Abstract Currently, gonadotropin products (follicle stimulating hormone, FSH, and luteinizing hormone, LH) used in animal reproduction are produced by extraction and purification from abattoir-derived pituitary glands. This method, relying on animal-derived materials, carries the potential risk of hormone contamination and pathogen transmission. Additionally, chorionic gonadotropins are extracted from the blood of pregnant mares (equine chorionic gonadotropin; eCG) or the urine of pregnant women (human chorionic gonadotropin; hCG). However, recent advancements have introduced recombinant gonadotropins for assisted animal reproduction therapies. The traditional use of FSH for superovulation has limitations, including labor requirements and variability in superovulation response, affecting the success of in vivo (SOV) and in vitro (OPU/IVEP) embryo production. FSH treatment for superstimulation before OPU can promote the growth of a homogenous follicular population and the recovery of competent oocytes suitable for IVEP procedures. At present, a single injection of a preparation of long-acting bovine recombinant FSH (rFSH) produced similar superovulation responses resulting in the production of good-quality in vivo and in vitro embryos. Furthermore, the treatment with eCG at FTAI protocol has demonstrated its efficacy in promoting follicular growth, ovulation, and P/AI, mainly in heifers and anestrous cows. Currently, treatment with recombinant glycoproteins with eCG-like activity (r-eCG) have shown promising results in increasing follicular growth, ovulation, and P/AI in cows submitted to P4/E2 -based protocols. Bovine somatotropin (bST) is a naturally occurring hormone found in cows. Recombinant bovine somatotropin (rbST), produced through genetic engineering techniques, has shown potential in enhancing reproductive outcomes in ruminants. Treatment with rbST has been found to improve P/IA, increase donor embryo production, and enhance P/ET in recipients. The use of recombinant hormones allows to produce non-animal-derived products, offering several advantages in assisted reproductive technologies for ruminants. This advancement opens up new possibilities for improving reproductive efficiency and success rates in the field of animal reproduction.

RESUMO: O objetivo deste estudo foi avaliar diferentes momentos para a realização da IATF em búfalas submetidas a um protocolo à base de P4/E2/eCG. Neste estudo, 204 búfalas foram distribuídas em um de dois grupos (IATF56, n=103 e IATF64, n=101). Em estágio aleatório do ciclo estral (Dia 0 = D0), pela manhã (IATF56, manhã) ou pela tarde (IATF64, tarde), as búfalas receberam um dispositivo intravaginal de progesterona (P4; 1,0 g) e BE (2,0 mg i.m.). No D9 pela manhã (IATF56) ou pela tarde (IATF64), a P4 foi removida e as búfalas receberam PGF2a (0,53 mg i.m. cloprostenol sódico) e eCG (400 UI i.m.). No D10 pela manhã (IATF56) ou pela tarde (IATF64), 24 h após a remoção da P4, as búfalas foram tratadas com BE (1,0 mg i.m.). As búfalas dos grupos IATF56 e IATF64 foram inseminadas 56 e 64 h após a remoção da P4 (D11, pela tarde e D12, pela manhã, respectivamente). Avaliações ultrassonográficas foram realizadas no D0 para verificar o status folicular ovariano, na IATF para medir o diâmetro do folículo dominante (FD) e no D42 para o diagnóstico de gestação. A análise estatística foi realizada utilizando o procedimento GLIMMIX do SAS®. Não houve diferença entre os grupos IATF56 e IATF64 no diâmetro do FD na IATF e na prenhez por IATF. Conclui-se que a IATF 56 ou 64 h após a remoção da P4 não afeta a fertilidade de búfalas submetidas à indução da ovulação com BE. A presente pesquisa evidencia que é possível realizar a IATF durante todo o dia em búfalas sincronizadas durante a estação reprodutiva desfavorável.

ABSTRACT: The aim of this study was to evaluate different times for timed artificial insemination (TAI) in buffalo submitted to a P4/E2/eCG-based protocol. In this study, 204 buffaloes were distributed into one of two groups (TAI56, n=103 and TAI64, n=101). At a random stage of the oestrous cycle (Day 0 = D0), in the morning (TAI56, a.m.) or afternoon (TAI64, p.m.), buffaloes received an intravaginal progesterone device (P4; 1.0 g) plus EB (2.0 mg i.m.). On D9 a.m. (TAI56) or p.m. (TAI64), the P4 was removed and buffaloes received PGF2a (0.53 mg i.m. sodium cloprostenol) and eCG (400 IU i.m.). On D10 a.m. (TAI56) or p.m. (TAI64), 24 h after P4 removal, buffaloes were treated with EB (1.0 mg i.m.). Buffaloes from TAI56 and TAI64 were inseminated 56 and 64 h after P4 removal (D11, p.m. and D12, a.m., respectively). Ultrasound examinations were performed on D0 to ascertain ovarian follicular status, at TAI to measure the diameter of the dominant follicle (DF) and D42 for pregnancy diagnosis. The statistical analysis was performed using the GLIMMIX procedure of SAS®. There was no difference between TAI56 and TAI64 for the diameter of the DF at TAI and the pregnancy per TAI. It was concluded that TAI 56 or 64 h after P4 removal did not affect fertility in buffaloes submitted to the induction of ovulation with EB. The present research supports that is possible to perform TAI at any time throughout the day in buffalo synchronized during the non-breeding season.

Abstract Ovulation synchronization protocols are well established in beef and dairy cows. However, the protocol response rate is around 70-90%. In beef cows, factors such as inadequate nutrition and calf presence negatively impact the response of progesterone (P4)/estradiol-based ovulation synchronization protocols by interfering with GnRH release and consequently reducing LH pulsatility and final follicular development. In dairy cows, protocols based on GnRH and prostaglandin (Ovsynch) are the most widely used in the world. However, the efficiency of Ovsynch is dependent on the presence of a large follicle at the time of administration of the first GnRH. In these ovulation synchronization protocols, pre-synchronization protocols (Prostaglandins, Double Ovsynch and P4synch) are usually attempted in an effort to increase responses. Thus, the objective of this review was to discuss pre-ovulation synchronization strategies (administration of injectable P4 or energetic/protein supplementation or pre-synchronization with intra-vaginal progesterone devices) aiming to increase the LH pulsatility in beef cows or induce the formation of a GnRH-responsive follicle at the beginning of the Ovsynch protocol in dairy cows.

RESUMOAlta produção leiteira, calor, estado fisiológico e manejo prejudicam a reprodução em vacas Holandesas nos trópicos. A utilização de embriões produzidos in vivo é uma alternativa para melhorar taxas de gestação; no entanto, existem limitações, especialmente pelas variações nas respostas superovulatórias. A produção in vitro de embriões evitaria tais variações, mas poucos estudos estão relatados. O objetivo é verificar os efeitos de variáveis relacionadas às receptoras sob um programa de transferência de embriões (TE) in vitroem fazenda comercial no sudeste do Brasil. Hipotetiza-se que taxas de gestação, após TE de embriões derivados de OPU em receptoras Hosltein em lactação, são influenciadas pelo GnRH aplicado no momento da TE, paridade, produção de leite e escore de condição corporal. As receptoras (267) foram alocadas para um de três tratamentos aplicados no momento da TE: Controle (92) - Salina 2,5 ml; Buserelina (86) - 10 µg de buserelina; Deslorelina (89) - 750 µg de deslorelina. Ultrassonografia e amostras de sangue realizadas no momento da TE e sete dias depois. O primeiro diagnóstico de gestação foi realizado entre 30-40 dias e o segundo entre 60-80 dias pós-TE. Dados foram analisados pelo GENMOD (SAS(r)). A proporção de vacas prenhes foi superior (P<0,05) nas receptoras tratadas com Buserelina (38,1%) comparadas com Controle (24,1%) no primeiro diagnóstico de gestação, mas semelhante às vacas tratadas com Deslorelina e Controle no segundo diagnóstico (13,0, 20,9 e 14,6% no Controle, Buserelina and Deslorelina, respectivamente). A Buserelina melhorou a taxa de gestação apenas transitoriamente, nas presentes condições experimentais.

ABSTRACTHigh milk production, heat, physiological status and management impair reproduction in Holstein cows. The use of in vivo-produced embryos has been reported as an alternative to enhance pregnancy outcome in the tropics; however there are several limitations for its production, especially from variations in superovulatory responses. The in vitro production of embryos would avoid such variations, but few studies have been reported. This study aims to verify the effects of variables related to recipients under a program of routine in vitro embryo transfer on a commercial dairy farm in southeastern Brazil. It was hypothesized that pregnancy rates after transfer of ovum pick up or OPUderived embryos (ET) to lactating Holstein recipients may be influenced by recipient GnRH-treatment at ET, parity, milk production and body condition score. Recipients (267) were allocated to one of three i.m. treatments given at ET: Control (92) - 2.5 ml saline; Buserelin (86) - 10 μg Buserelin acetate; Deslorelin (89) - 750 μg Deslorelin acetate. Ultrasound images and blood samples were taken at ET and seven days later. The first pregnancy diagnosis was performed between 30-40 days and the second between 60-80 days post ET. Data were analyzed by GENMOD (SAS(r)). The proportion of pregnant cows was greater (P<0.05) in Buserelin-treated recipients (38.3%) at the first pregnancy diagnosis than Controls (24.1%), but similar to Deslorelin and control cows at the second diagnosis (13.0, 20.9 and 14.6% in Control, Buserelin and Deslorelin, respectively). In conclusion, Buserelin improved pregnancy rate only transitorily, under the present conditions.

Dois experimentos foram conduzidos para avaliar o efeito da injeção intramuscular de betacaroteno associada ao tocoferol, na concentração plasmática de progesterona de novilhas Holandesas superovuladas (Experimento 1) e em novilhas cruzadas (Bos taurus x Bos indicus) submetidas à transferência embrionária em tempo fixo (TETF, experimento 2). No experimento 1, após a sincronização do estro e superovulação, os animais foram inseminados entre 12 a 24 h após o início do estro e os embriões recuperados após 7 dias. As novilhas foram divididas, aleatoriamente, em 1 de 3 tratamentos: Controle (duas injeções de soro); T800 (800 mg de betacaroteno mais 500 mg de tocoferol) e T1200 (1.200 mg de betacaroteno mais 750 mg de tocoferol). Os tratamentos foram administrados no dia da colocação do implante de progesterona e repetido no primeiro dia da superovulação. As colheitas de sangue foram realizadas no D0, D5, D9, D12 e D16. No experimento 2, os tratamentos foram realizados no momento da inserção do dispositivo de progesterona (D0). Utilizou-se o mesmo delineamento do experimento 1. A colheita de sangue para a determinação da progesterona por radioimunoensaio foi realizada no D17 (inovulação embrionária). No experimento 1, a concentração plasmática média, após formação do corpo lúteo (média da soma de D12 com D16) foi de 13,7±1,8 ng/ml, 14,5±2,3 ng/ml e 10,8±2,3 ng/ml para os grupos Controle, T800 e T1200, respectivamente, e não diferiram (P=0,44) entre os tratamentos. No experimento 2, a concentração de progesterona nas receptoras, no dia 17, foi semelhante (P=0,11) entre os grupos controle (8,88±0,57 ng/ml), T800 (7,48±0,64 ng/ml) e T1200 (5,90±1,33 ng/ml). Os resultados indicaram que as injeções de suplemento com betacaroteno e tocoferol não influenciaram na concentração plasmática de progesterona em novilhas Holandesas doadoras de embriões e em novilhas cruzadas, receptoras de embrião.

Two experiments were conducted to evaluate the effect of the intramuscular injection of betacarotene associated to tocopherol on the plasma concentration progesterone of superovulated Holstein heifers (experiment 1) and in crossbred (Bos taurus x Bos indicus) heifers submitted to fixed-time embryo transfer (FTET, experiment 2). In experiment 1, after estrus synchronization and superovulation animals were inseminated 12 and 24 hours after estrus onset and embryos flushed 7 days later. Heifers were allocated randomly to one of three treatments: Control; T800 (800 mg of betacarotene plus 500 mg of tocopherol) and T1200 (1,200 mg of betacarotene plus 750 mg of tocopherol). The treatments were given on the day of ear implant placement and repeated on the first day of superovulation. Blood samples were collected on D0, D5, D9, D12 and D16. In experiment 2, treatments were imposed at intravaginal device insertion (D0). The same experimental design, as in experiment 1, was used. Blood samples were collected on D17 (embryos implanted) for progesterone determination by radioimmunoassay. In experiment 1, average plasma progesterone concentrations after corpora lutea formation (D12 plus D16 means) were 13.7±1.8 ng/ml, 14.5±2.3 ng/ml and 10.8±2.3 ng/ml for control, T800 and T1200, respectively, and did not differ (P=0.44). In experiment 2, progesterone concentrations on D17 in Control (8.88±0.57 ng/ml), T800 (7.48±0.64 ng/ml) and T1200 (5.90±1.33 ng/ml) groups were similar (P=0.11). Results indicate that the supplemental betacarotene and tocopherol injections did not influence peripheral progesterone concentrations in superovulated Holstein donors and crossbreed recipients heifers.