Efficacy and limitations of different approaches to anticipate the diagnosis of pregnancy in cattle / Eficácia e limitações de diferentes abordagens para antecipar o diagnóstico de gestação em bovinos

The study evaluated sonographic and serologic exams performed for early (20 to 30d) diagnosis of pregnancy. One hundred-twenty (n= 120) bovine recipients were synchronized (estrous=D0) and timed embryo transferred (TET, D7) with fresh in vitro produced embryos. In the first trial (n= 46), diagnosis of pregnancy was performed on day 20 (D20) by detecting CL blood flow (BF) and by Pregnancy-Associated Glycoproteins (PAGs) serology. In the second trial (n= 30), pregnancy diagnosis was performed on D25 by ultrasound visualization of uterine contents and by PAGs serology. In the last trial, PAG's serology was performed on D30. Ultrasonographic detection of the uterine contents and embryo viability performed on D30 (DG30) was considered the gold standard. The PROC FREQ procedure was used to test the agreement between diagnostic methods. On D20, the Doppler ultrasonography of the CL had showed high sensitivity (100%), but only moderate specificity (53.3%). On the same day, serologic diagnostic had no agreement (k= -0.08, P< 0.46) with the gold standard, with very low sensitivity (6.3%). However, the sensitivity of the serologic exam increased dramatically (from 6.3 to 100%) from D20 to D25, and it contributed to detect false negatives from the ultrasound diagnosis, improving the overall accuracy from 90% to 96.7%.(AU)

O estudo foi planejado para correlacionar exames ultrassonográficos e sorológicos realizados para o diagnóstico precoce (20 a 30d) de gestação. Cento e vinte (n= 120) receptoras bovinas foram sincronizadas (estro=D0), e embriões frescos produzidos in vitro foram transferidos em tempo fixo (TETF, D7). No experimento 1 (n= 46), o diagnóstico de gestação foi realizado no D20, pela detecção do fluxo sanguíneo do CL e pela sorologia de glicoproteínas associadas à gestação (PAGs). No experimento 2 (n= 30), a detecção da gestação foi realizada por meio da visualização do conteúdo do útero e também pela sorologia para PAGs. No experimento 3, a sorologia para PAGs foi realizada no D30. Em todos os experimentos, a visualização ultrassonográfica da vesícula e da viabilidade embrionária, realizada no D30, foi considerada padrão-ouro. O procedimento PROC FREQ testou o nível de concordância dos métodos diagnósticos. No D20, o diagnóstico baseado na vascularização do CL mostrou alta sensibilidade (100%) e apenas moderada especificidade (53,3%). Nesse mesmo dia, o diagnóstico sorológico não apresentou concordância (k=-0,08, P<0,46) com o padrão-ouro, além de baixa sensibilidade (6,3%). No entanto, a sensibilidade do exame sorológico aumentou drasticamente (6,3 para 100%) do D20 para o D25, contribuindo para detectar falsos negativos diagnosticados pela ultrassonografia, melhorando a acurácia (90 para 96,7%).(AU)

Efficacy and limitations of different approaches to anticipate the diagnosis of pregnancy in cattle / Eficácia e limitações de diferentes abordagens para antecipar o diagnóstico de gestação em bovinos

The study evaluated sonographic and serologic exams performed for early (20 to 30d) diagnosis of pregnancy. One hundred-twenty (n= 120) bovine recipients were synchronized (estrous=D0) and timed embryo transferred (TET, D7) with fresh in vitro produced embryos. In the first trial (n= 46), diagnosis of pregnancy was performed on day 20 (D20) by detecting CL blood flow (BF) and by Pregnancy-Associated Glycoproteins (PAGs) serology. In the second trial (n= 30), pregnancy diagnosis was performed on D25 by ultrasound visualization of uterine contents and by PAGs serology. In the last trial, PAG's serology was performed on D30. Ultrasonographic detection of the uterine contents and embryo viability performed on D30 (DG30) was considered the gold standard. The PROC FREQ procedure was used to test the agreement between diagnostic methods. On D20, the Doppler ultrasonography of the CL had showed high sensitivity (100%), but only moderate specificity (53.3%). On the same day, serologic diagnostic had no agreement (k= -0.08, P< 0.46) with the gold standard, with very low sensitivity (6.3%). However, the sensitivity of the serologic exam increased dramatically (from 6.3 to 100%) from D20 to D25, and it contributed to detect false negatives from the ultrasound diagnosis, improving the overall accuracy from 90% to 96.7%.(AU)

O estudo foi planejado para correlacionar exames ultrassonográficos e sorológicos realizados para o diagnóstico precoce (20 a 30d) de gestação. Cento e vinte (n= 120) receptoras bovinas foram sincronizadas (estro=D0), e embriões frescos produzidos in vitro foram transferidos em tempo fixo (TETF, D7). No experimento 1 (n= 46), o diagnóstico de gestação foi realizado no D20, pela detecção do fluxo sanguíneo do CL e pela sorologia de glicoproteínas associadas à gestação (PAGs). No experimento 2 (n= 30), a detecção da gestação foi realizada por meio da visualização do conteúdo do útero e também pela sorologia para PAGs. No experimento 3, a sorologia para PAGs foi realizada no D30. Em todos os experimentos, a visualização ultrassonográfica da vesícula e da viabilidade embrionária, realizada no D30, foi considerada padrão-ouro. O procedimento PROC FREQ testou o nível de concordância dos métodos diagnósticos. No D20, o diagnóstico baseado na vascularização do CL mostrou alta sensibilidade (100%) e apenas moderada especificidade (53,3%). Nesse mesmo dia, o diagnóstico sorológico não apresentou concordância (k=-0,08, P<0,46) com o padrão-ouro, além de baixa sensibilidade (6,3%). No entanto, a sensibilidade do exame sorológico aumentou drasticamente (6,3 para 100%) do D20 para o D25, contribuindo para detectar falsos negativos diagnosticados pela ultrassonografia, melhorando a acurácia (90 para 96,7%).(AU)

Freezing goat embryos at different developmental stages and quality using ethylene glycol and a slow cooling rate / Congelação de embriões caprinos em diferentes estádios de desenvolvimento e qualidade utilizando etileno glicol e taxa de resfriamento lenta

The efficiency of an alternative freezing protocol for goat embryos of different morphology and quality was tested. Fifty-eight embryos on Day 6-7 stage were transferred as fresh or after freeze-thawing (n=29/group). For freezing, embryos were placed into 1.5M ethylene-glycol solution for 10min. During this time, they were loaded in the central part of 0.25mL straw, separated by air bubble from columns containing PBS/BSA 0.4% plus 20% BFS. Straws were then frozen using a freezing machine from 20ºC to -6ºC at a cooling rate of 3ºC/min, stabilization for 15min (seeding after 5min), from -6 C to -32ºC at 0.6 C/min,and plunged into liquid nitrogen. Frozen embryos were thawed for 30s at 37ºC in a water bath. Embryos subjected to fresh transfer were maintained in holding medium (37ºC). Fresh and frozen-thawed embryos were transferred at day 7 post-estrus to 30 recipients. Kidding and kid born rates were similar (P> 0.05), respectively, for recipients receiving fresh (66.7% or 10/15; 55.2% or 16/29) or frozen-thawed (60% or 9/15; 51.7% or 15/29) embryos. The cryopreservation of goat embryos using slow-freezing protocol and 1.5MEG resulted in similar efficiency rates of fresh embryos.(AU)

Este estudo testou a eficiência de protocolo alternativo de criopreservação de embriões caprinos de diferentes qualidades morfológicas. Foram utilizados 58 embriões, coletados entre o sexto e o sétimo dia do ciclo estral (n=29/grupo). Embriões congelados passaram por solução 1,5M etilenoglicol por 10min e foram aspirados durante esse tempo para parte central de palheta 0,25mL, separada por bolhas de ar de colunas contendo PBS 0,4% BSA e 20% SFB. As palhetas foram congeladas em máquina de congelação de 20ºC a -6ºC, com taxa de resfriamento de 3ºC/min, estabilização por 15min (seeding após 5min), -6ºC a -32ºC a 0,6ºC/min, e imersas em nitrogênio líquido. Os embriões foram descongelados por 30s a 37ºC, em água. Embriões frescos foram mantidos em solução de manutenção (37ºC). Embriões frescos e congelados/descongelados foram transferidos para 30 receptoras no sétimo dia do ciclo estral. A taxa de partos e a de crias nascidas (respectivamente) foram similares (P>0,05) para receptoras recebendo embriões frescos (66,7% ou 10/15; 55,2% ou 16/29) ou congelados/descongelados (60,0% ou 9/15; 51,7% ou 15/29). O protocolo de criopreservação de embriões utilizado no presente estudo resultou em índices de eficiência semelhantes aos de embriões frescos.(AU)

Freezing goat embryos at different developmental stages and quality using ethylene glycol and a slow cooling rate / Congelação de embriões caprinos em diferentes estádios de desenvolvimento e qualidade utilizando etileno glicol e taxa de resfriamento lenta

The efficiency of an alternative freezing protocol for goat embryos of different morphology and quality was tested. Fifty-eight embryos on Day 6-7 stage were transferred as fresh or after freeze-thawing (n=29/group). For freezing, embryos were placed into 1.5M ethylene-glycol solution for 10min. During this time, they were loaded in the central part of 0.25mL straw, separated by air bubble from columns containing PBS/BSA 0.4% plus 20% BFS. Straws were then frozen using a freezing machine from 20ºC to -6ºC at a cooling rate of 3ºC/min, stabilization for 15min (seeding after 5min), from -6 C to -32ºC at 0.6 C/min,and plunged into liquid nitrogen. Frozen embryos were thawed for 30s at 37ºC in a water bath. Embryos subjected to fresh transfer were maintained in holding medium (37ºC). Fresh and frozen-thawed embryos were transferred at day 7 post-estrus to 30 recipients. Kidding and kid born rates were similar (P> 0.05), respectively, for recipients receiving fresh (66.7% or 10/15; 55.2% or 16/29) or frozen-thawed (60% or 9/15; 51.7% or 15/29) embryos. The cryopreservation of goat embryos using slow-freezing protocol and 1.5MEG resulted in similar efficiency rates of fresh embryos.(AU)

Este estudo testou a eficiência de protocolo alternativo de criopreservação de embriões caprinos de diferentes qualidades morfológicas. Foram utilizados 58 embriões, coletados entre o sexto e o sétimo dia do ciclo estral (n=29/grupo). Embriões congelados passaram por solução 1,5M etilenoglicol por 10min e foram aspirados durante esse tempo para parte central de palheta 0,25mL, separada por bolhas de ar de colunas contendo PBS 0,4% BSA e 20% SFB. As palhetas foram congeladas em máquina de congelação de 20ºC a -6ºC, com taxa de resfriamento de 3ºC/min, estabilização por 15min (seeding após 5min), -6ºC a -32ºC a 0,6ºC/min, e imersas em nitrogênio líquido. Os embriões foram descongelados por 30s a 37ºC, em água. Embriões frescos foram mantidos em solução de manutenção (37ºC). Embriões frescos e congelados/descongelados foram transferidos para 30 receptoras no sétimo dia do ciclo estral. A taxa de partos e a de crias nascidas (respectivamente) foram similares (P>0,05) para receptoras recebendo embriões frescos (66,7% ou 10/15; 55,2% ou 16/29) ou congelados/descongelados (60,0% ou 9/15; 51,7% ou 15/29). O protocolo de criopreservação de embriões utilizado no presente estudo resultou em índices de eficiência semelhantes aos de embriões frescos.(AU)

Update and overview on assisted reproductive technologies (ARTs) in Brazil

The impressive increase in the use of assisted reproductive technologies (ARTs), especially in cattle, during the last few years in Brazil is well known worldwide. In 2015, there were over 13.7 million artificial inseminations (AI), of which, about 77% were carried out using fixed-time AI (FTAI). This technology has helped to substantially improve reproductive efficiency in beef and dairy cattle. In relation to embryo transfer, production of in vivo derived (IVD) embryos remained relatively stable, with average production of 30-40,000 embryos per year, whereas in vitro production (IVP) of embryos had a substantial increase, from about 12,500 IVP embryos in 2000 to more than 300,000 IVP embryos after 2010. The increasing availability and use of sex-sorted sperm was one of the factors responsible for a recent shift from the predominance of IVP embryos from beef breeds to dairy breeds in Brazil. Moreover, there was also an increase from 13% in 2014 to 29% in 2015 in the percentage of vitrified/frozen embryos. Moreover, the successful use of protocols for fixed-time ET (FTET) due to their high efficiency and ease of implementation, has facilitated the dissemination of ET programs all over Brazil. However, there is room for improvement, since there are several reports of high pregnancy loss and high peripartum loss, when IVP embryos are used. The production of healthy cattle by somatic cell nuclear transfer has also increased in the last few years in Brazil, but despite substantial progress in reducing postnatal losses, no drastic increase in cloning efficiency up to parturition has occurred.

Update and overview on assisted reproductive technologies (ARTs) in Brazil

The impressive increase in the use of assisted reproductive technologies (ARTs), especially in cattle, during the last few years in Brazil is well known worldwide. In 2015, there were over 13.7 million artificial inseminations (AI), of which, about 77% were carried out using fixed-time AI (FTAI). This technology has helped to substantially improve reproductive efficiency in beef and dairy cattle. In relation to embryo transfer, production of in vivo derived (IVD) embryos remained relatively stable, with average production of 30-40,000 embryos per year, whereas in vitro production (IVP) of embryos had a substantial increase, from about 12,500 IVP embryos in 2000 to more than 300,000 IVP embryos after 2010. The increasing availability and use of sex-sorted sperm was one of the factors responsible for a recent shift from the predominance of IVP embryos from beef breeds to dairy breeds in Brazil. Moreover, there was also an increase from 13% in 2014 to 29% in 2015 in the percentage of vitrified/frozen embryos. Moreover, the successful use of protocols for fixed-time ET (FTET) due to their high efficiency and ease of implementation, has facilitated the dissemination of ET programs all over Brazil. However, there is room for improvement, since there are several reports of high pregnancy loss and high peripartum loss, when IVP embryos are used. The production of healthy cattle by somatic cell nuclear transfer has also increased in the last few years in Brazil, but despite substantial progress in reducing postnatal losses, no drastic increase in cloning efficiency up to parturition has occurred.(AU)

Viable offspring after successful non-surgical embryo transfer in goats / Nascimento de fetos viáveis após transferência de embriões não-cirúrgica em caprinos

O objetivo do presente estudo foi avaliar a viabilidade da técnica de transferência não cirúrgica em cabras. Quatro cabras não-lactantes pluríparas da raça Toggenburg foram utilizadas como receptoras de embriões, sendo que duas receberam um embriões e duas receberam dois embriões coletados não cirurgicamente cabras doadoras. Os corpos lúteos das receptoras foram detectados um dia antes da transferência de embriões por ultrassonografia transretal. Uma seringa de 5mL contendo 2mL de meio holding foi acoplada em um cateter tomcat, no qual os embriões foram aspirados em uma coluna central a duas outras colunas. Um espéculo Colin número 2 foi inserido na vulva e na vagina, e com o uso de uma fonte de luz, a cerviz foi localizada e imobilizada com uma pinça de Allis. Um cateter uretral número seis acoplado a um mandril e lubrificado com meio PBS foi inserido na cérvix, e assim os aneis cervicais foram gradualmente transpostos. Após perder a resistência, o cateter uretral foi movido lateralmente para o corno uterino desejado. O mandril e a pinça de Allis foram retirados e o conjunto seringa e tomcat foi acoplado ao cateter uretral e o conteúdo injetado no corno uterino ipsilateral ao corpo lúteo com posterior retirada do cateter. Cabras que ovularam em apenas um ovário foram usadas para testar a eficiência da deposição do embrião. O tempo gasto entre a inserção do espéculo e a sua remoção foi inferior a três minutos. O tempo para transpor a cérvix foi inferior a um minuto. A ultrassonografia revelou a deposição de líquido no corno desejado. Receptoras que receberam dois embriões tornaram-se gestantes e pariram três crias. Estes primeiros resultados encorajam a técnica e demonstram que a transferência de embriões em caprinos pode ser feita totalmente por procedimentos não cirúrgicos.(AU)

Florfenicol associado ou não ao cloprostenol no tratamento de retenção de placenta em vacas leiteiras / Florfenicol and sodium cloprostenol for the treatment of retained placenta in dairy cows

The aim of this study was to compare the reproductive efficiency in dairy cows with retained placenta (RP), treated with Florfenicol associated or not with Cloprostenol. One hundred and eight cows with RP from dairy herds in southern Minas Gerais state and northern São Paulo state, were included for this purpose. The experiment was designed in factorial 2X2 . The four treatments encompassed two levels of each factor : 1 ) Florfenicol - 4 0mg/kg ( IM) divided in two injections or 40mg / kg ( SC) in one injection after calving; 2) Cloprostenol (0.530mg) - two injections associated with florfenicol treatment or without cloprostenol. The analysis of discrete variables used the Kruskal - Wallis (fou r groups) or Wilcoxon (two groups) statistic to compare groups. The Student "T" test was used to compare means of continuous variables. The overall occurrence of RP was 20.7%. The florfenicol groups had no effects on postpartum uterine health or subsequent reproductive efficiency of the cows. The cloprostenol injections of RP hastened the uterine involution and decreased the occurrence of uterine infection, and increased the percentage of animals with corpus luteum between 25 and 40 days after parturition. The intervals from parturition to first artificial insemination (AI) and to conception, as well as the number of AI per conception were reduced in the cloprostenol treatment.(AU)

Three-dimensional modeling of color Doppler images: a new approach to study follicular vascularization in cattle

Ovarian blood supply is directly related to follicle developmental potential and to oocyte quality, and color Doppler ultrasonography might be a valuable tool to predict in vitro fertilization outcomes. In most studies in large domestic animals, however, the evaluation of follicle blood flow is qualitative (presence or absence of color signal) or dependent on the analysis of a single image. The objective of the present study was to first describe the use of a three-dimensional (3D) modeling of color Doppler images for a quantitative assessment of vascularization in bovine ovarian follicles. Follicular wave emergence was synchronized in Holstein and Gir heifers (n = 20), and follicular dynamics were assessed every 12 h using a color Doppler ultrasound device. The recorded cine-loop of the dominant follicle was decomposed into frames and medical image processing software was used to isolate the Doppler signal, generate the 3D model and calculate the volume of vascularization. In experiment 1, the model was validated by comparing the expected and calculated volumes and was used to predict possible variations in the results of the 2D approach. In experiment 2, vascularization was analyzed during follicular development. In both breeds, the volume of vascularization increased after follicle deviation and was positively correlated (P < 0.05) to follicular diameter (r = 0.65 and 0.54 for Holstein and Gir heifers, respectively). Spatial analysis of the three-dimensional model showed an uneven distribution of vascularization in the follicular wall, with a more intense blood flow being detected in the basal (nearest the ovarian hilus) and lateral regions of the dominant follicles. These results demonstrate the potential of this technique as a new tool for in vivo studies of ovarian physiology in large animals.

Three-dimensional modeling of color Doppler images: a new approach to study follicular vascularization in cattle

Ovarian blood supply is directly related to follicle developmental potential and to oocyte quality, and color Doppler ultrasonography might be a valuable tool to predict in vitro fertilization outcomes. In most studies in large domestic animals, however, the evaluation of follicle blood flow is qualitative (presence or absence of color signal) or dependent on the analysis of a single image. The objective of the present study was to first describe the use of a three-dimensional (3D) modeling of color Doppler images for a quantitative assessment of vascularization in bovine ovarian follicles. Follicular wave emergence was synchronized in Holstein and Gir heifers (n = 20), and follicular dynamics were assessed every 12 h using a color Doppler ultrasound device. The recorded cine-loop of the dominant follicle was decomposed into frames and medical image processing software was used to isolate the Doppler signal, generate the 3D model and calculate the volume of vascularization. In experiment 1, the model was validated by comparing the expected and calculated volumes and was used to predict possible variations in the results of the 2D approach. In experiment 2, vascularization was analyzed during follicular development. In both breeds, the volume of vascularization increased after follicle deviation and was positively correlated (P < 0.05) to follicular diameter (r = 0.65 and 0.54 for Holstein and Gir heifers, respectively). Spatial analysis of the three-dimensional model showed an uneven distribution of vascularization in the follicular wall, with a more intense blood flow being detected in the basal (nearest the ovarian hilus) and lateral regions of the dominant follicles. These results demonstrate the potential of this technique as a new tool for in vivo studies of ovarian physiology in large animals.(AU)

Doppler ultrasonography as a tool for ovarian management

Doppler ultrasonography allows the characterization and measurement of blood flow, and can be used to indirectly make inference regarding the functionality of different organs, including the ovaries. Several studies highlighted the importance of an adequate blood flow for follicle development and acquisition of ovulatory potential, and for progesterone secretion by the corpus luteum. Due to some particularities of the ovarian vascular anatomy, however, different strategies had to be developed to measure the blood flow detected by Doppler imaging. Some of these approaches were successful to characterize the patterns of blood flow throughout the estrous cycle, pregnancy, and early postpartum, but require post acquisition image processing and do not allow real time decisions to be taken. The subjective evaluation of blood flow has been alternatively used in field conditions aiming to early detect non pregnant animals, select embryo recipients, or predict artificial insemination and in vitro fertilization outcomes. In summary, color Doppler imaging provides important information about the function of corpus luteum and follicles, supporting clinical diagnoses and management decisions. Nonetheless, the adoption of Doppler ultrasonography as a routine exam, instead of a limited use in an individual basis, requires further development of feasible blood flow evaluation procedures and the establishment of reference values.

Doppler ultrasonography as a tool for ovarian management

Doppler ultrasonography allows the characterization and measurement of blood flow, and can be used to indirectly make inference regarding the functionality of different organs, including the ovaries. Several studies highlighted the importance of an adequate blood flow for follicle development and acquisition of ovulatory potential, and for progesterone secretion by the corpus luteum. Due to some particularities of the ovarian vascular anatomy, however, different strategies had to be developed to measure the blood flow detected by Doppler imaging. Some of these approaches were successful to characterize the patterns of blood flow throughout the estrous cycle, pregnancy, and early postpartum, but require post acquisition image processing and do not allow real time decisions to be taken. The subjective evaluation of blood flow has been alternatively used in field conditions aiming to early detect non pregnant animals, select embryo recipients, or predict artificial insemination and in vitro fertilization outcomes. In summary, color Doppler imaging provides important information about the function of corpus luteum and follicles, supporting clinical diagnoses and management decisions. Nonetheless, the adoption of Doppler ultrasonography as a routine exam, instead of a limited use in an individual basis, requires further development of feasible blood flow evaluation procedures and the establishment of reference values.(AU)

Features and perspectives of the Brazilian in vitro embryo industry

In the last decade , in vitro fertilization emerged as an alternative to superovulation and has become the technique of choice for bovine embryo production, especially in zebu breeds. The recent growth in the commercial use of in vitro technologies in the Brazilian embryo industry is reviewed here in , highlighting the features and trends during different periods , as well as future challenges and perspectives . The data presented here w ere provided by the Statistics Committee of the Brazilian Embryo Techno logy Society and include reports from breeders associations, commercial IVF companies and ET practitioners . Three different periods were characterized for the use of IVF technologies in the Brazilian embryo industry: 1) the early years (1999 - 2003), when IV F growth was driven by the growing demand from the embryo market, although the technology was still labeled as elit ist ; 2) a period of exponential growth (2003 - 2006), when IVF overc a me conventional ET as the technique of choice for embryo production; and 3 ) a later period , when total numbers tend ed to stabilize but IVF started to increase in importance in dairy breeds. The whole picture shows IVF as an interesting example of innovation, since the development of these new embryo technologies provided new pro ducts, processes and possibilities to satisfy demands and remarkably change the scenario of the Brazilian embryo industry.

Features and perspectives of the Brazilian in vitro embryo industry

In the last decade , in vitro fertilization emerged as an alternative to superovulation and has become the technique of choice for bovine embryo production, especially in zebu breeds. The recent growth in the commercial use of in vitro technologies in the Brazilian embryo industry is reviewed here in , highlighting the features and trends during different periods , as well as future challenges and perspectives . The data presented here w ere provided by the Statistics Committee of the Brazilian Embryo Techno logy Society and include reports from breeders associations, commercial IVF companies and ET practitioners . Three different periods were characterized for the use of IVF technologies in the Brazilian embryo industry: 1) the early years (1999 - 2003), when IV F growth was driven by the growing demand from the embryo market, although the technology was still labeled as elit ist ; 2) a period of exponential growth (2003 - 2006), when IVF overc a me conventional ET as the technique of choice for embryo production; and 3 ) a later period , when total numbers tend ed to stabilize but IVF started to increase in importance in dairy breeds. The whole picture shows IVF as an interesting example of innovation, since the development of these new embryo technologies provided new pro ducts, processes and possibilities to satisfy demands and remarkably change the scenario of the Brazilian embryo industry.(AU)

Uso de procedimentos não cirúrgicos para a produção, recuperação e inovulação de embriões em pequenos ruminantes / In vivo embryo production, recovery and transfer in small ruminants by non surgical techniques

O objetivo deste artigo foi descrever técnicas não cirúrgicas associadas à recuperação e inovulação de embriões em caprinos e ovinos. Os procedimentos relativos a estas etapas da produção in vivo de embriões destes animais foram descritos de forma sequencial e detalhada, a fim de permitir sua execução por veterinários especialistas na área.(AU)

The objective of this article was to describe non surgical techniques related to recovery and transfer of sheep and goat embryos. The main procedures for these steps in the in vivo embryo production from these animals were described sequentially and with the requested details to support their performance by veterinarian experts.(AU)

Uso de procedimentos não cirúrgicos para a produção, recuperação e inovulação de embriões em pequenos ruminantes / In vivo embryo production, recovery and transfer in small ruminants by non surgical techniques

O objetivo deste artigo foi descrever técnicas não cirúrgicas associadas à recuperação e inovulação de embriões em caprinos e ovinos. Os procedimentos relativos a estas etapas da produção in vivo de embriões destes animais foram descritos de forma sequencial e detalhada, a fim de permitir sua execução por veterinários especialistas na área.

The objective of this article was to describe non surgical techniques related to recovery and transfer of sheep and goat embryos. The main procedures for these steps in the in vivo embryo production from these animals were described sequentially and with the requested details to support their performance by veterinarian experts.

Quantificação de transcritos maternos em oócitos bovinos submetidos a diferentes condições de maturação / Quantification of maternal transcripts in bovine oocytes under different maturation systems

Comparou-se a quantidade relativa de transcritos de origem materna entre oócitos bovinos maturados in vivo e maturados em diferentes condições in vitro. Avaliou-se também o efeito dos sistemas de maturação in vitro sobre a viabilidade das células do cumulus. Para a maturação in vivo, os oócitos foram coletados 19-20h após aplicação de gonadorelina em doadoras superestimuladas com FSH e sincronizadas com implante de progesterona. Para a maturação in vitro, oócitos imaturos, obtidos de ovários coletados em matadouro, foram maturados sob diferentes tensões de oxigênio e suplementação proteica. Avaliou-se a abundância dos transcritos de Zar1, MATER e GDF9 por PCR em tempo real. A viabilidade das células do cumulus de oócitos maturados in vitro foi analisada pela coloração de Azul de Tripan. Observou-se sub-regulação (P<0,05) dos transcritos em oócitos submetidos às diferentes condições de maturação in vitro em relação aos maturados in vivo. Não houve diferença (P>0,05) na viabilidade das células do cumulus. Conclui-se que o sistema de maturação influencia a quantidade de transcritos de origem materna armazenados no citoplasma de oócitos bovinos.(AU)

The relative abundance of maternal transcripts among bovine oocytes in vivo matured or under different in vitro conditions was compared. Viability of cumulus cells of in vitro matured oocytes was also evaluated. For in vivo maturation, oocytes were recovered from 19 to 20h after gonadorelin injection in donor cows, which were previously superestimulated with FSH and synchronized with progesterone implant. For in vitro maturation, immature cumulus-oocyte complexes, obtained from ovaries collected at slaughterhouse, were matured under different oxygen tensions and protein supplementation. Relative amount of Zar1, MATER, and GDF9 transcrispts were analyzed by real time PCR. Cumulus cell viability was analyzed by trypan blue. The expression of maternal effect genes were down-regulated (P<0.05) in oocytes matured under different in vitro conditions when compared to those in vivo matured. There was no difference (P>0.05) on cumulus cell viability among different in vitro maturation conditions. In conclusion, different maturation conditions affect the relative abundance of maternal transcripts stored into oocyte cytoplasm.(AU)

Influência do grupo genético, condição sexual e tratamento antiparasitário nas medidas de área de olho do lombo e espessura de gordura in vivo e na carcaça de bovinos de corte / Influence of breed, gender condition, and anti-parasitic treatment on the measurements of in vivo rib eye area and fat thickness and on the carcass of beef cattle

Foram estudados 48 bovinos machos oriundos de inseminação artificial, criados em pasto, sendo 24 (12 Nelore e 12 F1 ½ Red Angus-Nelore ) tratados com antiparasitários alopáticos e 24 (mesmo número de puros e cruzados) tratados com o antiparasitário bioterápico Fator C&MC. Os animais foram desmamados aos oito meses, metade de cada subgrupo genético (6) foi castrado aos 13 meses e todos abatidos aos 32 meses, com o objetivo de verificar a influência do tratamento antiparasitário, do grupo genético e da condição sexual sobre as medidas de área de olho de lombo (AOL) e espessura de gordura de lombo (EGL). Usaram-se medidas de ultrassonografia no animal vivo (AOLU e EGLU) e na carcaça, plástico quadriculado e paquímetro (AOLC e EGLC). Os animais F1, os inteiros e os tratados com alopatia apresentaram peso vivo maior quando comparados aos Nelores, castrados e tratados com bioterápicos. Não houve diferença da AOLU e AOLC entre os grupos genéticos. EGLC foi mais alta nos cruzados. Os animais inteiros apresentaram AOLU e AOLC maiores que os castrados, e EGLU e EGLC menores. Foram altamente significativos os coeficientes de correlação entre as medidas por ultrassom e na carcaça para área de olho de lombo (0,87) e espessura de gordura do lombo (0,95).(AU)

Forty-eight male bovines, products of artificial insemination and raised on pasture, were studied, being 24 (12 Nelore, 12 F1 ½ Nelore ½ Red Angus) treated with allopathic antiparasitic drugs and 24 (same number of pure and crossbred) treated with a biotherapic antiparasitic drug Factor C&MC. Animals were weaned at eight months of age and half of each genetic subgroup (six) was castrated at 13 months of age. All animals were slaughtered at 32 months of age, in an attempt to evaluate the influence of antiparasitic treatment, genetic group, and gender condition in the measurements of rib eye area (AOL) and fat thickness (EGL) of loin. Measurements of ultrasonography were used for live animals (AOLU and EGLU), whereas a direct plastic grid reading of the eye muscle and a pachymeter (AOLC and EGLC) were used for carcasses. F1 animals, non-castrated, and those treated with allopathic drugs showed higher live weight when compared with Nelore, castrated, and biotherapic treated animals. There were no differences between genetic groups of AOLU and AOLC. EGLC was higher in crossbred animals. Non-castrated animals showed higher AOLU and AOLC when compared with castrated animals, and lower EGLU and EGLC. Correlation coefficients for ultrasound and carcass measurements were highly significant for rib eye area (0.87) and fat thickness (0.95).(AU)

Efeito do citrato e taurina em meio CR2aa no desenvolvimento de embriões bovinos fecundados in vitro / Effect of citrate and taurine added to CR2aa medium on the development of in vitro-fertilized bovine embryos

Avaliou-se o efeito do citrato em meio CR2aa suplementado com soro fetal bovino (SFB) ou livre de proteínas séricas e sua associação com taurina no desenvolvimento de embriões bovinos fecundados in vitro. Embriões foram cultivados em CR2aa contendo 0, 0,5, 1,0 e 3,0mM citrato, suplementado com 10 por cento SFB (experimento 1) ou com álcool polivinil (PVA; experimento 2). No terceiro experimento, embriões foram cultivados em meio com 0,5mM citrato, ou 7mM taurina, ou com a associação de ambos, suplementado com SFB. Os cultivos foram realizados com células do cumulus em ambiente a 38,8ºC com 5 por cento de CO2 em ar atmosférico. Melhora no desenvolvimento embrionário foi observado no cultivo de embriões em CR2aa com 0,5 e 1,0mM citrato na ausência de SFB (P<0,05), 8,6 por cento e 11,3 por cento de blastocistos, respectivamente, porém com valores mais baixos (P<0,05) que embriões cultivados em CR2aa com SFB (31,9 por cento). Associação de citrato com taurina em meio com SFB não influenciou (P>0,05) a produção de embriões ou o número de células. Citrato em meio CR2aa pode ser uma alternativa para cultivo embrionário em condições atmosféricas com 5 por cento de CO2 em ar na ausência de proteína sérica.(AU)

The effect of citrate added to CR2aa medium supplemented with fetal calf serum (FCS) or serum-proteinfree and its association with taurine on the development of in vitro-fertilized bovine embryos was evaluated. Embryos were cultured with 0, 0.5, 1.0, and 3.0mM citrate, in CR2aa supplemented with 10 percent FCS (experiment 1), or polyvinyl alcohol (PVA; experiment 2). In experiment 3, embryos were cultured with 0.5mM citrate, 7.0mM taurine or with association of both, in medium supplemented with FCS. Embryo culture was performed with cumulus cells at 38.8ºC in 5 percent CO2 under air for all experiments. Positive effect on embryo development was only observed with 0.5 and 1.0mM citrate in FCS-free CR2aa (P<0.05; 8.6 percent and 11.3 percent blastocyst, respectively), however with lower embryo rate than CR2aa with FCS (31.9 percent). Association between citrate and taurine in medium supplemented with FCS did not affect (P>0.05) embryo rate nor total cell number. Citrate in CR2aa medium can be an alternative for serumfree embryo culture under 5 percent CO2 in air, absence of serum protein.(AU)

Parâmetros reprodutivos de cabras Toggenburg inseminadas com sêmen resfriado, após diluição em meio à base de gema de ovo / Reproductive parameters of Toggenburg goats inseminated with cooled semen diluted in egg yolk extender

Avaliaram-se a taxa de concepção, a resposta à prostaglandina, a duração do estro, a categoria reprodutiva e o tipo de muco de cabras inseminadas com sêmen diluído em meio à base de gema de ovo e resfriado a 5ºC, por 12 ou 24 horas. Foram utilizados dois reprodutores e 62 fêmeas da raça Toggenburg, que receberam duas doses de 22,5µg de PGF2α, em intervalos de 10 dias, para a sincronização do estro. A partir da primeira aplicação de PGF2α, o estro foi monitorado três vezes ao dia. Realizou-se uma única inseminação, 12 horas após o início do estro. As porcentagens de fêmeas em estro, após a primeira e segunda aplicações de PGF2α, foram de 85,5 por cento e 88,7 por cento, respectivamente. O intervalo de aplicação da primeira e segunda doses de PGF2α ao início do estro foi de 41,04±20,32 e 45,67±9,28 horas, e a duração do estro de 40,02±15,96 e 32,24±12,09 horas, respectivamente. A taxa de concepção total foi de 49,1 por cento. O período de armazenamento do sêmen e a categoria reprodutiva não influenciaram (P>0,05) a taxa de concepção. O tipo de muco observado no momento da inseminação influenciou (P<0,05) a fertilidade das fêmeas, sendo o de aspecto estriado associado aos maiores índices de concepção.(AU)

The conception rate, the prostaglandin response, the estrus duration, the reproductive class, and the mucous of goats inseminated with semen diluted in egg yolk extender and cooled at 5ºC, for 12 or 24 hours were evaluated. Sixty-two female goats and two sexually mature Toggenburg bucks were used. The females received two doses of 22.5µg of prostaglandine F2α, at 10-day intervals. After the first injection, the estrus was monitored three times a day (6:00, 12:00, and 18:00h), with a buck teaser. Only one insemination was used. The percentages of animals that showed estrus after the first and the second injection of PGF2α were 85.5 percent and 88.7 percent, respectively. The average intervals from first and second PGF2α injection to estrus were 41.04±20.32 and 45.67±9.28h, and the estrus durations for both injections were 40.02±15.96 and 32.24±12.09h, in this order. The interval from the PGF2α injection to the beginning of the estrus was longer (P<0.05), whereas the estrus duration was shorter (P<0.05) in the estrus induced after the second PGF2α injection. The total conception rate was 49.1 percent. Neither the semen storage length (12 or 24 hours) nor the reproductive class did not affect (P>0.05) the conception rate. The mucous observed at the insemination time influenced (P<0.05) the fertility of inseminated goats, with the striated aspect associated to higher fertility.(AU)