n = 36,X1X2X3Y1Y2Y3 were detected in the sample analyzed. Punctiform supernumerary chromosomes were present in the different karyotypes. The karyotypic evolution of Brazilian E. gigantea may have taken two directions, reduction in the diploid number of 2n = 36 to 24 through centric fusions or 2n = 24 to 36 due to chromosomal fissions. In addition, pericentric inversions were also involved. The complex multiple sex mechanism of this species seems to be old and well established since it is found in specimens from different populations. Small pericentromeric blocks of constitutive heterochromatin were located on the autosomes and terminal blocks were also found on some small pairs. The sex chromosomes showed larger constitutive heterochromatin blocks. Silver nitrate staining during prophase I of meiosis showed labeling of the sex chromosome chain. However, the rDNA sites could only be precisely determined by FISH, which permitted the identification of these ribosomal sites on chromosomes X1 and X2 of this species.

Cytogenetic analyses of two endemic fish species from the São Francisco River basin: Conorhynchus conirostris and Lophiosilurus alexandri (Siluriformes)

Two Siluriformes species endemic to the Sao Francisco River basin were characterized by conventional and differential cytogenetic analyses involving C-banding, Ag-nucleolar organizer region (NOR) and chromomycin A3 (CMA3) staining, and FISH (fluorescent in situ hybridization) with 18S and 5S rDNA probes. Conorhynchus conirostris presents a higher diploid number (2n = 60) than those detected in Pimelodidae representatives, whereas Lophiosilurus alexandri, with a karyotype of 2n = 54 chromosomes, presents a chromosomal constitution similar to that found in the family Pseudopimelodidae. Plesiomorphic characteristics such as single NORs at terminal positions are found in both species, as revealed by CMA3 and silver nitrate staining, and FISH with a 18S rDNA probe. C-banding evidenced centromeric and telomeric heterochromatic blocks distributed over most of the chromosomes with a conspicuous heterochromatin segment in a pair of submetacentric chromosomes in L. alexandri. Such karyotype data, if compared to the cytogenetic pattern of other Siluriformes species, can be partially related to their degree of endemism, favorable to the occurrence and fixation of chromosomal rearrangements. The present study in representatives from these two Siluriformes families from the Sao Francisco River contributes to a better understanding of the karyotype evolution in species of this important order of Neotropical fishes.

Cytogenetic markers as diagnoses in the identification of the hybrid between Piauçu (Leporinus macrocephalus) and Piapara (Leporinus elongatus)

The genetic monitoring of interspecific hybrids involves the application of methodologies able to provide an easy and indubitable genetic characterization of both parental and hybrid individuals. In the present work, cytogenetic techniques were used to identify a hybrid lineage of Piaupara in order to caracterize them in relation to the parental species, Leporinus macrocephalus (piaucu) and L. elongatus (piapara). The cytogenetic analysis revealed that L. macrocephalus presented 2n = 54 chromosomes and a nucleolar organizer regions (NOR) at the telomere of the long arm of the submetacentric chromosome pair 2. Analysis of constitutive heterochromatin (C-banding) revealed a conspicuous block at the pericentromeric region on the long arm of a submetacentric chromosome pair. L. elongatus presented the same diploid number, 2n = 54, and a karyotypic formula similar to that of L. macrocephalus. The NORs were also at the telomere of the long arm of the submetacentric pair 2, which was morphologically different from that of L. macrocephalus. Heterochromatic blocks were observed at both telomeres of a submetacentric chromosome pair. The hybrid Piaupara presented the same diploid number (2n = 54) and karyotypic formula as the parental species and there were no visible differences between parental and hybrid individuals. Differently from the Giemsa staining, NOR- and C-banding analysis showed marked differences which allowed the identification of the hybrids by the different morphology and/or size of the chromosomes carrying the NORs and patterns of heterochromatin distribution in their chromosomes. Such genetic studies are important for fish culture since they can provide tools for monitoring natural and artificial hybridization. They are also useful in biological conservation programmes and in the proper management of natural and reared fish stocks.

Banded karyotypes of the Persian squirrel Sciurus anomalus from Turkey

Abstract Karyotypes of seven males and three females of the Persian squirrel, Sciurus anomalus, were analyzed with G- and C-banding techniques and Ag-NOR (nucleolus organizer region) staining. All specimens had the diploid number (2n) of 40 and fundamental autosomal arm number (FN) of 76. The autosomal complement comprised 6 pairs of metacentrics and 13 pairs of submetacentrics. The X chromosome was a medium-sized metacentric. The Y chromosome was a small submetacentric. In the C-banded karyotype, except for centromeric heterochromatin, heterochromatin blocks were absent. Only the terminal region of one medium-sized metacentric pair had detectable Ag-NORs. The uniqueness of the chromosomal characteristics of S. anomalus suggests that S. anomalus is distantly related to the other Old World Sciurus species (S. lis and S. vulgaris).

Chromosome divergence and NOR polymorphism in Bryconamericus aff. iheringii (Teleostei, Characidae) in the hydrographic systems of the Paranapanema and Ivaí Rivers, Paraná, Brazil

Cytogenetic studies were carried out in three populations of Bryconamericus aff. iheringii from two hydrographic systems of the Paranapanema and Ivaí Rivers, separated by a watershed, both belonging to the upper Paraná River basin. Specimens had a constant diploid number 2n = 52 chromosomes. However, three karyotype formulae were identified in the three populations: B. aff. iheringii from the Maringá stream had 12M+18SM+8ST+14A (FN = 90); specimens from Keller River showed 8M+28SM+6ST+10A (FN = 94) and specimens from the Tatupeba stream had 8M+20SM+8ST+16A (FN = 88). Nucleolar organizer regions (NORs) were identified by silver nitrate staining and fluorescent in situ hybridization (FISH) with an 18S rDNA probe. Specimens from Tatupeba stream had a simple NOR system located in a terminal position of the short arm of a pair of large submetacentric chromosomes. Ag-NOR and FISH methodologies revealed multiple NORs in specimens of the Maringá stream and Keller River. Differences in chromosome structure and in NOR patterns in the three populations of B. aff. iheringii revealed fixed evolutionary chromosome divergence. Aspects related to karyotypic variations and to geographic isolation of these populations are discussed.

CONTEOS CROMOSOMICOS EN PHILESIA MAGELLANICA J.F.GMEL. (LILIALES: PHILESIACEAE)

Mitotic chromosome counts documented here for Philesia magellanica J.F.Gmel. (Philesiaceae) showed a diploid number 2n = 30 (x = 15), which is different to the numbers reported previously for the species. The 2n number of P. magellanica is similar to that documented previously for Lapageria rosea, which indicate a constancy of chromosome number within the family

Pronounced karyological divergence of the North American congeners Sphaerium rhomboideum and S. occidentale (Bivalvia: Veneroida: Sphaeriidae)

Chromosome sets of two North American sphaeriid species, Sphaerium rhomboideum Say, 1822 and S. occidentale Lewis, 1856, were studied using conventional Giemsa staining and karyometric analysis. Pronounced karyological divergence of congeners was revealed. The diploid number of 2n ¼ 44 was reported for S. rhomboideum and this is the first record of a diploid species in the highly polychromosomic Nearctic sphaeriid fauna. The karyotype was characterized by medium-sized and small chromosomes, which decreased in size gradually from 5.77 to 1.9 mm. Biarmed chromosomes with medially and submedially located centromeres predominated, but six pairs of subtelo-telocentric elements were also observed in the karyotype. The estimated mitotic chromosome number for S. occidentale ranges from 189 to 213, but most of the cells examined contained about 204– 209 chromosomes. A first attempt to karyotype a polyploid sphaeriid was made. It was revealed that the comparatively large and middle-sized chromosomes could be grouped in four, so the karyotype presumably evolved through tetraploidization. The small chromosomes formed the large fraction, about 137. Due to their similar and indistinct morphologies, it was impossible to arrange them into subgroups with confidence. Revealed karyological characteristics are discussed with reference to the existing phylogenetic interpretations of the evolutionary history of the Sphaeriinae.

Comparative chromosome painting map between two Ryukyu spiny rat species, Tokudaia osimensis and Tokudaia tokunoshimensis (Muridae, Rodentia)

Ryukyu spiny rats (genus Tokudaia) are indigenous species that are confined to three islands of the Nansei Shoto archipelago, Amami-Oshima, Tokunoshima and Okinawa-jima, Japan. Tokudaia tokunoshimensis from Tokunoshima Island and Tokudaia osimensis from Amami-Oshima Island are closely related taxonomically, although their karyotypes are quite different: the diploid chromosome numbers and sex chromosome constitution are 2n=45, X0/X0 for T. tokunoshimensis and 2n=25, X0/X0 for T. osimensis. We conducted comparative chromosome painting with chromosome-specific DNA probes of the laboratory mouse (Mus musculus) to molecularly examine the chromosome homology between T. tokunoshimensis and T. osimensis, and deduced a possible ancestral karyotype of Tokudaia species and the process of evolutionary chromosome rearrangements. The proposed ancestral karyotype with the diploid number of 2n=48, XX/XY was similar to the karyotype of T. tokunoshimensis, and the karyotype of T. osimensis would then have been established through at least 14 chromosomal changes, mainly centric fusion and tandem fusion, from the ancestral karyotype. The close karyological relationship between the ancestral karyotypes of Tokudaia and Apodemus also suggests that the chromosomal evolution in the Tokudaia-Apodemus lineage has been very slow and has accelerated only recently in the branch leading to T. osimensis.