Search

Submitted by DAVID JAVIER GALINDO HUAMÁN (david.galindo@unesp.br) on 2021-05-15T20:53:41Z No. of bitstreams: 1 galinndohuaman_dj_dr_jabo.pdf: 2488355 bytes, checksum: 3c726570b1835c52f524e7c1a5ef3b08 (MD5) Rejected by Alexandra Maria Donadon Lusser Segali null (alexmar@fcav.unesp.br), reason: Solicitamos que realize correções na submissão seguindo as orientações abaixo: Bom dia David, Vc deve fazer as correções naa refereência conformee-mail enviado ao repositório. Vi que teve financiamento da FAPESP também este deve ser colocado juntamente com o outro financiamento que teve durante a submissão, da seguinte maneira. FONDECYT: 116-2017 FAPESP: 17/07014-8 FAPESP: 19/06940-1 Agradecemos a compreensão. on 2021-05-17T13:18:45Z (GMT) Submitted by DAVID JAVIER GALINDO HUAMÁN (david.galindo@unesp.br) on 2021-05-17T15:39:45Z No. of bitstreams: 1 galindohuaman_dj_dr_jabo.pdf: 2491482 bytes, checksum: 4681f02327a42e6992a868e7b0adb1ef (MD5) Approved for entry into archive by Alexandra Maria Donadon Lusser Segali null (alexmar@fcav.unesp.br) on 2021-05-20T12:28:06Z (GMT) No. of bitstreams: 1 galindohuaman_dj_dr_jabo.pdf: 2491482 bytes, checksum: 4681f02327a42e6992a868e7b0adb1ef (MD5) Made available in DSpace on 2021-05-20T12:28:06Z (GMT). No. of bitstreams: 1 galindohuaman_dj_dr_jabo.pdf: 2491482 bytes, checksum: 4681f02327a42e6992a868e7b0adb1ef (MD5) Previous issue date: 2021-04-05 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) A família Cervidae destaca-se por possuir uma das maiores taxas de evolução cariotípica dentre os mamíferos, reflexo de uma fragilidade cromossômica acentuada, que facilita a ocorrência de quebras e rearranjos cromossômicos. Estas, quando vinculadas ao isolamento geográfico em um curto período, poderiam levar as populações a um processo de especiação. Isso é evidenciado na ampla diversificação cariotípica da família, apresentando exemplos extremos de baixo e alto número diploide, como Muntiacus muntjak (2n = 6/7) e Capreolus pygargus (2n = 70 + 1 – 14 Bs), respectivamente. Muitas espécies crípticas estão escondidas pela grande similaridade morfológica, acompanhada de ampla variação cariotípica, intra e interespecífica. Animais portadores de rearranjos cromossômicos podem apresentar diferentes padrões na segregação meiótica durante a espermiogênese, dando passo à formação de gametas desbalanceados. Isso pode estar relacionado com desordens reprodutivas, como evidenciado em animais domésticos, onde é observada a queda da capacidade reprodutiva. A técnica de hibridização in situ fluorescente (FISH) permite a marcação, identificação e localização dos cromossomos envolvidos nas translocações a partir de sondas de DNA com marcadores fluorescentes. Dada a falta de sondas específicas para espécies de cervídeos, diversos estudos apontam para o uso de sondas bovinas devido ao conhecido mapa genético bovino e à proximidade filogenética entre as famílias Cervidae e Bovidae. Dessa forma, a aplicação da FISH em células espermáticas (também chamada de sperm-FISH) permite estimar a proporção de gametas normais/balanceados e desbalanceados (portadores de aneuploidias). Assim, o presente estudo teve como objetivo: a) Estimar a proporção dos produtos da segregação meiótica em indivíduos portadores de translocações cromossômicas dentro do gênero Mazama, b) Avaliar os potenciais efeitos dos rearranjos cromossômicos na aptidão reprodutiva dos portadores, e c) Avaliar o papel dos rearranjos cromossômicos nos processos de especiação dentro do gênero Mazama. Na espécie M. gouazoubira foi possível avaliar uma translocação Robertsoniana (TR) em quatro animais, sendo que dois incluíam a presença de uma inversão paracêntrica (IPA). O valor médio dos espermatozoides desbalanceados nos portadores da TR/IPA (6,68%) quase dobrou em relação àquele dos portadores de TR (3,76%), mas não houve diferença significativa. Na espécie M. americana, foram avaliadas TRs em diferentes citótipos e fusões em tandem (FT) heterozigotas em híbridos entre citótipos da mesma linhagem cromossômica. Os portadores de TR apresentaram valor médio para a taxa de segregação adjacente de 1,80% e os portadores da FT apresentaram valor médio de 29,07% para os produtos equivalentes a aqueles da segregação adjacente na TR. Nossos resultados indicam um impacto de baixo a moderado do rearranjo cromossômico na aptidão reprodutiva dos machos de M. gouazoubira heterozigotos para TR e TR/IPA e um impacto baixo no caso das TRs em M. americana. No caso dos híbridos de M. americana portadores de FT em heterozigose, os resultados sugerem a formação de uma barreira pós-zigótica eficiente representada pela redução severa da fertilidade dos indivíduos. The family Cervidae stands out for showing one of the highest rates of karyotype evolution among mammals, reflecting a marked chromosomal fragility. Thus, the occurrence of chromosomal breaks and rearrangements is facilitated and lead populations to process of speciation when linked to geographic isolation in a short period. Besides, a wide karyotype diversification of the family has been observed in their extreme examples of low and high diploid numbers, such as Muntiacus muntjak (2n = 6/7) and Capreolus pygargus (2n = 70 + 1 – 14 Bs), respectively. Many cryptic species are hidden due to the great morphological similarity, accompanied by wide intra and interspecific karyotypic variation. Thus, animals with chromosomal rearrangements may present different patterns of meiotic segregation during spermiogenesis, giving way to the formation of unbalanced gametes. This can be related to reproductive disorders, as observed in domestic animals with reproductive fitness reduction. The fluorescent in situ hybridization (FISH) technique allows the marking, identification, and location of the chromosomes involved in translocations using DNA probes with fluorescent markers. Given the lack of specific probes for deer species, several studies point to the use of bovine probes due to the well-known bovine genetic map and the phylogenetic proximity between families Cervidae and Bovidae. Thus, the application of FISH in sperm cells (also called sperm-FISH) allows estimating the proportion of normal/balanced and unbalanced gametes (carriers of aneuploidies). The present study aimed to a) Estimate the mean rate of meiotic segregation products in carriers of chromosomal translocations within the genus Mazama, b) Assess the potential effects of chromosomal translocations on the reproductive fitness of carriers, and c) Assess the role of chromosomal polymorphisms in speciation processes within the genus Mazama. In the M. gouazoubira species, it was possible to evaluate a Robertsonian translocation (RT) in four animals, two of which included the presence of a paracentric inversion (PAI). The mean value of unbalanced gametes in carriers of the RT/PAI (6.68%) almost doubled in relation to that of carriers with the RT (3.76%); however, no significant difference was observed. Regarding M. americana, RTs were evaluated in different cytotypes and heterozygous tandem fusion (TF) was evaluated in hybrids from the same chromosomal lineage cytotypes. Carriers of RT showed a mean value for the adjacent segregation rate of 1.80% and carriers of TF showed a mean value of 29.07% for products equivalent to those of the adjacent segregation in RT. Our results indicate low to moderate impacts of chromosomal rearrangement on the reproductive fitness of M. gouazoubira heterozygotes for RT and RT/PAI and a low impact in the case of RTs in M. americana. In the case of M. americana hybrids with heterozygous TF, our results suggest an efficient postzygotic barrier represented by the severe reduction in the individuals' fertility. La familia Cervidae se destaca por tener una de las tasas más altas de evolución cariotípica entre los mamíferos, reflejo de una marcada fragilidad cromosómica, lo que facilita la aparición de rupturas y reordenamientos cromosómicos. Estos, cuando se vinculan con casos de aislamiento geográfico en un período corto, podrían llevar a las poblaciones por un proceso de especiación. Lo que se evidencia en la amplia diversificación cariotípica de la familia, presentando ejemplos extremos de número diploide bajo y alto, como Muntiacus muntjak (2n = 6/7) y Capreolus pygargus (2n = 70 + 1 - 14 Bs), respectivamente. Muchas especies crípticas quedan ocultas debido a la gran similitud morfológica, acompañada de una amplia variación cariotípica, intra e interespecífica. Los animales con reordenamientos cromosómicos pueden presentar diferentes patrones de segregación meiótica durante la espermiogénesis, dando paso a la formación de gametos desequilibrados. Esto puede estar relacionado con trastornos reproductivos, como se evidencia en animales domésticos, donde se observa una disminución de la capacidad reproductiva. La técnica de hibridación in situ fluorescente (FISH) permite el marcado, identificación y ubicación de los cromosomas implicados en translocaciones a mediante el uso de sondas de ADN con marcadores fluorescentes. Dada la falta de sondas específicas para especies de cérvidos, varios estudios apuntan al uso de sondas bovinas debido al gran conocimiento del mapa genético bovino y la proximidad filogenética entre las familias Cervidae y Bovidae. De esa forma, la aplicación de FISH en espermatozoides (también llamado sperm-FISH) permite estimar la proporción de gametos normales / equilibrados y desequilibrados (portadores de aneuploidía). Así, el presente estudio tuvo como objetivo: a) Estimar la proporción de productos de la segregación meiótica en individuos con translocaciones cromosómicas dentro del género Mazama, b) Evaluar los efectos potenciales de los reordenamientos cromosómicos en la aptitud reproductiva de los portadores, y c) Evaluar el rol de los reordenamientos cromosómicos en procesos de especiación dentro del género Mazama. En la especie M. gouazoubira, fue posible evaluar una translocación Robertsoniana (TR) en cuatro animales, dos de los cuales incluyeron la presencia de una inversión paracéntrica (IPA). El valor promedio de espermatozoides desequilibrados en pacientes con TR / IPA (6,68%) casi se duplicó con relación al de los pacientes con TR (3,76%), aunque no hubo diferencia significativa. En la especie M. americana, se evaluaron TR en diferentes citotipos y fusiones en tándem heterocigoto (FT) en híbridos entre citotipos del mismo linaje cromosómico. Los portadores de TR tuvieron un valor promedio de segregación adyacente de 1.80% y los portadores de FT tuvieron un valor promedio de 29.07% para productos equivalentes a los de segregación adyacente en TR. Nuestros resultados indican un impacto de bajo a moderado del reordenamiento cromosómico en la aptitud reproductiva de machos heterocigotos de M. gouazoubira para TR y TR / IPA y un impacto bajo en el caso de TR en M. americana. En el caso de los híbridos de M. americana con FT en heterocigosis, los resultados sugieren la formación de una barrera post-cigótica eficiente representada por la severa reducción de la fertilidad de los individuos. FONDECYT 116-2017 FAPESP 17/07014-8 FAPESP 19/06940-1

Chromosomal polymorphism plays a major role in speciation processes in mammals with high rates of karyotypic evolution, as observed in the family Cervidae. One remarkable example is the genus Mazama that comprises wide inter- and intra-specific chromosomal variability. To evaluate the impact of chromosomal polymorphisms as reproductive barriers within the genus Mazama, inter-specific hybrids between Mazama gouazoubira and Mazama nemorivaga (MGO &times, MNE) and intra-specific hybrids between cytotypes of Mazama americana (MAM) differing by a tandem (TF) or centric fusion (Robertsonian translocations&mdash, RT) were evaluated. MGO &times, MNE hybrid fertility was evaluated by the seminal quality and testicular histology. MAM hybrids estimation of the meiotic segregation products was performed by sperm-FISH analysis. MGO &times, MNE hybrids analyses showed different degrees of fertility reduction, from severe subfertility to complete sterility. Regarding MAM, RT, and TF carriers showed a mean value for alternate segregation rate of 97.74%, and 67.23%, and adjacent segregation rate of 1.80%, and 29.07%, respectively. Our results suggested an efficient post-zygotic barrier represented by severe fertility reduction for MGO &times, MNE and MAM with heterozygous TF. Nevertheless, RT did not show a severe effect on the reproductive fitness in MAM. Our data support the validity of MGO and MNE as different species and reveals cryptic species within MAM.

Made available in DSpace on 2021-06-25T10:27:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-07-01 Central European Institute of Technology ASCRS Research Foundation Consejo Nacional de Ciencia, Tecnología e Innovación Tecnológica Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Grantová Agentura České Republiky The genus Mazama stands out among the Neotropical deer due to their wide intra and interspecific karyotypic diversification, which is associated with an accentuated chromosomal fragility. There are reports of heterozygous Robertsonian translocation (RT) carriers in a free-range population of Mazama gouazoubira (brown brocket deer), as well as in captive animals of this and other species of the genus. To analyze possible negative impacts of heterozygous chromosome rearrangements on reproductive fitness of the carriers, we performed an analysis of sperm meiotic segregation in four brown brocket bucks, carriers of a rob(4;16), and compared the results with those of a normal buck. We established a reliable FISH and sperm-FISH protocol for the brown brocket deer using bovine (Bos taurus; diploid number, 2n = 60) whole chromosome painting (WCP) and BAC probes. Using BAC probes, we revealed the presence of a paracentric inversion (PAI) of the fused chromosome 4 in two of the four analyzed RT carriers. The mean frequency of normal/balanced sperm in the translocation carriers was significantly lower than in the normal buck (94.78% vs 98.40%). The mean value of total unbalanced spermatozoa was almost doubled in the RT/PAI carriers (6.68%) when compared to RT carriers (3.76%), but the difference was not statistically significant. This study demonstrated the efficiency of FISH with bovine WCP and BAC probes in the characterization of chromosome rearrangements and gametic segregation patterns in brown brocket deer. Our results indicate a low to moderate increase in the rates of unbalanced meiotic segregation products in brown brocket bucks heterozygous for RT and RT/PAIs. Núcleo de Pesquisa e Conservação de Cervídeos Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista – NUPECCE/FCAV/UNESP Central European Institute of Technology-Veterinary Research Institute Núcleo de Pesquisa e Conservação de Cervídeos Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista – NUPECCE/FCAV/UNESP Consejo Nacional de Ciencia, Tecnología e Innovación Tecnológica: 116-2017-FONDECYT FAPESP: 2017/07014-8 FAPESP: 2019/06940-1 Grantová Agentura České Republiky: 20–22517J

Robertsonian translocations are the most frequent chromosomal rearrangements detected in cattle. Here, we report on the detection of a new Robertsonian translocation between chromosomes BTA3 and BTA16. This rob(3;16) was dicentric, suggesting that its occurrence was recent. FISH analysis of decondensed sperm nuclei revealed a relatively low rate of unbalanced gametes produced by adjacent segregation (5.87%). In addition, and for the first time in bovines, a significant interchromosomal effect (ICE) was detected for 2 different autosomes: BTA17 (global disomy + nullisomy rate of 9%) and BTA20 (1.8%). These results suggest that ICE should be taken into consideration when assessing the putative effect of Robertsonian translocations on reproduction.

Bisphenol A (BPA) is a synthetic monomer widely used to polymerize polycarbonate plastics and resins. It is shown in vitro to interfere with microtubules, producing aberations in mitotic and meiotic spindles. An increase of meiotic abnormalities in untreated female mice from an experimental colony was temporally correlated with the accidental release of BPA from polycarbonate cages and bottles damaged by inadvertent treatment with harsh alkaline detergents [P.A. Hunt, K.E. Koehler, M. Susiarjo, C.A. Hodges, A. Ilagan, R.C. Voigt, S. Thomas, B.F. Thomas, T.J. Hassold, Bisphenol A exposure causes meiotic aneuploidy in the female mouse, Curr. Biol. 13 (2003) 546-553]. In the present study, potential aneugenic effects of BPA on mouse male and female germ cells and bone marrow cells have been evaluated after acute, sub-chronic or chronic in vivo exposure. Female mice were orally treated with a single BPA dose, with 7 daily administrations or exposed for 7 weeks to BPA in drinking water. No significant induction of hyperploidy or polyploidy was observed in oocytes and zygotes at any treatment condition. The only detectable effect was a significant increase of metaphase 11 oocytes with prematurely separated chromatids after chronic exposure; this effect, however, had no irreversible consequence upon the fidelity of chromosome segregation during the second meiotic division, as demonstrated by the normal chromosome constitution of zygotes under the same exposure condition. With male mice, no delay of meiotic divisions was found after six daily oral doses of BPA with the BrdU assay. Similarly, no induction of hyperploidy and polyploidy was shown in epydidimal sperm hybrized with probes for chromosomes 8, X and Y, 22 days after six daily oral BPA doses. Finally, two daily oral BPA doses did not induce any increase of micronucleus frequencies in polychromatic erythrocytes of mouse bone marrow. In conclusion, our results do not add evidence to the suspected aneugenic activity of BPA and suggest that other factors or co-factors should be considered to explain the unexpected burst of meiotic abnormalities previously attributed to accidental BPA exposure. (c) 2007 Elsevier B.V. All rights reserved.