GEOGRAPHIC POPULATION STRUCTURE AND SPECIES DIFFERENCES IN MITOCHONDRIAL DNA OF MOUTHBROODING MARINE CATFISHES (ARIIDAE) AND DEMERSAL SPAWNING TOADFISHES (BATRACHOIDIDAE)

Restriction-fragment length polymorphisms in mitochondrial DNA (mtDNA) were used to evaluate population-genetic structure and matriarchal phylogeny in four species of marine fishes that lack a pelagic larval stage: the catfishes Arius felis and Bagre marinus, and the toadfishes Opsanus tau and O. beta. Thirteen informative restriction enzymes were used to assay mtDNAs from 134 specimens collected from Massachusetts to Louisiana. Considerable genotypic diversity was observed in each species. However, major mtDNA phylogenetic assemblages in catfish and toadfish (as identified in Wagner networks and UPGMA phenograms) exhibited contrasting patterns of geographic distribution: in catfish, distinct mtDNA clades were widespread, while such clades in toadfish tended to be geographically localized. By both the criteria of species' ranges and the geographic pattern of intraspecific mtDNA phylogeny, populations of marine catfish in the western Atlantic have had greater historical interconnectedness than have toadfish. Results are also compared to previously published mtDNA data in freshwater and other marine fishes. Although mtDNA differentiation among conspecific populations of continuously distributed marine fishes is usually lower than that among discontinuously distributed freshwater species inhabiting separate drainages, it is apparent that historical biogeographic factors can importantly influence genetic structure in marine as well as freshwater species.     

Evolution of opercle bone shape along a macrohabitat gradient: species identification using mtDNA and geometric morphometric analyses in neotropical sea catfishes (Ariidae)

Transitions between the marine and freshwater macrohabitat have occurred repeatedly in the evolution of teleost fishes. For example, ariid catfishes have moved from freshwater to marine environments, and vice versa. Opercles, a skeletal feature that has been shown to change during such transitions, were subjected to 2D geometric morphometric analyses in order to investigate evolutionary shape changes during habitat transition in ariid catfishes and to test the influence of habitat on shape changes. A mtDNA marker, which proved useful in previous studies, was used to verify species identities. It greatly improved the assignment of specimens to a species, which are difficult to assign by morphology alone. The application of a mtDNA marker confirmed the occurrence of Notarius biffi in Central America, South of El Salvador. Molecular identification together with principal component analysis (PCA) and further morphological inspection of neurocrania indicated the existence of a cryptic species within Bagre pinnimaculatus. Principal component (PC) scores of individual specimens clustered in morphospace by genus rather than by habitat. Strong phylogenetic structure was detected using a permutation test of PC scores of species means on a phylogenetic tree. Calculation of Pagel's λ suggested that opercle shape evolved according to a Brownian model of evolution. Yet canonical variate analysis (CVA) conducted on the habitat groups showed significant differences in opercle shapes among freshwater and marine species. Overall, opercle shape in tropical American Ariidae appears to be phylogenetically constrained. This verifies the application of opercle shape as a taxonomic tool for species identification in fossil ariid catfishes. At the same time, adaptation to freshwater habitats shows characteristic opercle shape trajectories in ariid catfishes, which might be used to detect habitat preferences in fossils.     

Trends in chromosome evolution in the genus Hypostomus Lacépède, 1803 (Osteichthyes, Loricariidae): a new perspective about the correlation between diploid number and chromosomes types

Phylogenetic relationships and identification of species of the genus Hypostomus is still unclear. Considering this, cytogenetics may prove itself as an important tool in understanding the systematic of this genus. Reviews in Hypostomus indicate that the diploid number ranges from 54 to 84 chromosomes, and the increase in diploid number has been associated to higher percentages of subtelocentric and acrocentric chromosomes. Although there is a high number of species in the genus, there are relatively few papers concerning Hypostomus cytogenetics, and most of the data is published as grey literature. With the aim to understand the chromosomal evolution in the genus (correlation between diploid number x chromosomes types), H. ancistroides and H. topavae from the Piquiri River, Upper Paraná River basin, were cytogenetically analyzed, and the diploid number observed was 68 and 80 chromosomes, respectively. Additional data on the diploid number and chromosome formulae was compiled from papers (27 analyses) and abstracts from grey literature (77 analyses). Our analysis shows no correlation between chromosome numbers and percentages of subtelocentric and acrocentric chromosomes for most of the species, since there is considerable variation between these percentages even between species with the same diploid number, indicating that the proportion of chromosome types is not always associated to diploid numbers.     

Cytogenetic and Molecular Analyses Reveal a Divergence between Acromyrmex striatus (Roger, 1863) and Other Congeneric Species: Taxonomic Implications

The leafcutter ants, which consist of Acromyrmex and Atta genera, are restricted to the New World and they are considered the main herbivores in the neotropics. Cytogenetic studies of leafcutter ants are available for five species of Atta and 14 species of Acromyrmex, both including subspecies. These two ant genera have a constant karyotype with a diploid number of 22 and 38 chromosomes, respectively. The most distinct Acromyrmex species from Brazil is A. striatus, which is restricted to the southern states of Santa Catarina and Rio Grande do Sul. Several cytogenetic and phylogenetic studies have been conducted with ants, but the karyotypic characterization and phylogenetic position of this species relative to leafcutter ants remains unknown. In this study, we report a diploid number of 22 chromosomes for A. striatus. The phylogenetic relationship between A. striatus and other leafcutter ants was estimated based on the four nuclear genes. A. striatus shared the same chromosome number as Atta species and the majority of metacentric chromosomes. Nuclear data generated a phylogenetic tree with a well-supported cluster, where A. striatus formed a different clade from other Acromyrmex spp. This combination of cytogenetic and molecular approaches provided interesting insights into the phylogenetic position of A. striatus among the leafcutter ants and the tribe Attini.     

Reconstruction of the Doradinae (Siluriformes-Doradidae) ancestral diploid number and NOR pattern reveals new insights about the karyotypic diversification of the Neotropical thorny catfishes

Doradinae (Siluriformes: Doradidae) is the most species-rich subfamily among thorny catfishes, encompassing over 77 valid species, found mainly in Amazon and Platina hydrographic basins. Here, we analyzed seven Doradinae species using combined methods (e.g., cytogenetic tools and Mesquite ancestral reconstruction software) in order to scrutinize the processes that mediated the karyotype diversification in this subfamily. Our ancestral reconstruction recovered that 2n=58 chromosomes and simple nucleolar organizer regions (NOR) are ancestral features only for Wertheimerinae and the most clades of Doradinae. Some exceptions were found in Trachydoras paraguayensis (2n=56), Trachydoras steindachneri (2n=60), Ossancora punctata (2n=66) and Platydoras hancockii whose karyotypes showed a multiple NOR system. The large thorny catfishes, such as Pterodoras granulosus, Oxydoras niger and Centrodoras brachiatus share several karyotype features, with subtle variations only regarding their heterochromatin distribution. On the other hand, a remarkable karyotypic variability has been reported in the fimbriate barbells thorny catfishes. These two contrasting karyoevolution trajectories emerged from a complex interaction between chromosome rearrangements (e.g., inversions and Robertsonian translocations) and mechanisms of heterochromatin dispersion. Moreover, we believe that biological features, such as microhabitats preferences, populational size, low vagility and migratory behavior played a key role during the origin and maintenance of chromosome diversity in Doradinae subfamily.     

Repetitive DNA Sequences and Evolution of ZZ/ZW Sex Chromosomes in Characidium (Teleostei: Characiformes)

Characidium constitutes an interesting model for cytogenetic studies, since a large degree of karyotype variation has been detected in this group, like the presence/absence of sex and supernumerary chromosomes and variable distribution of repetitive sequences in different species/populations. In this study, we performed a comparative cytogenetic analysis in 13 Characidium species collected at different South American river basins in order to investigate the karyotype diversification in this group. Chromosome analyses involved the karyotype characterization, cytogenetic mapping of repetitive DNA sequences and cross-species chromosome painting using a W-specific probe obtained in a previous study from Characidium gomesi. Our results evidenced a conserved diploid chromosome number of 2n = 50, and almost all the species exhibited homeologous ZZ/ZW sex chromosomes in different stages of differentiation, except C. cf. zebra, C. tenue, C. xavante and C. stigmosum. Notably, some ZZ/ZW sex chromosomes showed 5S and/or 18S rDNA clusters, while no U2 snDNA sites could be detected in the sex chromosomes, being restricted to a single chromosome pair in almost all the analyzed species. In addition, the species Characidium sp. aff. C. vidali showed B chromosomes with an inter-individual variation of 1 to 4 supernumerary chromosomes per cell. Notably, these B chromosomes share sequences with the W-specific probe, providing insights about their origin. Results presented here further confirm the extensive karyotype diversity within Characidium in contrast with a conserved diploid chromosome number. Such chromosome differences seem to constitute a significant reproductive barrier, since several sympatric Characidium species had been described during the last few years and no interespecific hybrids were found.     

THE QUESTION OF POST-REDUCTION IN SPHAGNUM

The 19 spatially distinct chromosomal units at first meiotic metaphase in sporophytically diploid species of Sphagnum have usually been considered to be bivalents, but one investigator (Sorsa, 1956) has interpreted them as chromosomes from dissociated bivalents and meiosis as post-reductional. The present studies on diploid S. squarrosum (Pers.) Crome establish the chromosome number on the basis of the following evidence: there are in addition to m-chromosomes, 19 pairs of chromosomes in early prophase, 19 bivalents at diakinesis, 19 chromosomes in each of the two sets at second metaphase, 19 daughter chromosomes in each of the four sets at late second anaphase, and 19 chromosomes in gametophytic mitoses. The 19 bodies at first meiotic metaphase in diploid species are true bivalents in loose secondary association, which has led to their erroneous interpretation as chromosomes of dissociated bivalents. The gametic chromosome number in sporophytically diploid Sphagnum is therefore, without doubt, n = 19, and this evidence negates the claim for post-reduction in Sphagnum.     

Chromosome number and secondary chromosomal associations in wild populations of Geranium pratense L. from the cold deserts of Lahaul-Spiti (India)

In this work we studied the meiotic chromosome number and details of secondary chromosomal associations recorded for the first time in Geranium pratense L. from the alpine environments in the cold deserts of Lahaul-Spiti (India). All the presently studied individuals of the species existed at 4x level (x = 14). The present chromosome count of n = 28 in the species adds a new cytotype to the already existing diploid chromosome count of 2n = 28 from the Eastern Himalayas and outside of India. Out of the six accessions scored presently four showed normal meiotic course. However, two accessions investigated from Mud, 3800 m and Koksar, 3140 m depicted abnormal meiotic course due to the presence of multivalents and univalents, and secondary associations of bivalents/chromosomes. The secondary chromosomal associations in the species existed among bivalents/chromosomes were noticed in the PMCs at prophase-I (diakinesis) and persisted till the separation of sister chromatids at M-II. The variation in the number of bivalents/chromosomes involved in the secondary associations at M-I (2–8) and A-I/M-II (2–12) has also been recorded. The occurrence of such secondary associations of bivalents/chromosomes in G. pratense which existed at 4x level indicated the secondary polyploid nature of the species.     

Gametogenesis and the Chromosomes of Meloidogyne nataliei: Not Typical of Other Root-knot Nematodes

Studies of oogenesis and spermatogenesis revealed that Meloidogyne nataliei is a diploid, amphimictic species with four (n), relatively large chromosomes, and possibly with an XX ♀-XY ♂ mechanism of sex determination. It differs considerably from all other amphimictic, or meiotically parthenogenetic, species of Meloidogyne which have 13-18 smaller chromosomes and from Meloidogyne (Hypsoperine) spartinae which has seven. Consequently, the taxonomic position of M. nataliei needs to be re-evaluated. The chromosomes of M. nataliei and their behavior during gametogenesis resemble more closely chromosomes of the genus Heterodera than those of the genus Meloidogyne. This resemblance, however, may not imply a closer phyletic relationship of M. nataliei to heteroderid nematodes.     

Genomic analyses of the Formosan harvest mouse (Micromys minutus) and comparisons to the brown Norway rat (Rattus norvegicus) and the house mouse (Mus musculus)

The harvest mouse, Micromys minutus (MMIN), has a very wide range of distribution (from the British Isles across the Euroasian continent to Japan and Taiwan). We studied an isolated population of MMIN in Taiwan, which is at the southeastern margin of the species’ geographic distribution, and compared its genetic complement with those of the same species previously reported from other geographic locations and with two model rodent species, the house mouse (Mus musculus) and the brown Norway rat (Rattus norvegicus). The diploid number (2N) of MMIN was 68, consistent with that reported for other populations. However, variations were noted in the fundamental number (FN) and the shape and banding patterns of the individual chromosomes among populations. The FN of MMIN was estimated to be 72, including 2 bi-armed autosomes, 31 one-armed autosomes, and one pair of one-armed sex chromosomes. Here, we propose the first ideogram for MMIN. C-banding, Ag-NOR, and the locations of 18S rRNA gene sequences (MMIN chromosomes no. 10, 14, 19, 29, 31, 33, and X) mapped by fluorescence in situ hybridization (FISH) are also reported. Additionally, we compared the 18S rDNA sequences and performed cross-species X chromosome painting (FISH) for M. minutus, M. musculus, and R. norvegicus. The results indicate that both genetic elements are rather conserved across species. Thus, implications for the phylogenetic position of Micromys were limited.     

Single-copy gene fluorescence in situ hybridization and genome analysis: Acc-2 loci mark evolutionary chromosomal rearrangements in wheat

Fluorescence in situ hybridization (FISH) is a useful tool for physical mapping of chromosomes and studying evolutionary chromosome rearrangements. Here we report a robust method for single-copy gene FISH for wheat. FISH probes were developed from cDNA of cytosolic acetyl-CoA carboxylase (ACCase) gene (Acc-2) and mapped on chromosomes of bread wheat, Triticum aestivum L. (2n?=?6x?=?42, AABBDD), and related diploid and tetraploid species. Another nine full-length (FL) cDNA FISH probes were mapped and used to identify chromosomes of wheat species. The Acc-2 probe was detected on the long arms of each of the homoeologous group 3 chromosomes (3A, 3B, and 3D), on 5DL and 4AL of bread wheat, and on homoeologous and nonhomoeologous chromosomes of other species. In the species tested, FISH detected more Acc-2 gene or pseudogene sites than previously found by PCR and Southern hybridization analyses and showed presence/absence polymorphism of Acc-2 sequences. FISH with the Acc-2 probe revealed the 4A–5A translocation, shared by several related diploid and polyploid species and inherited from an ancestral A-genome species, and the T. timopheevii-specific 4A^t–3A^t translocation.     

Chromosome studies of the cultured cells of two species of side-necked turtles (Podocnemis unifilis and P. expansa)

The diploid chromosome number of two species of sidenecked turtles (Podocnemis unifilis and P. expansa) was found to be 28. Under normal culture conditions, half of the chromosomes of P. unifilis consistently show one or two clear secondary constrictions. In P. expansa, the incidence of cells with chromosomes bearing secondary constrictions and the number of such chromosomes per cell are less. Cells of two P. unifilis cell lines maintained a normal diploid karyotype for two years following their initiation. Then one cell line shifted to a hypodiploid mode of 27 and half of the population of the second line became pseudodiploid, the other half remaining diploid. A single six-month-old cell line from P. expansa has maintained a normal diploid mode through 10 passages.Supported in part by grant-CA 08737 from the National Cancer Institute.     

Comparative cytogenetics of eight species of Cycloramphus (Anura, Cycloramphidae)

Several aspects of the biology of Cycloramphus species of the Atlantic Forest are still poorly known, which makes it difficult to understand their historical relationships. Therefore, we were stimulated to promote a comparative cytogenetic analysis of several species of the genus Cycloramphus. The study of Cycloramphus acangatan, C. boraceiensis, C. brasiliensis, C. carvalhoi, C. eleutherodactylus, C. fuliginosus, C. lutzorum, and C. rhyakonastes, revealed that these eight species share a diploid number 2n = 26. Cycloramphus fuliginosus presented the most distinct karyotype, due to the presence of subtelocentric chromosomes in pairs 1 and 4. The main diagnostic feature observed in the other species was the presence of one pair of telocentric chromosomes in C. boraceiensis, C. carvalhoi, and C. eleutherodactylus, while the remaining species presented karyotypes composed exclusively of biarmed chromosomes. Constitutive heterochromatin was predominantly located in pericentromeric regions in all species, although additional C-bands detected on telomeric and/or interstitial regions were partially species-specific. Silver staining revealed Ag-NORs located on the pair 6 in six species, whereas C. acangatan presented it on pair 1 and a multiple pattern was observed in C. fuliginosus with three Ag-NOR bearing chromosomes. Fluorescent in situ hybridization using rDNA probe was performed in specimens of C. eleutherodactylus from Paraná, C. lutzorum, and C. rhyakonastes, which did not reveal inactive NOR. Despite the apparent highly conserved diploid number, data on the karyotype microstructure characterize the cytogenetic profile of the genus and may contribute to clarify the phylogenetic relationships among Cycloramphus, the Cycloramphinae, or even the family Cycloramphidae.     

The karyotype of Nodipecten nodosus (Bivalvia: Pectinidae)

Earlier karyotypical work on Nodipecten nodosus embryos indicated that this species has a diploid number of 38, with six pairs respectively of metacentric and submetacentric chromosomes and seven pairs of subtelocentric chromosomes, although there were some difficulties in obtaining complete metaphases. The present work provides additional results on specific regions of the chromosomes in N. nodosus and, by meiotic studies, confirms the chromosome number with more reliability. Active nucleolar organizer regions (NOR), detected in mitotic metaphases from embryos, can be characterized in N. nodosus by a high level of heteromorphism of NOR-sites, indicating that these regions are not appropriate as chromosomal markers in this species. The procedure for detecting constitutive heterochromatin of chromosomes allowed us to observe most of the heterochromatic blocks at a pericentromeric position and some at telomeric and interstitial positions. The analysis of meiotic chromosomes from gonad tissue revealed the presence of 19 bivalents during metaphase I, all homomorphic and isopicnotic, confirming the previously described diploid chromosomal number of 38 for N. nodosus. From these results, some evolutionary aspects of the Pectinidae are briefly discussed.     

Chromosome studies in species and hybrids ofPetrorhagia sect.Kohlrauschia (Caryophyllaceae)

Cytogenetic investigations have been made in the fourPetrorhagia species and hybrids of the sectionKohlrauschia. The three diploid species show close similarities in chromosome number, size and morphology, with the exception ofP. velutina, where one pair of metacentric chromosomes is represented by a pair of telocentrics. Meiotic studies in hybrids indicate close genomic homology between the diploid species and also between the two floral forms ofP. prolifera. The tetraploidP. nanteuilii behaves as an allotetraploid forming only bivalents at meiosis and results suggest thatP. velutina andP. prolifera are the diploid progenitors of this species. Since meiosis in diploid and triploid hybrids results in extensive intergenomic pairing it is concluded that the natural tetraploid has a bivalent promoting mechanism that prevents pairing between the genomes of its diploid progenitors.     

POLYPLOIDY,DYSPLOIDY, AND CHROMOSOME PAIRING IN ECHEVERIA (CRASSULACEAE) AND ITS HYBRIDS

The 140+ species of Echeveria have more than 50 gametic chromosome numbers, including every number from 12 through 34 and polyploids to n = ca. 260. With related genera, they comprise an immense comparium of 200+ species that have been interconnected in cultivation by hybrids. Some species with as many as 34 gametic chromosomes include none that can pair with each other, indicating that they are effectively diploid, but other species with fewer chromosomes test as tetraploids. Most diploid hybrids form multivalents, indicating that many translocations have rearranged segments of the chromosomes. Small, nonessential chromosomal remnants can be lost, lowering the number and suggesting that higher diploid numbers (n = 30–34) in the long dysploid series are older. These same numbers are basic to most other genera in the comparium (Pachyphytum, Graptopetalum, Sedum section Pachysedum), and many diploid intergeneric hybrids show very substantial chromosome pairing. Most polyploid hybrids here are fertile, even where the parents belong to different genera and have very different chromosome numbers. This seems possible only if corresponding chromosomes from a polyploid parent pair with each other preferentially, strong evidence for autopolyploidy. High diploid numbers here may represent old polyploids that have become diploidized by loss, mutation, or suppression of duplicate genes, but other evidence for this is lacking. Most species occur as small populations in unstable habitats in an area with a history of many rapid climatic and geological changes, presenting a model for rapid evolution.     

Chromosome relationships between Lolium and Festuca (Gramineae)

With a view to eclucidating chromosome relationships between Lolium perenne (Lp), L. multiflorum (Lm) and Festuca pratensis (Fp), chromosome pairing in different diploid (2n=14), auto-allotriploid (2n=3x=21), trispecific (2n=3x=21), amphidiploid (2n=4x=28) and auto-allohexaploid (2n=6x=42) hybrids between them was analysed. At all these levels of ploidy there was very good chiasmate pairing between the chromosomes of the three species and, on the whole, there was little evidence of preferential pairing of the chromosomes of a particular species in the triploid, tetraploid and hexaploid hybrids. A critical test for this also came from the synaptic ability of the chromosomes of the single genome with those of the duplicated genome in the auto-allotriploids which formed predominantly trivalents with 2, 3 or even 4 chiasmata. Moreover, the homology between the Lp and Lm chromosomes seems strong enough to pass the discrimination limits of the B-chromosomes which do not suppress homoeologous pairing in the Lp LmLm triploid and LpLm diploid hybrids. — The triploids having two genomes of a Lolium species and one of F. pratensis had some male and female fertility which suggested genetic compatibility of the parental chromosomes resulting, presumably, in compensation at the gametic level. Also, the occurrence of comparable chiasma frequencies in the auto-allotriploids and trispecific hybrids showed that they were not markedly affected whether two doses of one genome and one of the other or all the three different genomes from the three species were present. From the trend of chromosome pairing in all these hybrids it is concluded that there is little structural differentiation between the chromosomes of the three species, no effective isolation barrier to gene-flow between them, and that they are closely related phylogenetically, having possibly evolved from a common progenitor. Taxonomic revision of the two Lolium species is suggested.     

Chromosomal inheritance patterns of intergeneric hybrids of ictalurid catfishes: odd diploid numbers with equal parental contributions

Hybrid chromosomal compositions of channel catfish Ictalurus punctatus × black bullhead Ameiurus melas and channel catfish × flathead catfish Pylodictis olivaris were analysed by a computer-based method. The karyotype of each hybrid was highly asymmetric, and the diploid numbers and arm numbers were intermediate to the parental types. The hybrid offspring of channel catfish × black bullhead possessed a diploid number of 59 chromosomes, with an arm number estimate of 87. The hybrid offspring of the channel catfish × flathead catfish cross possessed a diploid number of 57 chromosomes, also with an arm number estimate of 87. Nucleolus organizer regions (NORs) were located on a single pair of chromosomes with symmetric staining intensity in channel catfish and in black bullhead, and on a single pair of chromosomes with asymmetric staining intensity in flathead catfish. The channel catfish × black bullhead hybrid had two unpaired chromosomes that stained positively for NORs. The channel catfish × flathead catfish had three unpaired chromosomes that stained positively for NORs. Specific marker chromosomes were identified in each hybrid. There was no evidence of androgenesis, gynogenesis, polyploidy or aneuploidy in the hybrids. Results of this study, plus information reported previously, indicate that chromosomes of ictalurid catfishes are inherited stably in a haploid pattern with an equal contribution to the genomes of F₁ hybrids, even in intergeneric crosses involving divergent numbers of parental chromosomes.     

B-chromosome frequency stability in <Emphasis Type="Italic">Prochilodus lineatus</Emphasis> (Characiformes,Prochilodontidae)

The genus Prochilodus includes individuals ranging in size from medium to large, being highly relevant for commercial and subsistence fishing. Prochilodus species have a diploid number of 2n = 54 chromosomes and up to seven supernumerary (B) microchromosomes. Previous research has shown that B frequency increased drastically in the Mogi-Guaçu river population of Prochilodus lineatus in the early 1980s, whereas it remained about constant in the 1990s. Here we analyses B frequency in this population during the 2003–2007 period and have found that frequency has not changed significantly since 1987, and that these B chromosomes do not show the intra-individual variation in number that characterized them in the 1980s. This indicates that these B chromosomes have been neutralized, after their invasion, through their mitotic stabilization.