ResumoOs estudos que envolvem os aspectos reprodutivos do tamanduá-bandeira (Myrmecophaga tridactyla) podem contribuir para a preservação da espécie auxiliando no desenvolvimento de um manejo reprodutivo mais eficiente em cativeiro. O presente estudo teve como objetivo obter uma melhor compreensão da fisiologia reprodutiva por meio da caracterização física e morfológica do sêmen de tamanduá-bandeira. Foram realizados três exames reprodutivos em dois tamanduás-bandeira mantidos no Zoológico da Universidade Federal de Mato Grosso, Cuiabá/MT/Brasil. O método de coleta do sêmen foi a eletroejaculação. O volume médio obtido foi de 2,62 ± 1,67 mL, com motilidade média de 50 ± 20% e vigor de 2,17 ± 0,82. O pH médio foi de 7,29 ± 0,40. Quanto à patologia espermática foram encontrados 16,5% de defeitos maiores, 20,5 % de defeitos menores, perfazendo 37 ± 4,2% de defeitos totais. Os testes preliminares de padronização da coloração eosina-nigrosina mostraram-se eficazes para avaliar a integridade da membrana plasmática de espermatozoides nesta espécie, que nesta pesquisa foi de 65 ± 7,77%.Palavras-chave: eletroejaculação, Pilosa, reprodução, Xenarthra. AbstractStudies involving the reproductive aspects of the giant anteater (Myrmecophaga tridactyla) can be contributed to the preservation of this specie. The aim of these study was accessed the reproductive physiology and describe physical and morphological giant anteater's sperm. We conducted three reproductive examination in two anteater maintained at the Federal University of Mato Grosso Zoo, Cuiaba/MT/Brazil. The method of semen collection used was electroejaculation. The volume average obtained was 2.62 ± 1.67 mL, the motility average was 50 ± 20% and the vigor was 2.17 ± 0.82. The pH mean was 7.29 ± 0.40. The sperm average concentration was 33.33 ± 12.52 x 10 6 sperm/mL. According sperm pathology we found 16.5% of larger defects, 20.5% of minor defects, amounting to 37 ± 4.2% of total defects. The preliminary tests of standardization of eosin-nigrosin staining proved to be effective to evaluate the integrity of plasma membrane of sperm in this species, in this research was 65 ± 7,77%.Keywords: eletroejaculation, Pilosa, reproduction, Xenarthra. IntroduçãoO tamanduá-bandeira (Myrmecophaga tridactyla), maior espécie de tamanduá do mundo, encontra-se distribuído por toda América Central e do Sul, vivendo em habitats variados, incluindo florestas e pastagens abertas (Miranda et al., 2003). Pertence à Ordem Xenarthra (Xenon = estranho e Arthros = articulação), atualmente desmembrada em duas ordens: Cingulata representada pelos tatus e Pilosa representada pelos tamanduás e preguiças (MEDRI et al., 2006). Não possuem dimorfismo sexual evidente, embora as fêmeas sejam menos corpulentas e apresentem peso menor comparada aos machos (Miranda, 2004).Os aspectos reprodutivos dos tamanduás ainda não foram profundamente estudados. Os machos apresentam seus testículos no interior da cavidade pélvica, não expõem o pênis e ambos os sexos apresentam uma fenda genital, dificultando a sexa...
Disturbances in oxidant/production in favour of oxidizing cause oxidative stress during the cryopreservation process. Vitamin C is an antioxidant non-enzymatic present in seminal plasma that protects sperm from oxidative stress. The aim of this study was to evaluate supplementation with ascorbate in a cryopreservation medium on oxidative stress and quality of cryopreserved sperm membranes. It was used in 24 bulls averaging 31 months and 732 kg, kept on pasture for this experiment. One ejaculate was collected from each breeder by electrostimulation. The treatments were CO-Control (no additives) and AS-ascorbate (0.45 mg mL–1). Ascorbate was added to diluted semen at the time of filling in 0.5-mL straws before the cryopreservation process. Thawing was performed in a water bath at 37°C for 30 s. After thawing the samples, aliquots for assessment of plasma and acrosome membrane integrity and evaluation of oxidative stress [TBARS concentration spontaneous (TE) and induced (TI)] were taken. The experiment was conducted in a completely randomised design. Data were analysed by ANOVA and compared by the Tukey average test with a significance level of 5%. No differences were observed (CO- 31.67 ± 2.81 v. AS- 31.08 ± 2.42, P > 0.05); the quality of the sperm membrane and oxidative stress (TECO: 5.98 ± 2.13 v. TEAS: 6.62 ± 2.33, P > 0.05 and TICO: 62.34 ± 8.22 v. TIAS: 58.52 ± 8.27, P > 0.05) in sperm cryopreserved with ascorbate were similar to the control group. Typically, animals under adequate nutritional conditions produce adequate amounts of ascorbate; perhaps because of that the treatment using exogenous ascorbate did not present a significant result. The supplementation of extender with ascorbate did not affect oxidative stress and the quality of the plasma membrane and acrosome sperm cryopreserved. Preliminary studies had shown that the addition of higher concentration (4,5 mg mL–1) of ascorbate has a beneficial effect on oxidative stress and membrane integrity (P < 0.05). Thus, further research should be done to better understand the effects of ascorbate in bovine semen.
SUMMARYThis study had objective to evaluate the integrity of the plasma membrane and semen quality of bulls supplemented with tocopherol. Bulls were used 16 with an average age of 24 months and average weight of 462.2 kg in two treatments: control group (CG) and group supplemented with tocopherol (GE-400 IU of tocoferol/animal / day) added to the concentrate supplement. The groups were kept in pasture supplemented with 4.5 kg / animal / day concentrate. Tocopherol supplementation was provided for 60 days. 4 semen samples were carried out: on days 0 , 30, 60 and 15 days after the end of supplementation. Semen was collected by electroejaculation , it has measured up the physical and morphological sperm characteristics. To assess the integrity of membrane used the hyposmotic test, eosin / nigrosine, Pope and trypan blue. The experiment was conducted in a completely
O presente estudo avaliou a adição da pentoxifilina, tocoferol, ascorbato e suas combinações sobre a proteção da célula espermática bovina contra os efeitos deletérios da criopreservação. Foram utilizados 24 touros Nelores (Bos taurus indicus), criados em sistema semi-intensivo. Foi coletado um ejaculado de cada reprodutor, diluídos em TRIS-citratogema-glicerol e divididos em seis partes. Cada parte foi suplementada da seguinte maneira: sem aditivos (controle), tocoferol (10 mmol/mL), tocoferol (10 mmol/mL) + pentoxifilina (1mg/mL), ascorbato (0,45mg/mL), ascorbato (0,45mg/mL) + pentoxifilina (1mg/mL) ou pentoxifilina (1mg/mL). Após o descongelamento, as amostras foram avaliadas quanto à motilidade e características do movimento, integridade da membrana plasmática e de acrossomo e atividade mitocondrial. Os níveis de peroxidação lipídica espontânea e induzida foram avaliados pela produção de substâncias reativas ao ácido tiobarbitúrico (TBARS). A suplementação com pentoxifilina, tocoferol, ascorbato e suas combinações, não alterou (P>0,05) a atividade mitocondrial, integridade acrossomal, e a concentração de TBARS espontâneo e induzido. A adição de tocoferol + pentoxifilina reduziu a motilidade progressiva quando comparado ao ascorbato e também a integridade da membrana espermática quando comparado ao controle e ao ascorbato (P˂0,05). Já a adição de ascorbato + pentoxifilina foi deletéria sobre linearidade em comparação ao tratamento ascorbato (P˂0,05). A adição de ascorbato, tocoferol e pentoxifilina individualmente ou em combinação, não foi eficiente em diminuir os danos causados pela criopreservação e estresse oxidativo em amostras pós descongelamento de sêmen bovino.
ResumoObjetivou-se avaliar o efeito da estação do ano sobre a qualidade do sêmen fresco e criopreservado de reprodutores Pantaneiros (Bos taurus) criados em condições tropicais. Foram utilizados 7 touros Pantaneiros e 3 Nelores (controle), dos quais foi aferido circunferência escrotal, consistência testicular e após a coleta e congelamento do sêmen realizada análise de motilidade, vigor, defeitos menores, maiores e totais, concentração, integridade de membrana plasmática e acrossomal para sêmen fresco e além destas, estresse oxidativo para sêmen criopreservado. O experimento foi conduzido em delineamento inteiramente casualizado, e arranjo fatorial 2x2 (2 raças e 2 estações do ano). A raça, estação do ano ou a interação entre eles, não alteraram significativamente as médias de circunferência escrotal, consistência testicular, motilidade, vigor, concentração, integridade de membrana acrossomal e porcentagem de defeitos menores. A integridade da membrana plasmática no sêmen fresco sofreu efeito da estação do ano e foi menor no inverno em ambas raças (95,76 ± 1,77% vs. 87,07 ± 4,78% P=0,03). A estação do inverno aumentou a porcentagem de defeitos maiores (29,15% vs. 16,44%, P<0,01) e totais (17,49% vs. 30,45%, P<0,01). Os parâmetros do sêmen congelado não foram influenciados pela raça, estação do ano ou interação entre elas. Portanto, nas condições edafoclimáticas estudadas, os reprodutores Pantaneiros apresentaram redução na sua qualidade seminal na estação do inverno.Palavras-chave: estresse oxidativo, temperatura ambiente, raças localmente adaptadas. AbstractThe aim of the present study was to evaluate the effect of seasonality on the quality of fresh and cryopreserved semen of Pantaneiro breed (Bos taurus) bulls raised under tropical conditions. Scrotal circumference and testicular consistency were performed in seven Pantaneiro and three Nellore (control) bulls. Sperm motility, vigor, minor, major and total defects, concentration, plasma and acrosomal membrane integrity were assessed in fresh and post-thawed semen. Additionally, oxidative stress was determined in post-thawed semen samples. The experiment was done in a complete randomized design, with a 2x2 factorial arrangement (2 breeds and 2 seasons). Breed, season of the year or the interaction of both did not alter scrotal circumference, testicular consistency, motility, vigor, concentration, acrosomal membrane integrity and percentage of minor defects (P<0.05). Plasma membrane integrity of fresh semen was affected by the season and was smaller in winter in both breeds (95.76 ± 1.77% vs 87.07 ± 4.78%, P=0.03). Nevertheless, also in the winter there was an increase in the percentage of major (29.15% vs 16.44%, P<0.01) and total defects (17.49% vs. 30.45%, P<0.01) in fresh semen samples. Breed, season or interaction of both did not influence the sperm parameters of cryopreserved semen. Thus, in the studied climatic conditions, the Pantaneiro breed bulls present decrease in semen quality mainly in the winter season.Keywords: oxidative stress, environment tempe...
Cottonseed and its derivatives are frequently used in cattle feed as an effective dietary fibre supply and high protein and energy food source. However, the cotton plant contains gossypol, which in its free form induces male and female infertility. Therefore, this study aimed to evaluate the effect of gossypol supplementation on bovine in vitro embryo production. Ovaries were retrieved from slaughterhouses, and cumulus-oocyte complexes (COC) were recovered by follicular puncture. Based on free gossypol concentration present on the in vitro maturation, sperm capacitation, IVF and in vitro culture media, grades I, II and III COC (n=646) were divided in 3 treatments: 0μg mL−1 (control), 5μg mL−1 (G5) and 10μg mL−1 (G10). The COC were matured under a humidified atmosphere of 5% CO2 in air at 38.5°C for 24h in 90-μL droplets containing TCM-199 supplemented with 10% FCS, 0.2mM sodium pyruvate, LH, FSH, 75μg mL−1 amikacin, 17β-oestradiol. Each droplet corresponded to one replicate (n=14) and contained 15 to 18 COC. Matured COC and sperm were co-incubated in droplets (8-13 COC per 90μL) of TALP-IVF media supplemented with 6mg mL−1 BSA, 0.2mM sodium pyruvate, 30μg mL−1 heparin, 20 μM penicillamine, 10 μM hypotaurine, 1 μM epinephrine, 75μg mL−1 amikacin under a 5% CO2 humidified atmosphere at 38.5°C, for 20h. For IVF, non-sexed frozen-thawed semen was selected with Percoll® gradient. The resulting pellet was subjectively evaluated for motility and concentration and then diluted to final concentration of sperm mL−1 with fertilization medium. Presumptive zygotes were then cultured in 90-μL droplets of SOFaaci medium supplemented with 2.7mM myo-inosytol, 0.2mM pyruvate, 2.5% FCS (v/v), 5mg mL−1 BSA, 75μg mL−1 amikacin, and maintained for 8 days at 38.5°C in a humidified atmosphere with 5% CO2 in air. Cleavage, blastocysts production and hatching rates were evaluated at Days 3, 7 and 8, respectively. Data were submitted to ANOVA for parametric data and Wilcoxon test for non-parametric variables using the SAS software (SAS Institute Inc., Cary, NC, USA). Significance level was set at 5%. Cleavage rates of the control (81.05%) and G5 (71.85%) were higher compared with G10 (19.64%; P < 0.0001). Blastocyst production was lower in G5 (12.18%) compared with control (30.35%), and the addition of 10μg mL−1 of free gossypol (G10) completely inhibited embryo development (0%; P < 0.0001). As for the percentage of hatched blastocysts, the control (66.75%) had greater values compared with G5 (34.52%; P < 0.0001). Thus, the addition of 5 and 10μg mL−1 of free gossypol are extremely hazardous for in vitro bovine embryo development. Whether these deleterious effects take place in a similar fashion during in vivo embryo production remains to be investigated.
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