Biotecnologias aplicadas à reprodução de ovinos e caprinos.

A criação de caprinos e ovinos é uma atividade agropecuária de elevada importância social, econômica e consolidada em determinadas regiões do Brasil, enquanto em outras, mostra-se em fase emergente. Diversos aspectos apontam para o desenvolvimento desta cadeia produtiva no país. Dentre eles, a produção nacional, especialmente de carne e leite, sequer abastece o mercado interno, mesmo considerando que o consumo ainda é reduzido em relação a outros países. Em função da qualidade e diferencial dos produtos de origem caprina e ovina, acredita-se que em pouco tempo a demanda aumentará expressivamente, requerendo sistemas de criação focados na produtividade. Assim, torna-se fundamental desenvolver e adequar tecnologias voltadas ao incremento produtivo. Para tanto, recursos humanos, meio ambiente, nutrição, sanidade, bem-estar animal, genética e reprodução devem estar e operar concomitantemente. Dentre eles, a eficiência reprodutiva desempenha papel de destaque. A reprodução animal é uma área de intensos estudos, avanços e que vem sendo cada vez mais explorada em seu aspecto aplicado. Neste cenário, as biotécnicas oferecem benefícios singulares ao incremento reprodutivo. Todavia, o sucesso apenas é alcançado quando são empregadas diante do conhecimento holístico da área. Esta obra poderá auxiliar na consolidação e domínio de estudantes, profissionais e técnicos acerca das biotecnologias aplicadas à reprodução em ovinos e caprinos. O material apresenta inicialmente informações de aspectos fisiológicos que possibilitam ao leitor o entendimento das biotécnicas aplicadas detalhadamente abordadas em sequência. São apresentados os métodos de sincronização e indução de estro e ovulação; inseminação artificial; múltipla ovulação e transferência de embriões; e tecnologia reprodutivas avançadas como a produção in vitro de embriões, transgênese e clonagem.bitstream/item/133578/1/Livro-Biotecnologias-aplicadas-completo.pd

Superovulation and transcervical embryo recovery in Lacaune ewes raised under tropical conditions.

This study assessed two superovulatory treatments and the feasibility of transcervical embryo recovery in Lacaune ewes. Ewes (n=23) received medroxyprogesterone acetate sponges (60mg, Progespon®, Syntex, (Prolise®, Tecnopec, São Paulo, Brazil) 24 h before sponge removal and were superovulated with 133 mg of porcine FSH i.m. (Folltropin®-V; Bioniche Animal Health, Belleville, Canada) in six decreasing doses (twice daily) at 60 h before sponge removal, under a crossover design. In conclusion, both treatments showed high variability in ovulatory response which might reduce the embryo yield average from donors. The protocol for cervical relaxation allowed the transcervical embryo recovery in high percentage of Lacaune ewes

Occurrence of hydrometra in dairy goats after estrous induction by either light program or hormonal protocol.

This study aimed to report the occurrence of hydrometra in different herds using multi-hormonal protocol (Herd 1) or light program (Herd 2). Saanen does, with ages ranging from eight months to nine years old from two different herds, both located in the Southeast region of Brazil, were used. Although hydrometra was significantly associated with estrous induction in Herd 1, this disease is more like to be associated with the fact of parturition during the onset of natural breeding season rather than estrous induction method. [Ocorrência de hidrometra em cabras leiteiras após indução de estro por programa de luz ou protocolo hormonal].Edição dos anais do XXII Congresso Brasileiro de Reprodução Animal (CBRA), Santos, SP, Brasil, maio 2017

Transcervical embryo recovery in Lacaune ewes superovulated with different doses of FSH.

This study assessed the effect of different FSH dosages for superovulation and the feasibility of transcervical embryo recovery in Lacaune ewes. Ewes (n = 2 5) received 60 mg medroxiprogesterone acetate sponge (Progespon®, Syntex, Buenos Aires, Argentina) for nine days, 37.5 μg d-cloprostenol i.m. (Prolise®, Tecnopec, São Paulo, Brazil) 24 h before sponge removal and 50 μg gonadorelin (GnRH analogue, Gestran®, Tecnopec, São Paulo, Brazil) 24 h after sponge removal. The protocol for cervical relaxation was efficient to allow the transcervical embryo recovery of Lacaune ewes

Viability of transcervical embryo transfer in goats.

Embryo transfer consists in the deposition of the embryo in the recipient uterus. The results of recipient?s fertility vary widely depending on the origin of these embryos: fresh or cryopreserved and produced in vivo or in vitro. Typically, pregnancy rates ranging from 40 to 80% are reported. The objective of this study was to test the feasibility of non-surgical embryo transfer of goat fresh embryos.Proceedings of the 29th Annual Meeting of the Brazilian Embryo Technology Society (SBTE); Gramado, RS, Brazil, August 20th to 23rd, 2015, and 31st Meeting of the European Embryo Transfer Association (AETE); Ghent, Belgium, September 11th and 12th, 2015. Abstract

The influence of different methods of frozen-thawed ovine spermatozoa selection on sperm capacitation and viability after incubation.

Abstract: Sperm capacitation is an essential event for fertilization; however, it decreases the sperm lifespan and viability. The aim of this study was to evaluate the effects of four sperm selection techniques on sperm capacitation and viability after incubation. A pool of frozen-thawed sperm from 10 Santa Inês rams was used. The samples were submitted to one of the following sperm selection techniques: sperm washing, Percoll gradient, mini-Percoll gradient, Swim-up and control group. At mini-Percoll technique, was used 400 microliters of 90% and 45% gradients and a centrifugation at 500 xg for 5 minutes. In Percoll, was used 1 mL of each gradient and a centrifugation at 700 xg for 10 minutos. During Swim-up, the sperm was incubated in 1 ml of SPERM-TALP for 45 minutos in humidified atmosphere at 37.5oC. Finaly, at sperm washing the sample suffered centrifugation at 300 xg for 8 minutes, using SPERM-TALP. At the end of each treatment, the selected spermatozoa were incubated at 37oC for 1 h, 2 h, and 3 h. Viability was assessed using acridine orange-propidium iodide combination by computer-assisted sperm analysis. Capacitation status was evaluated using chlortetracycline staining and observed under epifluorescence microscopy. Data were analyzed by ANOVA, followed by Tukey test (P0.05) among Percoll (36%), mini-Percoll (34%) and Swim-up (30%), and were lower (P0,05) entre Percoll (36%), mini-Percoll (34%) e Swim-up (30%), as quais foram inferiores (P<0,05) ao grupo controle (47%) e lavagem por centrifugação (41%). A taxa de espermatozoides não capacitados foi superior (P<0,05) no momento 0 h (12%) e diminuiu após 3 h (1,5%), independentemente do método. Houve interação (P<0,05) entre intervalo de incubação e método na taxa de espermatozoides reagidos. O índice de reagidos foi menor (P<0,05) em 0 h (50%) e 1 h (53%) e maior após 3 h (64%). Percoll (60%) e mini-Percoll (61%) apresentaram maiores valores para reagidos (P<0,05) enquanto o controle apresentou o menor (49%). Observou-se interação (P<0,05) entre intervalo de incubação e método na vitalidade espermática. A taxa de íntegros em 0 h foi a maior (17,5%; P<0,05), após os diferentes métodos. O Swim-up apresentou maior taxa de íntegros (17,4%; P<0,05), independentemente do intervalo de incubação. Em conclusão, o intervalo de incubação interfere nos padrões de capacitação e vitalidade de espermatozoides ovinos congelados. A seleção espermática aumenta a taxa de células reagidas e o Swim-up permite maior vitalidade durante a incubação.Proceedings of the 29th Annual Meeting of the Brazilian Embryo Technology Society (SBTE); Gramado, RS, Brazil, August 20 to 23, 2015. Abstracts

Goat incubator: can bovine oocytes be matured in the uterine horn of a goat?

Abstract We used a goat as a live incubator, along with associated nonsurgical embryo transfer techniques, to perform ex situ (in vivo) maturation of bovine oocytes. Immature bovine cumulus-oocyte complexes (COCs) aspirated from 3-8 mm follicles from slaughterhouse ovaries were randomly split into two groups for in vitro (IVM; n = 38) and ex situ maturation (ESM; n = 40). IVM was performed for a period of 24 h at 38.5 ºC and with 5% CO2 in the air of maximum humidity. For ESM, a presynchronized nulliparous goat (12 months old) received 40 immature COCs in the uterine horn apiece, via the transcervical route. After 24 h the structures were retrieved through uterine flushing. Analyses of nuclear maturation and lipid quantification were performed on oocytes from both groups. Fluorescent intensity was compared using the Student?s t-test. Forty-seven percent of the structures were recovered after uterine flushing (19/40). The nuclear maturation rate was 94.5% (18/19) and 81.6% (31/38) for the ESM and IVM groups, respectively. In vitro-matured COCs contained more lipid droplets, expressed as a higher amount (p < 0.05) of emitted fluorescent light than ex situ-matured COCs (858 ± 73 vs. 550 ± 64 arbitrary fluorescence units, respectively). This is the first report to associate nonsurgical embryo transfer techniques and a goat as a live incubator for the maturation of bovine oocytes. We conclude that bovine oocytes can progress meiotically in the uterus horn of a goat and that transcervical transfer of bovine oocytes to a goat?s uterus could present an alternative to nuclear maturation

GnRH potential to synchronize follicular emergence and ovulation prior to superovulatory day 0 protocol in sheep.

The role of GnRH to synchronize ovulation and follicular emergence previous to superovulatory protocol, started on the first day of the estrous cycle (Day 0), was assessed in Santa Inês ewes. For estrus synchronization, 60 mg medroxyprogesterone acetate sponges were used for 6 d plus 37.5 ?g d-cloprostenol and 300 IU eCG at fifth day. After sponge removal, ewes were assigned to three treatments: GControl ? saline at 12 h (n = 10); G24h ? GnRH at 24 h (n = 10); or G36h ? GnRH at 36 h (n = 9). In conclusion, considering the beneficial effects of G36h in synchronizing ovulation and to promote the absence of dominant follicles in the first days of estrous cycle. According to data obtained, the best time to start the superovulatory treatment, known as "Day 0", could be 80 h after sponge removal (56 h for the occurrence of ovulation plus 24 h to reset dominant follicles), in the induction of synchronized estrus, for Santa Inês ewes.Proceedings of the 29th Annual Meeting of the Brazilian Embryo Technology Society (SBTE); Gramado, RS, Brazil, August 20th to 23rd, 2015, and 31st Meeting of the European Embryo Transfer Association (AETE); Ghent, Belgium, September 11th and 12th, 2015

Synchronization of estrus with two doses of cloprostenol at different intervals and Artificial Insemination in dairy goats.

The present study evaluated the efficacy of two protocols for estrus synchronization with different intervals in order to use them in artificial insemination (AI) in Saanen and Alpine multiparous goats in June (end of the natural mating season). The animals were allocated into two groups, according to breeding, parity, and body condition score to receive two doses of 37.5 ?g d-cloprostenol (Prolise®, ARSA SRL, Buenos Aires, Argentina) latero-vulvar 7.5 (n=23) and 11.5 (n=25) days. No animal became pregnant after NM. In AI animals, conception rate was 69.2% (9/13) for T7.5 and 70.6% (12/17) for T11.5 (P> 0.05). Even at the end of the breeding season, the protocols were able to synchronize a significant percentage of animals in a short window of time, resulting in a high conception rate after AI

Goat incubator: the doe as a life incubator of bovine oocytes - first step.

Despite significant improvements in the in vitro production of cattle embryos, the suboptimal in vitro culture environment still limits the embryo quality and production. Techniques that associate the advantages of in vivo and in vitro systems, such as intrafollicular transfer of immature oocytes, have been proposed mainly to increase the embryo quality. In this context, we tried to use a goat as live incubator and associated nonsurgical embryo transfer techniques in small ruminants to perform ex situ (in vivo) maturation of bovine oocytes. For this, immature bovine cumulus-oocyte complexes (COCs) of grade 1 and 2 were randomly distributed into two groups for in vitro (IVM; n = 38) and ex situ (ESM; n = 40) maturation. The IVM was performed for a period of 24 h in TCM-199 medium (Gibco Life Technologies, Inc., Grand Island, NY, USA) supplemented with 20 mg/mL of FSH (Pluset, Calier, Barcelona, Spain), 0.36 mM sodium pyruvate (Sigma Chemical, St. Louis, MO, USA), 10 mM sodium bicarbonate (Sigma Chemical, St. Louis, MO, USA) and 50 mg/mL streptomycin/penicillin (Sigma Chemical, St. Louis, MO, USA) at 38.8 ºC in an atmosphere of 5% CO2 in air with maximum humidity. For ESM, a pre-synchronized nulliparous goat (12 months old) received 40 immature COCs in the uterine horn apice by transcervical route (Fonseca et al., 2014 Arq. Bras. Med.vet. Zootec) and 24 h after the procedure the structures were retrieved by the uterine flushing (Fonseca et al., 2013 Small Rumin Res). For analysis of the nuclear maturation rate and lipid quantification, the oocytes were denuded (0.1% hyaluronidase), fixed (4% paraformaldehyde) and stained with 10 ?g/mL Hoechst 33342 and 10 ?g/mL Nile Red (Molecular Probes, Inc., Eugene, OR, USA) dissolved in physiological saline (0.9% NaCl) with 1mg/mL polyvinylpyrrolidone. Oocytes displaying metaphase II plate were considered matured. The lipid amount was inferred by measuring the fluorescence intensity using the ImageJ program and fluorescence intensity were compared by Student's t-test. Forty-seven percent of the structures were recovered after uterine flushing (19/40). The nuclear maturation rate was 94.5% (18/19) and 81.6% (31/38) for ESM and IVM groups, respectively. In vitro-matured oocytes contained more lipid droplets, expressed as a higher (p < 0.05) amount of emitted fluorescence light (858 ± 73 arbitrary fluorescence units) than ex situ-matured oocytes (550 ± 64 arbitrary fluorescence units). This is the first report associating nonsurgical embryo transfer techniques with goat as live incubator for maturation of bovine oocytes. We conclude that transcervical transfer of bovine oocytes to uterine goat may be an alternative to in vitro maturation aiming the reduction of lipids without compromising nuclear maturation. Further studies are required to improve the oocyte recovery rate.Proceedings of the 31st Annual Meeting of the Brazilian Embryo Technology Society (SBTE); Cabo de Santo Agostinho, PE, Brazil, August 17th to 19th, 2017. Abstracts