Publications by authors named "Marcelo de Bello Cioffi"

77 Publications

Evolutionary breakpoint regions and chromosomal remodeling in Harttia (Siluriformes: Loricariidae) species diversification.

Genet Mol Biol 2022 20;45(2):e20210170. Epub 2022 May 20.

Universidade Estadual de Ponta Grossa, Departamento de Biologia Estrutural, Molecular e Genética, Ponta Grossa, PR, Brazil.

The Neotropical armored catfish genus Harttia presents a wide variation of chromosomal rearrangements among its representatives. Studies indicate that translocation and Robertsonian rearrangements have triggered the karyotype evolution in the genus, including differentiation of sex chromosome systems. However, few studies used powerful tools, such as comparative whole chromosome painting, to clarify this highly diversified scenario. Here, we isolated probes from the X1 (a 5S rDNA carrier) and the X2 (a 45S rDNA carrier) chromosomes of Harttia punctata, which displays an X1X1X2X2/X1X2Y multiple sex chromosome system. Those probes were applied in other Harttia species to evidence homeologous chromosome blocks. The resulting data reinforce that translocation events played a role in the origin of the X1X2Y sex chromosome system in H. punctata. The repositioning of homologous chromosomal blocks carrying rDNA sites among ten Harttia species has also been demonstrated. Anchored to phylogenetic data it was possible to evidence some events of the karyotype diversification of the studied species and to prove an independent origin for the two types of multiple sex chromosomes, XX/XY1Y2 and X1X1X2X2/X1X2Y, that occur in Harttia species. The results point to evolutionary breakpoint regions in the genomes within or adjacent to rDNA sites that were widely reused in Harttia chromosome remodeling.
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http://dx.doi.org/10.1590/1678-4685-GMB-2021-0170DOI Listing
May 2022

Chromosomal Rearrangements and Origin of the Multiple XX/XYY Sex Chromosome System in Species (Siluriformes: Loricariidae).

Front Genet 2022 21;13:877522. Epub 2022 Mar 21.

Departamento de Biologia Estrutural, Molecular e Genética, Universidade Estadual de Ponta Grossa, Paraná, Brazil.

The Neotropical genus comprises species with extensive chromosomal remodeling and distinct sex chromosome systems (SCSs). So far, three different SCSs with male heterogamety have been characterized in the group. In some species, the presence of the XX/XYY SCS is associated with a decrease in diploid numbers and several chromosomal rearrangements, although a direct relation to sex chromosome differentiation has not been shown yet. Here, we aimed to investigate the differentiation processes that have led to the establishment of the rare XX/XYY SCS and track its evolutionary history among other species. For that, four whole chromosome painting probes derived from chromosome 1 of (HTO-1), chromosomes 9 and X of (HCA-9 and HCA-X), and chromosome X from (HIN-X) were applied in nine species. Homeologous chromosome blocks were located in species and demonstrated that Robertsonian (Rb) fusions originated HTO-1, HCA-9, and HCA-X chromosomes, while Rb fissions explain Y and Y sex chromosomes. Specifically, in , HCA-X, HCA-9, and the NOR-bearing chromosome demonstrated that homeologous blocks were used in the HIN-X and metacentric pair 2 origins. Consequently, diploid numbers changed between the studied species. Overall, the data also reinforce the existence of unstable genomic sites promoting chromosomal differentiation and remodeling within the genus .
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http://dx.doi.org/10.3389/fgene.2022.877522DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8977651PMC
March 2022

Evolutionary Dynamics of Two Classes of Repetitive DNA in the Genomes of Two Species of Elopiformes (Teleostei, Elopomorpha).

Zebrafish 2022 02;19(1):24-31

Instituto de Estudos Costeiros, Universidade Federal do Pará, Laboratório de Evolução, Bragança, Brazil.

The order Elopiformes includes fish species of medium to large size with a circumglobal distribution, in both the open sea, coastal, and estuarine waters. The Elopiformes are considered an excellent model for evolutionary studies due to their ample adaptive capacity, which allow them to exploit a range of different ecological niches. In this study, we analyzed the karyotype structure and distribution of two classes of repetitive DNA (microsatellites and transposable elements) in two Elopiformes species ( and ). The results showed that the microsatellite sequences had a very similar distribution in these species, primarily associated to heterochromatin (centromeres and telomeres), suggesting these sequences contribute to the chromosome structure. In contrast, specific signals detected throughout the euchromatic regions indicate that some of these sequences may play a role in the regulation of gene expression. By contrast, the transposable elements presented a distinct distribution in the two species, pointing to a possible interspecific difference in the function of these sequences in the genomes of the two species. Therefore, the comparative genome mapping provides new insights into the structure and organization of these repetitive sequences in the Elopiformes genome.
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http://dx.doi.org/10.1089/zeb.2021.0027DOI Listing
February 2022

Satellitome analysis illuminates the evolution of ZW sex chromosomes of Triportheidae fishes (Teleostei: Characiformes).

Chromosoma 2022 Jun 31;131(1-2):29-45. Epub 2022 Jan 31.

Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos, São Paulo, Brazil.

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.
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http://dx.doi.org/10.1007/s00412-022-00768-1DOI Listing
June 2022

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

Comp Cytogenet 2021 1;15(4):429-445. Epub 2021 Dec 1.

Departament of Cell Biology and Genetics, Biosciences Center, Universidade Federal do Rio Grande do Norte, Natal, RN, 59078970, Brazil.

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, (Quoy et Gaimard, 1825) and (Valenciennes, 1883) (Naucratini), with circumtropical distributions, (Forsskål, 1775) (Carangini), widely distributed in the tropical and subtropical waters of the Indian and Pacific oceans, and (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.
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http://dx.doi.org/10.3897/CompCytogen.v15.i4.69638DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8654809PMC
December 2021

A new view on the scenario of karyotypic stasis in Epinephelidae fish: Cytogenetic, historical, and biogeographic approaches.

Genet Mol Biol 2021 15;44(4):e20210122. Epub 2021 Nov 15.

Universidade Federal do Rio Grande do Norte, Departamento de Biologia Celular e Genética, Centro de Biociências, Natal, RN, Brazil.

Epinephelidae (groupers) is an astonishingly diverse group of carnivorous fish widely distributed in reef environments around the world, with growing economic importance. The first chromosomal inferences suggested a conservative scenario for the family. However, to date, this has not been validated using biogeographic and phylogenetic approaches. Thus, to estimate karyotype diversification among groupers, eight species from the Atlantic and Indian oceans were investigated using conventional cytogenetic protocols and fluorescence in situ hybridization of repetitive sequences (rDNA, microsatellites, transposable elements). Despite the remarkable persistence of some symplesiomorphic karyotype patterns, such as all species sharing 2n=48 and most preserve a basal karyotype (2n=48 acrocentrics), the chromosomal diversification in the family revealed an unsuspected evolutionary dynamic, where about 40% of the species escape from the ancestral karyotype pattern. These karyotype changes showed a relation with the historical biogeography, likely as a byproduct of the progressive occupancy of new areas (huge diversity of adaptive and speciation conditions). In this context, oceanic regions harboring more recent clades such as those of the Indo-Pacific, exhibited a higher karyotype diversity. Therefore, the karyotype evolution of Epinephelidae fits well with the expansion and geographic contingencies of its clades, providing a more complex and diverse scenario than previously assumed.
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http://dx.doi.org/10.1590/1678-4685-GMB-2021-0122DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8608104PMC
November 2021

Evolutionary Tracks of Chromosomal Diversification in Surgeonfishes (Acanthuridae: ) Along the World's Biogeographic Domains.

Front Genet 2021 29;12:760244. Epub 2021 Oct 29.

Department of Cell Biology and Genetics, Biosciences Center, Federal University of Rio Grande do Norte, Natal, Brazil.

Fishes of the genus (Acanthuridae) are strongly related to reef environments, in a broad biogeographic context worldwide. Although their biological aspects are well known, cytogenetic information related to this genus remains incipient. In this study, species from populations inhabiting coastal regions of the Southwest Atlantic (SWA), South Atlantic oceanic islands (Fernando de Noronha Archipelago and Trindade Island), Greater Caribbean (GC), and Indo-Pacific Ocean (the center of the origin of the group) were analyzed to investigate their evolutionary differentiation. For this purpose, we employed conventional cytogenetic procedures and fluorescence hybridization of 18S rDNA, 5S rDNA, and H3 and H2B-H2A histone sequences. The Atlantic species (, and ) did not show variations among them, despite their vast continental and insular distribution. In contrast, from SWA and GC diverged from each other in the number of 18S rDNA sites, a condition likely associated with the barrier created by the outflows of the Amazonas/Orinoco rivers. The geminate species had a cytogenetic profile similar to that of . However, the chromosomal macrostructures and the distribution of rDNA and hisDNA sequences revealed moderate to higher rates of diversification when species from recently colonized areas (Atlantic Ocean) were compared to , a representative species from the Indian Ocean. Our cytogenetic data covered all species from the Western Atlantic, tracked phylogenetic diversification throughout the dispersive process of the genus, and highlighted the probable diversifying role of ocean barriers in this process.
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http://dx.doi.org/10.3389/fgene.2021.760244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8586516PMC
October 2021

Against the mainstream: exceptional evolutionary stability of ZW sex chromosomes across the fish families Triportheidae and Gasteropelecidae (Teleostei: Characiformes).

Chromosome Res 2021 12 25;29(3-4):391-416. Epub 2021 Oct 25.

Departamento de Genética e Evolução, Universidade Federal de São Carlos, Rod. Washington Luiz km 235, Sao Carlos, SP, 13565-905, Brazil.

Teleost fishes exhibit a breath-taking diversity of sex determination and differentiation mechanisms. They encompass at least nine sex chromosome systems with often low degree of differentiation, high rate of inter- and intra-specific variability, and frequent turnovers. Nevertheless, several mainly female heterogametic systems at an advanced stage of genetic differentiation and high evolutionary stability have been also found across teleosts, especially among Neotropical characiforms. In this study, we aim to characterize the ZZ/ZW sex chromosome system in representatives of the Triportheidae family (Triportheus auritus, Agoniates halecinus, and the basal-most species Lignobrycon myersi) and its sister clade Gasteropelecidae (Carnegiella strigata, Gasteropelecus levis, and Thoracocharax stellatus). We applied both conventional and molecular cytogenetic approaches including chromosomal mapping of 5S and 18S ribosomal DNA clusters, cross-species chromosome painting (Zoo-FISH) with sex chromosome-derived probes and comparative genomic hybridization (CGH). We identified the ZW sex chromosome system for the first time in A. halecinus and G. levis and also in C. strigata formerly reported to lack sex chromosomes. We also brought evidence for possible mechanisms underlying the sex chromosome differentiation, including inversions, repetitive DNA accumulation, and exchange of genetic material. Our Zoo-FISH experiments further strongly indicated that the ZW sex chromosomes of Triportheidae and Gasteropelecidae are homeologous, suggesting their origin before the split of these lineages (approx. 40-70 million years ago). Such extent of sex chromosome stability is almost exceptional in teleosts, and hence, these lineages afford a special opportunity to scrutinize unique evolutionary forces and pressures shaping sex chromosome evolution in fishes and vertebrates in general.
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http://dx.doi.org/10.1007/s10577-021-09674-1DOI Listing
December 2021

Tracking the Evolutionary Trends Among Small-Size Fishes of the Genus (Characiforme, Lebiasinidae): New Insights From a Molecular Cytogenetic Perspective.

Front Genet 2021 6;12:769984. Epub 2021 Oct 6.

Laboratorio de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos (UFSCar), São Carlos, Brazil.

Miniature fishes have always been a challenge for cytogenetic studies due to the difficulty in obtaining chromosomal preparations, making them virtually unexplored. An example of this scenario relies on members of the family Lebiasinidae which include miniature to medium-sized, poorly known species, until very recently. The present study is part of undergoing major cytogenetic advances seeking to elucidate the evolutionary history of lebiasinids. Aiming to examine the karyotype diversification more deeply in , here we combined classical and molecular cytogenetic analyses, including Giemsa staining, C-banding, repetitive DNA mapping, comparative genomic hybridization (CGH), and whole chromosome painting (WCP) to perform the first analyses in five species ( aff. , sp., , and cf. ). The diploid number (2n) ranged from 40 to 42 chromosomes among all analyzed species, but is strikingly differentiated by having 2n = 32 chromosomes and a karyotype composed of large meta/submetacentric chromosomes, whose plesiomorphic status is discussed. The distribution of microsatellites does not markedly differ among species, but the number and position of the rDNA sites underwent significant changes among them. Interspecific comparative genome hybridization (CGH) found a moderate divergence in the repetitive DNA content among the species' genomes. Noteworthy, the WCP reinforced our previous hypothesis on the origin of the XXY multiple sex chromosome system in . In summary, our data suggest that the karyotype differentiation in has been driven by major structural rearrangements, accompanied by high dynamics of repetitive DNAs.
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http://dx.doi.org/10.3389/fgene.2021.769984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8526856PMC
October 2021

Multiple sex chromosomes in teleost fishes from a cytogenetic perspective: state of the art and future challenges.

Philos Trans R Soc Lond B Biol Sci 2021 09 26;376(1833):20200098. Epub 2021 Jul 26.

Departamento de Genética e Evolução, Universidade Federal de São Carlos, Rod. Washington Luiz km 235 cep, 13565-905, São Carlos, Brazil.

Despite decades of cytogenetic and genomic research of dynamic sex chromosome evolution in teleost fishes, multiple sex chromosomes have been largely neglected. In this review, we compiled available data on teleost multiple sex chromosomes, identified major trends in their evolution and suggest further trajectories in their investigation. In a compiled dataset of 440 verified records of fish sex chromosomes, we counted 75 multiple sex chromosome systems with 60 estimated independent origins. We showed that male-heterogametic systems created by Y-autosome fusion predominate and that multiple sex chromosomes are over-represented in the order Perciformes. We documented a striking difference in patterns of differentiation of sex chromosomes between male and female heterogamety and hypothesize that faster W sex chromosome differentiation may constrain sex chromosome turnover in female-heterogametic systems. We also found no significant association between the mechanism of multiple sex chromosome formation and percentage of uni-armed chromosomes in teleost karyotypes. Last but not least, we hypothesized that interaction between fish populations, which differ in their sex chromosomes, can drive the evolution of multiple sex chromosomes in fishes. This underlines the importance of broader inter-population sampling in studies of fish sex chromosomes. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part II)'.
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http://dx.doi.org/10.1098/rstb.2020.0098DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8310710PMC
September 2021

Revisiting the Karyotypes of Alligators and Caimans (Crocodylia, Alligatoridae) after a Half-Century Delay: Bridging the Gap in the Chromosomal Evolution of Reptiles.

Cells 2021 06 5;10(6). Epub 2021 Jun 5.

Laboratório de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos 13565-905, Brazil.

Although crocodilians have attracted enormous attention in other research fields, from the cytogenetic point of view, this group remains understudied. Here, we analyzed the karyotypes of eight species formally described from the Alligatoridae family using differential staining, fluorescence in situ hybridization with rDNA and repetitive motifs as a probe, whole chromosome painting (WCP), and comparative genome hybridization. All Caimaninae species have a diploid chromosome number (2n) 42 and karyotypes dominated by acrocentric chromosomes, in contrast to both species of Alligatorinae, which have 2n = 32 and karyotypes that are predominantly metacentric, suggesting fusion/fission rearrangements. Our WCP results supported this scenario by revealing the homeology of the largest metacentric pair present in both spp. with two smaller pairs of acrocentrics in Caimaninae species. The clusters of 18S rDNA were found on one chromosome pair in all species, except for spp., which possessed three chromosome pairs bearing these sites. Similarly, comparative genomic hybridization demonstrated an advanced stage of sequence divergence among the caiman genomes, with standing out as the most divergent. Thus, although Alligatoridae exhibited rather low species diversity and some level of karyotype stasis, their genomic content indicates that they are not as conserved as previously thought. These new data deepen the discussion of cytotaxonomy in this family.
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http://dx.doi.org/10.3390/cells10061397DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8228166PMC
June 2021

Karyotype Evolution and Genomic Organization of Repetitive DNAs in the Saffron Finch, (Passeriformes, Aves).

Animals (Basel) 2021 May 19;11(5). Epub 2021 May 19.

School of Biosciences, University of Kent, Canterbury CT2 7NJ, UK.

The Saffron finch (), a semi-domestic species, is tolerant of human proximity and nesting in roof spaces. Considering the importance of cytogenomic approaches in revealing different aspects of genomic organization and evolution, we provide detailed cytogenetic data for . , including the standard Giemsa karyotype, C- and G-banding, repetitive DNA mapping, and bacterial artificial chromosome (BAC) FISH. We also compared our results with the sister groups, Passeriformes and Psittaciformes, bringing new insights into the chromosome and genome evolution of birds. The results revealed contrasting rates of intrachromosomal changes, highlighting the role of SSR (simple short repetition probes) accumulation in the karyotype reorganization. The SSRs showed scattered hybridization, but brighter signals were observed in the microchromosomes and the short arms of Z chromosome in . . BACs probes showed conservation of ancestral syntenies of macrochromosomes (except GGA1), as well as the tested microchromosomes. The comparison of our results with previous studies indicates that the great biological diversity observed in Passeriformes was not likely accompanied by interchromosomal changes. In addition, although repetitive sequences often act as hotspots of genome rearrangements, Passeriformes species showed a higher number of signals when compared with the sister group Psittaciformes, indicating that these sequences were not involved in the extensive karyotype reorganization seen in the latter.
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http://dx.doi.org/10.3390/ani11051456DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8160697PMC
May 2021

Comparative study of four species (Bagridae, Siluriformes) from Thailand: insights into their karyotypic diversity.

Comp Cytogenet 2021 26;15(2):119-136. Epub 2021 Apr 26.

Applied Science Program, Faculty of Interdisciplinary Studies, Nong Khai Campus, Khon Kaen University, Muang, Nong Khai 43000, Thailand.

Karyotypes of four catfishes of the genus Scopoli, 1777 (family Bagridae), Fowler, 1937, Roberts, 1992, (Bleeker, 1846) and (Bleeker, 1851), were analysed by conventional and Ag-NOR banding as well as fluorescence in situ hybridization (FISH) techniques. Microsatellite d(GC), d(CAA), d(CAT) and d(GAA) repeat probes were applied in FISH. The obtained data revealed that the four studied species have different chromosome complements. The diploid chromosome numbers (2n) and the fundamental numbers (NF) range between 52 and 102, 54 and 104, 56 and 98, or 58 and 108 in , , or , respectively. Karyotype formulae of , , and are 24m+26sm+4a, 26m+24sm+2a, 24m+18sm+14a and 30m+22sm+6a, respectively. A single pair of NORs was identified adjacent to the telomeres of the short arm of chromosome pairs 3 (metacentric) in , 20 (submetacentric) in , 15 (submetacentric) in , and 5 (metacentric) in . The d(GC), d(CAA), d(CAT) and d(GAA) repeats were abundantly distributed in species-specific patterns. Overall, we present a comparison of cytogenetic and molecular cytogenetic patterns of four species from genus providing insights into their karyotype diversity in the genus.
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http://dx.doi.org/10.3897/CompCytogen.v15i2.60649DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8093182PMC
April 2021

Evolution of a Multiple Sex-Chromosome System by Three-Sequential Translocations among Potential Sex-Chromosomes in the Taiwanese Frog .

Cells 2021 03 16;10(3). Epub 2021 Mar 16.

Amphibian Research Center, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima 739-8526, Japan.

Translocation between sex-chromosomes and autosomes generates multiple sex-chromosome systems. It happens unexpectedly, and therefore, the evolutionary meaning is not clear. The current study shows a multiple sex chromosome system comprising three different chromosome pairs in a Taiwanese brown frog (). The male-specific three translocations created a system of six sex-chromosomes, ♂XYXYXY-♀XXXXXX. It is unique in that the translocations occurred among three out of the six members of potential sex-determining chromosomes, which are known to be involved in sex-chromosome turnover in frogs, and the two out of three include orthologs of the sex-determining genes in mammals, birds and fishes. This rare case suggests sex-specific, nonrandom translocations and thus provides a new viewpoint for the evolutionary meaning of the multiple sex chromosome system.
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http://dx.doi.org/10.3390/cells10030661DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8002213PMC
March 2021

Meiotic synapsis of homeologous chromosomes and mismatch repair protein detection in the parthenogenetic rock lizard Darevskia unisexualis.

Mol Reprod Dev 2021 02 12;88(2):119-127. Epub 2021 Jan 12.

Vavilov Institute of General Genetics RAS, Moscow, Russia.

Parthenogenetic species of Caucasian rock lizards of the genus Darevksia are important evidence for reticulate evolution and speciation by hybridization in vertebrates. Female-only lineages formed through interspecific hybridization have been discovered in many groups. Nevertheless, critical mechanisms of oogenesis and specifics of meiosis that provide long-term stability of parthenogenetic species are still unknown. Here we report cytogenetic characteristics of somatic karyotypes and meiotic prophase I nuclei in the diploid parthenogenetic species Darevskia unisexualis from the new population "Keti" in Armenia which contains an odd number of chromosomes 2n = 37, instead of the usual 2n = 38. We revealed 36 acrocentric chromosomes and a single metacentric autosomal chromosome, resulting from Robertsonian translocation. Comparative genomic hybridization revealed that chromosome fusion occurred between two chromosomes inherited from the maternal species, similar to another parthenogenetic species D. rostombekowi. To trace the chromosome behaviour in meiosis, we performed an immunocytochemical study of primary oocytes' spread nuclei and studied chromosome synapsis during meiotic prophase I in D. unisexualis based on analysis of synaptonemal complexes (SCs). We found meiotic SC-trivalent composed of one metacentric and two acrocentric chromosomes. We confirmed that the SC was assembled between homeologous chromosomes inherited from two parental species. Immunostaining of the pachytene and diplotene nuclei revealed a mismatch repair protein MLH1 loaded to all autosomal SC bivalents. Possible mechanisms of meiotic recombination between homeologous chromosomes are discussed.
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http://dx.doi.org/10.1002/mrd.23450DOI Listing
February 2021

Looking for genetic effects of polluted anthropized environments on Caiman crocodilus crocodilus (Reptilia, Crocodylia): A comparative genotoxic and chromosomal analysis.

Ecotoxicol Environ Saf 2021 Feb 28;209:111835. Epub 2020 Dec 28.

Centro Universitário Dinâmica das Cataratas, Faculdade Anglo Americano, Foz do Iguaçu, Paraná, Brazil.

The Amazon aquatic ecosystems have been modified by the human population growth, going through changes in their water bodies and aquatic biota. The spectacled alligator (Caiman crocodilus crocodilus) has a wide distribution and adaptability to several environments, even those polluted ones. This study aimed to investigate if a Caiman species living in urban streams of Manaus city (Amazonas State, Brazil) is affected by environmental pollution. For that, it was used classical and molecular cytogenetic procedures, in addition to micronucleus and comet assays. Although the karyotype macrostructure remains unaltered (2 n = 42 chromosomes; 24 t + 18 m/sm; NF = 60), the genotoxic analysis and the cytogenetic mapping of repetitive DNA sequences demonstrated that polluted environments alter the genome of the specimens, affecting both the chromosomal organization and the genetic material.
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http://dx.doi.org/10.1016/j.ecoenv.2020.111835DOI Listing
February 2021

Chromosomal Analysis in (Aves, Cuculiformes) Reveals Extensive Genomic Reorganization and an Unusual Z-Autosome Robertsonian Translocation.

Cells 2020 12 22;10(1). Epub 2020 Dec 22.

School of Biosciences, University of Kent, Canterbury CT2 7NJ, UK.

Although cytogenetics studies in cuckoos (Aves, Cuculiformes) have demonstrated an interesting karyotype variation, such as variations in the chromosome morphology and diploid number, their chromosome organization and evolution, and relation with other birds are poorly understood. Hence, we combined conventional and molecular cytogenetic approaches to investigate chromosome homologies between chicken and the smooth-billed ani (). Our results demonstrate extensive chromosome reorganization in . , with interchromosomal rearrangements involving macro and microchromosomes. Intrachromosomal rearrangements were observed in some macrochromosomes, including the Z chromosome. The most evolutionary notable finding was a Robertsonian translocation between the microchromosome 17 and the Z chromosome, a rare event in birds. Additionally, the simple short repeats (SSRs) tested here were preferentially accumulated in the microchromosomes and in the Z and W chromosomes, showing no relationship with the constitutive heterochromatin regions, except in the W chromosome. Taken together, our results suggest that the avian sex chromosome is more complex than previously postulated and revealed the role of microchromosomes in the avian sex chromosome evolution, especially cuckoos.
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http://dx.doi.org/10.3390/cells10010004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7822047PMC
December 2020

Highly Rearranged Karyotypes and Multiple Sex Chromosome Systems in Armored Catfishes from the Genus (Teleostei, Siluriformes).

Genes (Basel) 2020 11 18;11(11). Epub 2020 Nov 18.

Laboratório de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos SP 13565-905, Brazil.

comprises an armored catfish genus endemic to the Neotropical region, including 27 valid species with low dispersion rates that are restricted to small distribution areas. Cytogenetics data point to a wide chromosomal diversity in this genus due to changes that occurred in isolated populations, with chromosomal fusions and fissions explaining the 2n number variation. In addition, different multiple sex chromosome systems and rDNA loci location are also found in some species. However, several species and populations remain to be investigated. In this study, and two still undescribed species, morphologically identified as sp. 1 and sp. 2, were cytogenetically analyzed. has 2n = 52 and 2n = 53 chromosomes, while sp. 1 has 2n = 56 and 2n = 57 chromosomes in females and males, respectively, thus highlighting the occurrence of an XX/XYY multiple sex chromosome system in both species. sp. 2 presents 2n = 62 chromosomes for both females and males, with fission events explaining its karyotype diversification. Chromosomal locations of the rDNA sites were also quite different among species, reinforcing that extensive rearrangements had occurred in their karyotype evolution. Comparative genomic hybridization (CGH) experiments among some species evidenced a shared content of the XYY sex chromosomes in three of them, thus pointing towards their common origin. Therefore, the comparative analysis among all species cytogenetically studied thus far allowed us to provide an evolutionary scenario related to the speciation process of this fish group.
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http://dx.doi.org/10.3390/genes11111366DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7698909PMC
November 2020

Chromosomes of Asian cyprinid fishes: Variable karyotype patterns and evolutionary trends in the genus Osteochilus (Cyprinidae, Labeoninae, "Osteochilini").

Genet Mol Biol 2020 6;43(4):e20200195. Epub 2020 Nov 6.

Universidade Federal de São Carlos, Departamento de Genética e Evolução, São Carlos, SP, Brazil.

The Cyprinidae family is a highly diversified but demonstrably monophyletic lineage of cypriniform fishes. Among them, the genus Osteochilus contains 35 recognized valid species distributed from India, throughout Myanmar, Laos, Thailand, Malaysia, Indonesian archipelago to southern China. In this study, karyotypes and other chromosomal characteristics of five Osteochilus species occurring in Thailand, namely O. lini, O. melanopleura, O. microcephalus, O. vittatus and O. waandersii were examined using conventional and molecular cytogenetic protocols. Our results showed they possessed diploid chromosome number (2n) invariably 2n = 50, but the ratio of uni- and bi-armed chromosomes was highly variable among their karyotypes, indicating extensive chromosomal rearrangements. Only one chromosome pair bearing 5S rDNA sites occurred in most species, except O. melanopleura, where two sites were detected. In contrast, only one chromosomal pair bearing 18S rDNA sites were observed among their karyotypes, but in different positions. These cytogenetic patterns indicated that the cytogenomic divergence patterns of these Osteochilus species were largely corresponding to the inferred phylogenetic tree. Similarly, different patterns of the distributions of rDNAs and microsatellites across genomes of examined species as well as their different karyotype structures indicated significant evolutionary differentiation of Osteochilus genomes.
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http://dx.doi.org/10.1590/1678-4685-GMB-2020-0195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7783954PMC
November 2020

Comparative chromosomal mapping of microsatellite repeats reveals divergent patterns of accumulation in 12 Siluridae (Teleostei: Siluriformes) species.

Genet Mol Biol 2020 6;43(4):e20200091. Epub 2020 Nov 6.

Universidade Federal de São Carlos (UFSCar), Departamento de Genética e Evolução, São Carlos, SP, Brazil.

The freshwater family Siluridae occurs in Eurasia and is especially speciose in South and Southeast Asia, representing an important aquaculture and fishery targets. However, despite the restricted cytogenetic data, a high diploid number variation (from 2n=40 to 92) characterizes this fish group. Considering the large genomic divergence among its species, silurid genomes have experienced an enormous diversification throughout their evolutionary history. Here, we aim to investigate the chromosomal distribution of several microsatellite repeats in 12 Siluridae species and infer about their possible roles in the karyotype evolution that occurred in this group. Our results indicate divergent patterns of microsatellite distribution and accumulation among the analyzed species. Indeed, they are especially present in significant chromosome locations, such as the centromeric and telomeric regions, precisely the ones associated with several kinds of chromosomal rearrangements. Our data provide pieces of evidence that repetitive DNAs played a direct role in fostering the chromosomal differentiation and biodiversity in this fish family.
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http://dx.doi.org/10.1590/1678-4685-GMB-2020-0091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7654372PMC
November 2020

Evolution of the parthenogenetic rock lizard hybrid karyotype: Robertsonian translocation between two maternal chromosomes in Darevskia rostombekowi.

Chromosoma 2020 12 30;129(3-4):275-283. Epub 2020 Oct 30.

Department of Zoology, Yerevan State University, Yerevan, Armenia.

Darevskia rostombekowi, the most outstanding of the seven known parthenogenetic species in the genus Darevskia, is the result of an ancestral cross between two bisexual species Darevskia raddei and Darevskia portschinskii. The chromosomal set of this species includes a unique submetacentric autosomal chromosome; the origin of this chromosome was unresolved as only acrocentric chromosomes are described in the karyotypes of Darevskia genus normally. Here, we applied a suite of molecular cytogenetic techniques, including the mapping of telomeric (TTAGGG) n repeats using fluorescence in situ hybridization (FISH), comparative genomic hybridization (CGH), and whole-chromosome painting (WCP) in both D. rostombekowi and parental (D. portschinskii and D. raddei) species. The obtained results in total suggest that a de novo chromosomal rearrangement via Robertsonian translocation (centric fusion) between two maternal (D. raddei) acrocentric chromosomes of different size was involved in the formation of this unique submetacentric chromosome present in the parthenogenetic species D. rostombekowi. Our findings provide new data in specific and rapid evolutional processes of a unisexual reptile species karyotype.
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http://dx.doi.org/10.1007/s00412-020-00744-7DOI Listing
December 2020

Revisiting the Karyotype Evolution of Neotropical Boid Snakes: A Puzzle Mediated by Chromosomal Fissions.

Cells 2020 10 10;9(10). Epub 2020 Oct 10.

Laboratory of Animal Genetics, Instituto Nacional de Pesquisas da Amazônia, Coordenação de Biodiversidade, Av. André Araújo 2936, Petrópolis, Manaus 69067-375, AM, Brazil.

The Boidae family is an ancient group of snakes widely distributed across the Neotropical region, where several biogeographic events contributed towards shaping their evolution and diversification. Most species of this family have a diploid number composed of 2n = 36; however, among Booidea families, the Boidae stands out by presenting the greatest chromosomal diversity, with 2n ranging between 36 and 44 chromosomes and an undifferentiated XY sex chromosome system. Here, we applied a comparative chromosome analysis using cross-species chromosome paintings in five species representing four Boidae genera, to decipher the evolutionary dynamics of some chromosomes in these Neotropical snakes. Our study included all diploid numbers (2n = 36, 40, and 44) known for this family and our comparative chromosomal mappings point to a strong evolutionary relationship among the genera , , , and . The results also allowed us to propose the cytogenomic diversification that had occurred in this family: a process mediated by centric fissions, including fission events of the putative and undifferentiated XY sex chromosome system in the 2n = 44 karyotype, which is critical in solving the puzzle of the karyotype evolution of boid snakes.
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http://dx.doi.org/10.3390/cells9102268DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7601083PMC
October 2020

Production of donor-derived eggs after ovarian germ cell transplantation into the gonads of adult, germ cell-less, triploid hybrid fish†.

Biol Reprod 2020 12;103(6):1289-1299

Noto Center for Fisheries Science and Technology, Faculty of Biological Science and Technology, Kanazawa University, Ishikawa, Japan.

In animals, spermatogonial transplantation in sterile adult males is widely developed; however, despite its utility, ovarian germ cell transplantation is not well developed. We previously showed that the interspecific hybrid offspring of sciaenid was a suitable model for germ cell transplantation studies as they have germ cell-less gonads. However, all these gonads have testis-like characteristics. Here, we tested whether triploidization in hybrid embryos could result in germ cell-less ovary development. Gonadal structure dimorphism and sex-specific gene expression patterns were examined in 6-month-old triploid hybrids (3nHybs). Thirty-one percent of 3nHybs had germ cell-less gonads with an ovarian cavity. cyp19a1a and foxl2, ovarian differentiation-related genes, were expressed in these gonads, whereas dmrt1 and vasa were not expressed, suggesting ovary-like germ cell-less gonad development. Some (26%) 3nHybs had testis-like germ cell-less gonads. Ovarian germ cells collected from homozygous green fluorescent protein (GFP) transgenic blue drum (BD) (Nibea mitsukurii) were transplanted into 6-month-old 3nHybs gonads via the urogenital papilla or oviduct. After 9 months, the recipients were crossed with wild type BD. Among the six 3nHyb recipients that survived, one female and one male produced fertile eggs and motile sperm carrying gfp-specific DNA sequences. Progeny tests revealed that all F1 offspring possessed gfp-specific DNA sequences, suggesting that these recipients produced only donor-derived eggs or sperm. Histological observation confirmed donor-derived gametogenesis in the 3nHyb recipients' gonads. Overall, triploidization reduces male-biased sex differentiation in germ cell-less gonads. We report, for the first time, donor-derived egg production in an animal via direct ovarian germ cell transplantation into a germ cell-less ovary.
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http://dx.doi.org/10.1093/biolre/ioaa168DOI Listing
December 2020

Landscape of snake' sex chromosomes evolution spanning 85 MYR reveals ancestry of sequences despite distinct evolutionary trajectories.

Sci Rep 2020 07 27;10(1):12499. Epub 2020 Jul 27.

Coordenação de Biodiversidade, Laboratory of Animal Genetics, Instituto Nacional de Pesquisas da Amazônia, Av. André Araújo 2936, Petrópolis, Manaus, AM, 69067-375, Brazil.

Most of snakes exhibit a ZZ/ZW sex chromosome system, with different stages of degeneration. However, undifferentiated sex chromosomes and unique Y sex-linked markers, suggest that an XY system has also evolved in ancestral lineages. Comparative cytogenetic mappings revealed that several genes share ancestry among X, Y and Z chromosomes, implying that XY and ZW may have undergone transitions during serpent's evolution. In this study, we performed a comparative cytogenetic analysis to identify homologies of sex chromosomes across ancestral (Henophidia) and more recent (Caenophidia) snakes. Our analysis suggests that, despite ~ 85 myr of independent evolution, henophidians and caenophidians retained conserved synteny over much of their genomes. However, our findings allowed us to discover that ancestral and recent lineages of snakes do not share the same sex chromosome and followed distinct pathways for sex chromosomes evolution.
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http://dx.doi.org/10.1038/s41598-020-69349-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7385105PMC
July 2020

Cytogenetic mechanisms of unisexuality in rock lizards.

Sci Rep 2020 05 26;10(1):8697. Epub 2020 May 26.

Vavilov Institute of General Genetics RAS, Moscow, Russia.

Darevskia rock lizards is a unique complex taxa, including more than thirty species, seven of which are parthenogenetic. In mixed populations of Darevskia lizards, tri- and tetraploid forms can be found. The most important issues in the theory of reticulate evolution of Darevskia lizards are the origin of parthenogenetic species and their taxonomic position. However, there is little data on how meiosis proceeds in these species. The present work reports the complex results of cytogenetics in a diploid parthenogenetic species - D. unisexualis. Here we detail the meiotic prophase I progression and the specific features оf mitotic chromosomes organization. The stages of meiosis prophase I were investigated by immunocytochemical analysis of preparations obtained from isolated primary oocytes of D. unisexualis in comparison with maternal species D. raddei nairensis. It has been shown that in D. unisexualis at the leptotene-zygotene stages the axial elements and the synaptonemal complex (SC) form typical "bouquets". At the pachytene-diplotene stage, 18 autosomal SC-bivalents and thickened asynapted sex Z and w univalents were observed. The presence of SYCP1 protein between the lateral elements of autosomal chromosomes proved the formation of assembled SCs. Comparative genomic hybridization (CGH) on the mitotic metaphase chromosomes of D. unisexualis was carried out using the genomic DNA isolated from the parental species D. raddei nairensis and D. valentini. In the pericentromeric regions of half of the mitotic chromosomes of D. unisexualis, specific regions inherited from maternal species have been found. Following our results, we suggest a model for diploid germ cells formation from diploid oocytes without premeiotic duplication of chromosomes in the oogenesis of diploid parthenogenetic lizards D. unisexualis. Taken as a whole, our findings confirm the hybrid nature of D. unisexualis and shed light on heterozygosity and automixis in diploid parthenogenetic forms.
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http://dx.doi.org/10.1038/s41598-020-65686-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7250862PMC
May 2020

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

Genes (Basel) 2020 04 28;11(5). Epub 2020 Apr 28.

Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, 277-21 Liběchov, Czech Republic.

The bighead carps of the genus ( and ) are important aquaculture species. They were subjected to extensive multidisciplinary research, but with cytogenetics confined to conventional protocols only. Here, we employed Giemsa-/C-/CMA- stainings and chromosomal mapping of multigene families and telomeric repeats. Both species shared (i) a diploid chromosome number 2 = 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 (CMA-positive) rDNA sites. Additionally, we applied here, for the first time, the conventional cytogenetics in , a species considered extinct in the wild and/or extensively cross-hybridized with . Its 2 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.
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http://dx.doi.org/10.3390/genes11050479DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7291238PMC
April 2020

Centric Fusions behind the Karyotype Evolution of Neotropical Pencilfishes (Characiforme, Lebiasinidae): First Insights from a Molecular Cytogenetic Perspective.

Genes (Basel) 2020 01 13;11(1). Epub 2020 Jan 13.

Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos, São Paulo 13565-905, Brazil.

Lebiasinidae is a Neotropical freshwater family widely distributed throughout South and Central America. Due to their often very small body size, Lebiasinidae species are cytogenetically challenging and hence largely underexplored. However, the available but limited karyotype data already suggested a high interspecific variability in the diploid chromosome number (2), which is pronounced in the speciose genus , a popular taxon in ornamental fish trade due to its remarkable body coloration. Aiming to more deeply examine the karyotype diversification in , we combined conventional cytogenetics (Giemsa-staining and C-banding) with the chromosomal mapping of tandemly repeated 5S and 18S rDNA clusters and with interspecific comparative genomic hybridization (CGH) to investigate genomes of four representative species: , , , and . Our data showed a remarkable variability in 2, ranging from 2 = 22 in (karyotype composed exclusively of metacentrics/submetacentrics) to 2 = 44 in (karyotype composed entirely of acrocentrics) On the other hand, patterns of 18S and 5S rDNA distribution in the analyzed karyotypes remained rather conservative, with only two 18S and two to four 5S rDNA sites. In view of the mostly unchanged number of chromosome arms (FN = 44) in all but one species (; FN = 36), and with respect to the current phylogenetic hypothesis, we propose Robertsonian translocations to be a significant contributor to the karyotype differentiation in (at least herein studied) species. Interspecific comparative genome hybridization (CGH) using whole genomic DNAs mapped against the chromosome background of found a moderate divergence in the repetitive DNA content among the species' genomes. Collectively, our data suggest that the karyotype differentiation in has been largely driven by major structural rearrangements, accompanied by only low to moderate dynamics of repetitive DNA at the sub-chromosomal level. Possible mechanisms and factors behind the elevated tolerance to such a rate of karyotype change in are discussed.
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http://dx.doi.org/10.3390/genes11010091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7017317PMC
January 2020

Cytogenetics of the small-sized fish, Copeina guttata (Characiformes, Lebiasinidae): Novel insights into the karyotype differentiation of the family.

PLoS One 2019 19;14(12):e0226746. Epub 2019 Dec 19.

Laboratório de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos, São Paulo, Brazil.

Lebiasinidae is a small fish family composed by miniature to small-sized fishes with few cytogenetic data (most of them limited to descriptions of diploid chromosome numbers), thus preventing any evolutionary comparative studies at the chromosomal level. In the present study, we are providing, the first cytogenetic data for the red spotted tetra, Copeina guttata, including the standard karyotype, C-banding, repetitive DNA mapping by fluorescence in situ hybridization (FISH) and comparative genomic hybridization (CGH), providing chromosomal patterns and novel insights into the karyotype differentiation of the family. Males and females share diploid chromosome number 2n = 42 and karyotype composed of 2 metacentric (m), 4 submetacentric (sm) and 36 subtelocentric to acrocentric (st-a) chromosomes. Blocks of constitutive heterochromatin were observed in the centromeric and interstitial regions of several chromosomes, in addition to a remarkably large distal block, heteromorphic in size, which fully corresponded with the 18S rDNA sites in the fourth chromosomal pair. This overlap was confirmed by 5S/18S rDNA dual-color FISH. On the other hand, 5S rDNA clusters were situated in the long and short arms of the 2nd and 15th pairs, respectively. No sex-linked karyotype differences were revealed by male/female CGH experiments. The genomic probes from other two lebiasinid species, Lebiasina melanoguttata and Pyrrhulina brevis, showed positive hybridization signals only in the NOR region in the genome of C. guttata. We demonstrated that karyotype diversification in lebiasinids was accompanied by a series of structural and numeric chromosome rearrangements of different types, including particularly fusions and fissions.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0226746PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6922430PMC
April 2020

Microdissection and whole chromosome painting confirm karyotype transformation in cryptic species of the Lariophagus distinguendus (Förster, 1841) complex (Hymenoptera: Pteromalidae).

PLoS One 2019 14;14(11):e0225257. Epub 2019 Nov 14.

Institute of Human Genetics, Jena University Hospital, Jena, Germany.

Karyotypes of two cryptic species of parasitoid Hymenoptera with n = 5 and 6 belonging to the Lariophagus distinguendus (Förster, 1841) complex, which includes cosmopolitan parasitoids of coleopteran stored-product pests, were studied using glass-needle based microdissection, reverse and cross-species fluorescence in situ hybridisation (FISH). This experiment strongly indicates that the largest metacentric chromosome in the karyotype with n = 5 originated from a particular fusion between the only acrocentric and a smaller metacentric chromosome of the set with n = 6, therefore confirming our previous hypothesis based on the karyotypic analysis using chromosome morphometrics. This study represents the first successful application of both microdissection and whole chromosome painting for the reconstruction of karyotypic rearrangements in closely related species of parasitoids, as well as in the order Hymenoptera in general.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0225257PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855445PMC
March 2020

Chromosome Microdissection on Semi-Archived Material.

Cytometry A 2019 12 18;95(12):1285-1288. Epub 2019 Sep 18.

Jena University Hospital, Institute of Human Genetics, Friedrich Schiller University, Jena, Germany.

Glass needle-based chromosome microdissection (midi) is a standard approach developed in the 1980s and remains more frequently applied in testing than the comparable technique using laser-based platforms. As the amount of DNA extracted by this technique is minimal and often in the range of picograms, the isolated DNA must be further amplified prior to use; the isolated amplified product can be readily utilized in multiple molecular research and diagnostic investigation. DNA libraries created by midi are either chromosome- or chromosome-region-specific. However, a critical component to this process is the need for timely chromosome preparation via the air-drying method not to exceed a ~2-3 h before midi is performed. Failure of this time-sensitive step often results in the chromosomes drying out after dropping, and upon initiation of the midi technique, the dissected material can jump away while touching by the needle, and collection of a suitable sample is inhibited. Herein, we demonstrate with a simple adaptation of the standard procedure, midi can be performed on semi-archived material stored for longer periods at -20°C. Thus, the critical step to obtain well-spread chromosome preparations can be completed under established conditions, for example, in the primary laboratory, stored at -20°C, and sent directly to specialized reference laboratories offering midi. In our study, we were able to obtain high-quality DNA libraries, as verified by gel electrophoreses and reverse fluorescence in situ hybridization, via midi extracted chromosome spreads derived from human, fish, snake, lampbrush, and insect stored for up to 6 months. © 2019 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.
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http://dx.doi.org/10.1002/cyto.a.23896DOI Listing
December 2019
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