A comparative cytogenetic study of Hypsibarbusmalcolmi and H.wetmorei (Cyprinidae, Poropuntiini).

Cyprininae are a highly diversified but demonstrably monophyletic lineage of cypriniform fishes. Here, the karyotype and chromosomal characteristics of Hypsibarbusmalcolmi (Smith, 1945) and H.wetmorei (Smith, 1931) were examined using conventional, nucleolus organizing regions (NORs) and molecular cytogenetic protocols. The diploid chromosome number (2n) of H.malcolmi was 50, the fundamental number (FN) was equal to 62, and the karyotype displayed 8m + 4sm + 38a with NORs located at the centromeric and telomeric positions of the short arms of chromosome pairs 1 and 2, respectively. 2n of H.wetmorei was 50, FN 78, karyotype 14m + 14sm + 22a with the NORs at the telomeric position of the short arm of chromosome pair 2. 2n and FN in males and females were identical. Fluorescence in situ hybridization using different microsatellite motifs as probes also showed substantial genomic divergence between both studied species. In H.wetmorei, (CAG)n and (CAC)n microsatellites accumulated in the telomeric regions of all chromosomes, while in H.malcolmi, they had scattered signals on all chromosomes. Besides, the (GAA)n microsatellites were distributed along all chromosomes of H.malcolmi, but there was a strong hybridization pattern in the centromeric region of a single pair in H.wetmorei. These cytogenomic difference across the genomes of these Hypsibarbus Rainboth, 1996 species are markers for specific evolutionary differentiation within these two species.

Karyotype description and comparative chromosomal mapping of rDNA and U2 snDNA sequences in Eigenmannialimbata and E.microstoma (Teleostei, Gymnotiformes, Sternopygidae).

The genus Eigenmannia Jordan et Evermann,1896 includes electric fishes endemic to the Neotropical region with extensive karyotype variability and occurrence of different sex chromosome systems, however, cytogenetic studies within this group are restricted to few species. Here, we describe the karyotypes of Eigenmannialimbata (Schreiner et Miranda Ribeiro, 1903) and E.microstoma (Reinhardt, 1852) and the chromosomal locations of 5S and 18S rDNAs (ribosomal RNA genes) and U2 snDNA (small nuclear RNA gene). Among them, 18S rDNA sites were situated in only one chromosomal pair in both species, and co-localized with 5S rDNA in E.microstoma. On the other hand, 5S rDNA and U2 snRNA sites were observed on several chromosomes, with variation in the number of sites between species under study. These two repetitive DNAs were observed co-localized in one chromosomal pair in E.limbata and in four pairs in E.microstoma. Our study shows a new case of association of these two types of repetitive DNA in the genome of Gymnotiformes.

The Satellite DNA Catalogues of Two Serrasalmidae (Teleostei, Characiformes): Conservation of General satDNA Features over 30 Million Years.

Satellite DNAs (satDNAs) are tandemly repeated sequences that are usually located on the heterochromatin, and the entire collection of satDNAs within a genome is called satellitome. Primarily, these sequences are not under selective pressure and evolve by concerted evolution, resulting in elevated rates of divergence between the satDNA profiles of reproductive isolated species/populations. Here, we characterized two additional satellitomes of Characiformes fish (Colossoma macropomum and Piaractus mesopotamicus) that diverged approximately 30 million years ago, while still retaining conserved karyotype features. The results we obtained indicated that several satDNAs (50% of satellite sequences in P. mesopotamicus and 43% in C. macropomum) show levels of conservation between the analyzed species, in the nucleotide and chromosomal levels. We propose that long-life cycles and few genomic changes could slow down rates of satDNA differentiation.

Comparative cytogenetic patterns in Carangidae fishes in association with their distribution range.

Carangidae are an important and widespreaded family of pelagic predatory fishes that inhabit reef regions or open ocean areas, some species occupying a vast circumglobal distribution. Cytogenetic comparisons among representatives of its different tribes help to understand the process of karyotype divergence in marine ecosystems due to the variable migratory ability of species. In this sense, conventional cytogenetic investigations (Giemsa staining, Ag-NORs, and C-banding), GC base-specific fluorochrome staining and FISH mapping of ribosomal DNAs were performed. Four species, Elagatisbipinnulata (Quoy et Gaimard, 1825) and Seriolarivoliana (Valenciennes, 1883) (Naucratini), with circumtropical distributions, Gnathanodonspeciosus (Forsskål, 1775) (Carangini), widely distributed in the tropical and subtropical waters of the Indian and Pacific oceans, and Trachinotuscarolinus (Linnaeus, 1766) (Trachinotini), distributed along the western Atlantic Ocean, were analyzed, thus encompassing representatives of three out its four tribes. All species have diploid chromosome number 2n = 48, with karyotypes composed mainly by acrocentric chromosomes (NF = 50-56). The 18S rDNA/Ag-NORs/GC+ and 5S rDNA loci were located on chromosomes likely homeologs. Karyotypes showed a pattern considered basal for the family or with small variations in their structures, apparently due to pericentric inversions. The migratory capacity of large pelagic swimmers, in large distribution areas, likely restricts the fixation of chromosome changes in Carangidae responsible for a low level of karyotype diversification.

Chromosomal Diversity in Two Allopatric Populations of Farlowella hahni Meinken 1937 (Teleostei: Siluriformes): Cytogenetics and Cytochrome b Analyses.

Farlowella is the second richest genus in Loricariinae, broadly distributed in freshwater streams and rivers of South America. In this article, we aimed to expand on the cytogenetic and molecular data available for two allopatric populations of Farlowella hahni. Both populations had diploid chromosome number 58, but with karyotype differences, indicative of chromosomal rearrangements. C-banding showed large heterochromatic blocks at telomeric regions in acrocentric chromosomes in both populations. Fluorescence in situ hybridization (FISH) revealed a single 18S rDNA site in both populations and a single 5S rDNA site for individuals from lower Paraná River basin (native region) and multiple 5S rDNA sites for individuals from upper Paraná River basin (non-native region). Mitochondrial sequence analyses did not separate the two F. hahni populations. The cytogenetic and molecular data obtained are relevant in a preliminary study and suggested the existence of cryptic diversity and the hypothesis that at least two Farlowella lineages may coexist in the Paraná basin.

Occurrence of euchromatic B chromosomes in natural populations of Moenkhausia bonita and M. forestii (Pisces: Characidae)

Moenkhausia is a highly specious genus among the Characidae, composed of 96 valid species. Only twelve species have a known karyotype. Thus, here are presented the first cytogenetic data of two allopatric populations of Moenkhausia bonita and one of M. forestii, both belonging to the upper Paraná River basin (PR) with discussion on the evolutionary and cytotaxonomic aspects of the genus. The two species presented 2n = 50 chromosomes but different karyotype formulas and occurrence of 1-2 B chromosomes. These elements are small metacentrics in M. bonita and small acrocentrics in M. forestii. In both species, B chromosomes were euchromatic. Ag-NOR sites were found in pair 3 (metacentric), coinciding with fluorescent in situ hybridization (FISH) by the 18S rDNA probe in both species. However, the species differed in terms of the number and position of 5S rDNA sites. Heterochromatic blocks, mapped in M. bonita showed the least amount of heterochromatin in the terminal and pericentromeric regions, while the M. forestii karyotype revealed a greater amount of interstitial heterochromatic blocks. The karyotype distinctions between the two species, including the morphology of B chromosomes, may contribute as a reference in the taxonomic studies in this group.(AU)

Moenkhausia é um gênero altamente especioso dentre os Characidae, composto por 96 espécies válidas, mas apenas doze espécies têm seus cariótipos conhecidos. Portanto, são apresentados aqui os primeiros dados citogenéticos de duas populações alopátricas de Moenkhausia bonita e uma de M. forestii, ambas pertencentes à bacia do alto rio Paraná (PR), com uma ampla discussão sobre os aspectos evolutivos e citotaxonômicos do gênero. As duas espécies apresentaram 2n = 50 cromossomos, mas diferentes fórmulas cariotípicas e ocorrência de 1-2 cromossomos B. Esses elementos são pequenos metacêntricos em M. bonita e acrocêntricos pequenos em M. forestii. Em ambas as espécies, os cromossomos B apresentaram-se eucromáticos. Sítios Ag-NOR foram encontrados no par 3 (metacêntrico), coincidindo com a hibridização fluorescente in situ (FISH) pela sonda 18S rDNA em ambas as espécies. No entanto, as espécies diferiram em termos de número e posição dos sítios de 5S rDNA. Blocos heterocromáticos mapeados em M. bonita revelaram pequena quantidade de heterocromatina nas regiões terminal e pericentromérica, enquanto o cariótipo de M. forestii revelou uma maior quantidade de blocos heterocromáticos intersticiais. As distinções cariotípicas entre as duas espécies, incluindo a morfologia dos cromossomos B, podem contribuir como uma referência em estudos taxonômicos neste grupo.(AU)

Cytomolecular investigations using repetitive DNA probes contribute to the identification and characterization of Characidium sp. aff. C. vidali (Teleostei: Characiformes)

Characidium sp. aff. C. vidali is a species found in coastal streams in southeastern Brazil, which has karyotypic explanatory elements as the occurrence of microstructural variations, keeping the chromosomal macrostructure of the genus. The objective of this study was to apply cytomolecular tools in the chromosomes of Characidium sp. aff. C. vidali to identify characteristics in their karyotype contributing to cytogenetic definition of this species, adding information about the evolution of the chromosomal structure of the group. The species showed 2n = 50 chromosomes and from 1 to 4 additional B microchromosomes. FISH technique showed histone H3 and H4 genes in the short arm of pair 10, and microsatellites (CA)15, (CG)15, (GA)15 and (TTA)10 clustered in the subtelomeric portions of all A chromosomes, with total accumulation by supernumerary. The telomeric probe marked terminal regions of all chromosomes, in addition to the interstitial portion of four pairs, called ITS sites, with these markings being duplicated in two pairs, hence the double-ITS classification. C-banding revealed that supernumerary chromosomes are completely heterochromatic, that ITS sites are C-banding positive, but double-ITS sites are C-banding negative. So, throughout the evolution to Characidium, genomic events are occurring and restructuring chromosomes in populations.(AU)

Characidium sp. aff. C. vidali é uma espécie encontrada em riachos costeiros do sudeste do Brasil, que apresenta elementos cariotípicos elucidativos quanto à ocorrência de variações microestruturais, conservando a macroestrutura cromossômica do gênero. O objetivo deste estudo foi aplicar ferramentas citomoleculares para identificar características no cariótipo de Characidium sp. aff. C. vidali, que contribuam para a definição citogenética desta espécie, agregando informações quanto à evolução da estruturação cromossômica do grupo. A espécie apresentou 2n = 50 cromossomos, além de 1 a 4 microcromossomos B por célula. A FISH mostrou os genes de histona H3 e H4 sintênicos no braço curto do par 10, e os microssatélites (CA)15, (CG)15, (GA)15 e (TTA)10 clusterizados nas porções subteloméricas de todos os cromossomos do complemento A, com grande acúmulo nos supranumerários. A sonda telomérica identificou marcações terminais em todos os cromossomos, além de quatro pares marcados intersticialmente, chamados de sítios ITS, e dois pares com duas marcações intersticiais, chamados de double-ITS. O bandamento C revelou que os cromossomos supranumerários são completamente heterocromáticos, que os sítios ITS são banda C positivos, mas os sítios double-ITS são banda C negativos. Então, ao longo da evolução de Characidium, eventos genômicos estão ocorrendo e reestruturando cromossomos nas populações.(AU)

Taxonomic Diversity Not Associated with Gross Karyotype Differentiation: The Case of Bighead Carps, Genus Hypophthalmichthys (Teleostei, Cypriniformes, Xenocyprididae).

The bighead carps of the genus Hypophthalmichthys (H. molitrix and H. nobilis) are important aquaculture species. They were subjected to extensive multidisciplinary research, but with cytogenetics confined to conventional protocols only. Here, we employed Giemsa-/C-/CMA3- stainings and chromosomal mapping of multigene families and telomeric repeats. Both species shared (i) a diploid chromosome number 2n = 48 and the karyotype structure, (ii) low amount of constitutive heterochromatin, (iii) the absence of interstitial telomeric sites (ITSs), (iv) a single pair of 5S rDNA loci adjacent to one major rDNA cluster, and (v) a single pair of co-localized U1/U2 snDNA tandem repeats. Both species, on the other hand, differed in (i) the presence/absence of remarkable interstitial block of constitutive heterochromatin on the largest acrocentric pair 11 and (ii) the number of major (CMA3-positive) rDNA sites. Additionally, we applied here, for the first time, the conventional cytogenetics in H. harmandi, a species considered extinct in the wild and/or extensively cross-hybridized with H. molitrix. Its 2n and karyotype description match those found in the previous two species, while silver staining showed differences in distribution of major rDNA. The bighead carps thus represent another case of taxonomic diversity not associated with gross karyotype differentiation, where 2n and karyotype structure cannot help in distinguishing between genomes of closely related species. On the other hand, we demonstrated that two cytogenetic characters (distribution of constitutive heterochromatin and major rDNA) may be useful for diagnosis of pure species. The universality of these markers must be further verified by analyzing other pure populations of bighead carps.

GC and Repeats Profiling along Chromosomes-The Future of Fish Compositional Cytogenomics.

The study of fish cytogenetics has been impeded by the inability to produce G-bands that could assign chromosomes to their homologous pairs. Thus, the majority of karyotypes published have been estimated based on morphological similarities of chromosomes. The reason why chromosome G-banding does not work in fish remains elusive. However, the recent increase in the number of fish genomes assembled to the chromosome level provides a way to analyse this issue. We have developed a Python tool to visualize and quantify GC percentage (GC%) of both repeats and unique DNA along chromosomes using a non-overlapping sliding window approach. Our tool profiles GC% and simultaneously plots the proportion of repeats (rep%) in a color scale (or vice versa). Hence, it is possible to assess the contribution of repeats to the total GC%. The main differences are the GC% of repeats homogenizing the overall GC% along fish chromosomes and a greater range of GC% scattered along fish chromosomes. This may explain the inability to produce G-banding in fish. We also show an occasional banding pattern along the chromosomes in some fish that probably cannot be detected with traditional qualitative cytogenetic methods.

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.

Paracentric Inversions Differentiate the Conservative Karyotypes in Two Centropomus Species (Teleostei: Centropomidae).

Centropomus is the sole genus of the Centropomidae family (Teleostei), comprising 12 species widely distributed throughout the Western Atlantic and Eastern Pacific, with 6 of them occurring in the Western Atlantic in extensive sympatry. Their life history and phylogenetic relationships are well characterized; however, aspects of chromosomal evolution are still unknown. Here, cytogenetic analyses of 2 Centropomus species of great economic value (C. undecimalis and C. mexicanus) were performed using conventional (Giemsa, Ag-NOR, and fluorochrome staining, C- and replication banding) and molecular (chromosomal mapping of 18S and 5S rDNA, H2A-H2B and H3 hisDNA, and (TTAGGG)n repeats) approaches. The karyotypes of both species were composed of 48 solely acrocentric chromosomes (2n = 48; FN = 48), but the single ribosomal site was located in varying positions in the long arms of the second largest chromosome pair. Replication bands were generally similar, although conspicuous differences were observed in some chromosome regions. In both species, the histone H3 genes were located on 3 apparently homeologous chromosome pairs, but the exact position of these clusters differed slightly. Interspecific hisDNA and rDNA site displacements can indicate the occurrence of multiple paracentric inversions during the evolutionary diversification of the Centropomus genomes. Although the karyotypes remained similar in both species, our data demonstrate an unsuspected microstructural reorganization between them, driven most likely by a series of paracentric inversions.

First chromosomal analysis of Gymnorhamphichthys britskii: the remarkable lowest diploid value within the family Rhamphichthyidae (Gymnotiformes)

Gymnorhamphichthys britskii is a Neotropical electric fish of family Rhamphichthyidae described from the Paraná-Paraguay system. This study reports the first karyotypic description of G. britskii collected from the upper Paraná river basin, which presented 2n=38 chromosomes, karyotype composed of 14 metacentric, 8 submetacentric, 2 subtelocentric and 14 acrocentric chromosomes, and fundamental number as 62 for both sexes. Heteromorphic sex chromosomes were absent. A single pair of nucleolar organizing regions (NORs) was detected in the submetacentric chromosome pair number 9 by silver staining and confirmed by the 18S rDNA probe. The 5S rDNA was located in a single chromosome pair. Heterochromatic regions were clearly observed in the short arms of the NOR-bearing chromosome pair and in the telomeric positions of most acrocentric chromosomes. Besides the present data are valuable to help in understanding karyotypic evolution in Rhamphichthyidae, data from NORs confirmed the tendency of this family in presenting simple NORs sites, similar to the other Gymnotiformes clades. Yet, the presence of a large heterochromatic block in the NOR-bearing chromosome can be used as cytogenetic markers for G. britskii, and that centric fusions appear to be an important mechanism in the karyotype evolution and differentiation among Gymnotiformes species.(AU)

Gymnorhamphichthys britskii é um peixe neotropical da família Rhamphichthyidae descrita no sistema Paraná-Paraguai. Este estudo relata a primeira descrição cariotípica de G. britskii coletado na bacia do alto rio Paraná, que apresentou 2n = 38 cromossomos, cariótipo composto por 14 metacêntricos, 8 submetacêntricos, 2 subtelocêntricos e 14 acrocêntricos, e número fundamental 62 para ambos sexos. Cromossomos sexuais heteromórficos estavam ausentes. Um único par de regiões organizadoras de nucléolos (RONs) foi detectado no par de cromossomos submetacêntricos número 9 por coloração com prata e confirmado pela sonda DNAr 18S. O DNAr 5S foi localizado em um único par cromossômico. Regiões heterocromáticas foram claramente observadas nos braços curtos do par de cromossomos que carrega a RON e nas posições teloméricas da maioria dos cromossomos acrocêntricos. Além dos dados presentes serem valiosos para auxiliar na compreensão da evolução cariotípica em Rhamphichthyidae, dados de RONs confirmaram a tendência desta família em apresentar sítios simples de RONs, semelhantes aos demais clados de Gymnotiformes. No entanto, a presença de um grande bloco heterocromático no cromossomo portador da RON, pode ser usado como marcador citogenético para G. britskii e as fusões cêntricas parecem ser um mecanismo importante na evolução e diferenciação cariotípica entre as espécies de Gymnotiformes.(AU)

Extensive karyotype variability of African fish genus Nothobranchius (Cyprinodontiformes).

Karyotypes of 65 species of the genus Nothobranchius Peters, 1868 were reviewed and of those 35 examined first time. The results of present study have shown that fishes of the genus Nothobranchius possessed highly diverse karyotypes. The diploid chromosome number (2n) ranged from 16 to 50. The most frequent 2n was 2n = 36 (in 35 species) while the second one 2n = 38 (in 13 species). Proportion of biarmed chromosomes varied from 0 to 95% between species. Diploid chromosome number variability apparently exists as a result of chromosomal fusions or fissions and extensive karyotypic formula alterations promoting by inversions. Multiple sex chromosomes of system X1X1X2X2/X1X2Y type were found only in karyotypes of 5 species. The extensive karyotype variability, unusual for teleosts, of genus Nothobranchius can be likely associated with the characteristics of its life cycle and inhabiting under unstable environment of East African savannah temporal pools.

Chromosomes of Asian cyprinid fishes: cytogenetic analysis of two representatives of small paleotetraploid tribe Probarbini.

BACKGROUND: Polyploidy, although still poorly explored, represents an important evolutionary event in several cyprinid clades. Herein, Catlocarpio siamensis and Probarbus jullieni - representatives of the paleotetraploid tribe Probarbini, were characterized both by conventional and molecular cytogenetic methods. RESULTS: Alike most other paleotetraploid cyprinids (with 2n = 100), both species studied here shared 2n = 98 but differed in karyotypes: C. siamensis displayed 18m + 34sm + 46st/a; NF = 150, while P. jullieni exhibited 26m + 14sm + 58st/a; NF = 138. Fluorescence in situ hybridization (FISH) with rDNA probes revealed two (5S) and eight (18S) signals in C. siamensis, respectively, and six signals for both probes in P. jullieni. FISH with microsatellite motifs evidenced substantial genomic divergence between both species. The almost doubled size of the chromosome pairs #1 in C. siamensis and #14 in P. jullieni compared to the rest of corresponding karyotypes indicated chromosomal fusions. CONCLUSION: Based on our findings, together with likely the same reduced 2n = 98 karyotypes in the remainder Probarbini species, we hypothesize that the karyotype 2n = 98 might represent a derived character, shared by all members of the Probarbini clade. Besides, we also witnessed considerable changes in the amount and distribution of certain repetitive DNA classes, suggesting complex post-polyploidization processes in this small paleotetraploid tribe.

Outcome of Patients With Newly Diagnosed Systemic Light-Chain Amyloidosis Associated With Deletion of 17p.

INTRODUCTION: Deletion 17p (del 17p) portends a poor prognosis in myeloma, but its significance in light-chain amyloidosis is unknown. PATIENTS AND METHODS: We identified patients with light-chain amyloidosis and del 17p at diagnosis, and analyzed presenting characteristics, treatments, and clinical outcomes. All had baseline biopsy results showing amyloid and serologic and marrow studies, including standard fluorescence in-situ hybridization determinations of del 17p using commercial probes. Consensus criteria for hematologic and organ involvement, progression, and response were used. Kaplan-Meier (log rank) analyses and Cox regression analysis of baseline variables were used to identify predictors of overall and progression-free survival (PFS). Six-month landmark analyses were performed to assess the impact of treatment-related variables. RESULTS: We identified 44 patients from 7 countries with median marrow and del 17p plasma cells of 22% (range, 3%-100%) and 30% (2%-93%). Ninety-five percent had cardiac involvement, including 44% stage III. Two-thirds of the patients initially received bortezomib-based therapy. Forty-nine percent of patients experienced complete response or very good partial response, with median time to best response of 4 months (range, 1-28 months). Median overall survival and PFS were 49 and 32 months. Cardiac stage and hematologic response were the key predictors of outcomes. Patients with > 50% and ≤ 50% del 17p in clonal plasma cells had median survivals of 28 and 52 months, respectively (P = .08). In landmark analyses, only hematologic response predicted both overall survival and PFS. CONCLUSION: Cardiac stage, hematologic response, and del 17p percentage impact outcomes in these cases. Emphasis should be placed on optimizing supportive care and achieving a deep hematologic response.

Chromosomal Mapping of Repeat DNA in Bergiaria westermanni (Pimelodidae, Siluriformes): Localization of 45S rDNA in B Chromosomes.

The occurrence of repetitive DNA in autosomes and B chromosomes of Bergiaria westermanni was examined using conventional and molecular cytogenetic techniques. This species exhibited 2n = 56 chromosomes, with intra- and interindividual variation in the number of heterochromatic B chromosomes (from 0 to 4). The 5S rDNA was localized in pairs 1 and 5, and histone probes (H1, H3, and H4) and U2 small nuclear RNA were syntenic with 5S rDNA in pair 5. Histone sequences were also located in chromosome pair 14. The (GATA)n sequence was dispersed throughout the autosomes and B chromosomes, with clusters (microsatellite accumulation) in some chromosome regions. The telomeric probe revealed no signs of chromosomal rearrangements in the genome of B. westermanni. The 45S rDNA sites were detected in the terminal region of pair 27; these sites corresponded to a GC-rich heterochromatin block. In addition, 3 of the 4 B chromosomes also contained 45S rDNA copies. Silver nitrate staining in interphase nuclei provided indirect evidence of the expression of these rRNA genes in B chromosomes, indicating the probable origin of these elements. This report shows plasticity in the chromosomal localization of repeat DNA in B. westermanni and features a discussion of genomic diversification.

Differential hypomethylation of the repetitive Tol2/Alu-rich sequences in the genome of Bodianus species (Labriformes, Labridae).

Representatives of the order Labriformes show karyotypes of extreme conservatism together with others with high chromosomal diversification. However, the cytological characterization of epigenetic modifications remains unknown for the majority of the species. In the family Labridae, the most abundant fishes on tropical reefs, the genomes of the genus Bodianus Bloch, 1790 have been characterized by the occurrence of a peculiar chromosomal region, here denominated BOD. This region is exceptionally decondensed, heterochromatic, argentophilic, GC-neutral and, in contrast to classical secondary constrictions, shows no signals of hybridization with 18S rDNA probes. In order to characterize the BOD region, the methylation pattern, the distribution of Alu and Tol2 retrotransposons and of 18S and 5S rDNA sites, respectively, were analyzed by Fluorescence In Situ Hybridization (FISH) on metaphase chromosomes of two Bodianus species, B. insularis Gomon & Lubbock, 1980 and B. pulchellus (Poey, 1860). Immunolocalization of the 5-methylcytosine revealed hypermethylated chromosomal regions, dispersed along the entire length of the chromosomes of both species, while the BOD regions exhibited a hypomethylated pattern. Hypomethylation of the BOD region is associated with the precise co-location of Tol2 and Alu elements, suggesting their active participation in the regulatory epigenetic process. This evidence underscores a probable differential methylation action during the cell cycle, as well as the role of Tol2/Alu elements in functional processes of fish genomes.

Sex Chromosome Evolution and Genomic Divergence in the Fish Hoplias malabaricus (Characiformes, Erythrinidae).

The Erythrinidae family (Teleostei: Characiformes) is a small Neotropical fish group with a wide distribution throughout South America, where Hoplias malabaricus corresponds to the most widespread and cytogenetically studied taxon. This species possesses significant genetic variation, as well as huge karyotype diversity among populations, as reflected by its seven major karyotype forms (i.e., karyomorphs A-G) identified up to now. Although morphological differences in their bodies are not outstanding, H. malabaricus karyomorphs are easily identified by differences in 2n, morphology and size of chromosomes, as well as by distinct evolutionary steps of sex chromosomes development. Here, we performed comparative genomic hybridization (CGH) to analyse both the intra- and inter-genomic status in terms of repetitive DNA divergence among all but one (E) H. malabaricus karyomorphs. Our results indicated that they have close relationships, but with evolutionary divergences among their genomes, yielding a range of non-overlapping karyomorph-specific signals. Besides, male-specific regions were uncovered on the sex chromosomes, confirming their differential evolutionary trajectories. In conclusion, the hypothesis that H. malabaricus karyomorphs are result of speciation events was strengthened.

Identification of a New Mullet Species Complex Based on an Integrative Molecular and Cytogenetic Investigation of Mugil hospes (Mugilidae: Mugiliformes).

Mullets are very common fishes included in the family Mugilidae, (Mugiliformes), which are characterized by both a remarkably uniform external morphology and internal anatomy. Recently, within this family, different species complexes were molecularly identified within Mugil, a genus which is characterized by lineages that sometimes show very different karyotypes. Here we report the results of cytogenetic and molecular analyses conducted on Mugil hospes, commonly known as the hospe mullet, from Ecuador. The study aims to verify whether the original described species from the Pacific Ocean corresponds to that identified in the Atlantic Ocean, and to identify species-specific chromosome markers that can add new comparative data about Mugilidae karyotype evolution. The karyotype of M. hospes from Ecuador is composed of 48 acrocentric chromosomes and shows two active nucleolar organizer regions (NORs). In situ hybridization, using different types of repetitive sequences (rDNAs, U1 snDNA, telomeric repeats) as probes, identified species-specific chromosome markers that have been compared with those of other species of the genus Mugil. Cytochrome c oxidase subunit I (COI) sequence analysis shows only 92-93% similarity with sequences previously deposited under this species name in GenBank, all of which were from the Atlantic Ocean. Phylogenetic reconstructions indicate the presence of three well-supported hospe mullet lineages whose molecular divergence is compatible with the presence of distinct species. Indeed, the first lineage includes samples from Ecuador, whereas the other two lineages include the Atlantic samples and correspond to M. brevirostris from Brazil and Mugil sp. R from Belize/Venezuela. Results here provided reiterate the pivotal importance of an integrative molecular and cytogenetic approach in the reconstruction of the relationships within Mugilidae.

Contrasting Evolutionary Paths Among Indo-Pacific Pomacentrus Species Promoted by Extensive Pericentric Inversions and Genome Organization of Repetitive Sequences.

Pomacentrus (damselfishes) is one of the most characteristic groups of fishes in the Indo-Pacific coral reef. Its 77 described species exhibit a complex taxonomy with cryptic lineages across their extensive distribution. Periods of evolutionary divergences between them are very variable, and the cytogenetic events that followed their evolutionary diversification are largely unknown. In this respect, analyses of chromosomal divergence, within a phylogenetic perspective, are particularly informative regarding karyoevolutionary trends. As such, we conducted conventional cytogenetic and cytogenomic analyses in four Pomacentrus species (Pomacentrus similis, Pomacentrus auriventris, Pomacentrus moluccensis, and Pomacentrus cuneatus), through the mapping of repetitive DNA classes and transposable elements, including 18S rDNA, 5S rDNA, (CA)15, (GA)15, (CAA)10, Rex6, and U2 snDNA as markers. P. auriventris and P. similis, belonging to the Pomacentrus coelestis complex, have indistinguishable karyotypes (2n = 48; NF = 48), with a peculiar syntenic organization of ribosomal genes. On the other hand, P. moluccensis and P. cuneatus, belonging to another clade, exhibit very different karyotypes (2n = 48, NF = 86 and 92, respectively), with a large number of bi-armed chromosomes, where multiple pericentric inversions played a significant role in their karyotype organization. In this sense, different chromosomal pathways followed the phyletic diversification in the Pomacentrus genus, making possible the characterization of two well-contrasting species groups regarding their karyotype features. Despite this, pericentric inversions act as an effective postzygotic barrier in many organisms, which appear to be also the case for P. moluccensis and P. cuneatus; the extensive chromosomal similarities in the two species of P. coelestis complex suggest minor participation of chromosomal postzygotic barriers in the phyletic diversification of these species.