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Ovarian tissue cryopreservation has been widely investigated for preserving female fertility. In the present study, we aimed to compare the effects of three concentrations (1, 1.5, and 3 M) of dimethylsulfoxide (DMSO) on the vitrification of ovarian tissue. The ovarian cortex was divided into control and vitrified groups: (i) 1 M-DMSO, (ii) 1.5 M-DMSO, and (iii) 3 M-DMSO. Follicles from all fragments were analyzed for DMSO-induced deleterious effects, morphological and morphometric aspects, and concentration of reactive oxygen species. Additionally, the fragments were cultured to assess the integrity and return of follicular development post-vitrification. All DMSO concentrations resulted in a higher percentage of degenerated preantral follicles than before the cryopreservation process. After vitrification, the cryopreserved ovarian fragments showed similar percentages of intact follicles; however, the 3 M DMSO concentration differed from the control. Analyzing free radical production, we found that the 3 M DMSO concentration had higher levels of oxidative stress than the lower DMSO. After in vitro cultivation of the vitrified/warmed fragments, the 1 M DMSO concentration exhibited higher percentages of morphologically intact follicles than the other concentrations. Therefore, we suggest that bovine preantral follicles can be cryopreserved in situ with greater efficiency in 1 M DMSO.

The microbiota plays an important role in numerous physiological processes, pathogenesis, development, and metabolism in different animal species. In humans, several studies have demonstrated an association between the vaginal microbiota and fertility rates, and even success in assisted reproduction techniques. In the context of cattle reproduction, although few studies have addressed the microbiota in a healthy state (which is not associated with diseases that affect the reproductive tract of cows), changes in its composition also seem to influence fertility. This review aims to explain the importance of the reproductive microbiota in female bovines and what is available in the literature regarding its possible role in increasing fertility. What are the challenges involved in this process? Future perspectives on its use and manipulation as a selection or intervention tool. Will it be possible to one day extrapolate the findings to reality and apply them in the field? In short, understanding the role of the reproductive microbiota of female bovines can signal the prospect of increasing production, whether of milk or meat, from the same number of animals, as it can optimize reproductive efficiency and perhaps become an allied tool for the economic profitability and sustainability of livestock farming.

Abstract The objective of this work was to investigate whether puberty and parturition alter the antral follicle count (AFC) in female Nelore (Bos indicus) beef cattle. Two experiments were carried out to compare AFC between the prepubertal and pubertal periods and between the prepubertal and postpartum periods. AFC and follicle concentrations did not differ between the prepubertal and pubertal periods. However, the ovarian area increased after puberty. In addition, AFC and the ovarian area were greater in the postpartum than in the prepubertal period. Therefore, AFC is not affected by the puberty of Nelore heifers, but increases significantly when the prepubertal heifers become cows.

In vitro embryo production (IVEP) is used to develop high-quality genetics associated with intergenerational genetic gain. It is characterized by acquisition (in vivo or post-mortem) and maturation (MIV) of oocytes from donors, followed by fertilization (FIV) of matured oocytes and culture (IVC) of embryos, which are then sent to transferred or cryopreserved. Even with extensive knowledge on IVEP, some biochemical and hormonal regulations that involve embryonic development are still unknown, leading to a low overall efficiency of the biotechnological process. Although in vitro developed embryos have a lower quality than that produced in vivo, in terms of resistance to challenging events, IVEP presents itself as a potential biotechnology. In cattle breeding, reproductive biotechnologies are key to increase and improve the genetic improvement of the herd, associated with productive and reproductive efficiency. In this article, the steps and strategies of IVEP and its contribution to reproduction in the cattle sector are discussed.

The application of Fixed Time Artificial Insemination (FTAI) protocols has strongly boosted artificial insemination in Brazilian bovine herds. The control of the estrous cycle with protocols based on progesterone and estrogen involves synchronizing the emergence of a follicular wave, controlling the progesterone phase, and inducing ovulation in a synchronized manner. The increase in pregnancy rates starts with the preparation of the breeding season and the choice of protocols and females. Thereafter, based on the characteristics of the property and the lots of animals, small adjustments can be made to the hormonal protocols as needed. Finally, resynchronization can increase usability by allowing multiple inseminations within short periods. The search for new strategies is of paramount importance for the development of the FTAI protocol and the increase in final pregnancy rates. Thus, this review sought to highlight the main critical points and adjustments of the FTAI protocols and their applicability in the reproduction of beef cattle.

The vaginal microbiota has been shown to be important in local immune regulation and may play a role in reproduction and fertility. Next-generation sequencing (NGS) technologies have been used to characterize the bovine vaginal microbiota, mainly using short-read sequencing (Illumina). However, the main limitation of this technique is its inability to classify bacteria at the species level. The objective of this study was to characterize the bovine vaginal microbiota at the species level using long-read sequencing (PacBio) and to compare it with the results of short-read sequencing. In addition, the vaginal microbiota of cows that became pregnant after artificial insemination (AI) was compared with that of infertile animals. Thirteen Holstein cows had vaginal swabs collected prior to AI. DNA was extracted and subjected to Illumina and PacBio sequencing to characterize the V4 region and the entire 16S rRNA gene, respectively. PacBio sequencing yielded 366,509 reads that were assigned to 476 species from 27 phyla. However, none of the most abundant reads (>1%) could be classified at the species level. Illumina sequencing yielded more reads and consequently was able to detect a more observed species, but PacBio sequencing was able to detect more unique and rare species. The composition of the vaginal microbiota varies according to the sequencing method used, which might complicate the interpretation of results obtained in the majority of the current studies. The present study expands on the current knowledge of bovine microbiota, highlighting the need for further efforts to improve the current databanks.

This study aimed to determine the effect of antral follicle count (AFC), and pubertal status on the fertility of beef heifers. In this study, 230 Nelore heifers, 20±2 months of age, were subjected to an estradiol progesterone-based timed artificial insemination (TAI) program. On Day 0 of the TAI protocol, the heifers were examined by transrectal ultrasound to record videos of the ovaries. Later, in the darkroom of the laboratory of images, the videos were analyzed for AFC (≥ 3 mm) of each ovary. Females who failed the first TAI were resynchronized with the same hormonal protocol. The pregnancy status was evaluated by ultrasonography 30 days after each FTAI. The general mean of the AFC was 22.0 follicles. Thus, the heifers were divided into 2 groups according to AFC: Low AFC (˂ 22 follicles, n = 114), and High AFC (≥ 22 follicles, n = 116). No differences (P > 0.05) in the pregnancy per AI (P/AI) were observed between the Low and High AFC groups, and between pubertal and prepubertal categories. The P/AI was not different between heifers that displayed or did not estrus (P = 0.2). However, considering the estrus response of each AFC group, High AFC heifers that displayed estrus had greater P/AI (P = 0.01) than High AFC heifers that did not display estrus. In summary, AFC and pubertal status did not affect the fertility of Nelore heifers. In contrast, the P/AI of heifers that did not display estrus was lower than heifers observed in estrus only in the High AFC group.

For this study, ½ blood nelore × aberdeen angus heifers (n = 40; 14 months) were divided into two groups: control (n=20; 310 kg) and pró-cio (n=20; 304 kg). for pró-cio group, 20g of homeopathic product (pró-cio®, real h, campo grande, brazil) was supplied together with corn eighteen days prior (d-18) to the beginning of the fixed-time artificial insemination protocol (ftai). on d0, all females received an intravaginal p4 device and 2 mg be. on d5, the afc was assessed by ultrasound and blood was collected for amh dosage. on d8, p4 device was removed; 300 iu ecg, 0.530 mg of cloprostenol sodium and 0.5 mg of ec were administered; an estrus-identifying adhesive was fixed at the tail insertion and the diameter of the dominant follicle (df) was measured. on d10, all heifers were inseminated, estrus manifestation was assessed and the preovulatory follicle (pof) was measured. on d20, the cl area was measured and blood flow was assessed using doppler ultrasound. data were analyzed by student, mann-whitney or friedman test. the rates of heat expression, ovulation, and blood flow score with doppler were analyzed using fisher’s exact test and pregnancy rate by binary logistic regression model (p ≤ 0.05). there was no difference in afc or amh dosage (p > 0.05). pró-cio group had greater diameters of the df (9.94 ± 0.42 mm) and pof (11.61 ± 0.56 mm) than the controls (df: 7.72 ± 0.34 mm and pof: 9.91 ± 0.37 mm), as well a larger cl area (3.26 ± 0.26 versus 2.35 ± 0.16 cm2) and a higher mean cl vascularization score (3.06 versus 2.26; p < 0.05). there was no significant difference (p > 0.05) between estrus manifestation or conception rate for control and pró-cio groups. heifers supplemented with the homeopathic product showed larger follicular diameters at the end of the ftai protocol and cl with better blood flow scores compared to the control group.

We evaluated the effects of nulliparous, primiparous, and multiparous conditions on the estrus and pregnancy rates in females that did not show estrus but were treated with gonadotropin-releasing hormone (GnRH) at the time of timed artificial insemination (TAI). Nelore females (n = 531) were allocated according to the following categories: nulliparous (n = 144), primiparous (n = 132), and multiparous (n = 255). The animals received a conventional TAI protocol, and estrus expression was identified by the absence of paint in the sacrococcygeal region on the day of TAI. Females that did not show estrus were treated with 10 µg of GnRH together with insemination. The rates of estrus and pregnancy were analyzed using a logistic regression model (P < 0.05). The estrus expression was lower (P = 0.006) in the primiparous (61.36%) group than in the nulliparous (76.39%) and multiparous (75.69%) groups. Similar pregnancy rates were observed in females that showed estrus (nulliparous 84.54%, primiparous 86.42%, and multiparous 80.31%; P = 0.39) and in females that did not show estrus and received GnRH (nulliparous 41.18%, primiparous 56.86%, and multiparous 58.06%; P = 0.24). The total pregnancy rates were also similar (P = 0.98) among the categories (nulliparous 74.3%, primiparous 75.0%, and multiparous 74.9%). The primiparous females had a lower rate of estrus, and the pregnancy rates were similar among the categories that received GnRH.

This study used Holstein cattle to evaluate the following: I) the productive and reproductive characteristics of cows with low, intermediate and high antral follicle counts (AFCs) that were subjected to artificial insemination (AI) and comparison of AFC variability at the time of AI (nonpregnant) and at two time points during pregnancy (30 and 60 days), and II) whether the pregnancy status and different pregnancy periods in heifers improves the efficiency at in vitro embryo production (IVEP). In study I, 75 high-milk production cows with body condition scores (BCSs) of 2.75 to 4.75 and ages of 23 to 99 months were selected. On the day of estrus, AFCs (follicles bigger, smaller 3 mm), BCS, body weight, and diameters of the dominant follicle, ovaries and the corpus luteum were evaluated. Posteriorly, AFC was monitored in pregnant cows at 30 days (n = 35) and at 60 days for comparison of AFC before pregnancy versus different periods. In study II, heifers (n = 9) with BCS of 2.5 to 3.5 and ages of 10 to 16 months were selected for ovum pick-up and an IVEP program before AI (nonpregnant) as well as at 0 to 30, 31 to 60, and bigger 60 days of pregnancy. Statistical analysis was performed using PROC GLM and binary logistic regression model (P bigger, smaller 0.05). In study I, AFC was not associated with variations in productive and reproductive parameters. However, AFCs in the low group increased following pregnancy (AI: 14.82 ± 1.36, 30 days: 23.45 ± 2.31 and 60 days: 35.18 ± 3.17 follicles). AFCs increased from AI to 60 days in the intermediate group but did not vary among time points in the high group. AFCs varied among AFC groups in AI and at 30 days of pregnancy but not at 60 days. In study II, the mean of total oocytes (17.43 ± 4.52 vs. 41.67 ± 3.79) and viable oocytes (13.77 ± 3.63 vs. 30.56 ± 3.45) increased from AI (nonpregnant) to the first 30 days of pregnancy. The mean number of embryos produced increased during pregnancy and during the first 30 days of pregnancy (4.55 ± 0.75) compared to the nonpregnant period (1.39 ± 0.85). In conclusion, pregnancy positively influences AFC, and the greatest effect is noted in low count cows. In addition, the OPU at the time of pregnancy increased the efficiency of IVEP in heifers.

Reproductive biotechnologies emerge as an important element for livestock, however, some strategies must be modified to adapt the different breeding systems, such as the use of follicular synchronization protocols. This study aimed to evaluate follicular synchronization using estradiol benzoate, with the presence of the corpus luteum, on the in vitro embryo production (IVEP) from Wagyu oocytes donors. Routines of IVEP were performed in heifers and cows (n=19), classified into three groups: G1/CL- animals with corpus luteum; G2/SCL- animals without corpus luteum, these groups being the control, and had the natural process of follicular dynamics and G3/CL + EB - animals with the presence of corpus luteum, submitted to follicular synchronization at D0, with estradiol benzoate. The results obtained showed that the synchronization of the follicular wave with the application of EB, in the presence of the corpus luteum (CL), reduced the number of small (6.05 ± 0.55) and large follicles (0.45 ± 0.15), but increased (P0.05). It was concluded that follicular synchronization using estradiol benzoate in Wagyu oocyte donors, presenting a corpus luteum, increased the number of medium follicles, however, there was no improvement in the efficiency of ovum pick-up, in vitro embryo production and pregnancy rate.

The objective of this study was to evaluate the efficiency and cost of two distinct hormonal protocols for induction of puberty in Bos indicus heifers in the western Amazon region. Nelore prepubertal heifers (n = 127) aged 20 to 24 months (21 ± 0.3 months) and weighing 300 to 340 kg (318 ± 7 kg) were submitted to two distinct protocols for induction of cyclicity. Females were previously submitted to two ovarian ultrasound examinations (12-day interval) to verify the absence of the corpus luteum (CL). Then, the heifers were divided into two experimental groups: G-CIDR/EC (n = 54) and G-P4/EC (n = 73). The G-CIDR/EC group utilized a CIDR® for 12 days and on the day of its withdrawal, 0.6 mg of estradiol cypionate (IM) was administered. The G-P4/EC group received 150 mg of injectable progesterone (IM) and one injection of 0.6 mg EC 12 days after the progesterone injection. Twelve days after the end of the hormonal protocols, the heifers were submitted to another ultrasound evaluation for identification of cyclicity (ovulatory follicle or CL). These females were then submitted to a conventional protocol of synchronization of ovulation for timed artificial insemination (TAI). Only heifers responsive to puberty induction (G-CIDR/EC (n=30) and G-P4/EC (n=51)) were included in the TAI protocol. After seven days of insemination, both groups were exposed to natural breeding. In addition, an economic analysis was performed to evaluate cyclicity induction, the TAI protocol, and pregnancy in both groups. The G-CIDR/EC group showed 81.48% (44/54) of females responsive to induction of cyclicity (presence of CL) while the G-P4/EC group obtained 86.3% (63/73; p=0.463). Regarding the conception rate to TAI and to bull (natural breeding), the G-CIDR/EC group displayed 43.33% (13/30) and 33.33% (10/30), respectively, and the G-P4/EC group exhibited 54.9% (28/51) and 39.22% (20/51), respectively. The overall pregnancy rate (TAI+Bull) was 76.67% (23/30) in the G-CIDR/EC group compared to 94.12% (48/51; P=0.023) in the G-P4/EC group. The cost of the hormonal protocol to cyclicity induction resulted in an economy of 42.8% for injectable P4 compared to the intravaginal P4 group. In addition, the final cost of cyclic induction associated with TAI was 34% lower in the G-P4/EC group compared to the G-CIDR/EC group. We concluded that both cycling induction protocols were efficient in Bos indicus heifers from the western Amazon. However, injectable P4 provided a higher conception rate and lower pregnancy cost at the end of the reproductive season.

Este estudo teve como objetivo avaliar o potencial de produção in vitro de embriões (PIVE) bovinos de vacas da raça Girolando (Bos taurus x Bos indicus) com alta e baixa contagem de folículos antrais (CFA), bem como a taxa de prenhez nas receptoras destes embriões. As vacas da raça Girolando (Bos taurus x Bos indicus) foram classificadas como de alta CFA quando apresentavam de 35 a 52 folículos (n = 13) e baixa CFA, quando apresentavam de 11 a 17 folículos (n = 15). Todos os animais foram submetidos a repetidas sessões de aspiração folicular (OPU) e subsequente PIVE. O protocolo de sincronização das receptoras foi realizado em um dia aleatório do ciclo estral (Dia 0) com implante de progesterona, benzoato de estradiol e prostaglandina. O grupo de alta CFA obteve uma maior média de oócitos aspirados / OPU (42,6 ± 5,2 vs. 14,6 ± 1,9; p < 0,01); oócitos cultivados / OPU (38,1 ± 6,6 vs. 12,3 ± 2,8; p < 0,01); porcentagem de blastocistos em D7 (23,0 ? 1,0% vs. 18,4 ? 1,5%; p < 0,05) e taxa de prenhez (42,7 ? 2,7% vs. 39,7 ? 4,6%; p < 0,05) maiores quando comparado ao grupo de baixa CFA. Desta maneira, podemos concluir que os animais de alta CFA apresentaram melhores taxas de PIVE que os animais de baixa CFA.

The aim of this study was to evaluate the dynamics of the scrotal temperature, sperm quality and testicular blood flow by using infrared thermography and Doppler ultrasonography in bulls submitted to scrotal insulation. Braford bulls (n = 8) at 18 months of age, were assigned into the following groups: insulated for 72 h (G 72; n = 2), 96 h (G 96, n = 2), 120 h (G 120, n = 2), and control animals (CON; n = 2) that remained without insulation. Infrared thermography and Doppler ultrasonography parameters were measured at four different time points: immediately after the scrotal insulation (M0), within 10 min (M10), 30 min (M30) and 60 min (M60) after scrotal insulation. The sperm quality was evaluated weekly (S1/S12). The data were analyzed by ANOVA, t-test (paired) and Pearson's correlation with a significance level of 5%. None of the observed variables were different between the insulated groups (P > 0.05). Insulated animals showed higher scrotal temperature in M0 compared to that in the M10, M30 and M60 periods (P < 0.05). There was no difference in the pulsatility and resistive indexes after scrotal insulation. However, blood flow velocity was higher in M10 compared to that in the M0, M30 and M60 periods (P < 0.05). The sperm quality was higher (P < 0.05) in all twelve collections from the control group compared with the insulated groups. Scrotal insulation resulted in changes in the sperm quality, scrotal temperature and testicular blood flow velocity.

We characterized the proliferative activity of multi-oocyte follicles with anti-nuclear antigen of proliferating cells (PCNA). Ovaries (n = 12) from heifers were processed for histology. From 789 multi-oocytes follicles observed, only 11 were considered appropriated for immunostaining, since they presented all nuclei of the oocytes clearly visible. All multi-oocyte follicles were positive for PCNA, but some oocytes showed no proliferative activity. We conclude that oocytes in multi-oocyte follicles seem to be in different stages of the cell cycle.

Abstract This study aimed to evaluate different concentrations of growth and differentiation factor-9 (GDF-9) on the development and maintenance of equine preantral follicle morphology during short-term in vitro culture. Ovaries (n=5) from five mares were collected from a local slaughterhouse and transported to the laboratory, where nine fragments (5x5x1mm) were procured from each ovary. One fragment from each was immediately fixed and submitted for histological analysis (control group; D0). The other eight fragments were cultured in situ for two (D2) or six (D6) days in MEM+ or MEM+ supplemented with GDF-9 at different concentrations (i.e., 50, 100 and 200 ng/mL the GDF-9). After culturing with different concentrations of GDF-9 for 2 or 6 days, the fragments were processed for histological analysis. After two days of cultivation, we observed an increase in the percentage of developing follicles for 0 (MEM+), 50, 100 and 200 ng/mL GDF-9 compared to control (D0; P

O objetivo deste estudo foi avaliar a reutilização de um dispositivo intravaginal de progesterona (P4) durante o diestro de vacas Nelore receptoras de embriões, a fim de melhorar a taxa de concepção, acelerar a ressincronização do ciclo estral de animais não prenhes e reduzir a quantidade de resíduos gerados por biotécnicas na produção animal. Foram realizados dois experimentos com 268 vacas Nelore multíparas. No experimento 1, todos os animais foram submetidos a um procedimento de transferência de embriões em tempo fixo (TETF), mas no momento da transferência embrionária foram estabelecidos dois grupos de tratamento: T1 - o tratamento controle (N = 132) e T2 - animais recebendo um dispositivo de segundo uso CIDR® por 12 dias (N = 136). O experimento 2 foi realizado em vacas que permaneceram vazias após o experimento 1 usando dois grupos: G1 - um grupo controle (N = 69) e vacas ressincronizadas (G2) que receberam um dispositivo P4 por 12 dias para a TETF (N = 74) No experimento 1, não foi observado efeito significativo do tratamento P4 na taxa de concepção (T1 = 37,9%, T2 = 39,7%; P = 0,50) e diâmetro do corpo lúteo (CL) (T1 = 17,5 ± 3,4 mm, T2 = 18,1 ± 3,4 mm; P = 0,61). No experimento 2, nenhum efeito significativo do tratamento foi observado na taxa de concepção (G1 = 22,2%, G2 = 35,7%; P = 0,24), taxa de utilização de receptoras (G1 = 75,4%, G2 = 70,3%; P = 0,86), e diâmetro do CL (G1 = 17,4 ± 3 mm, G2 = 18,1 ± 3,2 mm; P = 0,27). No entanto, o tratamento P4 (para ressincronização) aumentou significativamente a taxa de concepção (G1 = 22,2%, G2 = 35,7%; P = 0,04), semelhante ao de um protocolo TETF convencional realizado com dispositivo de primeiro uso (38,8%). Concluímos que dispositivos P4 intravaginais reutilizáveis podem ajudar a acelerar protocolos TETF, sugerindo um método de aplicação alternativo; além disso, este protocolo pode ajudar a reduzir a produção de resíduos nas biotécnicas de reprodução assistida.

This study evaluated the pregnancy rate in Nelore cows (Bos indicus) that were subjected to fixed-time artificial insemination (FTAI) using different protocols consisting of injectable progesterone (P4) or an intravaginal device (impregnated with P4). Multiparous cows 72-84 months in age, 30-45 days postpartum, were selected on the basis of the absence of a corpus luteum (CL) and follicles < 8 mm after transrectal palpation and ultrasound examinations. On a random day of the estrus cycle (D0), the selected animals (n = 135) were randomly assigned to one of three experimental groups (n = 45 each). Group I (injectable P4/FTAI 36 hours) received 250 mg of injectable P4 and 2 mg EB on D0; on D7, they received 500 µg of cloprostenol; on D8, 300 IU of eCG and 1 mg of EB were administered; and finally, FTAI was performed 36 hours after the application of EB. Group II (injectable P4/FTAI 48 hours) received the same protocol as Group I, except that the FTAI was performed 48 hours after ovulation induction. The animals of Group III (Control/CIDR) received a conventional protocol for FTAI using an intravaginal device (D0: P4 and 2 mg EB; D8: device removal, 500 µg cloprostenol, 300 IU eCG, 1 mg EB; and FTAI performed 48 hours after removal of the device). The results showed that cows synchronized with the conventional protocol for FTAI (Control/CIDR) had a higher pregnancy rate (60 %, 27/45) than those synchronized with an injectable P4/FTAI 36 hours (33.33 %; 15/45, P = 0.010). However, the group receiving injectable P4 group/FTAI 48 hours had a similar pregnancy rate (48.9 %; 22/45; P = 0.290) when compared to both the group receiving the conventional protocol and that receiving injectable P4/FTAI 36 hours (P = 0.134). Although the injectable P4 may affect pregnancy rate with the FTAI performed in 36 hours, we found similar pregnancy rates from cows inseminated 48 hours after induction ovulation, considering injectable or intravaginal P4. Therefore, we suggest that injectable P4 represents an alternative source of progesterone for synchronization of cattle for FTAI.

This study aimed investigate the relationship between epigenetics, follicular diameter and cleavage speed, by evaluating the developmental potential and occurence of H3K4 monomethylation of early-, intermediate- and late-cleaving Bos indicus embryos from in vitro fertilized oocytes originating from follicles up to 2 mm in diameter or between 4 and 8 mm in diameter. Oocytes (n = 699) from small follicles (? 2 mm) and 639 oocytes from large follicles (4-8 mm) were punched from 1,982 Bos indicus’ slaughterhouse ovaries. After maturation and in vitro fertilization (IVF), the cultured embryos were separated into early (? 28 h post-IVF), intermediate (> 28 h and ? 34 h post-IVF) and late (> 34 h and ? 54 h post-IVF) cleavage groups. Blastocysts were subjected to an immunofluorescence assessment for H3K4me investigation. The blastocyst rate for large follicles (36.3%) was higher than that for small follicles (22.9%, P < 0.05). In addition, blastocyst rates for early and intermediate cleavage groups (45.3% and 33.8%, respectively) were higher than that for late cleavage group (13.5%, P < 0.05). The blastocysts from all groups displayed H3K4me staining by immunofluorescence, particularly intense in what seemed to be trophectoderm cells and weak or absent in cells seemingly from the inner cell mass. For the first time for indicus embryos, data from this study demonstrate that higher blastocyst embryo rates are obtained from embryos that cleave within 34 h after fertilization and from those produced from follicles of 4-8 mm in diameter, indicating a greater ability of these embryos to develop to the stage of embryonic preimplantation. This is the first article demonstrating the occurrence of H3K4me in cattle embryos; its presence in all the evaluated blastocysts suggests that this histone modification plays a key role in maintaining embryo viability at preimplantation stage.

Most fixed-time artificial insemination (FTAI) protocols utilize progesterone (P4) as a hormonal source to achieve synchronization of estrus in cattle. The use of an injectable P4 source to control estrus would be an interesting pharmacological strategy owing to the practicality of parenteral application. However, the effects of injectable P4 on estrus cycle control in cattle remain poorly studied. In particular, no existing studies have investigated the effect of injectable P4 on the fertility of cows subjected to FTAI. The aim of this study was to evaluate the pregnancy rate of lactating and non-lactating Nelore cows subjected to FTAI with injectable P4. Of the 422 non-lactating cows in this study, 162 (38.3%) became pregnant by 60 days post-FTAI. In the lactating group (n = 516), 166 (32.1%) were pregnant by 60 days after treatment with injectable P4. The proportions of lactating and non-lactating cows becoming pregnant were compared using the chi-square test, adopting a significance level of P < 0.05. It was found that the pregnancy rate of the cows subjected to FTAI with injectable P4 was influenced by lactation status. Lactating cows had lower reproductive performance, possibly because of their higher nutritional requirements. However, the use of injectable P4 shows promising results and may prove to be a useful strategy in large-scale livestock production.

The aim of this study was to investigate the efficacy of the tissue fixatives Bouin, Carnoy and 10% Formaldehyde in equine ovarian fragments. Ovaries (n=4) from mares of mixed breeds were obtained at a local slaughterhouse and transported at 20 ºC in a thermo container. Immediately after collection, the ovaries were washed with a modified PBS solution (Cultilab®, Campinas-SP, Brazil) and divided into nine fragments with approximately 5x5x1 mm, removed from the parenchyma of each ovary. The ovarian fragments were then immersed in three different fixatives, Bouin (B) Carnoy (C) or 10% Formaldehyde (F) for 6, 12 or 24 hours. Each fragment was individually immersed in a 20 mL tube containing 20 times the volume of fixative solution. After this period, the fragments were held in 70% ethanol for 24 hours. Each procedure was performed in four replicates. For histological analysis, the specimens were dehydrated in increasing concentrations of alcohol, submitted to diaphanization in xylol and embedded in paraffin. Serial sections of 5 ?m were made with the use of a rotating microtome (Leica® type, Wetzlar, Germany), followed by slide mounting and staining with periodic acid-Schiff (PAS) and hematoxylin. A total of 540 slides with 1,620 sections were evaluated, which contained 465 preantral follicles that were classified as normal or degenerated. Follicles were considered as degenerated when presented at least one of the following aspects: cytoplasm retraction, pyknotic nucleus, cytoplasmic vacuoles, displacement of granulosa cells and/or disruption of the basal membrane. A logistic regression test was used for statistical analysis, and differences were considered significant when P

The aim of this study was to evaluate ovarian antral follicle populations (OAFP) of Nelore and Girolando breed heifers (12–18 months old) and cows (24–60 months old). Animals were assigned to four groups: (1) Nelore cows (n = 18), (2) Girolando cows (n = 20), (3) Nelore heifers (n = 7), and (4) Girolando heifers (n = 7). Cows were treated to synchronize follicular wave emergence by implantation of an intravaginal device containing 1.9 g of progesterone, as well as intramuscular administration of 2 mg of estradiol benzoate and 25 mg of dinoprost. This synchronization treatment was administered at a random day of the estrous cycle of each cow, designated D0. Intravaginal devices were removed on D7, and on D11, OAFP counts were performed by transvaginal ovarian ultrasound. For each cow, all follicles ?3 mm in diameter were counted in both ovaries and counts were performed three times at 35-day intervals. Counts were also obtained from heifers, but these animals were not treated for synchronization of follicular wave emergence. Analysis of variance (ANOVA) with Tukey’s test and Pearson’s correlation test were used to compare mean OAFPs between counts as well as mean OAFPs between breed and age groups. No differences were observed in mean OAFPs between Nelore and Girolando cows (30.9 vs. 26.7, respectively; P > 0.05) or heifers (16.2 vs. 18.1, respectively; P > 0.05). However, within each breed, there were differences in mean OAFPs between heifers and cows (for Nelore cattle: 16.2 and 30.9, respectively; for Girolando cattle: 18.1 and 26.7, respectively; both P < 0.05). In conclusion, OAFPs were similar between Nelore and Girolando breeds and were influenced by age. Furthermore, we observed a high correlation for individual animals between the mean numbers of follicles counted in both ovaries and total number of follicles counted in either the right or left ovary, indicating that the evaluation of a single ovary is sufficient to estimate the OAFP of an individual.

In follicular aspiration, physical aspects are of high significance for the technique to succeed, such as vacuum pressure, caliber of the needle and the way the follicular wall curettage is performed. The aim of this study was to investigate the recovery rate of equine oocytes aspirated by scraping of the follicular wall, testing different calibers of disposable needles, as well as the morphological evaluation of the cumulus oophorus complexes (COCs). Mares ovaries (n=447) obtained at a local slaughterhouse were transported to the laboratory in a thermal container (20 °C) and had the tunica albuginea and connective tissues dissected. The aspirated follicles had 10 to 25 mm in diameter, and 30x8 (21G 1 ¼) or 40x12 (18G 1 ½) needles were used for the aspiration, forming group A (G-A) and group B (G-B), respectively. In G-A and G-B, 480 and 548 follicles were aspirated, respectively. Under the stereomicroscope, the oocytes were evaluated according to the quality of the ooplasm and characteristics of the cumulus cells (grade I, II, III and denuded). The statistical analysis was performed using the Student’s t-test, logistic regression and test of proportions, and differences were considered significant when P

The objective of this study was to compare the number of follicles, oocytes and the recovery rate in sheep submitted to the one-shot protocol with or without ovarian priming with estradiol benzoate (EB). Pluriparous non-lactating sheep (n=33) with an average age of five years (range 4-6) and a body condition score of 3.0±0.3 were divided into three groups. The one-shot group (n=10) was treated with a subcutaneous implant containing 1.5 mg of norgestomet from D0 to D10. The animals in this group were administered 0.04 mg of D-cloprostenol, 200 IU of follicle stimulating hormone (FSH) and 300 IU of equine chorionic gonadotropin (eCG) on D8. Animals in the EB group (n=11) received the same treatment as one-shot plus the administration of 0.6 mg of EB on D0. In the untreated group (n=12), the animals received no hormone stimulation. The collection of the oocytes was performed by laparotomy 36 h after the administration of gonadotropins (D10). Oocytes were searched and classified based on morphology. An increase was observed (p

This study evaluated the follicular dynamics and the pregnancy rates of Nelore cows with or without a CL synchronized with FTAI employing injectable progesterone (P4). Multiparous cows (n=44), 45-65 days postpartum were allocated into a 2x2 factorial. Eleven cows with and 11 without a CL received a conventional protocol of FTAI (D0/1 g P4 + 2 mg EB; D8/500 ?g cloprostenol, 300 IU eCG and 24 h later 1 mg EB; D10/FTAI). Other 11 cows with and 11 without a CL received 350 mg of injectable P4 (300 mg sc and 50 mg im) and 2 mg of EB im on D0. On D6, 500 ?g of cloprostenol and 300 IU of eCG were administered im. After 24 h, 1 mg of EB was administered im and FTAI was performed on D8. The results were analyzed by ANOVA and Tukey or by Chi-square test (p?0.05). The presence of follicles ? 5mm on D4, follicular growth rate and CL diameter were similar between groups (50-55%, 1.1-1.2mm/ day and 16.1-17.6mm; respectively). The diameter of the dominant follicle (DF) at FTAI and ovulatory follicle (OF) were similar between groups with CL (10.5±1.8 and 11.4±1.3mm, respectively) and without CL (10.8±2.3 and 11.1±2.2mm, respectively) but device was higher than injectable (11.7±2.0 and 11.8±1.7 vs. 9.7±1.7 and 10.6±2.0mm, respectively). The ovulation and pregnancy rate were similar between groups with CL (59% and 36%, respectively) and without CL (77% and 27%, respectively) but device was higher than injectable (91% and 45% vs. 45.5% and 18%; respectively). The ciclicity did not affect the follicular dynamics and pregnancy rate. The device showed higher diameter of the DF at FTAI, OF and ovulation and pregnancy rates. However, the injectable P4 produced similar percentages of follicles ? 5 mm on D4, follicular growth rate and CL diameter, raising the possibility of further investigation.

This study aims to evaluate ovarian follicular dynamics of Nelore cows subjected to fixed-time artificial insemination (FTAI) with injectable progesterone (P4) or intravaginal device. In the device group, 20 cows received an intravaginal device with 1 g P4 and administration of 2 mg estradiol benzoate (EB) intramuscularly (im) on D0. The device was removed on D8, followed by administration of 500 ?g cloprostenol, 300 IU equine chorionic gonadotropin (eCG) and 1 mg estradiol cypionate (EC), im. The cows were inseminated 48h later. In the injectable group, 350 mg P4 (300 mg subcutaneously and 50 mg im) and 2 mg EB im were administered on D0 to 16 cows. On D6, 500 ?g cloprostenol, 300 IU eCG and 1 mg EC were administered im. The FTAI was performed 48h later. Sixty-five percent of cows from the device group (13/20) and 75% from the injectable group (12/16) showed follicles ? 5 mm on D4 (p>0.05). The average rate of follicular growth was 0.92±0.25 mm/day for the device group and 1.06±0.45 mm/day for the injectable group (p>0.05). The average diameter of the dominant follicle (DF) at FTAI was 11.4±1.4 mm for the device group and 10.8±1.8 mm for the injectable group (p>0.05). In the device group, ovulatory follicles (OF) had an average diameter of 12.1±1.4 mm, and in the injectable group 12.3±1.6 mm (p>0.05). Ninety percent of the device group (18/20) and 37.5% of the injectable group (6/16) demonstrated ovulation (p

This experiment was carried out with the objective of evaluating the effects of different levels of concentrate supplement in the pre and post-parturition periods on milk production and body score of dairy cow. 54 cows in a completely randomized design, distributed in 3 x 2 factorial arrangement, with 3 treatments, 2 periods and 9 repetitions for treatment, were used. The treatments with different levels of concentrate supplement (in dry basis) in the pre-parturition periods (PRE), were: PRE1 = 0% of the body weight (BW); PRE2 = 0.5% of the BW; PRE3 = 1.0% of the BW; and in the post-parturition (POS), were: POS1 = 1kg for each 2.5 kg of produced milk; POS2 = 1kg for each 2.0kg of produced milk and POS3 = 1kg for each 1.5kg of produced milk, above 6 kg of milk a day. The milk weight and body condition scores were evaluated every seven days. There was not significant difference among the treatments of PRÉ period for milk production, however, it was verified that the increase in the PRE supplementation, improved the average body condition of animals in post parturition period. In the treatments of POS period, there was positive and increasing effect with the increasing supplementation level, on milk production and medium body score of the animals. Improvement was not observed in the milk production with the increase of supplement in the PRE period, but in the POS period there was higher milk production with the increase supplement. There were not correlation among pre-parturition treatments and milk production, but there was correlation of these with body score to the parturition (r=0.66). There was correlation of supplement levels in the post parturition with medium body score (0.36) and with milk production (0.32). It can be concluded that the increasing supplement level in the pre-parturition period improves the body score but not the milk production of the animals. The increase of supplement in the post parturition period gets better so much the body score as the milk production. Este experimento teve como objetivo avaliar os efeitos de diferentes níveis de suplementação concentrada nos períodos pré-parto e pós-parto, sobre a produção de leite e o escore corporal de vacas leiteiras. Utilizaram-se 54 vacas em delineamento experimental inteiramente casualizado, distribuídas em esquema fatorial 3 x 2, sendo 3 tratamentos, 2 períodos e 9 repetições por tratamento. Os tratamentos compreenderam diferentes níveis de suplementação de concentrado (em base seca) nos períodos pré-parto (PRE): PRE1= 0% do PV; PRE2= 0,5% do PV; PRE3= 1,0% do PV; e no pós-parto (POS): POS1= 1kg para cada 2,5 kg de leite produzido; POS2= 1kg para cada 2,0kg de leite produzido e POS3= 1kg para cada 1,5kg de leite produzido, acima de 6 kg de leite por dia. A pesagem do leite e a avaliação de escore corporal foram realizadas a cada sete dias. Não houve diferença significativa entre os tratamentos do período PRE para produção de leite, no entanto verificou-se que o aumento na suplementação PRE melhorou a condição corporal média dos animais no período pós-parto (P < 0,05). Nos tratamentos do período POS, houve efeito positivo e crescente com o aumento do nível de suplementação, sobre a produção de leite e escore corporal médio dos animais (P < 0,05). Não se observou melhora na produção de leite com o aumento da suplementação no período PRE, mas nos tratamentos POS houve maior produção de leite com o aumento da suplementação (P < 0,05). Não houve correlação de tratamento pré-parto e produção de leite, mas houve correlação destes com escore corporal dos animais ao parto (r=0,66). Houve correlação de níveis de suplementação no pós-parto com escore corporal médio (0,36) e com produção de leite (0,32). Pode-se concluir que o aumento do nível de suplementação no período pré-parto melhora o escore corporal mas não a produção de leite dos animais, e o aumento de suplementação nos tratamentos pós-parto melhoram tanto o escore corporal quanto a produção de leite.

There are no efficient fixatives for histological analyses of ovarian tissue that permit an appropriate morphological analysis of ovary and their cells. The correct processing of the tissue is essential to avoid misinterpretation between follicular degeneration due to the physiological process of follicular atresia with signs of degeneration signals due to poor tissue fixation. This study aimed to identify the best fixative and better fixation period for morphological analysis of the functional unit of the ovary: follicle. Ovaries (n = 5) from five adult cows were collected from local abattoirs. Each ovary was divided into 12 fragments and submitted to six different fixatives in two different times (12 and 24h). It was analyzed 150 follicles per treatment. The best results were obtained with Carnoy’ and Bouin’s fixatives which had higher (P < 0.05) proportion of viable follicles after fixation for 12 or 24h (88 and 83% for Carnoy’s fixative vs. 79 e 73% for Bouin). It was concluded that Carnoy and Bouin fixatives are the best options to preserve for up to 24 h the morphology of preantral follicles in bovine ovaries.Na rotina de avaliação histológica, os fixadores mais adequados para o tecido ovariano para que se obtenha uma adequada análise morfológica do ovário e de suas células ainda não foram determinados. O correto processamento do tecido é fundamental para evitar erros de interpretação, como confundir degeneração folicular devido ao processo fisiológico de atresia folicular com sinais de degeneração devido à má fixação do tecido. Este estudo teve como objetivo identificar qual o melhor fixador e melhor período de fixação para análise morfológica da unidade funcional do ovário: o folículo. Para isso foram utilizados ovários (n=5) de cinco vacas adultas. Cada ovário foi divido em 12 fragmentos e submetidos a seis diferentes fixadores em dois tempos (12 e 24h). Foram analisados 150 folículos ovarianos por tratamento. Os melhores resultados foram obtidos com os fixadores Carnoy e Bouin, que apresentaram maior (P < 0,05) proporção de folículos viáveis após fixação por 12 ou 24h (88 e 83% para o fixador Carnoy vs. 79 e 73% para Bouin respectivamente). Conclui-se neste estudo que os fixadores Carnoy e Bouin são as melhores opções para preservar por até 24 h a morfologia de folículos pré-antrais em ovários bovinos.

In vitro embryo production is an important technique for facilitating the reproduction of animals with high genetic merit. The greatest challenge for the reproducibility of this technique is the quality of the oocyte that is submitted for in vitro maturation. The aim of this work was to evaluate the ultrastructural characteristics of oocytes from follicles of different diameters using transmission electron microscopy. The animals were divided into 2 groups and were given a single i.m. injection of 250 IU FSH (Pluset, Serono, Italy). To synchronize the follicular growth, all follicles > 2 mm were aspirated during the estrous cycle, which was considered day zero (D0). Group 1 (G1; n = 4) received FSH on day 1 (D1), and the 2- to 5-mm follicles were aspirated on day 2 (D2). The animals in group 2 (G2; n = 5) received FSH on day 2 (D2), and their 10- to 15-mm follicles were aspirated on day 5 (D5). After aspiration, the oocytes from both groups were fixed and prepared for ultrastructural analysis. The oocytes analyzed from both groups had similar ultrastructural characteristics. The presence and distribution of organelles in the cytoplasm of the oocytes did not differ between groups, suggesting that, in relation to the ultrastructural characteristics, oocytes from 2 to 5 mm and 10 to 15 mm follicles are similar.A produção in vitro de embriões é uma técnica importante para facilitar a reprodução de animais com elevado mérito genético. O maior desafio para a reprodutibilidade desta técnica é a qualidade do oócito destinado à maturação in vitro. O objetivo deste trabalho foi avaliar as características ultra-estruturais de oócitos provenientes de folículos com diferentes diâmetros por microscopia eletrônica de transmissão. Os animais foram divididos em dois grupos e receberam uma única injeção im de 250 UI de FSH (Pluset, Serono, Itália). Para sincronizar o crescimento folicular, todos os folículos > 2 mm foram aspirados durante o ciclo estral, que foi considerado o dia zero (D0). O Grupo 1 (G1, n = 4) recebeu FSH no dia 1 (D1), e os folículos de 2 a 5 mm foram aspirados no dia 2 (D2). Os animais do grupo 2 (G2, n= 5) receberam FSH no dia 2 (D2), e os folículos de 10 a 15 mm foram aspirados no dia 5 (D5). Após a aspiração, os oócitos de ambos os grupos foram fixados e preparados para análise ultra-estrutural. Os oócitos analisados de ambos os grupos apresentaram características ultra-estruturais semelhantes. A presença e distribuição de organelas no citoplasma dos oócitos não diferiram entre os grupos, sugerindo que, em relação às características ultra-estruturais, oócitos oriundos de folículos de 2-5 mm e 10-15 mm são similares.

The aim of this study was to evaluate two pre-aspiration protocols associated with PGF2a. Nelore heifers (n=14), not pregnant, body score of 2.5 (scale of 1 to 5) were used. All animals were submitted to two treatments (cross over) to control the follicular wave of growth, one by follicular ablation 3 days before the aspiration (PAF) and another by hormonal control with estrogen and injectable progesterone (PCH). Data were analyzed by ANOVA and significant differences were compared by Tukey test, with significance level of 5%. The hormonal control protocol (PCH) provided advantages as higher average of follicles and oocytes, reduction in the number of corpus luteum and one procedure less – the follicle ablation. We concluded that the hormone control protocol (PCH) is a viable alternative for pre-aspiration treatments, with additional benefits for the absence of corpus luteum at the time of follicular aspiration. O objetivo deste estudo foi comparar dois protocolos pré-aspiração folicular associados com PGF2a. Foram utilizadas 14 novilhas da raça Nelore, não gestantes, com escore corporal de 2,5 (escala de 1 a 5). Todos os animais foram submetidos aos dois tratamentos (cross over) para controle da onda de crescimento folicular, um através da ablação de todos os folículos 3 dias antes da aspiração folicular (PAF) e outro por controle hormonal (PCH) com benzoato de estradiol e progesterona injetável. Os dados foram analisados pelo ANOVA e as diferenças significativas pelo teste de Tukey, com nível de significância de 5 %. O protocolo controle hormonal (PCH) mostrou-se vantajoso por propiciar maior média de folículos e oócitos, além de redução no número de corpos lúteos evitando também o procedimento extra de aspiração prévia para ablação dos folículos. Conclui-se que o protocolo hormonal (PCH) apresentado mostra-se uma alternativa viável para tratamentos pré-aspiração, havendo benefícios adicionais pela ausência de corpo lúteo no momento da aspiração folicular.

The objective of the present study was to evaluate the effect of a second uterine flushing on number of recovered embryos. Cyclic Nelore cows (n= ) were used as donors. Animals were kept under grazing conditions in a farm at Mamborê, Paraná state. Cows were submitted to two sessions of multiple ovulation 80 days apart. At a random stage of estrus cycle (Day 0) donors received an intravaginal device with 1.9 g of progesterone (CIDR; Pfizer; Brasil). Twenty four hours later (Day 1) cows were injected with 2.5 mg/ IM of Estradiol Benzoate (Estrogin, Farmavet, Brasil). Between Day 5 and Day 9, donors received decreasing doses of 250 UI of FSH (Pluset; Calier, Brasil). On Days 7 and cows received 25 mg/ IM of dinoprost (Lutalyse, Pfizer, Brasil) and progesterone implants were removed 12 hs later. Artificial insemination was performed 12 and 24 hours after estrus detection. The first embryo recovery was performed 7 days after AI with the Foley catheter positioned in the uterine body. A total of 1.000 mL of PBS (Embriocare, Cultilab, Brasil) was used in a closed system of uterine flushing, with a 70 µm filter. Recovered uterine fluid was evaluated under a stereomicroscope (SMZ 654, Nikon, Japan) and embryos were classified accord to IETS (1998). Twenty four hours later, the second uterine flushing was performed following the same procedure. Results were analyzed by t-Student test (p < 0,0 ). After 12 superovulation procedures, we obtained a total of 131 ova/embryo, being 91.4% (120/131) from the first uterine flushing and 8.4% (11/131) from the second one. We conclude that the second uterine flushing provided a small improvement on the number of recovered embryos. The feasibility of this method is restricted to special conditions, when the value of the embryos supports the additional work.O objetivo do presente experimento foi verificar a viabilidade de uma segunda lavagem uterina sobre a recuperação de embriões. Doadoras cíclicas da raça Nelore (n= ) foram mantidas a pasto no município de Mamborê, Estado do Paraná. Os animais foram submetidos a tratamentos de superovulação com intervalo de 80 dias. Em um dia aleatório do ciclo estral (Dia 0) as doadoras receberam um dispositivo intravaginal contendo 1,9 g de progesterona (CIDR; Pfizer, Brasil). Vinte e quatro horas após (Dia 1) administrou-se , mg de benzoato de Estradiol (Estrogin; Farmavet, Brasil) por via intramuscular (IM). Entre Dia e Dia 9 realizou-se a superovulação com 0 UI de FSH IM em doses decrescentes (Pluset; Calier, Brasil). Administrou-se 25 mg de dinoprost IM (Lutalyse; Pfizer, Brasil) na manhã do D7 e 12 hs depois, simultaneamente à retirada do dispositivo de progesterona. As vacas foram inseminadas e horas após o início das manifestações estrais. A primeira lavagem uterina foi realizada no sétimo dia após a inseminação artificial, com o posicionamento das sondas de Foley caudal à bifurcação do útero. Em cada lavagem, foram usados .000 ml de DPBS (Embriocare; Cultilab, Brasil) em sistema fechado conectado a um filtro coletor de 70 µm. O lavado uterino foi avaliado com auxílio de um estereomicroscópio binocular (SMZ 654, Nikon, Japão) e os embriões foram classificados seguindo os padrões da IETS (1998). Vinte e quatro horas após, a segunda lavagem uterina foi realizada seguindo o mesmo procedimento. Os resultados foram analisados pelo teste t-Student com significância de 5%. Após 12 superovulações, foram obtidas 131 estruturas sendo 91,4% (120/131) na primeira lavagem e 8,4% (11/131) no segundo procedimento. A segunda lavagem uterina permitiu a recuperação de um número reduzido de embriões, condicionando o procedimento a embriões cujo valor justifique o acréscimo de trabalho.

This experiment was carried out with the objective of evaluating the effects of different levels of concentrate supplement in the pre and post-parturition periods on the reproductive performance of dairy cow. 54 cows in a completely randomized design, distributed in x factorial arrangement, being treatments, periods and 9 repetitions for treatment, were used. The treatments with different levels of concentrate supplement (in dry basis) in the pre-parturition periods (PRE), were: PRE1 = 0% of body weight (BW); PRE = 0.5% of BW; PRE = 1.0% of BW; and in the post-parturition (POS), were: POS1 = 1kg for each .5 kg of produced milk; POS = 1kg for each .0kg of produced milk and POS = 1kg for each 1.5kg of produced milk, above 6 kg of milk a day. The ultrasonography evaluation and body score were evaluated every seven days. 0 animals presented estrus and 1 animals became pregnant in the evaluated period. The average interval between parturition-first estrus was of 53.13 days, varying from 19 to 105 days. The amount of animals that entered in the estrus in the 1 0 days of evaluation increased with the post parturition supplementation level, being of , 4 and 7 animals for the treatments 1, and , respectively. For the conception, the only treatment in that all the animals became pregnant, was the treatment PRE and POS . There was higher number of conceptions among the animals that received treatment and , so much in the pre as in the post-parturition, in relation to animals that received the treatment 1. The increasing supplement level in the treatments pre and postparturition caused lineal decrease of the number of weeks for beginning of follicular activity (BFA) and beginning of follicular growth (BFG). There was negative correlation among BFA (-0. 4), BFG (-0. 1) and body score to the parturition. It can be concluded that the increasing supplement level in the pre and post-parturition treatments, increases the amount of animals that manifest estrus and become pregnant in the first 120 days post-parturition, as well as, in the pregnant maintenance. Lineal decrease occurred in the time for BFA and BFG as increases the supplement level in the pre and post- parturition, being these correlated with the body score to the parturition.Este experimento teve como objetivo avaliar os efeitos de diferentes níveis de suplementação concentrada nos períodos pré-parto e pós-parto, sobre o desempenho reprodutivo de vacas leiteiras mantidas a pasto. Utilizaram-se 54 vacas em delineamento experimental inteiramente casualizado, distribuídas em esquema fatorial 3x2 , sendo 3 tratamentos, 2 períodos e 9 repetições por tratamento. Os tratamentos compreenderam diferentes níveis de suplementação de concentrado (em base seca) nos períodos pré-parto (PRE): PRE1= 0% do PV; PRE = 0,5% do PV; PRE = 1,0% do PV; e no pós-parto (POS): POS1= 1kg para cada 2,5 kg de leite produzido; POS = 1kg para cada 2,0kg de leite produzido e POS = 1kg para cada 1,5kg de leite produzido, acima de 6 kg de leite por dia. Os animais foram submetidos à avaliação ultrasonográfica e de escore corporal a cada 7 dias. No período avaliado, 20 animais apresentaram cio e 13 animais conceberam. A média de intervalo entre parto-primeiro cio foi de 53 ,13 dias (p < 0,05). Com o aumento do nível de suplementação pós-parto, aumentou-se o número de animais que entraram em cio, sendo de 2, 4 e 7 animais para os tratamentos 1,2 e 3, respectivamente. O único tratamento em que todos os animais conceberam, foi o tratamento PRE e POS . Houve maior número de concepções entre os animais que receberam tratamento 2 e3 , tanto no pré quanto no pósparto, em relação aos que receberam o tratamento 1 (p < 0,05). O aumento do nível de suplementação nos tratamentos pré e pós-parto, provocaram diminuição linear do número de semanas para início de atividade folicular (IAF) e início de crescimento folicular (ICF). Houve correlação negativa entre o IAF (-0, 4), ICF (-0, 1) e escore corporal ao parto. Conclui-se que, o aumento no nível de suplementação tanto no pré quanto no pós-parto, aumenta a quantidade de animais que manifestam cio e concebem nos primeiros 120 dias pós-parto, assim como na manutenção da gestação. Ocorre diminuição linear no tempo para IAF e ICF à medida que se aumenta o nível de suplementação no pré e pós-parto, sendo estes correlacionados com o escore corporal ao parto.