Recent advances in early pregnancy loss diagnosis in dairy cows: New approaches.

Early pregnancy loss is a primary cause of low reproductive rates in dairy cows, posing severe economic losses to dairy farming. The accurate diagnosis of dairy cows with early pregnancy loss allows for oestrus synchronization, shortening day open, and increasing the overall conception rate of the herd. Several techniques are available for detecting early pregnancy loss in dairy cows, including rectal ultrasound, circulating blood progesterone, and pregnancy-associated glycoproteins (PAGs). Yet, there is a need to improve on existing techniques and develop novel strategies to identify cows with early pregnancy loss accurately. This manuscript reviews the applications of rectal ultrasound, circulating blood progesterone concentration, and PAGs in the diagnosis of pregnancy loss in dairy cows. The manuscript also discusses the recent progress of new technologies, including colour Doppler ultrasound (CDUS), interferon tau-induced genes (ISGs), and exosomal miRNA in diagnosing pregnancy loss in dairy cows. This study will provide an option for producers to re-breed cows with pregnancy loss, thereby reducing the calving interval and economic costs. Meanwhile, this manuscript might also act as a reference for exploring more economical and precise diagnostic technologies for early pregnancy loss in dairy cows.

The effectiveness of low-dose of eCG in timed AI Nelore (Bos indicus) heifers.

This study evaluated the efficacy of the administration of different doses of equine chorionic gonadotropin (eCG; 0 IU, 200 IU, or 300 IU) at the time of the progesterone device removal in 2-year-old Nelore (Bos indicus) heifers synchronized for fixed-timed artificial insemination (FTAI). On day 0 (D0), a total of 398 heifers received 2 mg of oestradiol benzoate i.m., 0.53 mg of cloprostenol i.m., and an eight-day previously used (second use) intravaginal device containing 1 g of progesterone (P4). Eight days later (D8), simultaneous with the P4 device removal, 0.5 mg of oestradiol cypionate i.m. and 0.53 mg of cloprostenol i.m. were administered. At the same time, heifers were randomly assigned to receive one of the following treatments: G-0 IU (n = 141; no eCG treatment), G-200 IU (n = 132; treated with 200 IU of eCG), and G-300 IU (n = 125; treated with 300 IU of eCG). FTAI was performed 48 h after the P4 device removal (D10). Ultrasonographic evaluations were performed at D0, D10, and D17. Heifers were scanned to measure the size of the largest follicle (LF), the presence, number, and size of the corpus luteum (CL), and the ovulation rate. Subsequently, at D40, the heifers underwent scanning to determine the pregnancy rate and identify any twin pregnancies. Additionally, at D70, scans were performed to assess pregnancy loss (PG). Data were analysed by orthogonal contrasts [C1 (eCG effect): control x (200 IU + 300 IU) and C2 (eCG dose effect): 200 IU × 300 IU]. On D0, CL presence was similar between the groups [G-0 IU = 65.2% (92/141), G-200 IU = 55.3% (73/132), and G-300 IU = 63.2% (79/125); p = .16]. No interactions between the presence of CL on D0 and eCG treatment were found for any of the variables (p > .05). The diameter of the LF at FTAI (D10) was not influenced by eCG treatment (p = .22) or eCG dose (p = .18). However, treatment with eCG increased the diameter of the CL at D17 (G-0 IU = 15.7 ± 0.3 mmb , G-200 IU = 16.6 ± 0.2 mma , and G-300 IU = 16.6 ± 0.3 mma ; p = .001), regardless of the dose used (p = .94). The ovulation rate was higher in heifers treated with eCG [G-0 IU = 79.4%b (112/141), G-200 IU = 90.2%a (119/132), and G-300 IU = 93.6%a (117/125); p = .002], but there was no effect of eCG dose (p = .36). Pregnancy per AI (P/AI) on D40 [G-0 IU = 32.6%b (46/141), G-200 IU = 42.4%a (56/132), and G-300 IU = 42.4%a (53/125); P = 0.05] and D70 [G-0 IU = 29.1%b (41/141), G-200 IU = 40.9%a (54/132), and G-300 IU = 40.8%a (51/125); p = .02] were higher on heifers that received eCG; however, no dose effect was observed for P/AI on D40 (p = .89) nor D70 (p = .98). Pregnancy loss between D40 and D70 tended to reduce (p = .07) in eCG-treated heifers without dose effect (p = .91). Heifers with CL at D0 presented a greater follicle diameter (LF) on D10 (With CL = 11.2 ± 0.2 mm and Without CL = 10.2 ± 0.2 mm; p = .05), CL diameter on D17 (With CL = 15.8 ± 0.03 mm and Without CL = 11.8 ± 0.6 mm; p = .01), and ovulation rate [With CL = 95.5% (233/244) and Without CL = 74.7% (115/154); p = .01]. However, no difference in pregnancy rate at D40 (p = .52) and D70 (p = .84) was found. In conclusion, eCG treatment increases ovulation and pregnancy rates of heifers submitted to a FTAI protocol. Furthermore, eCG treatment increases the diameter of the CL after FTAI and reduces pregnancy losses. No dose effect was observed, suggesting Nelore (Bos indicus) heifers respond to 200 IU of eCG treatment for FTAI.

Optimizing ReBreed21 I: Evaluation of endocrine and ovarian dynamics in non-bred Bos indicus heifers.

The present study evaluated follicular and endocrine dynamics during ReBreed21, a reproductive strategy that allows resynchronization of ovulation every 21 days in Bos indicus (Nelore) heifers. A synchronized estrous cycle was induced using a standard timed ovulation protocol (d -10: P4 implant inserted + 2 mg estradiol benzoate; d -2: P4 removed+ 0.5 mg cloprostenol + 0.6 mg estradiol cypionate + 200 IU equine chorionic gonadotropin (eCG); d0: 8.4 µg buserelin) without AI to ensure nonpregnancy in heifers. Day of GnRH was designated d0 of estrous cycle. On d12, heifers (n = 80) were randomized into three experimental groups: (1) ReBreed21 (n = 28) d12 P4 device inserted, d19 P4 device withdrawal plus 200 IU eCG, and d21 8.4 µg buserelin (GnRH); (2) ReBreed21+G (n = 26) same as ReBreed21 plus GnRH (16.8 µg) treatment on d12; and (3) Control (n = 26) no treatment. ReBreed21+G increased two-fold (62.9%; 18/26) percentage of heifers with synchronized follicular wave emergence compared to Control (34.6%; 9/26) whereas ReBreed21 (53.6%; 15/28) was intermediate. The ReBreeed21 groups (eCG on d19) increased (P < 0.01) follicular growth between d19 and d21 in ReBreed21 (2.3 ± 0.2 mm) and ReBreed21+G (3.4 ± 0.2 mm) compared with Control (1.2 ± 0.3 mm), resulting in greater (P < 0.01) follicle diameter on d21 for ReBreed21 (10.7 ± 0.4 mm) and ReBreed21+G (10.8 ± 0.4 mm) compared with Control (9.1 ± 0.5 mm). Structural luteolysis was similar among groups (P = 0.51), although the average day when P4 was <1 ng/mL was later (P < 0.01) for ReBreed21 (20.5 ± 0.2) and ReBreed21+G (20.7 ± 0.2) compared to Control (19.2 ± 0.4). Overall ovulation at the end of the estrous cycle was increased (P = 0.03) for ReBreed21 groups (83.3%; 45/54) compared with Control (57.7%; 15/26). Synchronized ovulation on day 22-23 was greater (P < 0.01) for ReBreed21 (78.6%; 22/28) and ReBreed21+G (76.9%; 20/26) compared with Control (30.8%; 8/26). Thus, the ReBreed21 resynchronization program produced acceptable endocrine and follicular dynamics, including synchronized ovulation at the end of the protocol in nonpregnant heifers providing good rationale for testing the fertility and practical implementation of this protocol under field conditions.

Ovarian lavage / artificial insemination method as an alternative approach for controlled reproduction of the northern pike (Esox lucius).

This study aimed to compare the effectiveness of the ovarian lavage / artificial insemination method with the traditional hormonal administration and fertilization methods over the artificial reproduction of the northern pike (Esox lucius). For this purpose, groups of five females were treated as follows: intraperitoneal injection of saline (C1); ovarian lavage with saline (C2); intraperitoneal injection of carp pituitary extract (CPE, T1); ovarian lavage with CPE (T2); intraperitoneal injection of CPE and ovarian lavage with semen after 72 h (T3); ovarian lavage with CPE and ovarian lavage with semen after 72 h (T4). According to the results, no fish ovulated in the control groups (C1 and C2). There were no significant differences (n.s.) among experimental treatments (P > 0.05, n.s.) in the reproductive parameters, such as latency time, ovulation rate, stripped egg amount, and pseudo-gonadosomatic index. The lowest fertilization rate (54.8%) was observed in the T4 treatment and significantly differed from the T1 and T2 treatments (P < 0.05). Moreover, the highest survival at swim-up stage was measured in the T4 treatment with a significant difference compared to the T1 group (P < 0.05). The survival at the eyed-egg stage, hatching rate, and malformations were similar (P > 0.05, n.s.) in all applied strategies. The results demonstrated that the ovarian lavage / artificial insemination method could be applied to control northern pike reproduction, like the traditional fertilization method. Consequently, this novel technique can be suggested as an alternative strategy to facilitate the hatchery operations in the controlled reproduction of this species.

Effect of follicle size on pregnancy outcomes in patients undergoing first letrozole-intrauterine insemination.

BACKGROUND: Letrozole has been proven to be an effective method for inducing ovulation. However, little attention has been paid to whether the lead follicle size will affect the success rate of intrauterine insemination (IUI) with ovulation induction with alone letrozole. Therefore, we hope to investigate the effect of dominant follicle size on pregnancy outcomes on human chorionic gonadotropin (hCG) day of the first letrozole-IUI. METHODS: A retrospective cohort study design was employed. We included patients with anovulation or unexplained infertility undergoing first IUI treatment with letrozole for ovarian stimulation. According to the dominant follicle size measured on the day of hCG trigger, patients were divided into six groups (≤ 18 mm, 18.1-19.0 mm, 19.1-20.0 mm, 20.1-21.0 mm, 21.1-22.0 mm, > 22 mm). Logistic models were used for estimating the odds ratios (ORs) with their 95% confidence interval (CIs) for achieving a clinical pregnancy or a live birth. A restricted cubic spline was drawn to explore the nonlinear relationship between follicle size and IUI outcomes. RESULTS: A total of 763 patients underwent first letrozole-IUI cycles in our study. Fisher exact test showed significant differences among the six follicle-size groups in the rates of pregnancy, clinical pregnancy and live birth (P < 0.05 in each group). After adjusting the potential confounding factors, compared with the follicles ≤ 18 mm in diameter group, 19.1-20.0 mm, 20.1-21.0 mm groups were 2.3 or 2.56 times more likely to get live birth [adjusted OR = 2.34, 95%CI (1.25-4.39); adjusted OR = 2.56, 95% CI (1.30-5.06)]. A restricted cubic spline showed an inverted U-shaped relationship between the size of dominant follicles and pregnancy rate, clinical pregnancy rate, and live birth rate, and the optimal follicle size range on the day of hCG trigger was 19.1-21.0 mm. When the E2 level on the day of hCG trigger was low than 200 pg/mL, the clinical pregnancy rates of 19.1-20.0 mm, 20.1-21.0 mm groups were still the highest. CONCLUSIONS: The optimal dominant follicle size was between 19.1 and 21.0 mm in hCG-triggered letrozole-IUI cycles. Either too large or too small follicles may lead to a decrease in pregnancy rate. Using follicle size as a predicator of pregnancy outcomes is more meaningful when estrogen on the day of hCG trigger is less than 200 pg/ml.

Sperm binding to oviduct epithelial spheroids varies among males and ejaculates but not among females in pigs.

The elimination of ejaculates and males with low fertility despite good sperm motility and morphology is crucial to maintain high pregnancy rates after artificial insemination (AI) in farm animals. The ability of sperm to survive in the female tract is particularly crucial in pigs due to the large variation in the timing between AI and ovulation and the high number of oocytes to fertilise. The objective of this study was to characterise a new in vitro model of oviduct sperm reservoir using porcine oviduct epithelial spheroids (OES) and to assess the variability in sperm binding to OES among gilts, boars and their ejaculates. Isthmic mucosa fragments were collected from gilt oviducts at a slaughterhouse, and after 48 h of culture, the OES that had spontaneously formed were sorted according to their vesicle shape and size (150-200 µm in diameter) for characterisation and sperm binding assays. The OES contained viable, cytokeratin-positive and vimentin-negative cells, of which 36.4 ± 2.0% were multiciliated. The average proportion of multiciliated cells per OES did not change among culture replicates. After co-incubation with boar fresh semen, only sperm of normal morphology were found to bind, by their head, to cilia of OES. The density of sperm bound to the OES surface increased linearly with sperm concentration. The bound sperm density on OES was used to assess the binding capacity of fresh ejaculates collected from Pietrain boars. For a given ejaculate, the bound sperm density did not vary among pools of OES female donors. The analysis of five successive ejaculates from nine boars indicated significant differences in bound sperm densities on the OES among individual boars and their ejaculates (P < 0.01). There was no correlation between the sperm bound density and sperm parameters measured by computer-assisted sperm analysis or the initial dilution of the ejaculate. In conclusion, the OES characterised in this study offered physiological conditions to study sperm binding to the isthmic reservoir and evidenced that sperm from different ejaculates and different boars vary in their ability to bind to these oviduct spheroids despite homogeneous motility and morphology.

The Effect of Reduced GnRH Dose on Ovulation and Follicular Dynamics in Ovsynch Programme of Pure and Bunaji-Crossbred Cows.

Our objective was to compare the efficacy of reducing GnRH dose from 100 µg to 50 µg on the formation of ovulation and sizes of ovarian structures following Ovsynch in apparently healthy Bunaji and Friesian × Bunaji Cows. Thirty female multiparous-apparently-healthy adult [Bunaji (n = 15) and Friesian × Bunaji (n = 15)] breeds of cattle were used. Five cows each were allocated randomly to three groups [control; full dose (FD), and half dose (HD)]. Cows in the control group were treated with 2 ml normal saline while FD-group received 100 µg lecirelin on day 0, with 500 µg clorprostenol on day 7 and with 100 µg lecirelin two days later. Furthermore, HD-group received the same treatment as FD-cows but the dose of lecirelin was reduced to 50 µg at both times of GnRH administration. Ovarian structures were monitored by ultrasound with a 5-MHZ linear transrectal probe on days - 1 to 12. The ovarian responses of the various groups to first GnRH administration showed (0%, 40% and 60%) ovulation rate for C, HD and FD groups respectively in Bunaji breeds while in Friesian × Bunaji, it was (0%, 60%, 60%). Following second GnRH administration ovulation rate for Bunaji was (20%, 60%, and 60%) for Control, HD and FD-groups, respectively, while for Friesian × Bunaji cows it was (20%, 60%, and 80%). There was no significant difference (p > 0.05) in the days of new follicular wave emergence following the first GnRH administration. It was concluded that 50 µg Lecirelin reduced the cost of drug without affecting the efficiency of Ovsynch protocol.

Estrogens improve the pregnancy rate in cattle: A review and meta-analysis.

Estrogens have proven to be effective in bovine estrus induction protocols. Considering the extensive use of these products in large-scale estrus synchronization, the primary objective of the present study was to assess their effects on pregnancy rate (PR) using a meta-analysis approach. A total of 797 papers were screened from three major databases (PubMed, Web of Science, Scopus). Sixty-one studies were eligible for inclusion in the meta-analysis. The pregnancy status (success or failure) at 30 days post-insemination was considered as the effect size data. The odds ratios (OR) of PR were evaluated by considering the effects of estrogens in groups with or without estrogen intervention. The impact of estrogen (including factors such as type, dose, and time of administration) and animal characteristics (such as breed, type, and parity) was taken into account when assessing the effectiveness of estrogen response as PR. The results showed an OR of 1.25 (95% CI: 1.15-1.36; P = 0.000) for PR in animals that received estrogen compared to cattle that did not receive estrogen. Estradiol benzoate (OR = 1.3) and estradiol cypionate (OR = 1.2), with doses ranging from 1 to 3 mg (OR = 1.13-1.7), significantly increased the OR of PR. In terms of PR, beef cattle exhibited a higher odds ratio (OR = 1.4; P = 0.000) compared to dairy cattle (OR = 1.1; P = 0.09). The administration of estrogens in the estrus synchronization protocol significantly improved PR in both artificial insemination (OR = 1.2; P = 0.000) and embryo transfer (OR = 1.3; P = 0.033) programs. In summary, incorporating estrogens into estrus induction protocols led to an enhancement of the OR of PR among cattle.

Antibiotics in semen extenders - a multiplicity of paradoxes.

Addition of antibiotics to semen extenders was taken for granted for many years, from the time that commercial artificial insemination in livestock first began many decades ago. However, there is now a growing realisation that this non-therapeutic utilisation of antibacterial agents is contrary to current recommendations for prudent use that medical and veterinary professionals are advised to follow. Furthermore, antibiotics are not benign, having negative effects on sperm samples, the inseminated female, personnel and potentially the environment. The purpose of this review is three-fold: to highlight the fact that antibiotics are used in semen extenders, with the result that considerable amounts are used globally in animal breeding, to review recent studies on the negative aspects of using antibiotics for this purpose, and to look at possible alternatives. Recent changes in the legislation regarding semen extenders occurred in some, but not all, countries, leaving question marks for semen producers as to whether antibiotics should be added to semen extenders or not.

Magnitude and persistence of higher estrus-associated temperatures in beef heifers and suckled cows.

Higher estrus-associated temperatures (HEAT) are a hallmark feature in sexually active females. The overarching aim of this study was to characterize the variability, magnitude, and persistence of HEAT in heifers and suckled beef cows as well as identify associated factors when occurring during thermoneutral conditions at the onset of the spring breeding season. In both heifers and cows, estrus was induced using a 7-d controlled internal drug release (CIDR)-PGF2α protocol. Vaginal temperature after prostaglandin F2α administration was recorded every 5 min using a Thermochron iButton affixed to a blank CIDR (containing no progesterone). Estrus was defined as when a heifer first stood to be mounted or when a cow had an Estrotect patch score of 3 or 4. Level of HEAT varied among individual animals. When comparing common HEAT variables using a mixed model with date nested within a year, maximum HEAT (39.9 ±â€…0.1 and 40.0 ±â€…0.1 °C) and duration (15.5 ±â€…0.8 and 15.4 ±â€…0.7) were similar in heifers and cows, respectively. However, the magnitude and persistence of HEAT differed. Total area under the HEAT curve was 117.1 ±â€…13.5 and 158.7 ±â€…12.3 for heifers vs cows, respectively (P = 0.0571). Further, 42.9% of heifers and 49% of cows had maximum HEAT ≥ 40 °C which persisted up to 6.5 and 10 h, respectively. When ambient conditions were predominantly thermoneutral, temperature humidity index had minimal impact on HEAT (mixed model, repeated measures over time). Toward identifying associated factors with different aspects of HEAT using best fit hierarchical linear regression models, baseline vaginal temperature and baseline duration were the most highly associated independent variables. Follicle size, estradiol and progesterone levels, and other available animal-related variables (e.g., age, weight, hair coat score) explained only a small amount of variation in HEAT. In summary, level of HEAT varies in estrus females even under thermoneutral conditions. Because HEAT can persist for an extended time, direct effects on fertility important components are unavoidable. Whether HEAT is a good or bad component of the periovulatory microenvironment is the basis of ongoing and future studies.

When striving for a pregnancy, estrus is a critically important event. Higher estrus-associated temperatures (HEAT) are a hallmark feature in sexually active females. The importance of HEAT for pregnancy, however, remains unclear. Toward filling this critical knowledge gap, efforts described in the current study focused on examining variability of HEAT in individual animals, 2) defining the magnitude and persistence of HEAT, 3) identifying HEAT-associated factors, and 4) examining the similarity of HEAT between heifers and suckled beef cows when occurring at the onset of a spring breeding season. Although the magnitude and persistence of HEAT varied, 42.9% of heifers and 49% of cows reached temperatures ≥ 40 °C which in some cases persisted up to 6.5 and 10 h, respectively. When attempting to identify factors that could explain why some females exhibiting estrus remained hot for an extended time, available animal and environmental data contributed little. Even so, because HEAT can persist for an extended time, direct effects on fertility important components are unavoidable. Whether too much HEAT is good or bad for pregnancy is the basis of ongoing and future studies.

Economic viability of using OvSynch and fixed timed artificial insemination protocol in breeding improvement of pastoral herds in the rangelands.

Though using Assisted Reproductive Technologies (ARTs) can improve oestrus detection, conception and pregnancy success, thus benefit breeding program implementation, empirical evidence of their economic viability is lacking to inform investment decisions in pastoral herds. This study assessed economic viability of using OvSynch and fixed Timed Artificial Insemination (TAI) protocol in Sahiwal upgrading breeding program under two hypothetical cases of best and worst in activity-based money allocations when pastoral herds deploy either optimal or low input husbandry practices. From herd owners' assessment of milk production, best-case scenarios attain on average 10 L/cow/day with optimal husbandry and 5 L/cow/day with low input husbandry. The worst-case scenarios attain 5 L/cow/day with optimal husbandry and 1 L/cow/day with low input husbandry. Benefit- Cost Analysis (BCA) estimated Net Present Value (NPV), Benefit-Cost ratio (BCR) and Internal Rate of Return (IRR) to establish economic viability of using OvSynch and TAI Protocol in pastoral breeding programs. Both best-case scenarios retuned positive NPVs (82,028 and 6,912), BCR values (1.68 and 1.08) and IRR (27.46% and 8.08%) while worst-case scenarios returned negative NPVs (-135,855 and -141,025), BCR values of below 1 (0.87 and 0.66) and IRR values below the minimum rate of returns. These economic parameters were sensitive to price changes in inputs and outputs, under both optimal and low input husbandry practices. Results indicate that using OvSynch and TAI Protocol is a profitable and economically viable investment under optimal husbandry practices but not under low input husbandry practices. By implications, use of OvSynch and TAI Protocol in Sahiwal upgrading breeding programs need be accompanied with improved husbandry practices and de-risking pastoral herd owners from price changes in input and output markets.

Supplementation with long-acting injectable progesterone 3 days after TAI impaired luteal function in buffaloes.

Three experiments were conducted to evaluate the effects of long-acting injectable progesterone (iP4) in buffalo cows. In Experiment 1, ovariectomized buffaloes received 300 mg (iP300) or 600 mg (iP600) of iP4, and serum P4 concentrations were evaluated. In experiment 2, three groups were compared: control or administration of 300 mg of iP4 3 (iP4-D3) or 6 days (iP4-D6) after timed artificial insemination (TAI). On day 16, reproductive tract was recovered for conceptus, endometrium, and corpus luteum (CL) analysis. In experiment 3, pregnancy per AI (P/TAI) and proportion of pregnancy losses were evaluated after administration of 300 mg of iP4 3 (iP4-D3) or 6 days (iP4-D6) after TAI in lactating buffaloes. In experiment 1, serum P4 concentrations remained over 1 ng/mL for ~ 3 days in both groups. The 300 mg dose was used in subsequent experiments. In experiment 2, CL weight and endometrial glands density were decreased, and conceptus length was increased in iP4-D3 compared to control and to iP4-D6 (P < 0.05). Transcript abundance of Prostaglandin F Receptor (FP) and ISG15 in CL and of ISG15 and MX1 in endometrium was greater in iP4-D3 when compared to control and to iP4-D6 (P < 0.05). In experiment 3, there was no difference among experimental groups for P/TAI at D30 and pregnancy losses (P > 0.1); however, iP4-D3 presented a lower P/TAI at day 60 (41.7%) when compared to control (56.8%) and iP4-D6 (57.7%; P = 0.07). In conclusion, administration iP4 at 3 days after TAI affects CL development and consequently decreases final pregnancy outcome in buffaloes.

Artificial insemination of all ejaculated sperm fractions accelerates embryo development and increases the uterine vascularity in the pig.

The semen of boar is characterized by ejaculation in well-differentiated fractions with specific concentration, composition, and volume. The 'sperm-rich fraction' (SRF), the most concentrated seminal fraction, is habitually collected in insemination centers to make artificial insemination (AI) doses. The absence of the other fractions in AI doses could alter the uterine reaction to AI and not trigger essential responses that could maximize fertility. Thus, there is an urge to ascertain the impact of different ejaculate fractions on the uterus after AI to optimize the semen doses. This work analyzed specific parameters related to fertility in pregnant artificially inseminated sows (n = 15) with ac-cumulative fractions of the semen of boars (n = 6): F1, composed of the sperm-rich fraction (SRF); F2, composed of F1 plus the intermediate fraction; F3, composed of F2 plus the post-SRF. Non-inseminated sows (n = 5) were included as control (C). The different types of seminal dose did not affect the number of ovulated follicles (CL; corpora lutea, p > 0.05) but did affect the embryo development (p < 0.05). The proportion of embryos in morula stages was significantly higher in AI-F1 sows (84.4%, p < 0.05). Morulas and blastocysts were balanced in AI-F2 or AI-F3 (p > 0.05). Independently of the type of seminal dose (F1, F2, or F3), we observed by immunohistochemistry that AI significantly increased uterine vascularization, although with some anatomical differences. The cranial region of the uterine horns was significantly more vascularized in AI-F1 or AI-F2 sows (26.7 ± 2.3 and 28.6 ± 2.0%, respectively), and AI-F3 showed significantly less vascularization at that point (17.8 ± 1.6%, p < 0.05). To summarize, the synergistic effect of all ejaculate fractions accelerates embryo development, at least during the preimplantation period, and increases the uterine reaction to AI in certain parts of the uterus.

Is estradiol valerate an alternative to estradiol benzoate in promoting the synchronization of ovulation and timed artificial insemination in suckled Bos indicus beef cows?

The objective of the present study was to evaluate the effect of estradiol valerate administered at the beginning of the ovulation synchronization protocol on the pregnancy rate of Bos indicus cows. In the experiments, the following products from MSD, Sao Paulo, Brazil were used: estradiol valerate (EV), estradiol benzoate (EB), intravaginal progesterone device (P4), estradiol cypionate (EC), equine chorionic gonadotropin (eCG) and cloprostenol (PGF). In Experiment 1, Bos indicus cows (n=899) with a body condition score (BCS) of 2.76 ± 0.01 were included in a 3 (device) × 2 (protocol: 5 mg of EV or 2 mg of EB) factorial design. There were three types of P4 devices: a new device (New), a device previously used for 9 days (1×), and a device previously used for 18 days (2×). Nine days later (D9), the P4 device was removed, and cows received 300 IU of eCG. In addition, cows in the EB group received 1 mg of EC and 265 µg of PGF. Timed artificial insemination (TAI) was performed 48 h after P4 device removal in the EB group (TAI48) and 54 h after P4 device removal in the EV group (TAI54). In Experiment 2, Bos indicus cows (n=434) with a BCS of 2.62 ± 0.01 received a new P4 device or one previously used for 9 days and 5 mg of EV. On D9, all cows received 300 IU of eCG, and the P4 devices were removed and distributed in TAI48 and TAI54 cows. In Experiment 3, Bos indicus cows (n=429) with a BCS of 2.80 ± 0.01 were divided into the control and EV/EC groups. All cows received a P4 device. In addition, cows in the control group received 2 mg of EB, and cows in the EV/EC group received 5 mg of EV on D0. On D9, all cows received 1 mg of EC and 300 IU of eCG, and the P4 devices were removed. Cows in the control group also received 265 µg of PGF. All cows were inseminated 48 h after the removal of the P4 device. In Experiment 1, there was no effect of the interaction between protocol and P4 device on the occurrence of estrus (P=0.45) or on the pregnancy per artificial insemination ratio (P/AI; P=0.30). In addition, the occurrence of estrus and P/AI were not different between in the two estradiol groups (P=0.12 and P=0.82) and across the types of intravaginal P4 device (P=0.91 and P=0.47). In Experiment 2, the pregnancy rate was lower (tendency) in TAI48 cows (P=0.07). In Experiment 3, the estrus rate (P=0.12) and P/AI (P=0.56) were similar between the experimental groups. In summary, protocols using estradiol valerate without exogenous ovulation induction require adjustments in the timing of AI from 48 to 54 h after P4 device removal. However, a combination of estradiol valerate at the beginning of the protocol and estradiol cypionate nine days later successfully induced ovulation in Bos indicus cows inseminated 48 h after P4 device removal.

Exposure to progesterone before an ovulation synchronization protocol increases the follicular diameter and fertility of multiparous suckled Bos taurus cows.

The objective of this study was to evaluate the effect of administering injectable progesterone (P4i) before a timed artificial insemination (TAI) protocol on the follicular growth, ovulation, and pregnancy rate of Bos taurus suckled cows. The effect of P4i administration before the TAI on the pregnancy rate (P/AI) was evaluated in 576 suckled Bos taurus cows at 30-90 days postpartum. In addition, the effect of P4i administration before TAI on follicular dynamics was evaluated in subgroup of 401 suckled Bos taurus cows. On Day -10 (D-10), cows were divided into two experimental groups (Control and P4i). In this moment, P4i cows received i.m. 150 mg of injectable long-action progesterone. After that, both experimental groups received a synchronization protocol (Day 0; D0) that consisted of administration i.m. of 2 mg of estradiol benzoate and a progesterone intravaginal insert on D0. On Day 8 (D8), the progesterone insert was removed, and the cows received 500 µg of cloprostenol, 400 IU of eCG, and 1 mg of estradiol cypionate. TAI was performed 48 h after the removal of the progesterone insert. The ultrasound exams were performed in a subgroup of cows on Days 0, 8, 10 and 12 to evaluate the diameter of the largest follicle, rate of follicular growth and risks of single and double ovulation. The pregnancy diagnosis was performed 30 days after TAI in all cows to determine the pregnancy rate. The diameter of the largest follicle, on D10 (P = 0.84), rate of follicular growth (P = 0.14), ovulation rate (P = 0.40) and double ovulation rates (P = 0.23) did not differ between experimental groups. The pregnancy rate was greater in the P4i group [Control 46.2 % (133/288) vs. P4i 55.6 % (160/288); P = 0.03]. The diameter of the largest follicles (LF) on D0 (Control 11.6 ± 0.2 vs. P4i 13.3 ± 0.3) was greater (P = 0.01) in the P4i group. In conclusion, injectable progesterone before the ovulation synchronization protocol increased the diameter of the largest follicle on the D0 and the pregnancy rate in multiparous Bos taurus suckled beef cows.

Effects of equine chorionic gonadotropin dosage and its splitting in different days on reproductive performance of Nellore cows synchronized for timed-artificial insemination.

This study evaluated the effects of dose of equine chorionic gonadotropin (eCG) and its splitting in different days of the synchronization protocol on reproductive performance of primiparous and multiparous Nellore cows. In the present study, 2,536 Nellore cows (1,634 primiparous and 902 multiparous) were assigned to receive in a 2 × 2 factorial design 1) an intravaginal progesterone (P4) device and 2.0 mg of estradiol benzoate (EB) on day -11, 12.5 mg (i.m.) of dinoprost tromethamine (PGF), 300 IU (i.m.) of eCG, 0.6 mg (i.m.) of estradiol cypionate (ECP), and P4 device withdrawal on day -2, followed by TAI on day 0 (n = 632 cows, being 409 primiparous and 223 multiparous; 300-2), 2) 300 IU (i.m) of eCG administered on days -4 and -2 (150 IU of eCG/day; n = 637 cows, being 412 primiparous and 225 multiparous; 300-4-2), 3) 400 IU (i.m.) of eCG administered on day -2 (n = 633 cows, being 406 primiparous and 227 multiparous; 400-2), and 4) 400 IU (i.m) of eCG administered on days -4 and -2 (200 IU of eCG/day; n = 634 cows, being 407 primiparous and 227 multiparous; 400-4-2). Individual cow BCS was assessed on days -11, 0 (timed-AI), and 31 of the study. Body condition score of the animals was classified into LOW or HIGH using the threshold of 2.75 (≤2.75 = LOW; >2.75 = HIGH). For primiparous cows, an eCG splitting effect was observed on follicle size, as cows receiving eCG on days -4 and -2 of the synchronization protocol had a larger follicle than cows administered eCG only on day -2. For day 31 P/AI, primiparous cows receiving 400-4-2, regardless of BCS, had a greater P/AI than cows from other treatments. Administering 400-4-2 to LOW BCS cows also resulted in greater P/AI than all other treatments assigned to LOW BCS cows. For multiparous cows, no treatment effect was observed for follicle size, estrus expression, and day 31 P/AI (P ≥ 0.21). In summary, increasing the dose and splitting the dose of eCG positively impacted the pregnancy rates of primiparous cows under a BCS ≤2.75, but no effects were detected on multiparous cows.

Analysis of factors associated with IUI pregnancy outcomes in elderly and young patients.

OBJECTIVE: The objective of this study was to investigate the correlation between various factors and the clinical outcomes of Intrauterine Insemination (IUI) in both young and aged patients, aiming to provide a theoretical basis for clinical consultations. METHODS: This retrospective analysis examined a total of 4,221 IUI cycles conducted at the Reproductive Center of Changzhou Maternal and Child Health Hospital between January 2016 and December 2020. The patients were categorized into two groups based on age: the elder group (≥ 35 years) and the young group (< 35 years). RESULTS: The findings of this study revealed a significant association between woman's age and BMI with pregnancy outcomes (0.93, 95% CI: 0.89-0.97) (1.04, 95% CI: 1.01-1.06). Moreover, in young women, both age and Body Mass Index (BMI)were found to be related to pregnancy outcomes (0.97, 95% CI: 0.89-0.97) (1.08, 95% CI: 1.01-1.06). Additionally, BMI and the number of cycles in aged IUI patients were found to be associated with pregnancy outcomes. The pregnancy rate in the second cycle was approximately 1.9 times higher than that in the first cycle (1.9, 95% CI: 0.97-3.77), and in the third cycle, it was approximately 3 times higher than that in the first cycle (3.04, 95% CI: 1.43-6.42). CONCLUSIONS: In conclusion, there is an association between woman's age and BMI and the clinical outcomes of IUI. However, the number of cycles did not affect the pregnancy outcomes in young women. Conversely, in elder women, the number of cycles was found to be related to the IUI pregnancy outcomes, with significantly higher pregnancy rates observed in the second and third cycles compared to the first cycle.

A prospective proof-of-concept trial on the effect of personalized dosages of follitropin delta in intrauterine insemination.

RESEARCH QUESTION: What is the efficacy and safety of individualized follitropin delta dosing for ovarian stimulation in intrauterine insemination (IUI)? DESIGN: This single-centre, prospective, open-label, single-cohort study involving 106 patients established an original dosing regimen based on body weight and anti-Müllerian hormone (AMH) concentrations, with adjustments based on the ovarian response from the previous IUI cycle. Each participant was enrolled in a maximum of three IUI cycles. RESULTS: Mean age was 34.5 ± 4.5 years, mean weight 69.2 ± 11.2 kg, mean AMH 15.7 ± 8.6 pmol/l, mean FSH 6.3 ± 2.6 IU/l and mean antral follicle count 16.4 ± 8.2. The percentage of patients who produced more than three mature follicles was 1.9%, 0% and 1.5%, respectively, for the three IUI cycles. The percentage of patients with two or three mature follicles was 34.0%, 36.9% and 47.1% for the three IUI cycles. The clinical pregnancy rate per IUI cycle was 17.9%, 14.3% and 17.6% for the three cycles, with a cumulative clinical pregnancy rate of 40.6%. Out of 258 cycles, 43 (16.7%) resulted in clinical pregnancy, with six of those resulting in multiple pregnancies (14.0%). Two resulted in spontaneous reduction within the first trimester and four resulted in live twin births, representing only 1.6% of the total cycles. CONCLUSIONS: This study is the first to utilize follitropin delta for stimulation in IUI. It demonstrates that individualized dosing is both effective and safe, resulting in satisfactory cumulative pregnancy rates and an acceptable multiple pregnancy rate, thus achieving the primary objectives of the research.

The fertility of dairy heifers and cows is not influenced by the follicular wave of the ovulatory follicle.

Two studies were conducted to evaluate the effects of the follicular wave on ovarian function and fertility in dairy heifers and lactating cows. In study 1, the estrous cycle of the selected Holstein heifers was initially synchronized using two intra-muscular prostaglandin F2α (PGF2α) administrations 11 days apart. Heifers in group FFW (n = 14) received an intra-muscular 500 µg PGF2α administration on day 7 after detecting standing estrus, while Heifers in group SFW (n = 14) were administered PGF2α 13 days after detecting standing estrus. The pregnancy rates of FFW (n = 98) and SFW (n = 100) heifers were also determined 35-37 days after artificial insemination (AI). In Study 2, healthy Holstein lactating cows (n = 28) were randomly assigned to either the FFW (n = 14) or SFW (n = 14) groups. The estrous cycles of the cows were presynchronized using two intra-muscular administrations of PGF2α given 14 days apart. Then, the emergences of the follicular waves were induced using an Ovsynch protocol. The pregnancy rate of FFW (n = 99) versus SFW (n = 98) cows was also determined 35-37 days after AI. The ovulatory follicle and corpus luteum (CL) resulting from the ovulatory follicle of FFW were larger than those of the dominant follicle and the CL of SFW in dairy heifers and lactating cows. However, the pregnancy rate did not differ between the FFW and SFW groups in heifers and lactating cows 35-37 days after AI. In conclusion, although the characteristics of the ovulatory follicles in FFW versus SFW animals differed, the follicular wave in dairy heifers or lactating cows did not affect fertility.

First successful artificial insemination of the endangered Louisiana pinesnake, Pituophis ruthveni.

The Louisiana pinesnake (Pituophis ruthveni) is considered one of the rarest snakes in North America and was federally listed under the Endangered Species Act in 2018. Captive breeding and reintroduction of zoo-bred hatchlings has been successful, however, limited founders in the captive population and the inability to bring new, wild genes into the captive colony presents a major concern for the conservation of this species. The use of artificial insemination (AI) was first applied to snakes in the 1980s but further development of the technique has since received little attention. Our goal was to develop a method of AI for use in breeding Louisiana pinesnakes to facilitate gene flow from wild to captive populations. We inseminated two captive Louisiana pinesnakes with semen collected from one donor male, novel to both females. Timing of AI occurred following the emergence of females from brumation, and when large, distinct follicles were detected using digital palpation. Females were inseminated four and five times over a period of 14 and 19 days, respectively, using fresh and 2-day refrigerator stored semen. One female laid seven eggs, which resulted in four fertile eggs and two viable hatchlings, while the second female produced three fertile of seven eggs laid but no viable hatchlings. Genetic analyses confirmed the donor male was the sire of hatchlings. This is the first successful AI of an endangered snake species and provides a framework for the use and optimization of assisted reproductive technologies for use in conservation breeding programs.