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Satellite DNA (satDNA) sequences are dynamic components of the eukaryotic genome that can play significant roles in species diversification. The Prochilodontidae family, which includes 21 Neotropical fish species, is characterized by a conserved karyotype of 2 n  = 54 biarmed chromosomes, with variation in some species and populations regarding the presence or absence of B chromosomes. This study aimed to investigate whether the chromosomal distribution of specific satDNA sequences is conserved among three Prochilodus species ( Prochilodus lineatus, Prochilodus costatus , and Prochilodus argenteus ) regarding organization and number of loci , and to compare their genomes using comparative genomic hybridization (CGH). Our results demonstrated that most satDNA sequences share a similar distribution pattern across the three species, and CGH analysis corroborated that their karyotypes are very similar in terms of repetitive DNA distribution. We also identified a potential CENP-B box sequence within PliSat01, a satDNA located in the pericentromeric region of all analyzed species. In contrast, PliSat04 and PliSat14 displayed differential locations and variations in the number of loci per genome, underscoring the dynamic nature of repetitive sequences even in species with otherwise highly conserved genomes. These findings represent the first evidence of karyotype diversification in Prochilodus , highlighting the evolutionary dynamism of satDNA sequences., Competing Interests: The authors declare there are no competing interests.

Do all birds' sex chromosomes follow the same canonical one-way direction of evolution? We combined cytogenetic and genomic approaches to analyze the process of the W chromosomal differentiation in two selected Passeriform species, named the Pale-breasted Thrush Turdus leucomelas and the Rufous-bellied thrush T. rufiventris. We characterized the full catalog of satellite DNAs (satellitome) of T. leucomelas, and the 10 TleSatDNA classes obtained together with 16 microsatellite motifs were in situ mapped in both species. Additionally, using Comparative Genomic Hybridization (CGH) assays, we investigated their intragenomic variations. The W chromosomes of both species did not accumulate higher amounts of both heterochromatin and repetitive sequences. However, while T. leucomelas showed a heterochromatin-poor W chromosome with a very complex evolutionary history, T. rufiventris showed a small and partially heterochromatic W chromosome that represents a differentiated version of its original autosomal complement (Z chromosome). The combined approach of CGH and sequential satDNA mapping suggest the occurrence of a former W-autosomal translocation event in T. leucomelas, which had an impact on the W chromosome in terms of sequence gains and losses. At the same time, an autosome, which is present in both males and females in a polymorphic state, lost sequences and integrated previously W-specific ones. This putative W-autosomal translocation, however, did not result in the emergence of a multiple-sex chromosome system. Instead, the generation of a neo-W chromosome suggests an unexpected evolutionary trajectory that deviates from the standard canonical model of sex chromosome evolution., (© 2024. The Author(s).)

Multiple sex chromosomes usually arise from chromosomal rearrangements which involve ancestral sex chromosomes. There is a fundamental condition to be met for their long-term fixation: the meiosis must function, leading to the stability of the emerged system, mainly concerning the segregation of the sex multivalent. Here, we sought to analyze the degree of differentiation and meiotic pairing properties in the selected fish multiple sex chromosome system present in the wolf-fish Hoplias malabaricus (HMA). This species complex encompasses seven known karyotype forms (karyomorphs) where the karyomorph C (HMA-C) exhibits a nascent XY sex chromosomes from which the multiple X 1 X 2 Y system evolved in karyomorph HMA-D via a Y-autosome fusion. We combined genomic and cytogenetic approaches to analyze the satellite DNA (satDNA) content in the genome of HMA-D karyomorph and to investigate its potential contribution to X 1 X 2 Y sex chromosome differentiation. We revealed 56 satDNA monomers of which the majority was AT-rich and with repeat units longer than 100 bp. Seven out of 18 satDNA families chosen for chromosomal mapping by fluorescence in situ hybridization (FISH) formed detectable accumulation in at least one of the three sex chromosomes (X 1 , X 2 and neo-Y). Nine satDNA monomers showed only two hybridization signals limited to HMA-D autosomes, and the two remaining ones provided no visible FISH signals. Out of seven satDNAs located on the HMA-D sex chromosomes, five mapped also to XY chromosomes of HMA-C. We showed that after the autosome-Y fusion event, the neo-Y chromosome has not substantially accumulated or eliminated satDNA sequences except for minor changes in the centromere-proximal region. Finally, based on the obtained FISHpatterns, we speculate on the possible contribution of satDNA to sex trivalent pairing and segregation., (© 2024. The Author(s).)

Background: Different patterns of sex chromosome differentiation are seen in Palaeognathae birds, a lineage that includes the ratites (Struthioniformes, Rheiformes, Apterygiformes, Casuariiformes, and the sister group Tinamiformes). While some Tinamiform species have well-differentiated W chromosomes, both Z and W of all the flightless ratites are still morphologically undifferentiated. Here, we conducted a comprehensive analysis of the ZW differentiation in birds using a combination of cytogenetic, genomic, and bioinformatic approaches. The whole set of satDNAs from the emu (Dromaius novaehollandiae) was described and characterized. Furthermore, we examined the in situ locations of these satDNAs alongside several microsatellite repeats and carried out Comparative Genomic Hybridizations in two related species: the greater rhea (Rhea americana) and the tataupa tinamou (Crypturellus tataupa)., Results: From the 24 satDNA families identified (which represent the greatest diversity of satDNAs ever uncovered in any bird species), only three of them were found to accumulate on the emu's sex chromosomes, with no discernible accumulation observed on the W chromosome. The W chromosomes of both the greater rhea and the emu did not exhibit a significant buildup of either C-positive heterochromatin or repetitive DNAs, indicating their large undifferentiation both at morphological and molecular levels. In contrast, the tataupa tinamou has a highly differentiated W chromosome that accumulates several DNA repeats., Conclusion: The findings provide new information on the architecture of the avian genome and an inside look at the starting points of sex chromosome differentiation in birds., (© 2024. The Author(s).)

Charadriiformes, which comprises shorebirds and their relatives, is one of the most diverse avian orders, with over 390 species showing a wide range of karyotypes. Here, we isolated and characterized the whole collection of satellite DNAs (satDNAs) at both molecular and cytogenetic levels of one of its representative species, named the wattled jacana ( Jacana jacana ), a species that contains a typical ZZ/ZW sex chromosome system and a highly rearranged karyotype. In addition, we also investigate the in situ location of telomeric and microsatellite repeats. A small catalog of 11 satDNAs was identified that typically accumulated on microchromosomes and on the W chromosome. The latter also showed a significant accumulation of telomeric signals, being (GA) 10 the only microsatellite with positive hybridization signals among all the 16 tested ones. These current findings contribute to our understanding of the genomic organization of repetitive DNAs in a bird species with high degree of chromosomal reorganization contrary to the majority of bird species that have stable karyotypes., Competing Interests: The authors declare there are no competing interests.

Background: Crocodilians are one of the oldest extant vertebrate lineages, exhibiting a combination of evolutionary success and morphological resilience that has persisted throughout the history of life on Earth. This ability to endure over such a long geological time span is of great evolutionary importance. Here, we have utilized the combination of genomic and chromosomal data to identify and compare the full catalogs of satellite DNA families (satDNAs, i.e., the satellitomes) of 5 out of the 8 extant Alligatoridae species. As crocodilian genomes reveal ancestral patterns of evolution, by employing this multispecies data collection, we can investigate and assess how satDNA families evolve over time., Results: Alligators and caimans displayed a small number of satDNA families, ranging from 3 to 13 satDNAs in A. sinensis and C. latirostris, respectively. Together with little variation both within and between species it highlighted long-term conservation of satDNA elements throughout evolution. Furthermore, we traced the origin of the ancestral forms of all satDNAs belonging to the common ancestor of Caimaninae and Alligatorinae. Fluorescence in situ experiments showed distinct hybridization patterns for identical orthologous satDNAs, indicating their dynamic genomic placement., Conclusions: Alligators and caimans possess one of the smallest satDNA libraries ever reported, comprising only four sets of satDNAs that are shared by all species. Besides, our findings indicated limited intraspecific variation in satellite DNA, suggesting that the majority of new satellite sequences likely evolved from pre-existing ones., (© 2024. The Author(s).)

Chromosomal rearrangements play a significant role in the evolution of fish genomes, being important forces in the rise of multiple sex chromosomes and in speciation events. Repetitive DNAs constitute a major component of the genome and are frequently found in heterochromatic regions, where satellite DNA sequences (satDNAs) usually represent their main components. In this work, we investigated the association of satDNAs with chromosome-shuffling events, as well as their potential relevance in both sex and karyotype evolution, using the well-known Pyrrhulina fish model. Pyrrhulina species have a conserved karyotype dominated by acrocentric chromosomes present in all examined species up to date. However, two species, namely P. marilynae and P. semifasciata, stand out for exhibiting unique traits that distinguish them from others in this group. The first shows a reduced diploid number (with 2 n = 32), while the latter has a well-differentiated multiple X 1 X 2 Y sex chromosome system. In addition to isolating and characterizing the full collection of satDNAs (satellitomes) of both species, we also in situ mapped these sequences in the chromosomes of both species. Moreover, the satDNAs that displayed signals on the sex chromosomes of P. semifasciata were also mapped in some phylogenetically related species to estimate their potential accumulation on proto-sex chromosomes. Thus, a large collection of satDNAs for both species, with several classes being shared between them, was characterized for the first time. In addition, the possible involvement of these satellites in the karyotype evolution of P. marilynae and P. semifasciata , especially sex-chromosome formation and karyotype reduction in P. marilynae, could be shown.

In this work, we trace the dynamics of satellite DNAs (SatDNAs) accumulation and elimination along the pathway of W chromosome differentiation using the well-known Triportheus fish model. Triportheus stands out due to a conserved ZZ/ZW sex chromosome system present in all examined species. While the Z chromosome is conserved in all species, the W chromosome is invariably smaller and exhibits differences in size and morphology. The presumed ancestral W chromosome is comparable to that of T. auritus , and contains 19 different SatDNA families. Here, by examining five additional Triportheus species, we showed that the majority of these repetitive sequences were eliminated as speciation was taking place. The W chromosomes continued degeneration, while the Z chromosomes of some species began to accumulate some TauSatDNAs. Additional species-specific SatDNAs that made up the heterochromatic region of both Z and W chromosomes were most likely amplified in each species. Therefore, the W chromosomes of the various Triportheus species have undergone significant evolutionary changes in a short period of time (15-25 Myr) after their divergence.

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.

B chromosomes are extra-genomic components of cells found in individuals and in populations of some eukaryotic organisms. They have been described since the first observations of chromosomes, but several aspects of their biology remain enigmatic. Despite being present in hundreds of fungi, plants, and animal species, only a small number of B chromosomes have been investigated through high-throughput analyses, revealing the remarkable mechanisms employed by these elements to ensure their maintenance. Populations of the Psalidodon scabripinnis species complex exhibit great B chromosome diversity, making them a useful material for various analyses. In recent years, important aspects of their biology have been revealed. Here, we review these studies presenting a comprehensive view of the B chromosomes in the P. scabripinnis complex and a new hypothesis regarding the role of the B chromosome in the speciation process.

Eukaryotic genomes are usually enriched in repetitive DNA sequences, which can be classified as dispersed or tandemly repeated elements. Satellite DNAs are noncoding monomeric sequences organized in a head-to-tail fashion that are generally located on the subtelomeric and/or pericentromeric heterochromatin. In general, a single species incorporates a diverse group of satellite DNA families, which collection is called satellitome. Here, we characterized three new satellitomes from distinct characid fish ( Psalidodon fasciatus, P. bockmanni, and Astyanax lacustris ) using a combination of genomic, cytogenetic, and bioinformatic protocols. We also compared our data with the available satellitome of P. paranae. We described 57 satellite DNA (satDNA) families of P. fasciatus (80 variants), 50 of P. bockmanni (77 variants), and 33 of A. lacustris (54 variants). Our analyses demonstrated that several sequences were shared among the analyzed species, while some were restricted to two or three species. In total, we isolated 104 distinctive satDNA families present in the four species, of which 10 were shared among all four. Chromosome mapping revealed that the clustered satDNA was mainly located in the subtelomeric and pericentromeric areas. Although all Psalidodon species demonstrated the same pattern of clusterization of satDNA, the number of clusters per genome was variable, indicating a high dynamism of these sequences. In addition, our results expand the knowledge of the As51 satellite DNA family, revealing that P. bockmanni and P. paranae exhibited an abundant variant of 39 bp, while P. fasciatus showed a variant of 43 bp. The majority of satDNAs in the satellitomes analyzed here presented a common library repetitive sequence in Psalidodon and Astyanax , with abundance variations in each species, as expected for closely related groups. In addition, we concluded that the most abundant satDNA in Psalidodon (As51) passed through a diversification process in this group, resulting in new variants exclusive of Psalidodon ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Goes, dos Santos, Aguiar, Alves, Silva, Foresti, Oliveira, Utsunomia and Porto-Foresti.)

Satellites are an abundant source of repetitive DNAs that play an essential role in the chromosomal organization and are tightly linked with the evolution of sex chromosomes. Among fishes, Triportheidae stands out as the only family where almost all species have a homeologous ZZ/ZW sex chromosomes system. While the Z chromosome is typically conserved, the W is always smaller, with variations in size and morphology between species. Here, we report an analysis of the satellitome of Triportheus auritus (TauSat) by integrating genomic and chromosomal data, with a special focus on the highly abundant and female-biased satDNAs. In addition, we investigated the evolutionary trajectories of the ZW sex chromosomes in the Triportheidae family by mapping satDNAs in selected representative species of this family. The satellitome of T. auritus comprised 53 satDNA families of which 24 were also hybridized by FISH. Most satDNAs differed significantly between sexes, with 19 out of 24 being enriched on the W chromosome of T. auritus. The number of satDNAs hybridized into the W chromosomes of T. signatus and T. albus decreased to six and four, respectively, in accordance with the size of their W chromosomes. No TauSat probes produced FISH signals on the chromosomes of Agoniates halecinus. Despite its apparent conservation, our results indicate that each species differs in the satDNA accumulation on the Z chromosome. Minimum spanning trees (MSTs), generated for three satDNA families with different patterns of FISH mapping data, revealed different homogenization rates between the Z and W chromosomes. These results were linked to different levels of recombination between them. The most abundant satDNA family (TauSat01) was exclusively hybridized in the centromeres of all 52 chromosomes of T. auritus, and its putative role in the centromere evolution was also highlighted. Our results identified a high differentiation of both ZW chromosomes regarding satellites composition, highlighting their dynamic role in the sex chromosomes evolution., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

B chromosomes are additional dispensable elements to the standard chromosomal set of an organism. In most cases, their transmission differs from Mendelian patterns, leading to their accumulation or extinction. The present study aimed to describe, for the first time, the transmission pattern of B chromosome in a population of Psalidodon paranae through directed crosses, as well as to analyze the populational dynamics of B chromosome. Our results revealed the possible elimination of B chromosome in crossings where only females were B-carriers, with a mean transmission rate ( k B ) of 0.149; however, k B was significantly higher in crossings involving male B-carriers ( k B  = 0.328-0.450). Moreover, we observed an increase in the frequency of B chromosomes in the natural population of P. paranae in the last two decades. These apparently contradictory results can make sense if the B chromosome provides adaptive advantages to their carriers. Here, we observed a differential transmission of B chromosomes in each sex of parental individuals, with higher transmission rates in crossing involving males B-carriers, in addition to describe the temporal changes of B chromosome frequency in P. paranae .

B or supernumerary chromosomes are dispensable elements that are widely present in numerous eukaryotes. Due to their non-recombining nature, there is an evident tendency for repetitive DNA accumulation in these elements. Thus, satellite DNA plays an important role in the evolution and diversification of B chromosomes and can provide clues regarding their origin. The characiform Prochilodus lineatus was one of the first discovered fish species bearing B chromosomes, with all populations analyzed so far showing one to nine micro-B chromosomes and exhibiting at least three morphological variants (Ba, Bsm, and Bm). To date, a single satellite DNA is known to be located on the B chromosomes of this species, but no information regarding the differentiation of the proposed B-types is available. Here, we characterized the satellitome of P. lineatus and mapped 35 satellite DNAs against the chromosomes of P. lineatus , of which six were equally located on all B-types and this indicates a similar genomic content. In addition, we describe, for the first time, an entire population without B chromosomes.

Supernumerary, or B, chromosomes are present in several eukaryotes, including characid fish of the genus Psalidodon. Notably, Psalidodon paranae carries the most studied B chromosome variant, a macro-B chromosome. The origin of this element was determined to be an isochromosome; however, data regarding its inheritance remain unavailable due to methodological barriers such as the lack of an efficient, non-invasive, and rapid protocol for identifying B-carrying individuals that would enable the design of efficient crossing experiments. Thus, in this study, we primarily aimed was to develop two non-invasive and fast (approximately 2 h) methods to identify the presence of B chromosomes in live specimens of P. paranae based on satellite DNA (satDNA) sequences known to be present in this element. The methods include fluorescence in situ hybridization in interphase nuclei and relative gene quantification of satDNAs using quantitative polymerase chain reaction. Our results reveal the efficiency of quick-fluorescence in situ hybridization and quantitative polymerase chain reaction for identifying B-carrying individuals using the proposed satDNA sequences and open up new possibilities to study B chromosomes.

Astyanax Baird et Girard, 1854, is one of the largest genera in the family Characidae and comprises 177 valid species. This genus has been the focus of cytogenetic studies primarily owing to the presence of B chromosomes and high karyotypic diversity among different populations. The intense genetic variability in Astyanax is one of the factors responsible for the occurrence of species complexes, which are groups (1) with certain difficulties in establishing common genetic pools or (2) belonging to different cryptic species. To evaluate cytogenetic marker inheritance and the possibility of the identification of these hybrids, this study aimed to describe cytogenetic hybrids from three strains of species of the genera Astyanax and Hyphessobrycon Eigenmann, 1908. A. lacustris Lütken, 1875, A. schubarti Britski, 1964, A. fasciatus Cuvier, 1819, and H. anisitsi Eigenmann, 1907 were used to generate three hybrid lineages. The diploid number, heterochromatin sites, and ribosomal genes (18S and 5S rDNA) of the parental strains and the hybrids were analyzed. The results indicated that the three hybrid lineages had cytogenetic markers of both parents, presenting Mendelian inheritance. However, differences in distribution of heterochromatic blocks were observed between the hybrids and the parent strains. Our results allowed the identification of the hybrid strains based on the cytogenetic markers applied, reinforcing the efficiency of cytogenetic markers as tools for identification and indicating that such events may increase the karyotypic diversity in the genera Astyanax and Hyphessobrycon ., (Caio Augusto Gomes Goes, Sandro Natal Daniel, Lucas Henrique Piva, George Shigueki Yasui, Roberto Ferreira Artoni, Diogo Teruo Hashimoto, Fausto Foresti, Fábio Porto-Foresti.)

Satellite DNAs (satDNAs) are tandemly repeated DNA sequences with great abundance in eukaryotic genomes. A single species may carry up to hundreds of satDNA families, which is collectively called as "satellitome," each showing its own dynamics and evolution rates. In this context, all live species contain a satDNA library that may be partially or totally shared with other related species/populations. In the late few years, next-generation sequencing (NGS) and novel bioinformatic tools facilitated the massive characterization of these sequences at low costs, and consequently, comparing satDNAs between species. In this study, we characterized two novel satDNAs (MsaSat03-80 and MsaSat04-142) in three characid fish ( Astyanax paranae and Astyanax fasciatus and two populations of Moenkhausia sanctaefilomenae ) and mapped their chromosomal location to unveil the evolutionary dynamics of satDNA repeats in those species. Our results evidenced that MsaSat03 is present in the genomes of all analyzed species, but is clustered only in the chromosomes of M. sanctaefilomenae , exhibiting a conserved number and location of sites. Conversely, MsaSat04 sequences is restricted to M. sanctaefilomenae and shows a differential distribution between the two analyzed populations. Altogether, our analyses point to a complex history of satDNA families in characid fish and the utility of NGS data for comparative satDNA analysis.

The accumulation of repetitive DNA sequences on the sex-limited W or Y chromosomes is a well-known process that is likely triggered by the suppression of recombination between the sex chromosomes, which leads to major differences in their sizes and genetic content. Here, we report an analysis conducted on the satellitome of Megaleporinus macrocephalus that focuses specifically on the satDNAs that have been shown to have higher abundances in females and are putatively located on the W chromosome in this species. We characterized 164 satellite families in M. macrocephalus, which is, by far, the most satellite-rich species discovered to date. Subsequently, we mapped 30 satellites, 22 of which were located on the W chromosome, and 14 were shown to exist only on the W chromosome. Finally, we report two simple, quick and reliable methods that can be used for sex identification in M. macrocephalus individuals using fin clips or scales, which could be applicable to future studies conducted in the field of aquaculture.

This work was aimed at developing an effective procedure to obtain sterile ideal host fish in mass scale with no endogenous germ cells in the germinal epithelium, owning permanent stem-cell niches able to be colonized by transplanted germ cells in surrogate technology experiments. Thus, triploids, diploid hybrids, and triploid hybrids were produced. To obtain hybrid offspring, oocytes from a single Astyanax altiparanae female were inseminated by sperm from five males (A. altiparanae, A. fasciatus, A. schubarti, Hyphessobrycon anisitsi, and Oligosarcus pintoi). Triploidization was conducted by inhibition of the second polar body release using heat shock treatment at 40 °C for 2 min. At 9-months of age, the offspring from each crossing was histologically evaluated to access the gonadal status of the fish. Variable morphological characteristics of the gonads were found in the different hybrids offspring: normal gametogenesis, gametogenesis without production of gametes, sterile specimens holding germ cells, and sterile specimens without germ cells, which were considered "ideal hosts". However, only in the hybrid derived from crossing between A. altiparanae and A. fasciatus, 100% of the individuals were completely sterile. Among them 83.3% of the male did not present germ cells inside germinal epithelium, having only somatic cells in the gonad. The other 16.7% also presented spermatogonia inside the niches. Such a methodology allows the production of sterile host in mass scale, opening new insights for application of surrogate technologies., (Copyright © 2017 Elsevier Inc. All rights reserved.)