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The main goals of this study were to document and compare the normal ranges of testicular haemodynamic parameters in pre- (aged 8-12 months; n = 4) and postpubertal (aged 24-259 months; n = 16) Miranda donkeys in the breeding season, and to correlate animal biometric data and testicular Doppler indices with basic semen quality parameters of sexually mature jacks. Colour and pulsed-Doppler ultrasonography were employed to assess blood flow in the left and right distal supratesticular artery (DsTA) and their marginal branches (marginal arteries-MA). Peak systolic velocity (PSV), end diastolic velocity (EDV), pulsatility index (PI) and resistive index (RI) were evaluated in both blood vessels, and TAMV (time-averaged mean velocity), TABF (total arterial blood flow) and TABF rate (TABF-R) were calculated for MA. The mean diameter of MA was greater (p < 0.05; 0.24 ± 0.05 vs. 0.19 ± 0.05 cm; mean ± SD) but TABF-R was less (p < 0.05; 0.004 ± 0.004 vs. 0.02 ± 0.01 mL/s/cm 3 ) in sexually mature donkeys compared with prepubertal animals. Apart from RI values for the right testicle of prepubertal donkeys, PI and RI were consistently greater (p < 0.05) in DsTA compared with MA. Significant correlations were found among select biometric and haemodynamic attributes of the testes (height, width and length, TV, TTV and PSV-ST) and ejaculate characteristics (volume, sperm defects-total, head and midpiece) in sexually mature donkeys (n = 8). The present results highlight the importance of scrotal ultrasonography for the reproductive assessment of jacks and provide reference values, based on the available subpopulation of Miranda donkeys that can be used in their clinical and reproductive management and research, or conservation programmes., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

At present, the success of non-surgical embryo recovery (NSER) and transfer (NSET) hinges upon the cervical passage of catheters, but penetration of the uterine cervix in ewes is problematic due to its anatomical structure (i.e., long and narrow cervical lumen with misaligned folds and rings). It is a major obstacle limiting the widespread application of NSER and NSET in sheep. While initial attempts to traverse the uterine cervix focused on adapting or re-designing insemination catheters, more recent studies demonstrated that cervical relaxation protocols were instrumental for transcervical penetration in the ewe. An application of such protocols more than tripled cervical penetration rates (currently at 90-95 %) in sheep of different breeds (e.g., Dorper, Lacaune, Santa Inês, crossbred, and indigenous Brazilian breeds) and ages/parity. There is now sufficient evidence to suggest that even repeatedly performed cervical passages do not adversely affect overall health and reproductive function of ewes. Despite these improvements, appropriate selection of donors and recipients remains one of the most important requirements for maintaining high success rates of NSER and NSET, respectively. Non-surgical ovine embryo recovery has gradually become a commercially viable method as even though the procedure still cannot be performed by untrained individuals, it is inexpensive, yields satisfactory results, and complies with current public expectations of animal welfare standards. This article reviews critical morphophysiological aspects of transcervical embryo flushing and transfer, and the prospect of both techniques to replace surgical methods for multiple ovulation and embryo transfer (MOET) programs in sheep. We have also discussed some potential pharmacological and technical developments in the field of non-invasive embryo recovery and deposition., Competing Interests: Declaration of Competing Interest I, Jeferson Ferreira da Fonseca, declare that I do not have any personal or financial interests that could influence or bias my actions, decisions, or judgments in any professional capacity. I have no affiliations, investments, or relationships that could create conflicts of interest. I acknowledge that conflicts of interest can compromise objectivity, fairness, and integrity, and I am committed to maintaining the highest level of ethical conduct in all my endeavors. I will disclose any potential conflicts of interest that may arise during the course of my work, and take appropriate steps to mitigate or address them transparently and in accordance with relevant laws, regulations, and organizational policies. Furthermore, I understand that any actions or decisions I take in my professional role should be based on merit, relevant expertize, and the best interests of the individuals, organizations, or entities that I serve. I will act in a fair, impartial, and transparent manner, and avoid any actions that could compromise my ability to fulfill my professional duties objectively and with integrity. I hereby affirm that the above declaration accurately reflects my current understanding and commitment to upholding high standards of integrity, transparency, and ethical conduct in all my professional endeavors., (Copyright © 2024 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

The effects of Mn 2+ -, Zn 2+ - or Cu 2+ -nanosuccinate added to freezing extender on select post-thaw semen characteristics were determined in six Texel rams (aged 2-4 years) during seasonal anestrus (April-May). Ejaculates (n = 6 per ram) collected into an artificial vagina were divided into ten isovolumetric fractions each. Semen was diluted in lactose-yolk-tris-citrate-glycerin medium and nanosuccinates (Mn 2+ - and Zn 2+ -nanosuccinate: 0.0 (control), 2.5, 5.0 and 7.5 μg/l; Cu 2+ -nanosuccinate: 0.0 (control), 1.25, 2.5 and 3.75 μg/l) were added to semen extender. Extended semen was loaded into 0.25-ml straws and frozen in liquid nitrogen. After thawing, sperm motility parameters were determined with computer assisted semen analysis (CASA), and the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) was measured with a spectrophotometric technique. The addition of 5.0 μg/l of Mn 2+ - and Zn 2+ -nanosuccinate significantly increased the sperm progressive motility and both 2.5 and 5.0 μg/l improved sperm motion kinetics. Further, both nanosuccinates at a dose of 5.0 μg/l significantly decreased SOD activity and stimulated an increase in GPx and CAT activity in semen samples. Alternatively, the addition of Cu 2+ -nanosuccinate (highest dose) significantly reduced the progressive motility and velocity of ram spermatozoa, increased the percentage of sperm with acrosomal/head defects and seminal SOD activity, and depressed CAT (highest dose) and GPx (all doses) activity. In summary, the addition of Mn 2+ - and Zn 2+ -nanosuccinate to semen extender had beneficial effects on sperm motility/motion kinetics and structural integrity, whereas Cu 2+ -nanosuccinate generally had debilitating effects on the post-thaw semen characteristics in rams., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to disclose. All co-authors have read and approved the contents of this manuscript, and there is no financial or other interest to report. We certify that this submission is an original work and is not under review at any other journal or with another publisher., (Copyright © 2024 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Ultrasonographic characteristics of skeletal muscles are related to their health status and functional capacity, but they still provide limited information on muscle composition during the inflammatory process. It has been demonstrated that an alteration in muscle composition or structure can have disparate effects on different ranges of ultrasonogram pixel intensities. Therefore, monitoring specific clusters or bands of pixel intensity values could help detect echotextural changes in skeletal muscles associated with neurogenic inflammation. Here we compare two methods of ultrasonographic image analysis, namely, the echointensity (EI) segmentation approach (EI banding method) and detection of selective pixel intensity ranges correlated with the expression of inflammatory regulators using an in-house developed computer algorithm (r-Algo). This study utilized an experimental model of neurogenic inflammation in segmentally linked myotomes (i.e., rectus femoris (RF) muscle) of rats subjected to lumbar facet injury. Our results show that there were no significant differences in RF echotextural variables for different EI bands (with 50- or 25-pixel intervals) between surgery and sham-operated rats, and no significant correlations among individual EI band pixel characteristics and protein expression of inflammatory regulators studied. However, mean numerical pixel values for the pixel intensity ranges identified with the proprietary r-Algo computer program correlated with protein expression of ERK1/2 and substance P (both 86-101-pixel ranges) and CaMKII (86-103-pixel range) in RF, and were greater ( p < 0.05) in surgery rats compared with their sham-operated counterparts. Our findings indicate that computer-aided identification of specific pixel intensity ranges was critical for ultrasonographic detection of changes in the expression of inflammatory mediators in neurosegmentally-linked skeletal muscles of rats after facet injury., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Ahmadi, Duarte, Srbely and Bartlewski.)

Exposure to heat stress (HS) in utero was postulated to trigger an adaptive molecular response that can be transmitted to the next generation. Hence, this study assessed the impact of HS exposure at different stages of the gestational period of mice on the female F1 population and their offspring. Heat stress exposure (41°C and 65% relative humidity-RH) occurred during the first half (FP), the second half (SP), or the entire pregnancy (TP). A control group (C) was maintained in normothermic conditions (25°C, 45% RH) throughout the experiment. Heat stress had a significant negative effect on intrauterine development, mainly when HS exposure occurred in the first half of pregnancy (FP and TP groups). Postnatal growth of FP and TP mice was hindered until 4 weeks of age. The total number of follicles per ovary did not vary (P > 0.05) between the control and HS-exposed groups. Mean numbers of primordial follicles were lower (P < 0.05) in the sexually mature FP than those in SP and TP F1 females. However, the mean number of viable embryos after superovulation was lower (P < 0.05) in TP compared with C group. The expression of genes associated with physiological and cellular response to HS, autophagy, and apoptosis was significantly affected in the ovarian tissue of F1 females and F2 in vivo-derived blastocysts in all HS-exposed groups. In conclusion, exposure to HS during pregnancy compromised somatic development and reproductive parameters as well as altered gene expression profile that was then transmitted to the next generation of mice., (© The Author(s) 2023. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

This article addresses morphokinetic changes and the extent of apoptosis in vitrified and non-vitrified in vitro-derived ovine blastocysts. Cumulus-oocyte complexes were collected after ovarian scarification obtain after slaughter and in vitro maturation was performed in TCM 199 medium supplemented with Earle's Salt, 10 % of FBS, and 5 µg/mL of LH/FSH at 38 °C for 24 h. After maturation, the oocytes were co-incubated with thawed ram semen (IVF) for 19 h.Embryo development was monitored with the aid of the Primo Vision Time-Lapse (TL) system. Twenty-five out of thirty-one ovine blastocysts that were vitrified using the Cryotop system at the early blastulation stage of development subsequently re-expanded. Both the vitrified (n = 25) and non-vitrified (control group: n = 28) blastocysts were examined for detection of apoptosis (TUNEL assay) and total blastomere counts at the time they attained the expanded blastocyst stage. Blastocyst formation occurred earlier in non-vitrified than in vitrified ovine embryos (147:49 ± 20:23 compared with 156:46 ± 19:24; hours:minutes post-insemination; mean ± SD; P < 0.05). The average number of blastocyst collapses was greater (2.45 ± 1.64 compared with 1.45 ± 1.64), but the number of weak contractions was less for vitrified than non-vitrified ovine blastocysts (P < 0.05). The mean number of blastomeres was greater (131.8 ± 38.6 compared with 91.5 ± 18.3; P < 0.05) while the number of TUNEL-positive cells (4.4 ± 1.6 compared with 6.3 ± 2.3) and apoptotic index (3.4 ± 1.2 % compared with 6.9 ± 2.6 %) were less (P < 0.05) in non-vitrified compared with vitrified blastocysts. Vitrification of ovine embryos was associated with a delayed blastocyst formation, greater numbers of apoptotic cells, significant reduction in the number of blastomeres, and higher/lower incidence of blastocyst collapse/weak contractions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Mammalian genome editing has utilized expensive and highly specialized electroporator devices. The "Gene Pulser XCell," a modular electroporation system for transfecting all cell types, has not been used extensively in mammalian embryo genome editing. The present experiment was undertaken to determine the usefulness of the Gene Pulser XCell for inserting the CRISPR/Cas9 system into intact zygotes in order to obtain the enhanced green fluorescent protein reporter rats (eGFP-R). An electroporation pulse response test using mCherry mRNA was performed to optimize the settings of the electroporator. Forty-five combinations of five pulse voltages (15, 25, 30, 35 and 40 V), three pulse durations (5, 10 and 25 ms), and three pulse frequencies (2, 5 and 6 pulses) applied at a constant 100-ms pulse interval and temperature of 37.5 °C were evaluated. The test revealed that the 35 V was the only voltage suitable for insertion of mCherry mRNA into intact rat zygotes and the only one that resulted in the production of embryos attaining the blastocyst stage. The incorporation of mCherry mRNA increased but the survival of the electroporated embryos declined with an increment in the number of pulses. Subsequent transfer of 1112 surviving Sprague Dawley rat embryos (after 8 h of incubating 1800 zygotes electroporated with the CRISPR/Cas9) resulted in the production of 287 offspring (25.8%). Ensuing PCR and phenotypic evaluation confirmed that twenty animals (6.96%) expressed eGFP in all body organs/tissues except for blood and blood vessels. The mortality of males and females before the attainment of puberty was 2 and 3 pups, respectively, and the final number/ratio of male to female of offspring was 9:11. All the surviving rats mated naturally and successfully transmitted the GFP transgene to their progeny. The Gene Pulser XCell total system with the settings predetermined in the present experiment can effectively be used to produce transgenic rats through the CRISPR/Cas9-mediated genome editing of zygotes., (Copyright © 2023 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Artificial insemination (AI) and in vivo embryo production (or multiple ovulation and embryo transfer, MOET) programs are both instrumental in accelerating the propagation of genetically and economically superior goats and sheep. The aim of this review was to present the current gestalt of non-surgical AI and embryo recovery (NSER) procedures in small ruminants. Small body size, precluding rectal palpation, and highly limited penetrability of the uterine cervix in ewes are the major reasons for the scarce use of non-surgical assisted reproduction techniques in this species. As a result, AI and embryo recovery techniques in sheep mainly involve laparoscopy or laparotomy (LAP). In does, however, the Embrapa method of AI allows for successful intrauterine deposition of semen, resulting in pregnancy rates from 50 to 80% under field conditions (>3 000 goats inseminated) when frozen-thawed semen is used. After the administration of prostaglandin F 2α (PGF 2α ), non-surgical (transcervical) embryo recovery is also feasible in goats, with the cervical penetration rate approaching 100%. There is a paucity of information on the efficacy of non-surgical AI using frozen semen in sheep, but the results are satisfactory with fresh, cooled, or chilled ram semen. An application of the NSER technique in ewes has greatly improved over the last decade, and cervical penetration rates of ∼90% can be achieved when a hormonal cervical dilation protocol using PGF 2α , oxytocin, and/or estradiol ester (e.g., estradiol benzoate) is applied. In some genotypes of sheep, sufficient cervical dilation can be induced without estradiol ester included in the protocol. Several studies indicated that recovery of transferable quality ovine embryos using NSER is comparable to that employing a ventral midline laparotomy, and NSER is evidently a method of choice when animal welfare is concerned. Considering both the number of retrievable embryos and animal well-being, the NSER is a viable alternative for surgical procedures. With further developments, it has the makings of a primary, if not exclusive, embryo recovery technique in small ruminants worldwide., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Human chorionic gonadotropin (hCG) has been used to improve goats reproductive efficiency. This study aimed to (i) evaluate if hCG administered by the intramuscular (i.m.) or intravaginal (i.vag.) route can be detected by a rapid β-hCG test in blood plasma samples and (ii) document ovarian effects of hCG administered by both routes at the time of artificial insemination (AI) performed 60 h after oestrus synchronization in goats. Twenty-two Alpine goats received two i.m. injections of 30 μg of d-cloprostenol (Prolise®, Tecnopec, São Paulo, Brazil) 7.5 days apart. One day after the onset of oestrus (at the time of AI), the goats were randomly allocated to one of the three groups that received: control (n = 7): 0.3 ml of saline solution intravaginally; hCG i.m. (n = 7): 300 IU of hCG (Vetecor®; Hertape-Calier, São Paulo, Brazil) i.m. and hCG i.vag. (n = 8): 300 IU of hCG deposited intravaginally. Blood samples were drawn at -1, 3, 6, 9 and 24 h after as well as on days 3, 7, 10, 13, 17 and 21 after hCG treatment/AI. All animals tested negative for hCG (ECO Diagnóstica, Corinto, Brazil) at -1 h, and all control animals tested negative throughout the entire blood collection period. All hCG i.m. animals tested positive from 3 h until D3 post-AI but only 50% of hCG i.vag. goats tested positive during the present study. In all animals studied, mean circulating P 4 concentrations increased (p < .05) from D3 to D7 after AI and then declined (p < .05) from D10 to D17 in control and hCG i.m. groups and from D17 to D21 in the hCG i.vag. group. Total cross-sectional luteal area (CLA), mean colour Doppler area (DA), DA/CLA, mean high-velocity Doppler area and HVDA/CLA all declined (p < .05) by D17-D21 in all animals studied. In summary: (i) human chorionic gonadotropin could consistently be detected in blood samples using the rapid β-hCG test only in the hCG i.m. group; and (ii) there were no significant differences in the mean pregnancy rate, circulating P 4 concentrations and various luteal parameters studied among Control, hCG i.m. and hCG i.vag. dose., (© 2022 Wiley-VCH GmbH.)

Premature regression of corpora lutea (PRCL) may adversely affect the outcome of hormonal ovarian superstimulation in small ruminants, and the total dose of exogenous gonadotropins used may be one of the causes of this condition. There were two major objectives of the present study: (1) to evaluate the effects of different superovulatory doses of porcine follicle-stimulating hormone (pFSH) on the biometry, blood perfusion (Doppler), and echotextural characteristics of luteal structures; and, (2) to determine the usefulness of biometric, vascular, and echotextural luteal variables, as well as measurements of circulating progesterone (P 4 ) concentrations for early detection of PRCL in superovulated Santa Inês ewes. Twenty-seven Santa Inês ewes received an intravaginal P 4 -releasing device (CIDR) from Days 0 to 8 (Day 0 = random day of the anovulatory period). An IM injection of d-cloprostenol (37.5 μg) was given at the time of the CIDR insertion and withdrawal. On Day 6, all the ewes received 300 IU of eCG IM and were divided into three treatment groups (each n = 9): G100 (100 mg); G133 (133 mg); and G200 (200 mg of pFSH) administered IM every 12 h in eight injections. Transrectal ovarian ultrasonography and jugular blood sampling for serum P 4 measurements were performed on Days 11 to 15. On the day of embryo recovery (Day 15), all the ewes underwent diagnostic videolaparoscopy and were classified, based on their luteal characteristics, into three response groups: nCL (ewes with normal CL only); rCL (ewes with regressing CL only); and ewes with both nCL and rCL following the superovulatory regimen. Our present results indicate that the total pFSH doses of 100 mg and 200 mg result in similar ovulatory responses and luteal function/biometrics, although the percentage of donor ewes with nCL was greater ( p < 0.05) for G100 compared with the G200 animals. An application of 133 mg of pFSH was associated with diminished luteogenesis. Lastly, circulating P 4 concentrations, ultrasonographic estimates of total luteal area, and CL pixel heterogeneity (standard deviation of numerical pixel values) are promising markers of luteal inadequacy in superovulated ewes.

This study set out to examine ultrasonographic attributes of non-neurosegmentally (pectoral-forelimb) and neurosegmentally linked (hindlimb) myotomes in an experimental model that leads to neurogenic inflammation in segmentally linked myotomes, and to evaluate quantitative correlations among ultrasonographic attributes of the muscles, relative content of various inflammatory mediators, and nociceptive thresholds (hot and mechanical) in rats. Twelve male Wistar Kyoto rats were randomly divided into two equinumerous groups: surgery group, in which the left lumbar (L4-L6) facet joints were compressed for 3 min with modified Kelly forceps under general anesthesia, and sham-operated rats. All ultrasonograms were obtained with the Vevo 2100 Visual Sonic scanner connected to a 24-MHz transducer at four different time points: pre-surgery and 7, 14, and 21 days after surgical procedures. Digital ultrasonographic images of quadriceps femoris, hamstring, and pectoral-brachial muscle groups were analyzed using a polygonal meter region of interest placed on the largest cross-sectional area of the muscles displayed in Image ProPlus ® analytical software to compute numerical pixel values and pixel heterogeneity (standard deviation of mean pixel values). On day 21, pain behavior tests (hot plate and von Frey) were performed and then all animals were euthanized. Protein expression of inflammatory mediators in biceps brachii and rectus femoris muscles was measured by Western blot. The most prominent differences in muscle echotextural attributes between the two subsets of rats occurred 14 days post-surgery in pectoral-brachial and quadriceps femoris muscles. The expression of calcitonin-gene-related peptide was directly related to both echotextural variables only in biceps brachii (pixel intensity: r  = 0.65, P  = 0.02; and heterogeneity: r  = 0.66, P  = 0.02, respectively). Our findings have revealed the occurrence of echotextural changes in skeletal muscles of rats during myositis; however, the accumulation of inflammatory mediators and the outcomes of sensory tests did not relate to the changes in first-order echotextural characteristics of affected hindlimb muscles.

The present experiment employed time-lapse (TL) imaging to assess the effects of vitrification on the development of ovine blastocysts and to see if the timing of blastocyst formation and expansion was correlated with the numbers of embryo- and trophoblasts determined through differential staining of the expanded blastocysts. Ovaries were obtained after slaughter from cycling (October-March) Polish Longwool ewes aged 1-3 years and cumulus-oocyte complexes were collected by ovarian scarification. In vitro maturation was performed in TCM 199 medium supplemented with Earle's Salt, 10% of FBS, and 5 μg/mL of LH/FSH at 38 °C for 24 h. After maturation, the oocytes were incubated with thawed ram semen (IVF) for 19 h and all presumptive zygotes were transferred to a 16-well dish containing Cult medium for monitoring with the Primo Vision TL system. A portion of ovine embryos were vitrified (Cryotop system) at the early cavitation stage and TL observations of warmed (n = 30) and non-vitrified (n = 32) blastocysts continued until the attainment of the expanded blastocyst stage, at which point they were differentially stained with bisbenzimide and propidium iodide for microscopic enumeration of embryoblasts (inner cell mass blastomeres) and trophoblast-cells. There were no statistically significant differences in the timing of blastulation (tB) and formation of expanded blastocysts (tEB) between vitrified and non-vitrified ovine embryos, but non-vitrified blastocysts exceeded (P < 0.05) their vitrified and warmed counterparts in the mean number of embryo- and trophoblasts. In addition, the number of trophoblasts was negatively and moderately correlated with tB and tEB, for both vitrified and non-vitrified embryos. It can be concluded that even though vitrification of ovine embryos is associated with a significant reduction in the number of blastomeres, the rate of blastocyst development remains closely linked to the numbers of trophoblastic cells., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

This review addresses the influence of homebox A10/a10 (HOXA/Hoxa10) gene on reproductive tract anatomy and functional fertility in mammalian species, and discusses major endocrine and environmental regulators of HOXA/Hoxa10 expression. Female reproductive efficiency or success is a function of several factors including the ovulation and fertilization rate, and uterine receptivity. A family of HOX/Hox genes establishes the segmental identity of the reproductive tract during embryogenesis and retains its physiological plasticity in sexually mature animals and humans. In particular, the HOXA/Hoxa10 gene is an intrinsic component of implantation, decidualization, and immunomodulation in the adult uterus. It was, therefore, suggested that knowledge of HOXA/Hoxa10 regulation might be essential in navigating molecular mechanisms with the aim of enhancing female reproductive potential. However, a recent study in pigs revealed a lack of associations between endometrial HOXA10 expression and reproductive tract morphology, and very poor correlations with sows' fertility metrics. Retinoic acid mainly regulates 3' HOX/Hox paralogs but may also modify the expression of downstream HOX/Hox genes, including HOXA/Hoxa10. Sex steroids directly regulate HOXA/Hoxa10 expression. The vitamin D receptor pathway modulates HOXA/Hoxa10 expression in the adult reproductive tract. Lastly, endocrine disruptors such as diethylstilbestrol, methoxychlor, bisphenol A, and isoflavones were shown to alter HOXA/Hoxa10 expression, thus affecting reproductive competence of the female., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The assessment of morphology and digital image opacity may provide valuable information on the present embryo quality. Time-lapse imaging has been employed in research to establish a means of monitoring the dynamic nature of preimplantation embryo development. The aim of present study was to use time-lapse imaging for assessing various prospective morphometric and phototextural markers of the developmental potential of in vitro -derived ovine embryos. Oocytes were obtained by scarification of ovaries from nine Polish Longwool ewes. After in vitro maturation (IVM) and fertilization (IVF) of oocytes with fresh ram semen, the development of embryos to the blastocyst stage was monitored and evaluated using Primo Vision time-lapse imaging technology. Commercially available Image-Pro ® Plus software was used to measure zona pellucida thickness, embryo diameter, total area of the perivitelline space, cellular grey-scale pixel intensity and cellular pixel heterogeneity. Statistical assessment of all attributes was done at various time points during embryo development (i.e., presumptive zygote stage: t(0); first cleavage detected at t(2) or t(3); and second cleavage detected at t(4) or t(6)). Out of thirty-seven zygotes analyzed in this study, five did not divide, 26 arrested before and six developed to the blastocyst stage. Our present results indicate that most parameters analyzed did not differ among embryos varying in their developmental fate except for the perivitelline space area that was greater (P<0.05) for non-dividing zygotes than future blastocysts at the presumptive zygote stage (4040±1850 vs. 857±262 µm 2 , respectively; means±SEM). Consequently, the measurement of perivitelline space at t(0) can potentially be used to prognosticate developmental potential of in vitro -produced ovine embryos albeit further confirmational studies are needed., Competing Interests: Conflict of interest: The authors have no conflict of interest to declare.

The present study compared the outcomes of in vivo embryo production in Morada Nova ewes subjected to either 9-day (G-9 SOV , n = 21) or 12-day (G-12 SOV , n = 21) progesterone (P 4 )-based estruses synchronization protocol coupled with superovulatory treatment with decreasing doses of porcine follicle-stimulating hormone (133 mg of pFSH given over 3 days). Non-surgical embryo recovery (NSER) was performed 6-7 days after the onset of oestrus. Total antral follicle count doubled from the first to the sixth pFSH dose in both groups (p < .05). Oestrus responses did not vary between the two groups of animals (95.2%). Corpora lutea (CL) were detected in 85.0% and 60.0% of ewes that previously manifested oestrus behaviour in G-9 SOV and G-12 SOV respectively. NSER was successfully completed in 86.2% of ewes that had CL (p > .05). The mean number of CL per ewe/successfully flushed donor ewe was greater (p < .05) in G-12 SOV (12.3 ± 1.7/12.1 ± 1.9) than in G-9 SOV (7.9 ± 1.4/8.2 ± 1.6). Mean numbers of retrieved blastocysts and viable embryos were greater (p > .05) in G-12 SOV (5.8 ± 1.9 and 3.7 ± 1.7) than G-9 SOV (3.5 ± 1.1 and 0.8 ± 0.3 respectively). The total follicle count (all follicles ≥2 mm in diameter) at the sixth pFSH dose (at P 4 -device removal) was positively correlated (p < .05) with the number of CL (r = .95) and viable embryos (r = .91) in G-12 SOV . The ewes with ≥10 Cl (48% of all flushed donors) yielded 80.5% of viable embryos. In summary: (a) Morada Nova ewes from G-12 SOV group had better superovulatory responses compared with G-9 SOV group; (b) total follicle count at the last pFSH dose was a good predictor of superovulatory responses only in the ewes primed with P 4 for 12 days; and (c) animals with ≥10 ovulations are main contributors to viable embryo production in Morada Nova ewes., (© 2021 Wiley-VCH GmbH.)

Two experiments were conducted in acyclic Alpine (A) and Saanen (S) goats that received intravaginal sponges containing 60 mg of medroxyprogesterone acetate for 6 days, as well as 200 IU of eCG and 30 μg d-cloprostenol i.m. 24 h before sponge removal. On day 7 (day 0 = onset of synchronized estrus), all goats were randomly divided into two groups: animals treated with 300 IU of hCG i.m. (hCG; Exp.1: n = 8A; Exp.2: n = 75A + S) and untreated controls (Control; Exp.1: n = 8A; Exp. 2: n = 70A + S). In Exp.2, all goats were artificially inseminated. Transrectal ovarian ultrasonography and blood collection were done on days 7, 10, 13, 17, and 21 (Exp.1), and pregnancy detection on day 60 (Exp.2). Estrus and ovulations occurred in five hCG and seven Control animals. Accessory CL (aCL) were detected in all hCG does. The total luteal area of ovulatory corpora lutea (oCL) increased (P < 0.05) on day 10 in hCG does and remained greater (P < 0.05) than in Control until day 21. Total and high-velocity color Doppler area were greater (P < 0.05) for oCL of hCG does on days 13 and 17. Progesterone concentrations were greater (P < 0.05) in hCG does from days 13 to 21 and related directly to the total luteal and oCL area for the duration of the study in all does. The pregnancy rate was higher (P < 0.05) in hCG than in Control by 22.5 %. Human chorionic gonadotropin given on day 7 of the synchronized estrous cycle positively affected CL function and pregnancy rates in seasonally anovular dairy goats., Competing Interests: Declaration of Competing Interest The authors report no declarations of interest., (Copyright © 2021 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

The aim of this study was to analyze the factors that can influence pork prices, particularly the effects of various types of fluctuations on the volatility of pork prices in the European Union as a whole market and individual EU countries. The research material consisted of monthly time series of pork prices collected from 2009 to 2020. These data originated from the Integrated System of Agricultural Information coordinated by the Polish Ministry of Agriculture. Information on global pork production volumes is from the Food and Agriculture Organization Statistics (FAOSTAT) database. Time series of prices were described by the multiplicative model, and seasonal breakdown was performed using the Census X-11 method. The separation of the cyclical component of the trend was performed using the Hodrick-Prescott filter. In 2019, pork production in the European Union totaled 23,954 thousand tonnes, which accounted for 21.8% of global pork production. The largest producers were Germany, Spain, and France, supplying more than half of the pork to the entire European Union market. Pork prices in the EU, averaged over the 2009-2020 period were Euro (EUR) 154.63/100 kg. The highest prices for pork were recorded in Malta, Cyprus, Bulgaria, and Greece, whereas the lowest prices in Belgium, the Netherlands, Denmark, and France. The breakdown of the time series for pork prices confirmed that, in the period from 2009 to 2020, pork prices exhibited considerable fluctuations of both a long-term and medium-term nature as well as short-term seasonal and irregular fluctuations. Prices were higher than average in summer (with a peak in June-August) and lower in winter (January-March). Overall, the proportions of different types of changes in pork prices were as follows: random changes-7.9%, seasonal changes-36.6%, and cyclical changes-55.5%.

This study set out to examine associations among echotextural, physicochemical and sensory attributes of the pectoralis major muscles in 17-week-old organic turkeys (B.U.T. Big-6) varying in the amount of wheat and oat grain in daily feed rations (Group C: complete feed only; Group Exp1: 5-30% of wheat and 0-20% of oat; and Group Exp2: 5-50% of wheat and 0-50% of oat; n = 15 turkeys/group). Digital ultrasonograms of the left pectoral muscle in four different planes (longitudinal-L, transverse-T, and two oblique planes-O1 and O2) were obtained with a 5.0-MHz linear-array transducer just before slaughter. Mean numerical pixel intensity (MPI) and pixel heterogeneity (MPH) of the muscle parenchyma were computed using the ImageProPlus ® analytical software. Ten significant correlations between echotextural attributes and various meat characteristics were recorded in Group C, one in Group Exp1, and eight in Group Exp2. When data were pooled for all birds studied, there were twelve significant correlations ( p < 0.05); all but one correlation (between MPH and moisture) were for physical and sensory characteristics of meat samples. Computer-assisted analysis is a potential method to determine moisture as well as physical (e.g., coloration) and sensory (e.g., aroma) characteristics of pectoralis major muscles in organic turkeys.

The present study set out to determine the effects of incorporating cDDGS into starter, grower, and finisher diets (containing 5%, 10%, and 15% of cDDGS, respectively) on growth performance, carcass and meat quality, and cost effectiveness of pig fattening. Sixty-four pigs (mean body weight of 15.0 ± 2.1 kg) were divided into two groups ( n = 32) and fed a control diet (cereal-soybean meal-based) or cDGGS-containing diets (with soybean meal partially replaced with cDDGS). Live weights of pigs as well as weight gains/daily weight gains across all fattening phases did not differ between the two groups of fattener pigs studied ( p > 0.05). Addition of cDDGS decreased feed intake per pig during the grower ( p < 0.05) and finisher ( p < 0.01) phases, and, as a result, throughout the entire fattening period (254 vs. 245 kg for control and cDDGS groups, respectively; p < 0.01). The feed conversion ratio (FCR) for the entire fattening period was significantly less for cDDGS-fed fatteners (2.77) than for controls (2.91; p < 0.05). Carcass weights, fat thickness, and meatiness did not vary between the two groups of animals ( p > 0.05). Loin depth was greater in the cDDGS group by ~5 mm ( p < 0.05). Slaughter value was higher for the cDDGS group (76.1% vs. 77.0%, p < 0.05). The total cost of fattening and total cost of 1 kg of body weight decreased in cDDGS compared with the control subset of fatteners by ~7% and 8% during the grower and finisher phases, respectively ( p < 0.01). The simplified direct surplus per pig was approximately 63% higher for the cDDGS group. Our results indicate that even moderate inclusion of cDDGS to concentrate mixtures (or a partial replacement of soybean meal with cDDGS) may improve FCR without any substantial changes in meat and back fat characteristics as well as significantly decrease the cost of feeding and increase the profitability of pig production.

This study assessed the feasibility of in vivo embryo production and nonsurgical embryo recovery (NSER) in Morada Nova ewes (an endangered native Brazilian breed of sheep) subjected to different estrus synchronization and/or superovulation protocols. Ewes received intravaginal sponges soaked with 60 mg medroxyprogesterone acetate (MAP), which were kept in place for six (G6; n  = 12), nine (G9; n  = 12), or 12 (G12; n  = 12) days. Half of the ewes in each group remained estrus synchronized only ( SYNCH ) and the other half was superovulated ( SOV ) with 133 mg porcine follicle-stimulating hormone (pFSH). There were no differences ( p  > 0.05) in antral follicle counts determined with ultrasonography 60 hours before MAP sponge removal (or at the time of the first pFSH dose) among G6 (6.4 ± 0.9), G9 (6.2 ± 0.7), and G12 (5.5 ± 0.6). Estrus responses and NSER success rates did not vary ( p  > 0.05) among the three progestin-treatment groups of ewes for either estrus-induced or superovulated animals. The onset of estrus occurred 10-12 hours later ( p  < 0.01) in G9 SYNCH ewes compared with G6 SYNCH and G12 SYNCH, and the duration of estrus was ∼19 hours greater ( p  < 0.01) in G9 SOV than in G6 SOV . The average duration of the NSER procedure was 32.6 ± 1.3 minutes. At least one structure was recovered in 85.7% of synchronized and in 87.5% of superovulated ewes. Viable embryo recovery rates were also similar ( p  > 0.05) for G6 (1.0 ± 0.3 and 2.5 ± 1.5), G9 (1.3 ± 0.5 and 4.8 ± 2.0), and G12 groups (1.0 ± 0.3 and 4.8 ± 2.3; estrus-synchronized and superovulated ewes, respectively). In conclusion, progestogen pretreatment of different durations and NSER can be employed in Morada Nova ewes, resulting in reasonable viable embryo recovery rates in both estrus-synchronized and superovulated animals. Therefore, both techniques are suitable for use in commercial settings as well as small ruminant conservation programs.

Economic potential of the swine industry hinges upon the reproductive performance of sows, which may be enhanced by improving uterine capacity, a component trait of litter size and piglet productivity. Previous attempts at characterizing morphological traits indicative of high uterine volume have not been completely successful, resulting in the continued need for a reliable method of predicting reproductive value to improve production efficiency of the sow. Hence, the main objective of this study was to scrutinize macro- and micro-morphology of the sow's reproductive tract for quantitative correlations with fertility indices. Reproductive records from Polish Landrace × Polish Large White sows were used to examine the associations between fertility and ovarian/uterine morphology (n = 34) or uterine histomorphometry (n = 10). Several measures related to the ovary, including right and left ovarian weight (r = 0.50, p = 0.005 and r = 0.49, p = 0.006, respectively), were positively correlated with the litter size, while left ovarian number of corpora lutea (r = -0.38, p = 0.04) was negatively correlated with the mean litter size. Analysis of histomorphological characteristics of the uterine wall collected during the luteal phase of the estrous cycle revealed correlations between mean litter size and myometrial vascular content (r = 0.75, p = 0.03), the proportion of myometrial stroma (r = -0.68, p = 0.03), and the variability of endometrial thickness (r = -0.72, p = 0.02) in sows. Eight ovarian, vaginal and uterine characteristics were significantly correlated with mean lifetime numbers of live born and stillborn piglets/litter or the last litter size before slaughter. In conclusion, several anatomical and histomorphological metrics that relate to reproductive performance of swine may be used to inform production protocols and as a tool for selection of elite breeding sows, warranting future research into non-invasive or minimally invasive techniques for obtaining such measures., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Selection of appropriate housing conditions for sows is critical for their physical health and long-term reproductive success. The present objective was to evaluate the influences of housing system postweaning (i.e., individual stalls (IS) or group pens (GP)), season and parity on piglet productivity of sows in a commercial setting. This study utilized 3 053 Polish Large White×Polish Landrace sows that were weaned at a rate of 20-30 animals per week at the median age of 4 weeks; 1 474 sows were moved into GP of seven to eight animals each, while 1 579 were placed in IS after weaning. Starting 2 days postweaning all animals were checked for estrus with a teaser boar and then artificially inseminated using 3×10 9 spermatozoa per dose of an inseminate at the onset of heat and 24 h later. The proportion of sows showing the signs of standing heat at or before 6 days postweaning was greater (P < 0.05) for sows moved to GP compared with IS; this difference manifested mainly in second parity sows weaned in the summer and fall. Conception and farrowing rates were significantly higher (P < 0.01) and the weaning-to-estrus interval shorter in GP compared with IS sows in every season but autumn. Mean litter size was lower (P < 0.05) in IS groups in summer, autumn, and winter, and the number of live-born piglets/sow was lower (P < 0.05) for IS sows in the summer and fall. Beneficial effects of group housing on piglet productivity manifested up until the seventh consecutive farrowing and then began to wane. In summary, there was a significantly greater proportion of sows going estrus "on time" (i.e., <7 days) in group housing compared to single stalls but this effect was confined to the second parity sows during the summer and fall months; these results suggest the existence of a seasonal and age-related aspect to sow fertility worthy of further investigation. While both housing systems have their pros and cons, our present results indicate that, in commercial settings, group housing postweaning improved nearly all reproductive parameters of sows., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Time-lapse (TL) imaging provides a practical and safe tool to constantly monitor the development of in vitro-derived embryos. TL may help develop novel methods of predicting the timing of embryo cleavage that will lead to optimizing blastocyst cryopreservation or transfer. The primary objective of the present study was to employ TL imaging to examine associations among the division kinetics of ovine embryos, their quality and rates of development to the blastocyst stage. Oocytes were collected by ovary scarification from 78 Longwool ewes slaughtered in the breeding season (November-March). Cumulus oocyte complexes (COCs) were matured for 24 h in TCM 199 media containing 0.1 IU/mL LH/FSH and 10% FBS. In-vitro fertilization was carried out by co-incubation of semen and COCs for 19 h. Presumptive zygotes were placed in microwells, in droplets of Cult medium (Gynemed, Lensahn, Germany). Digital images of developing embryos were captured every 10 min by Primo Vision TL system (EVO+; Vitrolife, Göteburg, Sweden). The following time intervals were recorded: from IVF to the attainment of two-cell (t2), three-cells (t3) or four-cell (t4) stage, to morula detection (tM), blastulation (tSB) and blastocyst formation (tB). Lastly, the duration of the second cell cycle (cc2; t3-t2) and complete synchronous cell division (s2; t4-t3) were calculated, and the incidence of developmental anomalies noted. Out of 147 embryos selected for TL observations, 55 (37.4%) developed to the blastocyst stage (normally developing embryos, NE) and 92 (62.6%) failed to reach the blastocyst stage (arrested embryos, AE; P < 0.05). Mean t2, tM, s2 and cc2 were all less (P ≤ 0.02) in NE compared with AE. Approximately 61.9% of embryos exhibited developmental anomalies (35.5% in the NE group and 78.2% in the AE group; P < 0.05) and AE exceeded (P < 0.05) NE in the proportion of FRG (blastomeric fragmentation), IRR (blastomeres of irregular size after cleavage), DC (direct cleavage) and MA (multi-morphological aberrations). Of all NE, 63.6% were classified as good quality and 36.4% as poor quality blastocysts (P < 0.05). Good quality ovine blastocysts attained t2, t3, t4, tSB and tB stages earlier (P ≤ 0.03) than poor quality blastocysts and none of the poor quality blastocysts was seen to hatch. To recapitulate, the present results indicate that the kinetics of early ovine embryo development are significant predictors of their potential to develop to the blastocyst stage and the markers of blastocyst quality. Time-lapse imaging may serve as a useful technique for predicting the outcome and enhancing efficacy of in vitro embryo production in sheep., (Copyright © 2020 Elsevier Inc. All rights reserved.)

This study examined the effects of exogenous hCG administration on ovarian function and pregnancy rates in estrous-induced dairy goats during the transition into the breeding season. Eighty-six Toggenburg does received 60 mg of medroxyprogesterone acetate intravaginal sponge for 6 d plus 200 IU of equine chorionic gonadotropin and 30 μg of d-cloprostenol i.m. 24 h before sponge removal, and were then bred for 96 h. Seven days (D7) after first mating the does received either 1 mL of saline (the control group, n = 43) or 300 IU of hCG (the hCG-treated group, n = 43) i.m. Transrectal ovarian ultrasonography (B-mode and color Doppler) was performed on D7, D13, D17, and D21 and ultrasonographic pregnancy detection on D30. Pregnancy rate was higher (P < 0.05) in hCG-treated goats (90.7%; 39/43) than that in control animals (74.4%; 32/43). Accessory luteal structures (ALSs) were detected in 46.5% (20/43) of hCG-treated does. All hCG-treated does that had ALSs and 82.6% of goats without ALS post-treatment remained pregnant. The total luteal area increased (P < 0.05) from D7 to D13 in pregnant animals of both groups, whereas mean vascular area declined (P < 0.05) by D21 in all nonpregnant does. Serum progesterone concentrations increased (P < 0.05) on D21 in pregnant goats of both groups, but they were related to changes in luteal tissue content only in control does throughout the present study. Mean daily numbers of small- and medium-sized antral follicles decreased (P < 0.05) only in pregnant animals of both groups with a decline in medium follicle numbers occurring earlier in hCG-treated (D13) compared with control does (D17). To summarize, a single dose of hCG given on D7 after estrus was followed by a decrease in the number of medium-sized antral follicles in gestating hCG-treated does, induced the formation of ALSs in ~47% of all hCG-treated does, and significantly increased the pregnancy rate in estrous-induced Toggenburg goats in the transition to the breeding season., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The aim of this study was to compare the efficiency of in vitro embryo production (IVP) following the collection of bovine ovaries and 22-h in vitro maturation (IVM) of oocytes in media prepared with Milli-Q® Water (n = 509 oocytes) or nanowater (NW; n = 304 oocytes). The mean cleavage (63.8 ± 4.6 % vs. 63.6 ± 6.1 %, respectively; mean ± SEM) and blastocyst formation rate (16.3 ± 3.4 % vs. 16.7 ± 6.7 % of presumptive zygotes, respectively) did not vary (P > 0.05; Student t-test) between the two types of media diluents. NW is a safe substitute for Milli-Q® Water for IVM of bovine oocytes., Competing Interests: Declaration of Competing Interest The authors report no declarations of interest., (Copyright © 2020 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Thirty clinically healthy Holstein-Friesian cows underwent twice daily machine milking and ultrasonographic examinations of the udder just prior to and after milking. Digital ultrasonographic images of each udder quarter were subjected to computer-assisted echotextural analyses to obtain mean numerical pixel values (NPVs) and pixel heterogeneity (PSD) of the mammary gland parenchyma. The average milk yield and pH were higher ( p < 0.05) in the morning, whereas crude fat, total solids, solids non-fat and citric acid content were higher ( p < 0.05) during the evening milking period. Mean NPVs and PSDs of the mammary gland parenchyma were greater ( p < 0.05) after than before milking. There were significant correlations among echotextural characteristics of the udder and protein percentage, lactose content and freezing point depression determined in the milk samples collected in the morning and crude protein, casein, lactose and solids non-fat in the evening. Our results can be interpreted to suggest that computerized analysis of the mammary gland ultrasonograms has the makings of a technique for estimating non-fat milk constituents in cows. However, future validating studies are necessary before this method can be employed in commercial settings and research. Moreover, significant inter-quarter differences in udder echogenicity may necessitate further echotextural studies of separate quarters.

Outcomes of short- (6.5 days) and long-term (14.5 days) estrous synchronization for 6.5 d (G-6.5d) or 14.5 d (G-14.5d) and followed by the 4-day or 3-day declining-dose follicle-stimulating hormone superovulatory regimen, respectively, were compared using 16 estrous-cycling Santa Inês ewes. Non-surgical embryo recovery (NSER) procedures were performed 60 d apart starting 6 or 7 d after the onset of estrus; an i.m. injection of estradiol benzoate and of d-cloprostenol at 16 h was followed by an i.v. oxytocin injection administered 20 min before NSER. There was a longer (P < 0.05) period before estrous onset in ewes during the second (September) compared with the first study replicate (July) by approximately 14 h. The NSER could be performed in 11 of 15 ewes that were in estrus, with an average of three viable-embryos/donor and the mean duration of the procedure being 29 min. There were no differences in superovulatory responses between the two groups of ewes, but there were only degenerated embryos in ewes of the G-6.5d group. In summary: i. the duration of progestin-priming and of multiple-dose pFSH treatment had a limited effect on superovulatory responses in estrous-cycling Santa Inês ewes; ii. NSER is a safe and repeatable method of embryo collection in ewes subsequent to superovulation; and iii. duration of the superovulatory treatment regimen may alter the effects of endogenous steroids on oocyte/embryo quality in ewes., Competing Interests: Declaration of Competing Interest Nothing to declare., (Copyright © 2020. Published by Elsevier B.V.)

The main goal of this study was to examine the utility of measuring systemic concentrations of steroid hormones, namely progesterone (P 4 ) and estrone sulfate (E 1 S), for monitoring the progression of porcine pregnancy and predicting sow fertility. There were 3 subsets of artificially inseminated (AI'd) sows used in the present experiments: (i) animals sacrificed on gestational day 20 (gd20; n = 16) or (ii) gd50 (n = 16; Experiment 1), and (iii) animals maintained throughout pregnancy (n = 24; Experiment 2). Blood samples (10 mL) were drawn from the orbital sinus and the endocrine data determined at different time points around ovulation/artificial insemination (gd0 (first AI), gd1 (second AI), and gd2) and maternal recognition of pregnancy (gd11), as well as on gd20 and gd50 (during 2 periods of increased embryonic/fetal mortality in swine) were examined for correlations with the numbers of healthy, arrested, and reabsorbing embryos (Experiment 1) or with the number of live, stillborn, and mummified piglets recorded at farrowing (Experiment 2). No correlations were recorded between circulating concentrations of both steroids and the numbers of healthy, arresting, or reabsorbing conceptuses on gd20 or 50 (Experiment 1). The number of corpora lutea (CL) was directly related to the number of healthy embryos/conceptuses on gd20 and 50 (r = 0.71, P = 0.007 and r = 0.76, P = 0.0007, respectively) and the number of arresting embryos on gd20 (r = 0.54, P = 0.05), and negatively correlated with the number of reabsorbing embryos on gd20 (r = -0.53, P = 0.05). In Experiment 2, circulating P 4 concentrations on gd11 related directly to the number of live-born piglets (r = 0.46, P < 0.04). Systemic E 1 S concentrations on gd0, gd1, gd2 and gd50 were correlated with the number of mummified conceptuses recorded at farrowing (r = 0.50, P = 0.03; r = 0.59, P = 0.01; r = 0.48, P = 0.04; and r = 0.56, P = 0.01, respectively) and plasma concentrations of E 1 S on gd20 related directly to the number of stillborn piglets (r = 0.60, P = 0.02). In summary, the number of CL on gd20 and 50 is a reliable marker of embryonic/fetal pig status. Measurements of P 4 and E 1 S on gd20 and 50 showed limited diagnostic value (ie, were not indicative of the number of healthy and abnormally developing embryos/fetuses). However, measurements of circulating P 4 and E 1 S concentrations during the periconceptional period and in the early/mid-pregnancy of sows have the makings of a practical method to predict gestational outcomes., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Nineteen ewes received 200 mg of pFSH administered in eight decreasing doses from Days 1 to 4, starting three days before CIDR® device removal. Ten ewes received an injection of 350 μg of estradiol benzoate at CIDR® device insertion (Group E) and nine animals served as controls (Group C). B-mode and spectral Doppler ultrasonographic examinations were performed daily throughout superovulatory treatment to enumerate ovarian antral follicles and to determine ovarian blood flow indices, respectively. There were no differences (P > 0.05) in superovulatory responses between left and right ovaries/uterine horns or the two groups of animals. End-diastolic velocity (EDV) and mean velocity (Vm) values were greater (P < 0.05) on Days 1 and 2, and peak systolic velocity (SVp) was greater (P < 0.05) on Day 3 in Group C than in Group E. In Group E 15 correlations was recorded among indices (SVp, Vm, EDV, flow velocity integral-FVI, and pulsatility index-PI) and follicles numbers in different size classes on Days 1, 2 and 4, and seven correlations among indices (SVp, EDV, Vm, and vascular resistance index-RI) and superovulatory/embryo results (numbers of regressing corpora lutea, numbers/percentages of degenerated embryos and viability rates) on Days 1, 2 and 3. In Group C, there were three correlations among EDV and RI and medium-sized/large follicle numbers on Days 1 and 3, and five correlations among indices (EDV, RI and PI) and superovulatory/embryo results (numbers of luteinized unovulated follicles, degenerated embryos and unfertilized eggs) on Days 2 or 4. There was a lack of consistency in the velocimetric correlates of antral follicle numbers and superovulatory responses between the left and right side. Therefore, the usefulness of ovarian arterial indices to predict ovine superovulatory outcomes remains equivocal and requires further confirmatory studies., (Copyright © 2019 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

The aim of this study was to assess the usefulness of nanowater (NW; water declusterized using cold plasma treatment) as a diluent for a commercial boar semen extender during the 15-day storage (Days 1 to 15) at 16-18 °C. Ejaculates collected from 8 boars were subjected to the standard evaluation and then diluted in the extender prepared with deionized water (DW) or NW to a final concentration of 3×10 9 spermatozoa/ml. The proportion of defective spermatozoa increased (P<0.05) from Day 10 to Day 15 of storage (22.8±16.6% to 41.8±26.4% in DW group and 18.6±11.7% to 34.8±25.4% in NW group) and it was significantly greater in DW group compared with NW group on Days 5 and 10 due mainly to a greater (P<0.05) number of mid-piece defects in semen stored in the DW-containing extender. Sperm progressive motility decreased (P<0.05) in both groups between Days 2 and 6, Days 6 and 10, and Days 10 and 12, whereas the percentage of motile spermatozoa declined (P<0.05) to Day 14 only in NW group. Sperm motility was greater (P<0.05) in NW group compared with DW group from Day 5 to Day 13. A decline in sperm progressive motility below 40% in all semen samples occurred by Day 11 in DW group and by Day 12 in NW group. The mean survival time of sperm at 37 °C ex situ was greater in NW group than in DW group on Day 5 (314±87 min compared with 284±87 min) and Day 10 (223±34 min compared with 182±27 min; NW group compared with DW group, respectively). There were no differences (P>0.05) between the two groups in the concentrations of alkaline phosphatase and aspartate aminotransferase in semen extender. To summarize, the use of NW as an extender diluent exerts cytoprotective effects on boar spermatozoa and delays a decline in sperm progressive motility., Competing Interests: Conflicts of interest: The authors have no conflict of interest to declare., (Copyright © The Author(s).)

The main goal of this preliminary study was to determine and compare ultrasonographic characteristics of the mammary gland in two genotypes of ewes varying in milk productivity at 2, 3 and 4 weeks after lambing. Ultrasonographic images of the udder were obtained using the 5.0- and 7.5-MHz transducers, in axial and coronal planes, in four low milk-yielding Polish Mountain sheep and six high milk-yielding Olkuska ewes. All ultrasonograms were subjected to computerized image analyses using commercially available image analytical software (Image ProPlus ® ; Media Cybernetics Inc., San Diego, CA, USA) to determine numerical pixel values (NPVs) and heterogeneity (pixel standard deviation-PSD) of the mammary gland parenchyma. During the 28-day period post-partum, the Olkuska sheep exceeded (P < 0.05) Polish Mountain ewes in milk productivity (31.6 ± 2.7 l and 25.0 ± 4.2 l, respectively; means ± SEMs) as estimated by the mean weight gains of suckling lambs. In animals examined with the 5.0-MHz transducer, mean NPVs of the mammary gland parenchyma in Olkuska ewes and mean PSD in both genotypes of ewes were lower (P < 0.05) before than after milking. In addition, PSD recorded both before and after milking were lower (P < 0.05) in the Polish Mountain compared with Olkuska breed. Mean PSD values for the mammary gland were less (P < 0.05) before than after milking in Polish Mountain ewes and they were greater (P < 0.05) in Olkuska compared with Polish Mountain ewes examined with the 7.5-MHz probe after milking. It can be concluded that milk quantity, histomorphology of the udder and ultrasound transducer frequency may all impinge on the echotextural characteristics of the mammary parenchyma in different breeds of sheep. Our observations warrant future studies of correlations between milk composition, mammary gland histophysiology and ultrasonographic image attributes of the mammary gland in ruminants., Competing Interests: Conflicts of interest:The authors have no conflict of interest to declare., (Copyright © The Author(s).)

This study examined the effects of bovine oviductal fluid (bOF) obtained during the follicular or luteal phase of the estrous cycle on ram sperm kinematics, capacitation status and plasma membrane (PM) integrity at various time points during the 24-h incubation period. Fresh ram spermatozoa were selected using the swim-up technique and then incubated separately with either follicular phase (FbOF) or luteal phase (LbOF) bovine oviductal fluid added to Fert-TALP medium (positive control - POSControl) or in Fert-TALP medium without capacitating agents (negative control - NEGControl) at 38 °C under 5% CO 2 . Incubation with FbOF or LbOF for 2 h and 4 h promoted an increase (P <  0.05) in most of the sperm motility parameters as compared with the NEGControl group, and bOF-induced changes in sperm kinematics were similar (P >  0.05) to those seen in the POSControl group. After 6 h of incubation, the stimulatory effect of FbOF or LbOF on ram sperm kinematics was no longer observed (P >  0.05). Sperm PM integrity was not affected (P >  0.05) by incubation in bOF-supplemented media or in absence of capacitating factors (NEGControl). Although neither FbOF nor LbOF had any effect on sperm capacitation rates, the proportion of acrosome-reacted spermatozoa was greater (P < 0.05) for bOF-containing media compared with the NEGControl group during the long incubation periods (18 h and 24 h). In conclusion, bOF from either follicular or luteal phase of the estrous cycle enhances ram sperm motility for up to 4 h and the rate of acrosome reaction after long (18-24 h) incubation periods without affecting sperm viability., (Copyright © 2019 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

This study compared the effects of intravaginal and intravenous routes of oxytocin (OT) administration in 46 oestrous-induced Santa Inês ewes (6-day treatment with progestin-releasing intravaginal sponges and a single injection of 200 IU of eCG at the time of sponge removal) that underwent transcervical embryo recovery 6-7 days after oestrous onset and mating. All ewes received 37.5 μg of d-cloprostenol via latero-vulvar route, and 1 mg of oestradiol benzoate i.m. 16 hr before and 50 IU of OT 20 min before non-surgical embryo recovery (NSER), with OT being administered intravenously (n = 21) or intravaginally (n = 21). An overall oestrous response was 95.6% (44/46), and adequate cervical retraction could be accomplished in 78.6% (33/42) of ewes. The percentage of successful NSER procedures was 57% (24/42) or 72.7% (24/33) of animals with sufficient cervical retraction. The duration of NSER procedure averaged 28 min (range: 17-40 min) and ~96% of flushing fluid could be recovered (range: 85%-100%). Out of 18 ewes that could not undergo NSER, 12 (66.6%) presented various anatomical barriers, whilst the other 33.4% did not present these barriers and still could not be traversed. Excluding the ewes with those anatomical features, the overall success rate of NSER was 80% (24/30). The route of OT administration had no effect on NSER efficiency or the ease with which transcervical embryo flushing was performed. Both routes of OT administration can be used for cervical dilation protocol. Discarding ewes with anatomical features precluding cervical penetration is highly recommended to increase the efficacy of NSER in sheep., (© 2019 Blackwell Verlag GmbH.)

This study examined the relationships among physicochemical properties and ultrasonographic image attributes of pectoralis major muscles in broiler chickens. Forty male Ross 308 chicks were randomly assigned to four equinumerous fat-supplementation groups (Group SO: soybean oil; Group FO: flax oil; Group SO + FO: soybean oil + flax oil; and Group BF: beef fat). Ultrasonograms of birds' pectoral muscles were obtained just before slaughter at 6 weeks of age and were subjected to digital image analyses to determine the mean pixel intensity (MPI) and pixel heterogeneity values (standard deviation of numerical pixel values; MPH). A total of 2, 4, 2, and 6 significant correlations were recorded in Groups SO, FO, SO + FO, and BF, respectively; there were no correlations with the chemical composition of the muscles in Groups SO and SO + FO. The strongest correlations were found between muscle lightness (L*) and MPH in Group BF (physical characteristic; r = -0.82, p = 0.003), and between crude fat/protein content and MPI/MPH of pectoral the major muscles in Groups FO/BF (chemical characteristics; r = 0.72, p = 0.02). There exists a potential application of ultrasonographic imaging and computerized image analysis for predicting certain physicochemical properties of pectoralis major muscles in broiler chickens.

The effects of estradiol esters, d-cloprostenol and oxytocin on induction of cervical dilation prior to non-surgical embryo recovery in Santa Inês ewes (Days 6-7 estrous cycle) were assessed in this study. In Trial 1, transcervical embryo flushing was performed in estrous-induced ewes administered 37.5 μg of d-cloprostenol i.m. 10 h before and 50 IU of oxytocin i.v. 20 min before uterine flushing with (EB-PGF-OT; n = 13) or without (PGF-OT; n = 11) 1 mg of estradiol benzoate i.m. administered concurrently with d-cloprostenol injection. In Trial 2, the estrous-synchronized animals were treated with 1 mg of estradiol benzoate (EB-PGF-OT; n = 12) or estradiol cypionate (EC-PGF-OT; n = 12) i.m. along with 37.5 μg of d-cloprostenol i.m. 16 h before and 50 IU of oxytocin i.v. 20 min before uterine flushing. In Trial 1, uterine flushing could be accomplished in 38% of ewes in the EB-PGF-OT and 27% those in the PGF-OT (P>0.05) group. Flushing fluid recovery averaged 90% and there were 1.0 ± 1.1 embryos/ewe collected with mean duration of the flushing procedure being ˜36 min. In Trial 2, uterine flushing was accomplished in 78% of ewes in the EB-PGF-OT and 44% of those in the EC-PGF-OT group (P>0.05) with mean flushing fluid recovery rate being 88% and time elapsing to complete flushing being ˜33 min. Within the subsets of animals treated with EB, the percentages of successful transcervical penetrations were 38% compared with 78% in Trials 1 and 2, respectively (i.e., with EB administered 10 h compared with 16 h before uterine flushing: P<0.05). The interval from EB administration to the beginning of transcervical penetration can affect the efficacy of embryo recovery procedures utilizing a combined EB/d-cloprostenol/oxytocin pre-treatment., (Copyright © 2019 Elsevier B.V. All rights reserved.)

There is a paucity of information on the relationships of testicular morphology, echotextural attributes, and blood flow dynamics with pubertal development of rams raised in a subtropical climate. Forty-five Dorper rams (24 rams aged 8-11 months and 21 rams aged 12-24 months) were examined using a portable ultrasound scanner connected to a 7.5-MHz transducer. Computer-assisted analyses of testicular ultrasonograms utilized commercially available Image ProPlus ® analytical software. Spectral Doppler scans of testicular arteries were performed immediately after scrotal (B-mode) ultrasonography to determine peak systolic velocity (PSV), end-diastolic velocity (EDV), resistive index (RI = [PSV-EDV]/PSV), and pulsatility index (PI = [SPV-EDV]/mean velocity) of the blood vessels. The length of the testes (9.7 ± 0.3 compared with 9.0 ± 0.2 cm) and scrotal circumference (33.3 ± 0.5 compared with 31.8 ± 0.4 cm) were greater (p < 0.05) but testicular depth (4.5 ± 0.1  compared with 4.9 ± 0.08 cm) was less (p < 0.05) in sexually mature compared with peripubertal rams. [Corrections added on 9 Jan 2019 after initial online publication: The testicular size values in the sentence were corrected.] There were no differences (p > 0.05) between the two age groups of Dorper rams in blood flow indices of testicular arteries. Mean numerical pixel values (100.5 ± 4.1 compared with 89.2 ± 4.8) and pixel heterogeneity (25.6 ± 0.6 compared with 23.6 ± 0.5) of testicular parenchyma were greater (p < 0.05) in peripubertal than in postpubertal rams. Semen volume was negatively correlated with PI of testicular arteries (r = -0.57, p = 0.04). In summary, the attainment of sexual maturity in the rams of the present study was associated with significant changes in testicular length and depth, scrotal circumference, and parenchymal echogenicity/hetrogeneity but not in testicular volume and blood perfusion rates. Testicular artery PI can be used to predict the volume of ejaculate in rams., (© 2018 Blackwell Verlag GmbH.)

This study examined the feasibility of transcervical embryo recovery after the hormonal treatment to induce cervical dilation, following the 7-day oestrous synchronization protocol in multiparous Santa Inês ewes. A total of 23 cyclic ewes received two doses of 37.5 μg of d-cloprostenol by latero-vulvar route 7 days apart. After the second injection of d-cloprostenol, the ewes were checked for oestrus (every 12 hr) and then mated by fertile rams throughout the oestrous period. All ewes received 37.5 μg of d-cloprostenol (latero-vulvar) and 1 mg of oestradiol benzoate by either intramuscular (EBim group; n = 12) or intravaginal (EBivg group; n = 11) route 16 hr before embryo flushing. Twenty minutes before the flushing, 50 IU of oxytocin were administered intravenously. The oestrous response (i.e., the percentage of ewes that showed signs of oestrous behaviour after the second d-cloprostenol injection) was 91.3% (21/23). The proportion of successfully penetrated ewes (81.8% compared with 80.0%), the mean duration of embryo flushing (24.7 ± 2.0 min compared 26.2 ± 1.9 min), the flushing fluid recovery rate (94.8 ± 1.3% compared with 91.0 ± 2.9%) and the average number of structures recovered per ewe (0.5 ± 0.4 compared with 0.8 ± 0.4) did not vary (p > 0.05) between the EBim and EBivg groups. Viable embryos were recovered from 41.2% (7/17) of successfully penetrated ewes. It can be concluded that nonsurgical (i.e., transcervical) embryo collection can be performed in oestrous-synchronized Santa Inês ewes pretreated with d-cloprostenol, oxytocin and oestradiol benzoate, with the latter hormone administered by either the intramuscular or intravaginal route., (© 2018 Blackwell Verlag GmbH.)

Developmental potential of cryopreserved in vitro matured oocytes is very low in nearly all mammalian species studied to date. Despite relatively high cleavage rates, the vitrified/warmed metaphase II oocytes have a decreased rate of blastocyst formation, which can be attributed to the elevated cytoplasmic lipid content and lipid droplet fragmentation. Secretory products of ampulla oviductal epithelial cells (AECs) at the periovulatory stage of the ovarian cycle enhance the viability of in vitro matured oocytes. The present study was undertaken to determine if co-culture of cumulus-oophorus complexes (COCs) with conspecific AECs or reducing the lipid content of in vitro matured ovine oocytes would improve their cryotolerance and ensuing developmental competence. Ovine COCs aspirated from the slaughterhouse ovaries were matured in the following media or culture conditions: TCM199 + FBS + AECs (T1); TCM199 + FBS (T2); TCM199 + BSA (T3); TCM199 + 0.6 mg/mL of l-carnitine (T4); TCM199+ l-carnitine + FBS (T5), or TCM199 only (Ctr). Subsequently, the oocytes were vitrified and used for in vitro fertilization (IVF). The lowest degree of zona pellucida (ZP) hardening following vitrification of in vitro matured sheep oocytes was observed in T1 and T5 (P < 0.05). Cleavage, blastocyst formation and ensuing development (i.e., total cell numbers) as well as blastocyst hatching rates were all greater (P < 0.05) in T1 compared with the remaining groups; in vitro matured COCs in T4 and Ctr did not develop beyond the cleavage stage. The inner cell mass: trophectoderm cell ratio in T1 (1:3.29) was significantly greater compared with T2 (1:3.39), T3 (1:3.40) and T5 (1:3.44). The present results indicate that the ovine COCs/AECs co-culture system had the most positive influence on cryotolerance, ZP hardening, and developmental competence of in vitro matured oocytes., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Objective was to evaluate the echotexture and characteristics during terminal development of canine foetal respiratory and hepatic systems through elastographic examinations. Fifteen pregnant bitches were evaluated by ultrasonography twice daily, from the 53rd gestational day until whelping, and images obtained from 120 to 0 h before parturition were analysed. Images of foetal lungs and liver were recorded and then used for computer-assisted analyses to determine quantitative attributes. Acoustic Radiation Force Impulse (ARFI) elastographic of internal organs were classified as 'soft' (white areas) or 'hard' (dark areas) and quantitative analyses determined the mean shear wave velocities (SWV) of foetal lungs and liver. After delivery, canine neonates were clinically evaluated, and their health status was monitored weekly until 60 days post-partum. Sonographic parameters over time were compared by ANOVA and Pearson's correlations were used to determine associations between SWVs and echotextural variables. Foetal lungs and liver had a homogeneous echotexture and pulmonary parenchyma appeared hyperechoic when compared with that of the liver. Mean numerical pixel values (NPVs) of lungs decreased from 120 to 24 h and subsequently increased until parturition (P = 0.04). Lungs and liver mean (± SD) SWVs (0.98 ± 0.12 and 0.84 ± 0.11 m/s, respectively) didn't vary (P >  0.05) over time. Fluctuations in pulmonary NPVs indicated there was a pattern corresponding to structural and functional changes that occur during the terminal stage of pre-natal canine development and hence can be a useful diagnostic tool in veterinary. Foetal lung and liver SWVs were relatively consistent and there was no detectable changes during the pre-partum period for this variable or in echotexture., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Azaperone can reduce stress caused by weaning and relocation of breeding sows, but its effects on reproductive processes are still poorly understood. The primary aim of this study was to describe and compare the endocrine and ovarian activity in ultrasonographically monitored second parity sows, with or without azaperone treatment at weaning (2 mg/kg BW i.m.). The intervals from weaning to the onset of estrus and ovulation were both greater (P<0.05) in azaperone-treated (n=12) than in control sows (n=12) by ~12 h. Mean daily growth rates of identified antral follicles were less (P<0.05) in azaperone-treated than in control sows (1.08±0.17 v.1.23±0.18 mm/day; mean±SD) and treated animals exceeded (P<0.05) controls in the mean ovulation rate (13.7±1.3 v. 12.6±1.2). A transient suppression of cortisol release was observed in the treatment group (at 10 and 30 min after azaperone injections) but circulating cortisol concentrations were variable in both groups of sows for the remainder of the study. The preovulatory rise in LH and estradiol secretion was delayed (P<0.05), and the duration of the LH surge was greater (P<0.001) in azaperone-treated sows compared with their control counterparts. The amplitude of episodic fluctuations in serum cortisol concentrations was correlated with the number of stillborn piglets in control sows (r=0.63, P=0.04). The amplitude and concentration of the preovulatory rise in estradiol secretion were negatively correlated with ovulatory response and litter size (r=-0.63 to -0.82, P<0.05), whereas the time at which the LH surge ended was directly related to the number of live-born piglets (r=0.82, P=0.002) in azaperone-treated animals. The present results indicate that administration of azaperone at weaning had a profound effect on preovulatory LH secretion as well as growth kinetics and estrogenicity of ovarian antral follicles. However, the causative associations among various characteristics of the preovulatory LH discharge, ovarian and adrenal steroid secretion post-weaning, and reproductive variables in sows remain equivocal.

Azaperone treatment can control aggression and decrease stress due to weaning, re-grouping and hierarchical fighting of gilts and sows. However, the effects of this butyrophenone neuroleptic and sedative administered at weaning on pig reproductive function are poorly characterized. In this year-long study, a total of 619 cross-bred sows (Polish Large White×Polish Landrace) kept on a commercial farm received an i.m. injection of azaperone (Stresnil®; 2 mg/kg BW) just before weaning and were artificially inseminated during the ensuing estrus with 3×109 spermatozoa per dose of an inseminate; 1180 sows served as untreated controls. Immediately after weaning, the sows were moved to four pens of seven to nine animals each. A teaser boar was used twice daily to check for estrus and sows were bred at heat detection. Subsequently, all sows stayed in individual stalls until pregnancy testing on day 30 post-artificial insemination and were then re-grouped until farrowing. The proportion of pigs that were in estrus within 6 days post-weaning was significantly lower in azaperone-treated groups of animals than in controls (71.4% v. 84.2%). Overall, the azaperone-treated sows had a significantly longer weaning-to-estrus interval (WEI; 8.7±10.1 v. 6.3±8.1 days; mean±SD) and a significantly larger litter size (LS: 11.8±3.0 v.11.3±3.2; azaperone-treated v. control sows). Treatment of the winter-farrowing sows was associated with increased LS (12.8±2.6 and 11.3±3.1 piglets/sow, respectively; P<0.05) and longer (P<0.05) weaning-to-effective-service intervals (11.7±19.3 and 8.4±12.3 days, respectively) as well as farrowing intervals (155.7±19.7 and 152.2±16.1 days, respectively) compared with untreated controls. In the summer months, significantly longer WEIs (12.1±21.0 v. 8.4±16.9 days) were accompanied by a significant decline in LS only in azaperone-treated sows that were inseminated within 6 days post-weaning (10.8±2.9 v. 11.5±3.3 piglets/sow; azaperone-treated v. controls). Azaperone-treated second parity sows had greater LS (P<0.001) along with prolonged WEIs (P<0.05) in comparison to their respective controls, regardless of the timing of estrus. An application of azaperone at weaning increased the annual piglet productivity of winter-farrowing animals and of second parity sows but depressed it significantly in summer. The extra cost and labor due to delayed onset of estrus may cancel out any reproductive benefits of azaperone treatment.