Spectrophotometric method for determination of Colossoma macropomum (tambaqui) sperm concentration / Determinação da concentração espermática de tambaqui (Colossoma macropomum) por espectrofotometria

To assist the reproductive management of tambaqui (Colossoma macropomum) males in laboratory and commercial fish farming, a linear regression model was obtained from concentration curves using the spectrophotometric method. Twenty-two tambaqui males with an average age of three years old were selected and divided into two groups containing 11 animals each. Both groups alternately received a single dose of carp pituitary extract (CPE; 2.0 mg/kg body weight, intracoelomic). Sperm was collected 14 h after hormonal treatment and diluted (1:4000; sperm:formaldehyde saline). The concentration was estimated by counting spermatozoa in a Neubauer chamber and by using a spectrophotometer (λ=540 nm). Individual sperm concentration ranged from 11.40 to 71.13 × 109 sperm/mL. The degree of transmittance ranged from 62.1% to 95.0%. There was a significant correlation (r2 = 0.966; p < 0.0001) between sperm concentration analyzed in a Neubauer chamber and transmittance at 540 nm. Analysis by spectrophotometry and the prediction provided by the equation Y=100.293 - 0.509X proved to be an efficient and fast method for estimating sperm concentration in tambaqui and can be used in routine procedures in artificial fish reproduction laboratories.

Visando auxiliar o manejo reprodutivo de machos de tambaqui (Colossoma macropomum) em piscicultura de laboratório e comercial, obteve-se um modelo de regressão linear a partir de curvas de concentração por método espectrofotométrico. Foram selecionados 22 machos de tambaqui com idade média de três anos. Eles foram divididos em dois grupos contendo 11 animais cada. Ambos os grupos receberam alternadamente uma única dose de extrato de hipófise de carpa (EHC; 2,0 mg/kg de peso corporal, intracelomático). O esperma foi coletado 14 horas após o tratamento hormonal e diluído (1:4000; esperma: solução salina formaldeído). A concentração foi estimada por contagem de espermatozoides em câmara de Neubauer e por espectrofotômetro (λ=540 nm). A concentração espermática individual variou de 11,40 a 71,13 × 109 espermatozoides/mL. O grau de transmitância variou de 62,1 a 95,0%. Houve correlação significativa (r2 = 0,966; p < 0,0001) entre a concentração espermática analisada em câmara de Neubauer e a transmitância em 540 nm. A análise por espectrofotometria e a predição pela equação Y=100,293 - 0,509X mostrou ser um método eficiente e rápido para estimar a concentração espermática de tambaqui, podendo ser utilizado em procedimentos de rotina em laboratórios de reprodução artificial de peixes.

Assessment of coconut water added to Numida meleagris egg yolk as cryoprotectant for goat semen

Effects were assessed of the dilutants TRIS and ACP - 101c® with the addition of different guinea fowl (Numida meleagris) egg yolk concentrations. Fifteen ejaculates were collected from five goats of the Anglo Nubian breed. The ejaculates were pooled and then divided into 12 groups, two control groups (GC1 TRIS, with 2.5% Gallus gallus domesticus hen egg yolk GOGD), (GC2 Control Group ACP - 101c®, with the addition of 2.5% Gallus gallus domesticus hen egg yolk GOGD) and ten experimental groups (EG), containing TRIS and ACP added with different concentrations of egg yolk from guinea hen (Numida meleagris) (TRIS 2,5% GONM; TRIS 5% GONM; TRIS 10% GONM; TRIS 15% GONM; TRIS 20% GONM; ACP® 2,5% GONM; ACP® 5% GONM; ACP® 10% GONM; ACP® 15% GONM; ACP® 20% GONM). Then cryopreservation was carried out and the samples stored in liquid nitrogen (-196 °C). After seven days, the samples were thawed and assessed for spermatic kinetics, immunofluorescence and sperm morphology. Analysis of GOMN by the CASA system showed that the various parameters were similar to those of GOGD (P>0.05). The membrane integrity, mitochondrial potential and the acrosome were not influenced by the treatment (P>0.05) nor by the dilutant used for cryopreservation (P>0.05). The spermatic morphology was also preserved by the different GOGD and GONM concentrations in the ACP® and TRIS dilutants, with no statistically significant differences (P<0.05). It was concluded that Numida meleagris egg yolk, as external membrane cryoproctant added to the dilutants ACP-101c® and TRIS, improved goat semen quality.(AU)

Effect of adding anethole to the cryopreservation medium (powdered coconut water) on morphology, kinetics, and oxidative stress of buck sperm / Efeito da adição de anethole ao meio de criopreserção (água de coco em pó) na morfologia, cinética e estresse oxidativo do sêmen de bode

The quality of post-thawing goat sperm is critical to the success of artificial insemination protocols and may be influenced by extenders, cryoprotectants, and antioxidant substances. Therefore, the objective of this study was to evaluate the effects of the antioxidant anethole on goat sperm diluted in preservation medium based on powdered coconut water (ACP-101c) and frozen. For that, each ejaculate was submitted to the following treatments: ACP-101c (control); control plus supplementation with 30, 300, or 2000 µg/ mL anethole. The samples were thawed and evaluated for morphology, kinetics, membrane integrity, and reactive oxygen species (ROS). The addition of anethole increased morphological abnormalities (P < 0.05), however, it did not affect sperm kinetics. Flow cytometry analysis showed that sperm cells cryopreserved with 300 µg/mL anethole had lower acrosome integrity than those cryopreserved in other treatments. Evaluation of oxidative stress revealed that cells stored in the presence of 2000 µg/mL anethole had small amounts of ROS when compared to those preserved in the control medium alone or supplemented with 300 µg/mL anethole (P < 0.05). After cryopreservation of sperm with 2000 µg/mL anethole, the highest percentage of viable sperm without ROS was observed (P < 0.05). In conclusion, despite reducing ROS levels, the supplementation of anethole in ACP-101c did not affect sperm kinetics or membrane integrity post-thawing, however, it did cause morphological damage to sperm.(AU)

A qualidade do espermatozoide caprino pós-descongelação é crítica para o sucesso dos protocolos de inseminação artificial e pode ser influenciada por extensores, crioprotetores e substâncias antioxidantes. Portanto, o objetivo deste estudo foi avaliar os efeitos do antioxidante anetole sobre espermatozoides caprinos diluídos em meio de conservação à base de água de coco em pó (ACP-101c) e congelados. Para tanto, cada ejaculado foi submetido aos seguintes tratamentos: ACP-101c (controle); controle mais suplementação com 30, 300 ou 2000 µg / mL de anetole. As amostras foram descongeladas e avaliadas quanto à morfologia, cinética, integridade de membranas e espécies reativas de oxigênio. A adição de anetole aumentou as anormalidades morfológicas (P < 0,05), no entanto, não afetou a cinética dos espermatozoides. A análise da citometria de fluxo mostrou que as células de esperma criopreservadas com 300 µg / mL anethole tinham integridade acrosma menor do que aquelas criopreservadas em outros tratamentos. A avaliação do estresse oxidativo revelou que as células armazenadas na presença de 2000 µg / mL anethole apresentaram pequenas quantidades de ROS quando comparadas às preservadas em meio de controle isoladamente ou suplementadas com 300 µg / mL anethole (P < 0,05). Após a criopreservação de espermatozoides com 2000 µg / mL anethole, observou-se a maior porcentagem de espermatozoides viáveis sem ROS (P < 0,05). A população com espermatozoides viáveis sem ROS foi maior quando utilizado 2.000 µg / mL (P < 0,05). Em conclusão, apesar de reduzir os níveis de ROS, a suplementação de anetole em ACP-101c não afetou a cinética espermática e a integridade da membrana pós-descongelação, entretanto, causou danos morfológicos nos espermatozoides.(AU)

Sêmen ovino e caprino imunossexado e diluído em meio de conservação à base de água de coco em pó (ACP101/102c) / Ram and goat semen immunosexed and diluted in powdered coconut water-based preservation medium (ACP101/102c)

Sperm sexing aims to separate sperm populations in carriers of the "X" or "Y" chromosome. Currently, flow cytometry is a technique that allows greater accuracy; however, it causes structural changes in sperm, reduces viability, and has a high cost. As a result, other methods have been researched, including immunosexing, which uses monoclonal antibodies to detect sex-specific surface antigens. Thus, the objective of this study was to evaluate the immunosexing technique using a monoclonal antibody against sex-specific protein (HY) in the conservation of ram and goat semen in ACP101/102c. Ejaculates from five rams and five goats were collected with the aid of an artificial vagina; they were evaluated and submitted to the immunosexing protocol, according to the manufacturer's recommendations, using the Monoclonal Antibody Kit specific for mammalian sperm with "Y" chromosomes (HY; HY Biotechnology, Rio de Janeiro, RJ, Brazil). After sexing, the supernatant was resuspended in the cryopreservation diluent: ACP ram (ACP101/102c + 20% egg yolk + 7% glycerol) and ACP goat (ACP101/102c + 2.5% egg yolk + 7% glycerol), packaged in 0.25 mL straws, refrigerated at 4°C, stabilized for 30 min, frozen in liquid nitrogen vapor (-60°C) for 15 min, immersed in liquid nitrogen, and stored in cryogenic cylinders. The samples were evaluated in natura (T1), after immunosexing (T2) and after thawing (T3) for sperm motility subjectively using conventional microscopy (40x). Plasma membrane integrity (IMP) and sperm cell morphology were evaluated by the smear staining technique using eosin-nigrosine dye, and the percentages of healthy and morphologically defect spermatozoa were determined. In the evaluation of ram semen regarding sperm motility and IMP, no statistically significant differences were observed between treatments after sexing in the evaluation of absolute data (P > 0.05), with the difference being observed only between T1 and T2, and T3 (P < 0.05). Regarding the relative percentage and sperm morphology, no statistically significant differences were observed (P > 0.05). Regarding the evaluation of goat semen samples, the motility parameters were consistent with the technique submitted; however, the IMP data did not appear as expected, requiring further evaluation for a better assessment of the technique for this species. The data obtained from ram semen submitted to the immunosexing protocol, regarding the absolute evaluation of motility and IMP, demonstrated that the non-sexed semen (T1) was superior to the sexed treatments (T2 and T3); however, it is noteworthy that freezing started with approximately 50% of the cells, since the immunosexing technique results in a loss of viability of approximately 50% of the sperm, which corresponds to the ratio of sperm carrying the X chromosome. In addition, when the data in this study were transformed into relative values, no statistical differences were observed, indicating that the immunosexing protocol, as well as the freezing protocol, did not significantly affect the quality of ram sperm cells. In relation to the immunosexing of goat semen, future studies should be conducted in vitro to define a more appropriate protocol for the species and, in addition, in vivo studies should be performed to prove the quality of the technique. It was concluded that the immunosexing process using a monoclonal antibody against sex-specific protein (HY) associated with the use of powdered coconut water diluent (ACP101/102c) in the cryopreservation of semen proved to be efficient in the in vitro evaluation of ovine species.(AU)

Biotecnologias como ferramentas para o desenvolvimento do Nordeste do Brasil / Biotechnologies as tools for the development of northeast Brazil

A biotecnologia tem sido um ramo de estudo diferencial para diversos setores da sociedade, apresentando, através de bioprodutos e bioprocessos, melhorias para o avanço e desenvolvimento da região Nordeste do Brasil. Vale pontuar que um importante meio que traz anualmente acréscimos inovadores à área biotecnológica é o setor acadêmico, que através de cursos stricto sensu a nível de mestrado e doutorado, promovem pesquisas relevantes para vários setores como economia, agroindústria, saúde, dentre outros. Exemplos disso são: o curso de Doutorado em Biotecnologia da RENORBIO) e o Programa Profissional de Pós-Graduação em Biotecnologia em Saúde Humana e Animal (PPGBiotec). O presente artigo se sobre o percurso trilhado pelos cursos stricto sensu mencionados, bem como ressalta a relevância da Biotecnologia para a região Nordeste do Brasil, em seus diferentes campos de investigação, com ênfase nos Recursos Naturais, Agropecuária e Saúde. Exemplificamos as biotecnologias utilizando a água de coco que vêm sendo trabalhadas desde os anos 1980’s e sua evolução até os dias atuais. Com base em toda a potencialidade da Região Nordeste para a geração de bioprodutos e bioprocessos, ressaltamos que os mesmos só serão úteis se realmente forem tratados como inovação tecnológica, gerarem nota fiscal e impactarem positivamente para o bem estar da sociedade.

Biotechnology has been a branch of differential study for various sectors of society, presenting, through bioproducts and bioprocesses, improvements for the advancement and development of the Northeast region of Brazil. It is worth noting that an important means that annually brings innovative additions to the biotechnological area is the academic sector, which through stricto sensu courses at the master's and doctoral level, promote relevant research for various sectors such as economics, agribusiness, health, among others. Examples of this are: the Doctorate course in Biotechnology (RENORBIO) and the Professional Graduate Program in Biotechnology in Human and Animal Health (PPGBiotec). This article is about the path taken by the stricto sensu courses mentioned, as well as emphasizing the relevance of Biotechnology for the Northeast region of Brazil, in its different research fields, with emphasis on Natural Resources, Agriculture and Health. We exemplify biotechnologies using coconut water that has been worked since the 1980's and its evolution to the present day. Based on all the potential of the Northeast Region for the generation of bioproducts and bioprocesses, we emphasize that they will only be useful if they are really treated as technological innovation, generate invoices and have a positive impact on society's well-being.

Biotecnologias como ferramentas para o desenvolvimento do Nordeste do Brasil / Biotechnologies as tools for the development of northeast Brazil

A biotecnologia tem sido um ramo de estudo diferencial para diversos setores da sociedade, apresentando, através de bioprodutos e bioprocessos, melhorias para o avanço e desenvolvimento da região Nordeste do Brasil. Vale pontuar que um importante meio que traz anualmente acréscimos inovadores à área biotecnológica é o setor acadêmico, que através de cursos stricto sensu a nível de mestrado e doutorado, promovem pesquisas relevantes para vários setores como economia, agroindústria, saúde, dentre outros. Exemplos disso são: o curso de Doutorado em Biotecnologia da RENORBIO) e o Programa Profissional de Pós-Graduação em Biotecnologia em Saúde Humana e Animal (PPGBiotec). O presente artigo se sobre o percurso trilhado pelos cursos stricto sensu mencionados, bem como ressalta a relevância da Biotecnologia para a região Nordeste do Brasil, em seus diferentes campos de investigação, com ênfase nos Recursos Naturais, Agropecuária e Saúde. Exemplificamos as biotecnologias utilizando a água de coco que vêm sendo trabalhadas desde os anos 1980s e sua evolução até os dias atuais. Com base em toda a potencialidade da Região Nordeste para a geração de bioprodutos e bioprocessos, ressaltamos que os mesmos só serão úteis se realmente forem tratados como inovação tecnológica, gerarem nota fiscal e impactarem positivamente para o bem estar da sociedade.(AU)

Biotechnology has been a branch of differential study for various sectors of society, presenting, through bioproducts and bioprocesses, improvements for the advancement and development of the Northeast region of Brazil. It is worth noting that an important means that annually brings innovative additions to the biotechnological area is the academic sector, which through stricto sensu courses at the master's and doctoral level, promote relevant research for various sectors such as economics, agribusiness, health, among others. Examples of this are: the Doctorate course in Biotechnology (RENORBIO) and the Professional Graduate Program in Biotechnology in Human and Animal Health (PPGBiotec). This article is about the path taken by the stricto sensu courses mentioned, as well as emphasizing the relevance of Biotechnology for the Northeast region of Brazil, in its different research fields, with emphasis on Natural Resources, Agriculture and Health. We exemplify biotechnologies using coconut water that has been worked since the 1980's and its evolution to the present day. Based on all the potential of the Northeast Region for the generation of bioproducts and bioprocesses, we emphasize that they will only be useful if they are really treated as technological innovation, generate invoices and have a positive impact on society's well-being.(AU)

Aspectos econômicos e avaliação do período de viabilidade de espermatozoides ovino imunossexados e diluídos em meio de conservação à base de água de coco em pó (ACP-102c) / Economic aspects and evaluation of the viability period of immunosexed and diluted ram sperm in powdered coconut water conservation extender (ACP-102c)

Background: Sperm sexing is increasing in use because pre-determining the sex of the calf allows greater profitability and promotes significant gains in the productive systems that utilize the technique. Deployment of a low-cost and practical preservation methodol-ogy may further favor the cost-benefit ratio. Flow cytometry, the most commonly used sexing technique, has high costs and is very restricted. As an alternative, immunosexing has been studied, which uses sex-specific monoclonal antibodies. Thus, the objective of this study was to evaluate the immunosexing technique in conjunction with cryopreservation in ACP-102c and examine its economic aspects with regard to ram semen.Materials, Methods & Results: Ejaculates from two ram individuals were collected with the aid of an artificial vagina, evaluated, and submitted to the immunosexing protocol, according to the manufacturer’s recommendations, using the Monoclonal Antibody Kit specific for mammalian sperm with “Y” chromosomes (HY; HY Biotechnology, Rio de Janeiro-RJ, Brazil). After sexing, the supernatant was resuspended in the cryopreservation diluent: ACP (ACP-102c + 20% egg yolk + 7% glycerol), packaged in 0.25 mL straws, refrigerated to 4°C, stabilized for 30 min, frozen in liquid nitrogen vapors (-60°C) for 15 min, immersed in liquid nitrogen, and stored in cryogenic cylinders. The samples were thawed and evaluated for sperm kinetics both by using computerized semen analysis with SCA® software (Sperm Class Analyzer version 5.0) and subjectively comparing specimens from the two animals using conventional microscopy (40x). Plasma membrane integrity (IMP) and sperm cell morphology were evaluated by the smear staining technique...

Uso da polpa desidratada do fruto de Mauritia flexuosa como suplemento ao diluente de congelação do sêmen caprino / Use of dehydrated pulp of Mauritia flexuosa fruit as supplement to freezing extender of goat semen

Objetivou-se avaliar a qualidade in vitro do sêmen caprino descongelado utilizando diluentesuplementado com a polpa desidratada do fruto de Mauritia flexuosa. O experimento foi dividido emduas etapas. Na primeira, foram utilizados 15 pools,fracionados em 13 tratamentos com diferentesconcentrações do extrato bruto. Os melhores resultados de viabilidade espermática obtidos na primeiraetapa foram utilizadas na segunda etapa (criopreservação). Para isto, foram formados dois gruposutilizando 15 pools, sendo um diluente constituído (TRIS + 7% glicerol + melhores concentrações doextrato bruto) e outro pelo diluente (TRIS + 2,5% gema de ovo + 7% glicerol + melhores concentraçõesdo extrato bruto). Na primeira etapa os grupos contendo baixa quantidade do extrato não diferiram dogrupo controle (P≤0,05). Todavia na segunda etapa, após descongelação, os grupos TRIS contendo 2,5%ou 0% de gema de ovo apresentaram diferença significativa, onde o grupo TB06GLGE foi superior aogrupo controle. Portanto, a polpa desidratada do fruto de Mauritia flexuosa,nas concentrações de 0,25% a1%, não atuou de forma benéfica sobre parâmetros espermáticos do sêmen caprino após acriopreservação/descongelação.(AU)

The objective was to evaluate the in vitro quality of the thawed goat semen using diluentsupplemented with the dehydrated pulp of the Mauritia flexuosa fruit. The experiment was divided intotwo stages. In the first, 15 fractionated pools were used in 13 treatments with different concentrations ofthe crude extract. The best results of sperm viability obtained in the first experimental stage were used inthe second experimental stage (cryopreservation). Afterwards, two groups were formed using 15 pools,one constituent (TRIS + 7% glycerol + best concentrations of the crude extract) and another by thediluent (TRIS + 2,5% egg yolk + 7% glycerol + best concentrations of the crude extract). In the firststage, the groups containing low amount of extract did not differ from the control group (P≤0.05).However, in the second stage, after thawing, TRIS groups containing 2.5% or 0% egg yolk presented asignificant difference, where the TB06GLGE group was superior to the control group. Therefore, thedehydrated fruit pulp of Mauritia flexuosa at concentrations of 0.25% to 1% did not benefit goat semenparameters after cryopreservation/thawing.(AU)

Uso da polpa desidratada do fruto de Mauritia flexuosa como suplemento ao diluente de congelação do sêmen caprino / Use of dehydrated pulp of Mauritia flexuosa fruit as supplement to freezing extender of goat semen

Objetivou-se avaliar a qualidade in vitro do sêmen caprino descongelado utilizando diluentesuplementado com a polpa desidratada do fruto de Mauritia flexuosa. O experimento foi dividido emduas etapas. Na primeira, foram utilizados 15 pools,fracionados em 13 tratamentos com diferentesconcentrações do extrato bruto. Os melhores resultados de viabilidade espermática obtidos na primeiraetapa foram utilizadas na segunda etapa (criopreservação). Para isto, foram formados dois gruposutilizando 15 pools, sendo um diluente constituído (TRIS + 7% glicerol + melhores concentrações doextrato bruto) e outro pelo diluente (TRIS + 2,5% gema de ovo + 7% glicerol + melhores concentraçõesdo extrato bruto). Na primeira etapa os grupos contendo baixa quantidade do extrato não diferiram dogrupo controle (P≤0,05). Todavia na segunda etapa, após descongelação, os grupos TRIS contendo 2,5%ou 0% de gema de ovo apresentaram diferença significativa, onde o grupo TB06GLGE foi superior aogrupo controle. Portanto, a polpa desidratada do fruto de Mauritia flexuosa,nas concentrações de 0,25% a1%, não atuou de forma benéfica sobre parâmetros espermáticos do sêmen caprino após acriopreservação/descongelação.

The objective was to evaluate the in vitro quality of the thawed goat semen using diluentsupplemented with the dehydrated pulp of the Mauritia flexuosa fruit. The experiment was divided intotwo stages. In the first, 15 fractionated pools were used in 13 treatments with different concentrations ofthe crude extract. The best results of sperm viability obtained in the first experimental stage were used inthe second experimental stage (cryopreservation). Afterwards, two groups were formed using 15 pools,one constituent (TRIS + 7% glycerol + best concentrations of the crude extract) and another by thediluent (TRIS + 2,5% egg yolk + 7% glycerol + best concentrations of the crude extract). In the firststage, the groups containing low amount of extract did not differ from the control group (P≤0.05).However, in the second stage, after thawing, TRIS groups containing 2.5% or 0% egg yolk presented asignificant difference, where the TB06GLGE group was superior to the control group. Therefore, thedehydrated fruit pulp of Mauritia flexuosa at concentrations of 0.25% to 1% did not benefit goat semenparameters after cryopreservation/thawing.

Influência do meio de conservação à base de água de coco em pó (ACP-102c) na manutenção da atividade mitocondrial de espermatozoides ovinos criopreservados / Influence of coconut powder water-based conservation medium (APC-102c) for maintaining mitochondrial activity of cryopreserved ram sperm

Background: Semen extenders are required to protect and preserve semen, and the development of suitable extenders iskey for artificial insemination. Although the use of Tris-based diluent is widespread, new diluents such as powdered coconut water have been developed for better sperm protection. One way to evaluate the effectiveness of diluents is throughmicroscopic analyses that evaluate sperm motility, vigor, and concentration. However, these analyses are limited, and maynot provide accurate results. New evaluation techniques have been studied, and one of the tests that can be used to addreliability to these analyses is mitochondrial activity evaluation, which can sum all the parameters, and provide a moreaccurate evaluation. Thus, the present study aimed to evaluate the efficacy of ACP-102c in cryopreserved ram semen.Materials, Methods & Results: Five semen samples were collected from two ram breeders using artificial vagina (n = 10).Each ejaculate was divided into the following two treatments: T1 - ACP-102c + 20% egg yolk + 7% glycerol and T2 - TRIS+ 20% egg yolk + 7% glycerol. Extended semen samples were then packed in 0.5 mL plastic straws, subjected through therefrigeration curve up to 4°C (0.35°C/min), and equilibrated for 2 h at 4°C. Subsequently, the straws were placed at 4 cmabove liquid nitrogen level (-60°C) for 15 min, immersed, and then finally stored in the liquid nitrogen at -196°C. Bothfresh and thawed samples were evaluated for total and progressive sperm motility using conventional microscopy (40x),and the same evaluator on each occasion. For plasma membrane integrity (IMP), the smear staining technique with theEosin-Nigrosin staining was used; 200 sperms were counted and classified as whole (unstained) and unhealthy (stained).Mitochondrial activity was evaluated using a cytochemical technique based on the oxidation of 3,3’-diaminobenzidine(DAB); 200 sperms were counted, and classified into four...

Influência do meio de conservação à base de água de coco em pó (ACP-102c) na manutenção da atividade mitocondrial de espermatozoides ovinos criopreservados / Influence of coconut powder water-based conservation medium (APC-102c) for maintaining mitochondrial activity of cryopreserved ram sperm

Background: Semen extenders are required to protect and preserve semen, and the development of suitable extenders iskey for artificial insemination. Although the use of Tris-based diluent is widespread, new diluents such as powdered coconut water have been developed for better sperm protection. One way to evaluate the effectiveness of diluents is throughmicroscopic analyses that evaluate sperm motility, vigor, and concentration. However, these analyses are limited, and maynot provide accurate results. New evaluation techniques have been studied, and one of the tests that can be used to addreliability to these analyses is mitochondrial activity evaluation, which can sum all the parameters, and provide a moreaccurate evaluation. Thus, the present study aimed to evaluate the efficacy of ACP-102c in cryopreserved ram semen.Materials, Methods & Results: Five semen samples were collected from two ram breeders using artificial vagina (n = 10).Each ejaculate was divided into the following two treatments: T1 - ACP-102c + 20% egg yolk + 7% glycerol and T2 - TRIS+ 20% egg yolk + 7% glycerol. Extended semen samples were then packed in 0.5 mL plastic straws, subjected through therefrigeration curve up to 4°C (0.35°C/min), and equilibrated for 2 h at 4°C. Subsequently, the straws were placed at 4 cmabove liquid nitrogen level (-60°C) for 15 min, immersed, and then finally stored in the liquid nitrogen at -196°C. Bothfresh and thawed samples were evaluated for total and progressive sperm motility using conventional microscopy (40x),and the same evaluator on each occasion. For plasma membrane integrity (IMP), the smear staining technique with theEosin-Nigrosin staining was used; 200 sperms were counted and classified as whole (unstained) and unhealthy (stained).Mitochondrial activity was evaluated using a cytochemical technique based on the oxidation of 3,3-diaminobenzidine(DAB); 200 sperms were counted, and classified into four...(AU)

Supplemented powdered coconut water (ACP-406®) promotes growth of goat secondary follicles and oocyte meiotic resumption

The objective of this study was to test the efficiency of powdered coconut water (ACP-406®) base-medium without or with the addition of supplements on in vitro culture of isolated goat secondary follicles. Follicles were cultured for 18 days in α-MEM or in ACP-406®, both without supplements (referred to as α-MEM and ACP, respectively), or both supplemented with BSA, insulin, transferrin, selenium, glutamine, hypoxanthine, and ascorbic acid (referred to as α-MEM+ and ACP+). Follicular morphology, antrum formation, follicular and oocyte diameter, levels of glutathione (GSH), and chromatin configuration after in vitro maturation were evaluated. At the end of culture, ACP-406® base-medium (without or with supplements) showed a higher (P < 0.05) percentage of normal follicles than α-MEM (without or with supplements). Antrum formation was similar among α-MEM+, ACP and ACP+, and significantly higher than α-MEM without supplements. The follicular diameter was greater in ACP+ than α-MEM, and similar to other treatments. Moreover, fully and daily grown rates were higher (P < 0.05) in ACP-406® base-medium (without or with supplements) than α-MEM (without or with supplements). Levels of GSH were similar between ACP+ and α-MEM+ treatments. Both ACP+ and α-MEM+ allowed meiotic resumption without a significant difference between the two groups. In conclusion, supplemented ACP-406® base-medium maintained follicular survival and promoted the development as well as meiotic resumption of isolated goat secondary follicles cultured in vitro for 18 days.(AU)

Supplemented powdered coconut water (ACP-406®) promotes growth of goat secondary follicles and oocyte meiotic resumption

The objective of this study was to test the efficiency of powdered coconut water (ACP-406®) base-medium without or with the addition of supplements on in vitro culture of isolated goat secondary follicles. Follicles were cultured for 18 days in α-MEM or in ACP-406®, both without supplements (referred to as α-MEM and ACP, respectively), or both supplemented with BSA, insulin, transferrin, selenium, glutamine, hypoxanthine, and ascorbic acid (referred to as α-MEM+ and ACP+). Follicular morphology, antrum formation, follicular and oocyte diameter, levels of glutathione (GSH), and chromatin configuration after in vitro maturation were evaluated. At the end of culture, ACP-406® base-medium (without or with supplements) showed a higher (P < 0.05) percentage of normal follicles than α-MEM (without or with supplements). Antrum formation was similar among α-MEM+, ACP and ACP+, and significantly higher than α-MEM without supplements. The follicular diameter was greater in ACP+ than α-MEM, and similar to other treatments. Moreover, fully and daily grown rates were higher (P < 0.05) in ACP-406® base-medium (without or with supplements) than α-MEM (without or with supplements). Levels of GSH were similar between ACP+ and α-MEM+ treatments. Both ACP+ and α-MEM+ allowed meiotic resumption without a significant difference between the two groups. In conclusion, supplemented ACP-406® base-medium maintained follicular survival and promoted the development as well as meiotic resumption of isolated goat secondary follicles cultured in vitro for 18 days.

Congelação do sêmen de pequenos ruminantes sem uso de gema de ovo utilizando bases vegetais em substituição à gema de ovo / Small ruminants sperm freezing without egg yolk using vegetable bases instead of egg yolk

Na conservação seminal é necessária a utilização de diluentes que forneçam nutrientes e protejam as células espermáticas contra o choque térmico. Os aditivos de origem animal, como a gema de ovo e o leite, amplamente empregados na preservação do sêmen, representam um risco potencial de contaminação. Com o intuito de reduzir esses impactos quanto ao uso de substâncias de origem animal, esta revisão teve como objetivo abordar os principais pontos acerca da utilização de diluentes para congelação do sêmen de pequenos ruminantes livres de gema de ovo, um diluente constituído 100% por produtos de origem vegetal.(AU)

For semen conservation, it is necessary to use extenders that provide nutrients and protect the spermatozoa against thermal shock. Animal source additives, such as egg yolk and milk, widely used in semen preservation, represent a potential contamination risk. To reduce impacts related to the use of animal origin substances, this review aimed to address the main points about the use of extenders for small ruminant sperm freezing free of egg yolk, an extender composed by 100% vegetable origin products.(AU)

Nova biotécnica de imunosexagem de espermatozoides de ruminantes / New biotech of immunosexing of ruminants spermatozoa

A sexagem espermática é uma biotecnologia que consiste na separação de espermatozoides portadores do cromossomo “X” ou “Y”. Esta técnica tem se destacado na área de produção animal, uma vez que tem despertado um grande interesse comercial, devido à possibilidade de aumentar a rentabilidade dos empreendimentos. Assim, esta revisão teve como objetivo abordar as principais técnicas utilizadas para sexagem espermática e seus avanços em relação a aplicabilidade no sistema pecuário. Todavia, diversos aspectos relacionados à técnica de sexagem de células espermáticas ainda necessitam ser elucidados, a fim de aprimorar ou desenvolver novas tecnologias que servirão para o desenvolvimento de metodologias mais eficientes e acessíveis aos produtores.(AU)

The sperm sexing is a biotechnology that consists of the separation of spermatozoa bearing the X or Y chromosome. This technique has been outstanding in animal production, since it has aroused a great commercial interest, due to the possibility of increasing the profitability of the enterprises. Thus, this review aimed to approach the main techniques used for sperm sexing and its advances regarding applicability in the livestock system. However, several aspects related to the sexing technique of sperm cells still need to be elucidated to improve or develop new technologies that will serve to develop more efficient and accessible methodologies for producers.(AU)

Nova biotécnica de imunosexagem de espermatozoides de ruminantes / New biotech of immunosexing of ruminant’s spermatozoa

A sexagem espermática é uma biotecnologia que consiste na separação de espermatozoides portadores do cromossomo “X” ou “Y”. Esta técnica tem se destacado na área de produção animal, uma vez que tem despertado um grande interesse comercial, devido à possibilidade de aumentar a rentabilidade dos empreendimentos. Assim, esta revisão teve como objetivo abordar as principais técnicas utilizadas para sexagem espermática e seus avanços em relação a aplicabilidade no sistema pecuário. Todavia, diversos aspectos relacionados à técnica de sexagem de células espermáticas ainda necessitam ser elucidados, a fim de aprimorar ou desenvolver novas tecnologias que servirão para o desenvolvimento de metodologias mais eficientes e acessíveis aos produtores.

The sperm sexing is a biotechnology that consists of the separation of spermatozoa bearing the X or Y chromosome. This technique has been outstanding in animal production, since it has aroused a great commercial interest, due to the possibility of increasing the profitability of the enterprises. Thus, this review aimed to approach the main techniques used for sperm sexing and its advances regarding applicability in the livestock system. However, several aspects related to the sexing technique of sperm cells still need to be elucidated to improve or develop new technologies that will serve to develop more efficient and accessible methodologies for producers.

Congelação do sêmen de pequenos ruminantes sem uso de gema de ovo utilizando bases vegetais em substituição à gema de ovo / Small ruminant’s sperm freezing without egg yolk using vegetable bases instead of egg yolk

Na conservação seminal é necessária a utilização de diluentes que forneçam nutrientes e protejam as células espermáticas contra o choque térmico. Os aditivos de origem animal, como a gema de ovo e o leite, amplamente empregados na preservação do sêmen, representam um risco potencial de contaminação. Com o intuito de reduzir esses impactos quanto ao uso de substâncias de origem animal, esta revisão teve como objetivo abordar os principais pontos acerca da utilização de diluentes para congelação do sêmen de pequenos ruminantes livres de gema de ovo, um diluente constituído 100% por produtos de origem vegetal.

For semen conservation, it is necessary to use extenders that provide nutrients and protect the spermatozoa against thermal shock. Animal source additives, such as egg yolk and milk, widely used in semen preservation, represent a potential contamination risk. To reduce impacts related to the use of animal origin substances, this review aimed to address the main points about the use of extenders for small ruminant sperm freezing free of egg yolk, an extender composed by 100% vegetable origin products.

Avaliação da capacidade reprodutiva de caprinos tratados com testosterona bioidêntica por via transdérmica / Libido evaluation, serum testosterone and sperm quality goats treated with bioidentical testosterone transdermally

O objetivo deste estudo foi avaliar o efeito da aplicação de testosterona bioidêntica, por via transdérmica escrotal em caprinos, sobre o comportamento sexual, a produção quanti-qualitativa dos espermatozoides e, as concentrações de testosterona, FSH (Hormônio Folículo Estimulante) e LH (Hormônio Luteinizante), para o tratamento de reprodutores temporariamente inaptos à reprodução, uma vez que não existem dados na literatura referentes à associação do hormônio bioidêntico com a via de aplicação transdérmica em caprinos. Foram utilizados quatro machos caprinos (Saanen=03 e Anglo Nubiano=01), apresentando baixa libido e qualidade seminal insatisfatória, comprovados por andrológico. Semanalmente, ao longo de dois meses, foram realizadas colheitas e avaliação do sêmen; mensuração da circunferência escrotal (CE); aplicação da testosterona bioidêntica por via transdérmica escrotal e colheita de sangue, para determinar concentração sérica de testosterona, FSH e LH; além da verificação do comportamento sexual através do teste de libido. Para efeito de análise dos dados utilizou-se o programa estatístico SAS v.8 (2000). As concentrações séricas de testosterona apresentaram diferença significativa em função do período de tratamento, entretanto, não foi observada diferença significativa para os valores de FSH e LH. Na análise, do sêmen fresco, e resfriado, os parâmetros seminais avaliados não foram influenciados significativamente ao longo do período experimental. Conclui-se que o método de aplicação da testosterona bioidêntica por via transdérmica em caprinos, foi eficaz, considerando-se que tornou mais explícita a exibição da libido dos animais tratados, embora apesar do aumento dos níveis séricos de testosterona, não houve nenhuma interferência na qualidade seminal.

The objective of this study was to evaluate the effect of testosterone bioidentical by transdermal route of applicantion in goat scrotal, sexual behavior, quantitative and qualitative production of sperm, testosterone concentrations, FSH (Follicle Stimulating Hormone) and LH (Luteinizing Hormone) for the treatment of reproductive temporarily unable to play, since there is no data in the literature concerning association with hormone bioidêntico transdermal route of application in goats. We used four male goats (Saanen=03 and Anglo Nubian=01), with low libido and poor sperm quality, proven by andrological exam. Every week, over two months, it were taken and evalued sperm samples the scrotal circumference (SC) was mensure the bioidentical testosterone was applicated in the scrotal by transdermal route; blood samples were collected to determine serum testosterone, FSH and LH; beyond sexual behavior verification by testing libido. For purposes of data analysis used the statistical program SAS v.8 (2000). Serum testosterone showed significant differences depending on the treatment period, however, no significant difference was observed for FSH and LH between the different treatment periods. At the fresh and cooled sperm, the seminal parameters evaluated were not affected significantly throughout the experimental period. It can be concluded that the transdermal bioidentical testosterone application method in goats was effective, considering that it made the libido of treated animals more explicit, although despite the increase in serum testosterone levels, there was no interference in the seminal quality.

Avaliação da capacidade reprodutiva de caprinos tratados com testosterona bioidêntica por via transdérmica / Libido evaluation, serum testosterone and sperm quality goats treated with bioidentical testosterone transdermally

O objetivo deste estudo foi avaliar o efeito da aplicação de testosterona bioidêntica, por via transdérmica escrotal em caprinos, sobre o comportamento sexual, a produção quanti-qualitativa dos espermatozoides e, as concentrações de testosterona, FSH (Hormônio Folículo Estimulante) e LH (Hormônio Luteinizante), para o tratamento de reprodutores temporariamente inaptos à reprodução, uma vez que não existem dados na literatura referentes à associação do hormônio bioidêntico com a via de aplicação transdérmica em caprinos. Foram utilizados quatro machos caprinos (Saanen=03 e Anglo Nubiano=01), apresentando baixa libido e qualidade seminal insatisfatória, comprovados por andrológico. Semanalmente, ao longo de dois meses, foram realizadas colheitas e avaliação do sêmen; mensuração da circunferência escrotal (CE); aplicação da testosterona bioidêntica por via transdérmica escrotal e colheita de sangue, para determinar concentração sérica de testosterona, FSH e LH; além da verificação do comportamento sexual através do teste de libido. Para efeito de análise dos dados utilizou-se o programa estatístico SAS v.8 (2000). As concentrações séricas de testosterona apresentaram diferença significativa em função do período de tratamento, entretanto, não foi observada diferença significativa para os valores de FSH e LH. Na análise, do sêmen fresco, e resfriado, os parâmetros seminais avaliados não foram influenciados significativamente ao longo do período experimental. Conclui-se que o método de aplicação da testosterona bioidêntica por via transdérmica em caprinos, foi eficaz, considerando-se que tornou mais explícita a exibição da libido dos animais tratados, embora apesar do aumento dos níveis séricos de testosterona, não houve nenhuma interferência na qualidade seminal.(AU)

The objective of this study was to evaluate the effect of testosterone bioidentical by transdermal route of applicantion in goat scrotal, sexual behavior, quantitative and qualitative production of sperm, testosterone concentrations, FSH (Follicle Stimulating Hormone) and LH (Luteinizing Hormone) for the treatment of reproductive temporarily unable to play, since there is no data in the literature concerning association with hormone bioidêntico transdermal route of application in goats. We used four male goats (Saanen=03 and Anglo Nubian=01), with low libido and poor sperm quality, proven by andrological exam. Every week, over two months, it were taken and evalued sperm samples the scrotal circumference (SC) was mensure the bioidentical testosterone was applicated in the scrotal by transdermal route; blood samples were collected to determine serum testosterone, FSH and LH; beyond sexual behavior verification by testing libido. For purposes of data analysis used the statistical program SAS v.8 (2000). Serum testosterone showed significant differences depending on the treatment period, however, no significant difference was observed for FSH and LH between the different treatment periods. At the fresh and cooled sperm, the seminal parameters evaluated were not affected significantly throughout the experimental period. It can be concluded that the transdermal bioidentical testosterone application method in goats was effective, considering that it made the libido of treated animals more explicit, although despite the increase in serum testosterone levels, there was no interference in the seminal quality.(AU)

Avaliação do sêmen fresco e criopreservado de caprinos autóctones no Estado do Piauí: Resultados preliminares / Evaluation of cryopreserved sperm of autochthonous goats in the State of Piauí: Preliminary results

This work aimed to evaluate the quantitative parameters of fresh and thawed sperm of Canindé goats in theState of Piauí. Ten sperm samples were collected, using artificial vagina, from four goats at reproductive age. Eachejaculate was evaluated for volume, sperm concentration, mass motility, total and progressive motility, percentage ofmotile spermatozoa and vigor, for further dilution in ACP-101. Then, the sperm was filled in 0.25 mL straws,cryopreserved using the TK3000 machine and stored in nitrogen cylinders at -196 °C, for further analysis at SCA. Thesperm parameters were submitted to the T test for comparison between the fresh and thawed groups. The values forfresh sperm relative to total and progressive motility, vigor and sperm viability were higher (P < 0.05) whencompared to thawed, although both fresh and thawed sperm of these animals could be used in programs of artificial insemination (IA).(AU)