Chromosomal Evolution and Evolutionary Relationships of Lebiasina Species (Characiformes, Lebiasinidae)

We present the first cytogenetic data for Lebiasina bimaculata and L. melanoguttata with the aim of (1) investigating evolutionary events within Lebiasina and their relationships with other Lebiasinidae genera and (2) checking the evolutionary relationships between Lebiasinidae and Ctenoluciidae. Both species have a diploid number 2n = 36 with similar karyotypes and microsatellite distribution patterns but present contrasting C-positive heterochromatin and CMA3 + banding patterns. The remarkable interstitial series of C-positive heterochromatin occurring in L. melanoguttata is absent in L. bimaculata. Accordingly, L. bimaculata shows the ribosomal DNA sites as the only GC-rich (CMA3 +) regions, while L. melanoguttata shows evidence of a clear intercalated CMA3 + banding pattern. In addition, the multiple 5S and 18S rDNA sites in L. melanogutatta contrast with single sites present in L. bimaculata. Comparative genomic hybridization (CGH) experiments also revealed a high level of genomic differentiation between both species. A polymorphic state of a conspicuous C-positive, CMA3 +, and (CGG)n band was found only to occur in L. bimaculata females, and its possible relationship with a nascent sex chromosome system is discussed. Whole chromosome painting (WCP) and CGH experiments indicate that the Lebiasina species examined and Boulengerella maculata share similar chromosomal sequences, thus supporting the relatedness between them and the evolutionary relationships between the Lebiasinidae and Ctenoluciidae families

An Insight into the Chromosomal Evolution of Lebiasinidae (Teleostei, Characiformes)

Lebiasinidae fishes have been historically neglected by cytogenetical studies. Here we present a genomic comparison in eleven Lebiasinidae species, in addition to a review of the ribosomal DNA sequences distribution in this family. With that, we develop ten sets of experiments in order to hybridize the genomic DNA of representative species from the genus Copeina, Copella, Nannostomus, and Pyrrhulina in metaphase plates of Lebiasina melanoguttata. Two major pathways on the chromosomal evolution of these species can be recognized: (i) conservation of 2n = 36 bi-armed chromosomes in Lebiasininae, as a basal condition, and (ii) high numeric and structural chromosomal rearrangements in Pyrrhulininae, with a notable tendency towards acrocentrization. The ribosomal DNA (rDNA) distribution also revealed a marked differentiation during the chromosomal evolution of Lebiasinidae, since both single and multiple sites, in addition to a wide range of chromosomal locations can be found. With some few exceptions, the terminal position of 18S rDNA appears as a common feature in Lebiasinidae-analyzed species. Altogether with Ctenoluciidae, this pattern can be considered a symplesiomorphism for both families. In addition to the specific repetitive DNA content that characterizes the genome of each particular species, Lebiasina also keeps inter-specific repetitive sequences, thus reinforcing its proposed basal condition in Lebiasinidae

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

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. © 2019 Toma et al

Chromosomal evolution in Lebiasinidae family (Teleostei, Characiformes), with emphasis on Lebiasina species

The Lebiasinidae family is composed by seven genus of small freshwater fishes that can be found in small rivers of Central and South America. Members of this family presents a great diversity in form and body color, but their evolutionary relationships are still unclear. Our objective was to realize the first cytogenetical analysis in two species of Lebiasina genus, aiming to clarify their karyotypic characteristics and their chromosomal evolution. For this, Lebiasina bimaculata and L. melanoguttata presents 2n = 36 as their diploid number, exclusively composed by bi-armed chromosomes. The C-positive heterochromatin was located in centromeric and telomeric regions of several chromosomes in both species. But, in L. melanoguttata there is a remarkable interstitial series of c-positive heterochromatin, absent in L. bimaculata. The rDNA 5S is located in the interstitial region on q-arm of the first chromosomic pair in both species, with an additional site on telomeric region of 13 chromosome of L. melanoguttata. The 18S rDNA can be found in telomeric region of the third pair in L. bimaculata, while L. melanoguttata presents multiple marks located in telomeric region on long arm of pairs 1 and 3, in telomeric region on short arm of 7 and 9 pairs and in both regions of the second chromosomic pair. The whole chromosome painting with a probe from the first chromosomic pair showed a share of genomic content between the two Lebiasina species and Boulengerella (Ctenoluciidae), evidencing the close relationship between this two families. Additionally, the comparative genomic hybridization among the two Lebiasina species reveals a high level of genomic differentiation amid them. Another experiment of comparative genomic hybridization reveals a telomeric sex-specific region on the third chromosomic pair of L. bimaculata females. This result with the differential pattern of c-banding, CMA3+ differential stain and the fluorescent in situ hybridization with the microsatellite probe (CGG)n indicates two possibilities (i) the presence of a copy number variation process, generating differences between the homologous pair, or (ii) a nascent sex chromosome system of ZZ/ZW type. Besides that, two chromosomal evolutionary pathways were recognized in Lebiasinidae family, being the first responsible for a probable diploid number conservation in Lebiasininae and, in contrast, an acrocentrization tendency in Pyrrhulininae.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)A família Lebiasinidae é composta por sete gêneros de pequenos peixes de água doce, sendo encontrados em pequenos riachos da América Central e do Sul. Os membros desta família apresentam uma grande diversidade de formas corpóreas e coloração, mas suas relações evolutivas se mantêm inexplicadas. Nosso objetivo é realizar as primeiras análises citogenéticas em duas espécies do gênero Lebiasina, permitindo investigações acerca das características cariotípicas e de sua evolução cromossômica. Para isso, Lebiasina bimaculata e L. melanoguttata apresentaram 2n = 36 cromossomos em seu cariótipo, exclusivamente composto por cromossomos meta/submetacêntricos. A heterocromatina C-positiva se localizou nas regiões centroméricas e teloméricas de diversos cromossomos em ambas as espécies. Em L. melanoguttata, uma série intersticial de heterocromatina C-positiva é obervada, sendo ausente em L. bimaculata. O rDNA 5S se encontra na região intersticial do braço q do primeiro par cromossômico em ambas as espécies, com um sítio adicional na região telomérica do cromossomo 13 em L. melanoguttata. O rDNA 18S está localizado na região telomérica do terceiro par em L. bimaculata, enquanto L. melanoguttata apresenta múltiplas marcações localizadas na região telomérica dos braços longos dos pares 1 e 3, na região telomérica do braço curto dos pares 7 e 9 e em ambos os telômeros do segundo par cromossômico. A pintura cromossômica total com sonda do primeiro par cromossômico revelou o compartilhamento total do conteúdo genômico deste par entre ambas as espécies de Lebiasina e Boulengerella (Ctenoluciidae), evidenciando o relacionamento próximo destas famílias. Adicionalmente, a hibridização genômica comparativa entre as duas espécies de Lebiasina revelou um alto grau de diferenciação genômica entre elas. Outro experimento de hibridização genômica comparativa revelou uma região telomérica sexo-específica no terceiro par cromossômico de fêmeas de L. bimaculata. Este resultado, assim como o padrão diferencial de bandas C-positivas, coloração diferencial CMA3+ e a hibridização fluorescente com o microssatélite (CGG)n indica duas possibilidades i) a presença de uma variação de número de cópias, gerando diferenças entre os pares homólogos, ou ii) um sistema cromossômico sexual nascente do tipo ZZ/ZW. Além disso, é possível observar duas tendências na evolução cromossômica da família Lebiasinidae, sendo a primeira responsável por uma provável conservação do número diploide em Lebiasininae e, contrariamente, a tendência de acrocentrização em Pyrrhulininae.CAPES: 88882.426716/2019-0

Uso do Role-Playing Game (RPG) como complemento didático no ensino de Imunologia

O Role-Playing Game (RPG) é definido como um jogo de interpretação de papéis, no qual os participantes podem jogar uns contra os outros, ou trabalharem juntos em prol de um objetivo. Assim, elaborou-se um jogo de RPG para trabalhar o conteúdo de Imunologia com crianças e adolescentes (5a a 8a série) onde as batalhas medievais, envolvendo criaturas fantásticas, se associam a elementos que compõem o Sistema Imunológico, como um xamã que representa um linfócito T. O mestre, neste caso o professor, guiará os participantes para ensinar o conteúdo de forma lúdica, ao longo de uma “guerra” dentro de uma pessoa. Os alunos controlarão os personagens responsáveis por atacar e defender o corpo (agentes infecciosos e glóbulos brancos, respectivamente) dispondo de um dado para a rolagem de suas ações no jogo. Como resultado, espera-se com este trabalho que o conteúdo de Imunologia seja fixado mais facilmente, de forma agradável e divertida.RPG is defined as a Role-Playing Game, in which participants can play against each other, or work together towards a objective. Thereby, an RPG game was developed to work on the content of Immunology with children and adolescents (5th to 8th grade) where medieval battles, involving fantastic creatures, are associated with elements that make up the Immune System, such as a shaman representing an T lymphocyte. To teach Immunology through the RPG is primarily needed a master, will guide the participants to teach the content in a playful way, throughout a "war" within a person. The students who will control the characters responsible for attacking and defending the body (infectious agents and white blood cells, respectively) having a dice for the scrolling of their actions in the game. As a result, it is expected with this work that Immunology content will be more easily fixed in a fun and enjoyable way

Occurrence of Sex Chromosomes in Fish of the Genus <i>Ancistrus</i> with a New Description of Multiple Sex Chromosomes in the Ecuadorian Endemic <i>Ancistrus clementinae</i> (Loricariidae)

Ancistrus Kner, 1854, is the most diverse genus among the Ancistrini (Loricariidae) with 70 valid species showing a wide geographic distribution and great taxonomic and systematic complexity. To date, about 40 Ancistrus taxa have been karyotyped, all from Brazil and Argentina, but the statistic is uncertain because 30 of these reports deal with samples that have not yet been identified at the species level. This study provides the first cytogenetic description of the bristlenose catfish, Ancistrus clementinae Rendahl, 1937, a species endemic to Ecuador, aiming to verify whether a sex chromosome system is identifiable in the species and, if so, which, and if its differentiation is associated with the presence of repetitive sequences reported for other species of the family. We associated the karyotype analysis with the COI molecular identification of the specimens. Karyotype analysis suggested the presence of a ♂ZZ/♀ZW1W2 sex chromosome system, never detected before in Ancistrus, with both W1W2 chromosomes enriched with heterochromatic blocks and 18S rDNA, in addition to GC-rich repeats (W2). No differences were observed between males and females in the distribution of 5S rDNA or telomeric repeats. Cytogenetic data here obtained confirm the huge karyotype diversity of Ancistrus, both in chromosome number and sex-determination systems

Occurrence of Sex Chromosomes in Fish of the Genus Ancistrus with a New Description of Multiple Sex Chromosomes in the Ecuadorian Endemic Ancistrus clementinae (Loricariidae)

Ancistrus Kner, 1854, is the most diverse genus among the Ancistrini (Loricariidae) with 70 valid species showing a wide geographic distribution and great taxonomic and systematic complexity. To date, about 40 Ancistrus taxa have been karyotyped, all from Brazil and Argentina, but the statistic is uncertain because 30 of these reports deal with samples that have not yet been identified at the species level. This study provides the first cytogenetic description of the bristlenose catfish, Ancistrus&nbsp;clementinae Rendahl, 1937, a species endemic to Ecuador, aiming to verify whether a sex chromosome system is identifiable in the species and, if so, which, and if its differentiation is associated with the presence of repetitive sequences reported for other species of the family. We associated the karyotype analysis with the COI molecular identification of the specimens. Karyotype analysis suggested the presence of a &#9794;ZZ/&#9792;ZW1W2 sex chromosome system, never detected before in Ancistrus, with both W1W2 chromosomes enriched with heterochromatic blocks and 18S rDNA, in addition to GC-rich repeats (W2). No differences were observed between males and females in the distribution of 5S rDNA or telomeric repeats. Cytogenetic data here obtained confirm the huge karyotype diversity of Ancistrus, both in chromosome number and sex-determination systems

Small Body, Large Chromosomes: Centric Fusions Shaped the Karyotype of the Amazonian Miniature Fish <i>Nannostomus anduzei</i> (Characiformes, Lebiasinidae)

Miniature refers to species with extraordinarily small adult body size when adult and can be found within all major metazoan groups. It is considered that miniature species have experienced severe alteration of numerous morphological traits during evolution. For a variety of reasons, including severe labor concerns during collecting, chromosomal acquisition, and taxonomic issues, miniature fishes are neglected and understudied. Since some available studies indicate possible relationship between diploid chromosome number (2n) and body size in fishes, we aimed to study one of the smallest Neotropical fish Nannostomus anduzei (Teleostei, Characiformes, Lebiasinidae), using both conventional (Giemsa staining, C-banding) and molecular cytogenetic methods (FISH mapping of rDNAs, microsatellites, and telomeric sequences). Our research revealed that N. anduzei possesses one of the lowest diploid chromosome numbers (2n = 22) among teleost fishes, and its karyotype is entirely composed of large metacentric chromosomes. All chromosomes, except for pair number 11, showed an 18S rDNA signal in the pericentromeric region. 5S rDNA signals were detected in the pericentromeric regions of chromosome pair number 1 and 6, displaying synteny to 18S rDNA signals. Interstitial telomeric sites (ITS) were identified in the centromeric region of pairs 6 and 8, indicating that centric fusions played a significant role in karyotype evolution of studied species. Our study provides further evidence supporting the trend of diploid chromosome number reduction along with miniaturization of adult body size in fishes

Integrating Genomic and Chromosomal Data: A Cytogenetic Study of <i>Transancistrus santarosensis</i> (Loricariidae: Hypostominae) with Characterization of a ZZ/ZW Sex Chromosome System

The plecos (Loricariidae) fish represent a great model for cytogenetic investigations due to their variety of karyotypes, including diploid and polyploid genomes, and different types of sex chromosomes. In this study we investigate Transancistrus santarosensis a rare loricariid endemic to Ecuador, integrating cytogenetic methods with specimens’ molecular identification by mtDNA, to describe the the species karyotype. We aim to verify whether sex chromosomes are cytologically identifiable and if they are associated with the accumulation of repetitive sequences present in other species of the family. The analysis of the karyotype (2n = 54 chromosomes) excludes recent centric fusion and pericentromeric inversion and suggests the presence of a ZZ/ZW sex chromosome system at an early stage of differentiation: the W chromosome is degenerated but is not characterized by the presence of differential sex-specific repetitive DNAs. Data indicate that although T. santarosensis has retained the ancestral diploid number of Loricariidae, it accumulated heterochromatin and shows non-syntenic ribosomal genes localization, chromosomal traits considered apomorphic in the family

Homeology of sex chromosomes in Amazonian Harttia armored catfishes supports the X-fission hypothesis for the X1X2Y sex chromosome system origin

Abstract The Neotropical monophyletic catfish genus Harttia represents an excellent model to study karyotype and sex chromosome evolution in teleosts. Its species split into three phylogenetic clades distributed along the Brazilian territory and they differ widely in karyotype traits, including the presence of standard or multiple sex chromosome systems in some members. Here, we investigate the chromosomal rearrangements and associated synteny blocks involved in the origin of a multiple X1X2Y sex chromosome system present in three out of six sampled Amazonian-clade species. Using 5S and 18S ribosomal DNA fluorescence in situ hybridization and whole chromosome painting with probes corresponding to X1 and X2 chromosomes of X1X2Y system from H. punctata, we confirm previous assumptions that X1X2Y sex chromosome systems of H. punctata, H. duriventris and H. villasboas represent the same linkage groups which also form the putative XY sex chromosomes of H. rondoni. The shared homeology between X1X2Y sex chromosomes suggests they might have originated once in the common ancestor of these closely related species. A joint arrangement of mapped H. punctata X1 and X2 sex chromosomes in early diverging species of different Harttia clades suggests that the X1X2Y sex chromosome system may have formed through an X chromosome fission rather than previously proposed Y-autosome fusion