Demi-embryo reconstitution, a factor to consider for the success of embryo bisection. Review

Durante muchos años se ha intentado incrementar la eficiencia reproductiva del ganado utilizando biotecnologías como la bisección de embriones. Sin embargo, pese a su potencial en el ganado, su nivel de adopción es limitado. Este trabajo reseña la importancia de la reconstitución de los demi-embriones después de la bisección y los principales factores que limitan su éxito en el ganado. El uso de esta técnica podría ser más generalizado si se incrementara su eficiencia, lo cual podría lograrse mediante una selección más precisa de los embriones que se someterán a bisección. La calidad de los embriones es uno de los factores más importantes que determinan su potencial de reconstituirse en demi-embriones viables después de la bisección, permitiendo obtener resultados más confiables en los programas de transferencia de embriones.For many years it has been sought to increase the reproductive efficiency of livestock using biotechnologies such as embryo bisection. However, despite its potential in livestock, its level of adoption is limited. The present work reviews the importance of demi-embryo reconstitution, after bisection, and the main factors that limit its success in livestock. It is possible to increase its level of adoption if it is possible to increase the efficiency currently obtained with this technique, this can be achieved by making a more precise selection of the embryos subjected to bisection. Embryo quality is one of the most important factors related to the potential to reconstitute into viable demi-embryos after bisection, which can be used with greater reliability in embryo transfer programs

Compromising between European and US allergen immunotherapy schools: Discussions from GUIMIT, the Mexican immunotherapy guidelines

Background: Allergen immunotherapy (AIT) has a longstanding history and still remains the only disease-changing treatment for allergic rhinitis and asthma. Over the years 2 different schools have developed their strategies: the United States (US) and the European. Allergen extracts available in these regions are adapted to local practice. In other parts of the world, extracts from both regions and local ones are commercialized, as in Mexico. Here, local experts developed a national AIT guideline (GUIMIT 2019) searching for compromises between both schools. Methods: Using ADAPTE methodology for transculturizing guidelines and AGREE-II for evaluating guideline quality, GUIMIT selected 3 high-quality Main Reference Guidelines (MRGs): the European Academy of Allergy, Asthma and Immunology (EAACI) guideines, the S2k guideline of various German-speaking medical societies (2014), and the US Practice Parameters on Allergen Immunotherapy 2011. We formulated clinical questions and based responses on the fused evidence available in the MRGs, combined with local possibilities, patient's preference, and costs. We came across several issues on which the MRGs disagreed. These are presented here along with arguments of GUIMIT members to resolve them. GUIMIT (for a complete English version, see Supplementary data) concluded the following: Results: Related to the diagnosis of IgE-mediated respiratory allergy, apart from skin prick testing complementary tests (challenges, in vitro testing and molecular such as species-specific allergens) might be useful in selected cases to inform AIT composition. AIT is indicated in allergic rhinitis and suggested in allergic asthma (once controlled) and IgE-mediated atopic dermatitis. Concerning the correct subcutaneous AIT dose for compounding vials according to the US school: dosing tables and formula are given; up to 4 non-related allergens can be mixed, refraining from mixing high with low protease extracts. When using European extracts: the manufacturer's indications should be followed; in multi-allergic patients 2 simultaneous injections can be given (100% consensus); mixing is discouraged. In Mexico only allergoid tablets are available; based on doses used in all sublingual immunotherapy (SLIT) publications referenced in MRGs, GUIMIT suggests a probable effective dose related to subcutaneous immunotherapy (SCIT) might be: 50–200% of the monthly SCIT dose given daily, maximum mixing 4 allergens. Also, a table with practical suggestions on non-evidence-existing issues, developed with a simplified Delphi method, is added. Finally, dissemination and implementation of guidelines is briefly discussed, explaining how we used online tools for this in Mexico. Conclusions: Countries where European and American AIT extracts are available should adjust AIT according to which school is followed

Investigations into procedures for the implementation of a multiple ovulation and embryo transfer scheme using ewe lambs: a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Animal Science at Massey University

Three trials using 6-7 month-old Romney ewe lambs and adult ewes were conducted to evaluate the feasibility of implementing a juvenile MOET scheme. Older lambs were used to evaluate ways of improving the ovulation rates in 3 additional trials, one of them involved Booroola-cross lambs. Studies were also carried out to develop and evaluate a technique for splitting sheep embryos, and these were transferred to recipient ewes. In the three trials involving superovulation and transfer a range of gonadotrophin treatments were used. The more highly purified preparations (FSH-P, Ovagen) gave lower responses compared with PMSG (1.78, 1.06 vs 4.18 respectively in Trial 3, P<0.01). Ovulation rate of ewe lambs relative to adult ewes in the 3 trials were respectively 1.20 vs 2.00 (P<0.01), 4.18 vs 6.35 and 1.61 vs 3.15. Overall, administration of GnRH did not significantly increase the ovulation rates, although lambs treated with PMSG + GnRH tended to give higher responses. Egg recovery was higher in most cases in adults than in lambs (91% vs 55%, P<0.01; 72% vs 47%. P<0.01; 34% vs 43%, for trials 1, 2 and 3 respectively). Fertilization rate in trial 1 was lower in lambs than in adults (50% vs 82%, P<0.05) but not different in trials 3 and 5, probably due to the use of intrauterine insemination. Ovulation rate was not improved by treating lambs at 8-9 months of age or by administering PMSG 2 or 4 days before sponge removal. Ovulation rates following gonadotrophin stimulation were higher in androstenedione-immunized ewe lambs than in non-immunized ewe lambs (2.22 vs 1.59. P<0.05). Booroola-cross lambs treated at 6-7 months of age gave considerably higher ovulation rates compared to those recorded in Romney lambs from another experiment conducted at the same time (3.79 vs 1.61). Lower embryo survival and incidence of twins were found for demi-embryos generated from 9-10 month-old lambs compared to those obtained from 24 months or older animals. Higher pregnancy rate and demi-embryo survival were found when bisecting blastocysts rather than morulae harvested in the same flush. A deterministic model incorporating parameters provided by the study showed that the limited number of lambs generated from lamb donors restricted the improvement in genetic progress

Studies on embryo transplantation procedures using Booroola-Merino x Perendale ewes : a thesis presented in partial fulfilment of the requirements for the degree of Master of Agricultural Science at Massey University

A study was carried out to develop an embryo transfer programme under farm conditions. For this embryos were collected from donor ewes 5-6 days after single-sire mating by rams of the same crossbreed as the ewes. Then the embryos were transplanted soon after collection or after they had been initially frozen and later on thawed before their transfer. To enable this study to be carried out, one hundred and sixty-one Booroola-Merino x Perendale ewes aged between 3 and 6 years and of 34-69 kg live weight were examined. These animals were classified as either carriers (F+) or non-carriers (++) of the fecundity gene on the basis of the number of ovulations recorded at laparoscopy carried out a few days after progestagen sponge treatment to induce oestrus synchronisation, and in a few cases on the basis of previous lambing data. The aims of the study therefore were also to identify ewe carriers or non-carriers of the fecundity gene, and to evaluate their ovulatory response to several doses of Pregnant Mare Serum Gonadotrophin (Tl=O, T2=350, T3=700 and T4=1050 i.u. of PMSG) given during the embryo transfer programme. On the basis of the criteria used to classify F+ and ++ ewes, 76 ewes were considered as carriers and 85 as non-carriers of the fecundity gene. Immediately after progestagen synchronisation treatment the incidence of oestrus and the distribution of onset of oestrus was similar in both groups of ewes, and also following PMSG treatment given in the succeeding cycle. Ovulation rate after the progestagen sponge treatment was significantly affected by genotype and age of the ewes, but it was not associated with the live weight of the ewes. The least-squares means for number of ovulations in F+ and ++ ewes and in 3 and 4-6 years old animals were 3.02, 1.73, 2.14 and 2.45 respectively. Higher sensitivity to PMSG treatment was observed from the right ovary than from the left ovary (3.40 v. 2.82 ovulations respectively). No significant effect of dose of PMSG was found on the percentage of embryos recovered, the overall recovery rate being 66%. Of the ova or potential embryos recovered 78% had been fertilised and had developed to embryos. There were no significant differences between the 3 rams in the fertilisation rate in the ewes compared on a per ewe basis. The reproductive performance of donor ewes which had been flushed, was considerably influenced by the efficiency of the embryo recovery procedures, since every embryo not recovered represents a potential pregnancy and such a situation sometimes is not desirable. In this study 30% of the ewes that were flushed, subsequently became pregnant and produce lambs as a result of one or more embryos not being recovered at surgery. The number of ovulations after PMSG treatment was significantly affected by the dose-level of PMSG and the genotype, age and live weight of the ewes. F+ ewes recorded a significantly higher ovulation rate than ++ ewes. Their respective least-squares means were 3.61 and 2.31 corpora lutea. No significant difference was found between treatments 1 and 2 and between treatments 3 and 4. However, the response from the last two treatments was significantly higher than that from the first two treatments. Their respective least-squares means were 2.10, 2.01, 3.74 and 4.37 corpora lutea. Ewes 4-6 years old recorded a significantly higher number of ovulations than younger ewes (3 years old). Small significant effect of live weight was found on the ovulatory response of the ewes. Analysis within each genotype showed a similar trend in both genotypes, but small significant effect of weight was only detected in the ++ ewes. The pregnancy rate that occurred after the transfer of two fresh embryos (86%) was significantly affected by the genotype of the recipient ewe (F+ ewes 75% v. ++ ewes 96%). The number of ovulations in the recipients and whether or not they had received PMSG before transfer had no effect on the incidence of pregnancy. The pregnancy rate achieved after the transfer of frozen embryos (35%) was significantly influenced by the number of embryos transferred, but not by the ovulation rate in the recipient ewe or the time elapsing from flushing until freezing. Pregnancy rate was significantly higher after the transfer of 2 embryos compared to the transfer of single embryos (43% v. 0%), but only 8 transfers were made in the later category. On the basis of the number of ovulations, it can be concluded that ewe carriers of the fecundity gene recorded significantly higher ovulation rate after oestrus synchronisation and were more sensitive to PMSG stimulation than non-carrier ewes. There was an encouraging pregnancy rate obtained with frozen embryos, even although the small number of transfers carried out limits conclusions that can be drawn. However, the eighty-six percent pregnancy rate achieved after the transfer of two fresh embryos per recipient, shows the feasibility of the embryo transfer programmme under conditions where suitable recipients are available. Where recipients are limited then additional embryos might be frozen and stored until ready for transplantation

Superovulatory response and embryo quality of Holstein heifers treated with one or two injections of somatotropin

Background: recombinant bovine somatotropin (rbST) is used in bovine embryo donors to improve superovulatory response and embryo quality. Objective: this study evaluated the effect of applying one versus two injections of 500 mg of rbST to donor Holstein heifers on estrus incidence (IE), diameter of the largest preovulatory follicle (dLPF), superovulation response (SR), embryo yield, and pregnancy rate in recipient Holstein cows (PRR). Methods: two superstimulation programs were conducted. Heifers were assigned to one of two treatments: 1) rbST-I, n = 5: heifers received one injection of rbST on day 0 (day of CIDR (controlled internal drug release) removal); 2) rbST-II, n = 5: heifers received the first rbST injection on day -8 and the second one on day 0. Thirty-eight cows were used as recipients and were assigned to receive one embryo from one of the two treatments. Results: there was no effect of treatment (p˃0.05) on PRR, dLPF, IE, and SR. The number of oocytes increased (p˂0.05) in the rbST-I treatment (1.3 ± 0.4 vs 0.2 ± 0.2), but there was no difference in the number (p˃0.05) of degenerated (1.0 ± 0.5 vs 4.5 ± 3.0) or transferable (1.0 ± 0.5 vs 1.4 ± 0.7) embryos between heifers in the rbST-I and rbST-II treatments, respectively. Moreover, no oocytes or embryos were recovered from 36.8% of donor heifers in either treatment. Conclusion: the application of 500 mg of rbST on days -8 and 0 of the follicular wave synchronization program did not increase the superovulatory response but significantly reduced the number of oocytes recovered from superovulated Holstein heifers.Antecedentes: a somatotropina bovina recombinante (rbST) tem sido aplicada em vacas doadoras de embriões com o objetivo de melhorar a resposta superovulatória e a qualidade embrionária. Objetivo: o presente trabalho avaliou o efeito da aplicação de uma contra duas injeções de 500 mg de rbST em novilhas de raça Holandesa doadoras de embriões na incidência do cio (IE), diâmetro do maior folículo pré-ovulatorio (dLFP), resposta superovulatória (SR), produção de embriões e taxa de gestação em vacas receptoras de raça Holandesa (PRR). Métodos: foram feitos dois programas de superestimulação, em cada superstimulação as novilhas foram distribuídas em um de dois tratamentos: 1) rbST-I, n = 5: as novilhas receberam uma injeção de rbST no dia zero (dia de remoção do CIDR -dispositivo intravaginal de liberação controlada de progesterona). 2) rbST-II, n = 5: as novilhas receberam a primeira injeção de rbST no dia -8 e a segunda injeção no dia zero. Trinta e oito vacas foram utilizadas como receptoras e receberam um embrião proveniente de uma doadora de um dos dois tratamentos. Resultados: a aplicação de uma ou duas injeções de rbST não afetou (p˃0,05) a PRR, dLFP, IE nem a SR. O número de oócitos foi maior (p˂0,05) no tratamento rbST-I (1,3 ± 0,4 vs 0,2 ± 0,2), o número de embriões não-viáveis (1,0 ± 0,5 vs 4,5 ± 3,0) ou viáveis (1,0 ± 0,5 vs 1,4 ± 0,7) não foram diferentes (p˃0,05) entre as novilhas do tratamento rbST-I e rbST-II. Além disso, do 36.8% das doadoras de ambos os tratamentos não foi possível coletar nenhum oócito ou embrião. Conclusões: A aplicação de 500 mg de rbST o dia -8 e zero do protocolo de sincronização da onda folicular não aumentou a resposta superovulatória, mas diminuiu significativamente o número de oócitos coletados de novilhas de raça Holandesa superovuladas.Antecedentes: la somatotropina bovina recombinante (rbST) ha sido aplicada en vacas donadoras de embriones con el objetivo de mejorar la respuesta superovulatoria y la calidad embrionaria. Objetivo: este trabajo de investigación evaluó el efecto de aplicar una versus dos inyecciones de 500 mg de rbST a novillas Holstein donadoras de embriones sobre la incidencia de celo (IE), diámetro del folículo preovulatorio de mayor tamaño (dLFP), respuesta superovulatoria (SR), producción de embriones y tasa de concepción en vacas Holstein receptoras de embriones (PRR). Métodos: se llevaron a cabo dos programas de superestimulación. En cada programa las novillas fueron asignadas a uno de dos tratamientos: 1) rbST-I, n = 5: las novillas recibieron una inyección de rbST el día cero (día de remoción del CIDR (dispositivo intravaginal liberador de progesterona); 2) rbST-II, n = 5: las novillas recibieron la primera inyección de rbST el día -8 y la segunda el día cero. Treinta y ocho vacas fueron utilizadas como receptoras. Cada receptora recibió un embrión proveniente de una donadora de uno de los dos tratamientos. Resultados: la aplicación de una o dos inyecciones de rbST no afectó (p˃0,05) la PRR, dLFP, IE ni la SR. El número de ovocitos fue mayor (p˂0,05) en el tratamiento rbST-I (1,3 ± 0,4 vs 0,2 ± 0,2), el número de embriones degenerados (1,0 ± 0,5 vs 4,5 ± 3,0) o transferibles (1.0 ± 0,5 vs 1,4 ± 0,7) no fue diferente (p˃0,05) entre las vaquillas del tratamiento rbST-I y rbST-II. Además, del 36,8% de donadoras de ambos tratamientos no se colectó ningún ovocito o embrión. Conclusión: la aplicación de 500 mg de rbST los días -8 y cero del protocolo de sincronización de la onda folicular no incrementó la respuesta superovulatoria pero redujo significativamente el número de ovocitos recolectados de vaquillas Holstein superovuladas

Antioxidant supplementation in female ruminants during the periconceptional period: A review

Abstract Oxidative stress is the result of an imbalance between free radicals and antioxidants. Under normal physiological conditions, free radicals are involved in reproductive events such as cell cycle activation, ovulation and luteolysis. However, when an overproduction of free radicals surpasses antioxidant capacity, oxidative damage, reproductive anomalies and diminished fertility occur. Supplementation with antioxidants prevents oxidative damage and can be incorporated into reproductive management to improve fertility in females. Selection of the preovulatory follicle, ovulation, fertilization, embryo development and formation of the corpus luteum occur during the periconceptional period. This is a dynamic period and the events are susceptible to oxidative stress damage. Therefore, the objective of this review is to discuss the effect of oxidative stress on reproductive events during the periconceptional period, as well as to address antioxidant supplementation during this period.Resumo O stress oxidativo é gerado por um desequilíbrio entre radicais livres e antioxidantes. Sob condições fisiológicas normais, os radicais livres participam de eventos reprodutivos, como ativação do ciclo estral, ovulação e luteólise. No entanto, quando é produzido em quantidades que excedem a capacidade antioxidante do organismo, produzem danos oxidativos e distúrbios reprodutivos que diminuem a fertilidade da fêmea. A suplementação com antioxidantes previne o dano oxidativo e sua incorporação em programas de gerenciamento reprodutivo pode ser uma opção para melhorar a fertilidade da fêmea. A seleção do folículo pré-ovulatório, ovulação, fecundação, desenvolvimento embrionário e formação do corpo lúteo ocorrem durante o período periconcepcional. Este é um período dinâmico e os eventos que ocorrem são suscetíveis ao dano por estresse oxidativo. Portanto, o objetivo dessa revisão é fornecer ao leitor conhecimento sobre o efeito do estresse oxidativo em eventos reprodutivos durante o período periconcepcional, e também discutir a suplementação com antioxidantes em ruminantes durante este período.Resumen El estrés oxidativo es generado por un desbalance entre radicales libres y antioxidantes. Bajo condiciones fisiológicas normales, los radicales libres participan en eventos reproductivos tales como activación del ciclo celular, ovulación y luteólisis. Sin embargo, cuando estos son producidos en cantidades que sobrepasan la capacidad antioxidante del organismo producen daño oxidativo y trastornos reproductivos que disminuyen la fertilidad de la hembra. La suplementación con antioxidantes previene el daño oxidativo y su incorporación a programas de manejo reproductivo puede ser una opción para mejorar la fertilidad de la hembra. La selección del folículo preovulatorio, ovulación, fecundación, desarrollo embrionario y formación del cuerpo lúteo ocurren durante el periodo periconcepcional. Este es un periodo dinámico y los eventos que ocurren en él son susceptibles a daño por estrés oxidativo. Por tanto, el objetivo de esta revisión es discutir el efecto del estrés oxidativo en los eventos reproductivos durante el periodo periconcepcional, así como la suplementación de antioxidantes en rumiantes durante este periodo

Supplementation of ascorbic acid to improve fertility in dairy cattle. Review

Ascorbic acid (vitamin C: VC) is an antioxidant that participates in the regulatory processes involved in the development of ovarian structures and fertility. However, supplementation of VC to dairy cattle to improve fertility has received little attention. However, reduced fertility in dairy cattle associated with high genetic merit for milk production and heat stress, which also diminish blood VC concentrations, suggest a potentially beneficial role for VC supplementation. The objectives of this review are to contribute to the current knowledge regarding the relationship between VC and fertility and to share many experiences that support the relevance of VC supplementation to improve dairy cattle reproductive performance

Effect of milk yield, number of calving and calving season on the resumption of postpartum ovarian activity in Hoistein cows

El objetivo fue evaluar el efecto de la producción de leche acumulada hasta el día 50 posparto (PL50), el número (NP) y época (EP) de parto sobre los intervalos para la aparición del primer folículo dominante (FD), la aparición del primer cuerpo lúteo (CL), el tiempo para completar la involución uterina (¡U), y la probabilidad de ovulación del primer FD. En vacas Hoistein con diferente NP que parieron en invierno o primavera en Tlahualilo, Durango, México. Ni la EP ni el NP afectaron los días a la aparición del primer FD (α =0,05). Sin embargo, el primer FD en vacas con alta PL50 se presentó dos días más tarde que en las de baja PL50 (13,6 vs 11,9 días; P<0,05). El NP, la EP y la PL50 no afectaron (α =0,05) la probabilidad de ovulación del primer FD. Sin embargo, las vacas con mejor condición corporal al parto fueron más aptas (OR = 9,0; P<0,05) que las de condición corporal pobre para desencadenar la ovulación del primer FD. La PL50 no afectó la aparición del primer CL después del parto (α=0,05); sin embargo, las vacas de cuatro o más partos presentaron un primer CL más tarde (P<0,05) que las de menor NP. Las vacas que parieran en invierno presentaron un primer CL más tarde que las que parieron en primavera (20 vs 16 días; P<0,01). Sin embargo, las vacas paridas en primavera requirieron más días que las paridas en invierno para completar la IU (36 vs 32 días; P<0,05). Se concluye que el crecimiento folicular ovárico no es el principal factor limitarte para el reinicio de la actividad ovárica posparto en vacas Holsteir manejadas en las condiciones del presente experimento.548 - 558BimestralThe objective of the study was to evaluate the effect of accumulated milk yield from calving to day 50 of lactation (MY50), number (NC) and season (SG) of calving on the intervals from calving to the appearance of the first dominant follicle (DF), the appearance of the first corpus luteum (CL), the time to completion of uterine involution (UI), and the probability of ovulation of the first DF in Holstein cows calving in spring or winter in a commercial dairy farm in Tlahualilo, Durango, México. Neither SO nor NO affected the appearance of the first DF (α =0.05). However, cows with higher MY50 presented the first DF two days latter than cocas with low MY50 (13.6 vs 11.9 days; P<0.05). The effects of NO, SO, and MY50 did not affect the probability of ovulation of the first DF (α =0.05). However, cows with higher body condition score at calving (BCS X3.6 units) were more likely (OR = 9.0; P<0.05) to achieve ovulation in the first DF than cows with lower BCS. The appearance of the first CL after calving was not affected by MY50; however, cows with four or more calvings showed their first CL later (P<0.05) than younger cows. Likewise, cows calving in winter showed their first CL later than those calving in spring (20 vs 16 days; P<0.01). However, cows calving in spring required longer time intervals to complete their UI (36 vs 32 days; P<0.05). It is concluded that follicular growth is not the main limiting factor to the resumption of postpartum ovarian activity in Holstein cows managed under the conditions of the present experiment

Application of equine follicular fluid (EFF) in creole synchronized ewes

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