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The quality of porcine oocytes for use in IVF is commonly graded according to the number of layers of cumulus cells (CCs) surrounding the oocyte; together these form the cumulus oocyte complex (COC). At least three compact layers of CCs is regarded as important for efficient IVP. To test this, oocytes were scored according to cumulus investment, with grade A representing COCs with three or more cumulus layers including granulosa cell‐cumulus oocyte complexes, grade B those with an intact corona radiata surrounded by another layer of cumulus cells and grades C and D representing COCs with lower CC investment. These oocytes were then monitored for in vitro maturation (IVM), as assessed by tubulin immunostaining for meiotic progression, the development of a cortical granule ring, and by glutathione levels. Results indicate that grading correlates closely with nuclear maturation and cytoplasmic maturation, suggesting that grading oocytes by cumulus investment is a reliable method to predict IVM success. Importantly, Grade A and B oocytes showed no significant differences in any measure and hence using a cut‐off of two or more CC layers may be optimal. We also determined the effect of assisted aspiration for oocyte retrieval, comparing the effect of needle size and applied pressure on the retrieval rate. These data indicated that both variables affected oocyte recovery rates and the quality of recovered oocytes. In combination, these experiments indicate that grade A and B oocytes have a similar developmental potential and that the recovery of oocytes of these grades is maximised by use of an 18‐gauge needle and 50 mmHg aspiration pressure. [ABSTRACT FROM AUTHOR]

Introduction: Embryo cryopreservation is a valuable technique used for preserving genetic resources for long periods. However, the survival rate of embryos is dependent on the method used. Therefore, in this study, we evaluated the efficiency of slow freezing method but with an additional dehydration step prior to freezing to overcome the formation of ice crystals., Methods: Oocytes collected from the ovaries of native Korean cattle subjected to in vitro fertilization were cultured for 7 days until the formation of expanded blastocysts. Before freezing, the blastocysts were placed in four pre-equilibration media: a control medium with no addition of sucrose, and three experimental media with the addition of 0.1, 0.25, and 0.5 M sucrose, respectively. Then, the pre-equilibrated embryos were frozen. Embryo survival and hatching rates were evaluated morphologically at 24, 48, and 72 h after thawing. Immunofluorescence staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and gene expression analysis of the re-expanded blastocytes were examined 24 h after freeze-thawing., Results: The survival rate was significantly higher in the 0.1 M group than in the control group ( p  < 0.05), and the hatching rate at 72 h was significantly higher in the 0.25 and 0.5 M groups than in the control group ( p  < 0.05). TUNEL-positive cells were significantly lower in the 0.25 M group than in the control group (12.5 ± 0.9 vs. 8.3 ± 0.8; p  < 0.05). The gene expression of BCL2 associated X, heat shock protein 70 kDa, and aquaporin 3 in the 0.25 M group was significantly lower than that in the control group ( p  < 0.05)., Conclusion: Our study revealed that treatment with 0.25 M sucrose before slow freezing improved the viability of bovine embryos after freeze-thawing., Competing Interests: SJ, HS, and Y-GJ were employed by ET Biotech Co. Ltd. The remaining 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 Jung, Sul, Oh, Jung, Lee and Hyun.)

The use of in vitro embryo production (IVP) has increased globally, particularly in the United States. Although maternal factors influencing embryo development have been extensively studied, the influence of the sire is not well understood. Sperm plays a crucial role in embryo development providing DNA, triggering oocyte maturation, and aiding in mitosis. Current sire fertility measurements do not consistently align with embryo production outcomes. Low-fertility sires may perform well in IVP systems but produce fewer pregnancies. Testing sires in vitro could identify characteristics affecting embryo development and pregnancy loss risk in IVP and embryo transfer programs., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The objective of this study was to evaluate the effects of different concentrations of antifreeze protein (AFP) extracted from the larva of the beetle, Tenebrio molitor ( Tm AFP), on vitrification of in vitro -produced bovine embryos. In vitro -produced blastocysts were divided into three experimental groups and vitrified using a cryotop. Tm AFP was added to the equilibrium solution (ES) and vitrification solution (VS) at a concentration of 0 ng/mL (control), 500 ng/mL (500 Tm AFP), or 1000 ng/mL (1000 Tm AFP). Vitrification was carried out by first placing the blastocysts in ES for 2 minutes (7.5% ethylene glycol [EG] and 7.5% dimethyl sulfoxide [DMSO]). The blastocysts were then transferred to VS (15% EG and 15% DMSO) and promptly deposited on a cryotop stem and submerged in liquid nitrogen. Warming was carried out in three steps with decreasing sucrose concentrations. After warming, the blast cells were cultured for 24 hours for subsequent survival analysis and ultrastructural evaluation. There was a significant difference in the survival rate and expansion in the 500 Tm AFP group compared with the other groups. The ultrastructural analysis revealed intracellular lesions in all vitrified embryos; however, the embryos of the 500 Tm AFP and 1000 Tm AFP groups showed fewer cytoplasmic lesions compared with the control group. Taken together, addition of Tm AFP can mitigate cellular changes that involve organelles and cellular components essential for proper functioning and improve the viability of warmed and vitrified in vitro -produced bovine embryos.

Assisted reproduction techniques (ARTs) provide access to early stage embryos whose analysis and assessment deliver valuable information. The handling of embryos, including the in vitro production of bovine embryos, is a rapidly evolving area which nonetheless exposes the embryos to unnatural conditions for a period of time. The Fallopian tube provides innumerable quantitative and qualitative factors, all of which guarantee the successful development of the embryo. It is well known that the Fallopian tube can be bypassed, using embryo transfer, resulting in successful implantation in the target recipient animal and the birth of calves. However, the question arises as to whether such circumvention has a negative impact on the embryo during this sensitive development period. First crosstalk between the embryo and its environment confirms mutual recognition activities and indicate bilateral effects. Nowadays, in vitro production of bovine embryos is a well-established technology. However, it is still evident that in vitro generated embryos are not qualitatively comparable to embryos obtained ex vivo. To counteract these differences, comparative studies between in vitro and ex vivo embryos are advantageous, as embryos grown in their physiological environment can provide a blueprint or gold standard against which to compare embryos produced in vitro. Attempts to harness the bovine oviduct were sometimes very invasive and did not result in wide acceptance and routine use. Long-term development and refinement of transvaginal endoscopy for accessing the bovine oviduct has meanwhile been routinely applied for research as well as in practice. Comparative studies combining in vitro development with development in the cattle oviduct revealed that the environmental conditions to which the embryo is exposed before activation of the embryonic genome can have detrimental and lasting effects on its further development. These effects are manifested as deviations in gene expression profiles and methylation signatures as well as frequency of whole chromosomal or segmental aberrations. Furthermore, it was shown that hormonal superstimulation (multiple ovulation and embryo transfer), varying progesterone concentrations as well as metabolic disorders caused by high milk production, markedly affected embryo development in the postpartum period. Assisted reproductive techniques that allow the production and handling of extra numbers of generated embryos promise to have a very high impact on scientific and practical application. Any influence on the early embryonic life, both in animals and in vitro, is accompanied by a sensitive change in embryonic activity and should be assessed in vivo on the basis of physiological conditions before being used for ART.

This study aimed to evaluate the effect of the force and duration of centrifugation and the impact of cushioned centrifugation on sperm selection by Percoll gradient, on sperm quality and development kinetics of in vitro produced bovine embryos. Two experiments were performed. In Experiment I, a pool of semen was selected by Percoll gradients and the pellet was divided into four groups and distributed in a 2 × 2 factorial, with two forces (2200 × g or 9000 × g) and two durations (1 min or 3 min) of centrifugation. In Experiment II, semen was divided into two groups and selected by Percoll gradient with Cushion Fluid (CF) or without CF (Control) in the second centrifugation. The morphofunctionality, biochemical characteristics and fertilizing capacity of the selected sperms were evaluated. In addition, the development of the resulting bovine embryos was monitored for 48 h post-insemination. Duncan and Chi-square tests (P < 0.05) were used to compare the means. In Experiment I, there was a significant increase in sperm vigor (P < 0.05) after sperm selection in all treatments. The force and duration of centrifugation did not have any effect on sperm motility, vigor, and recovery rate among the different treatments (P > 0.05). In Experiment II, the recovery rate and reactive oxygen species (ROS) production in semen were similar among treatments (P > 0.05) although a higher ROS production was observed in the CF fertilization medium. Total fertilization rate was superior in the CF group (65.4 ± 5.3%) compared to that in Control (39.6 ± 4.9%). However, the normal fertilization and cleavage rate did not differ between the Control (94 ± 6.3% and 58.3 ± 8.3%) and CF (89 ± 7.1% and 75.0 ± 7.3%) groups. The reduction in the force and duration of centrifugation did not decrease the sperm recovery during selection by the Percoll gradient and the use of CF in the second centrifugation did not affect the normal fertilization and development of bovine IVF embryos up to 48 h.

Abstract: The objective of this work was to evaluate the response of an in vitro host range to Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV), a pathogenic virus to the fall armyworm (Spodoptera frugiperda, Lepidoptera: Noctuidae), for the further development of a biopesticide based on cell culture systems. The cell lines from Bombyx mori (BM-5), Lymantria dispar (IPLB-LD-625Y), Trichoplusia ni (BTI-Tn-5B1-4), Anticarsia gemmatalis (UFL-AG-286), and S. frugiperda (IPLB-SF-21AE and Sf9) were tested for their susceptibility to a highly-virulent Brazilian isolate of SfMNPV. The cytopathic effects induced by the virus, the production of viral particles, and the synthesis of viral polypeptides were examined and compared. Both S. frugiperda cell lines showed hypertrophy of cell nuclei and production of many polyhedra. The SDS-Page of radiolabed proteins showed that the cell protein synthesis was shutoff, while an intense band of about 30 kDa, recognized as polyhedrin, was synthesized. The other cell lines did not show polyhedra production, although some of them underwent morphological changes and protein synthesis shutdown in response to virus infection. The SF-21 and Sf9 cell lines are recommended for further in vitro production of SfMNPV.

Many factors influence the final oocyte maturation, fertilisation, and early embryo development, and there are both similarities and differences between species. When comparing the advancement of assisted reproductive technologies (ARTs), the development in the bovine species is not far behind the medical front, with around one million in vitro-produced bovine embryos each year. This rate of progress is not seen in the other domestic species. This review aims to give an overview of the development and specific difficulties of in vitro embryo production in various domestic animal species, with the main focus on cows, pigs, and cats. In production animals, the aim of ARTs is commonly to increase the genetic progress, not to treat reproductive failure. The ARTs are also used for preservation of genetic diversity for the future. However, specifically for oocyte maturation, fertilisation, and early embryonic development, domestic mammals such as the cow and pig can be used as models for humans. This is particularly attractive from an animal welfare point of view since bovine and porcine oocytes are available in large numbers from discarded slaughterhouse material, thereby decreasing the need for research animals. Both for researchers on the animal and human medical fronts, we aim for the development of in vitro production systems that will produce embryos and offspring that are no different from those conceived and developed in vivo. Species-comparative research and development can provide us with crucial knowledge to achieve this aim and hopefully help us avoid unnecessary problems in the future. [ABSTRACT FROM AUTHOR]

Assisted reproduction techniques (ARTs) provide access to early stage embryos whose analysis and assessment deliver valuable information. The handling of embryos, including the in vitro production of bovine embryos, is a rapidly evolving area which nonetheless exposes the embryos to unnatural conditions for a period of time. The Fallopian tube provides innumerable quantitative and qualitative factors, all of which guarantee the successful development of the embryo. It is well known that the Fallopian tube can be bypassed, using embryo transfer, resulting in successful implantation in the target recipient animal and the birth of calves. However, the question arises as to whether such circumvention has a negative impact on the embryo during this sensitive development period. First crosstalk between the embryo and its environment confirms mutual recognition activities and indicate bilateral effects. Nowadays, in vitro production of bovine embryos is a well-established technology. However, it is still evident that in vitro generated embryos are not qualitatively comparable to embryos obtained ex vivo. To counteract these differences, comparative studies between in vitro and ex vivo embryos are advantageous, as embryos grown in their physiological environment can provide a blueprint or gold standard against which to compare embryos produced in vitro. Attempts to harness the bovine oviduct were sometimes very invasive and did not result in wide acceptance and routine use. Long-term development and refinement of transvaginal endoscopy for accessing the bovine oviduct has meanwhile been routinely applied for research as well as in practice. Comparative studies combining in vitro development with development in the cattle oviduct revealed that the environmental conditions to which the embryo is exposed before activation of the embryonic genome can have detrimental and lasting effects on its further development. These effects are manifested as deviations in gene expression profiles and methylation signatures as well as frequency of whole chromosomal or segmental aberrations. Furthermore, it was shown that hormonal superstimulation (multiple ovulation and embryo transfer), varying progesterone concentrations as well as metabolic disorders caused by high milk production, markedly affected embryo development in the postpartum period. Assisted reproductive techniques that allow the production and handling of extra numbers of generated embryos promise to have a very high impact on scientific and practical application. Any influence on the early embryonic life, both in animals and in vitro , is accompanied by a sensitive change in embryonic activity and should be assessed in vivo on the basis of physiological conditions before being used for ART. [ABSTRACT FROM AUTHOR]

Some antioxidants and growth factors play an important role in promoting oocyte maturation and embryo development in many mammalian species, but there is little information about the yak (Bos grunniens). Therefore, the objective of this study was to evaluate Cys, insulin-like growth factor-1 (IGF-1) and epidermis growth factor (EGF) on yak oocyte maturation, cleavage and blastocyst rates after in vitro fertilized with Jersey sperm. A single or different combination of Cys, EGF and IGF-1 was added to in vitro maturation (IVM) and in vitro culture (IVC) media. The results showed that a single addition of Cys and IGF-1 increased oocyte maturation and blastocyst rates (p <.05), but did not increase cleavage rate; EGF or IGF-1 + EGF increased oocyte maturation, cleavage and blastocyst rates (p <.05) compared with the control. A combination of IGF-1 + EGF + Cys could have a beneficial effect (p <.05). These results indicated that supplementation of IVM and IVC media with Cys, IGF-1, EGF and their combinations could improve in vitro production efficiency of yak-cattle crossbred embryos. [ABSTRACT FROM AUTHOR]

The saponins are a diverse class of natural products, with a broad scale distribution across different plant species. Chemically characterized as triterpenoid glycosides, they posses a 30C oxidosqualene precursor-based aglycone moiety (sapogenin), to which glycosyl residues are subsequently attached to yield the corresponding saponin. Based on the chemically distinct aglycone moieties, broadly, they are divided into triterpenoid saponins (dammaranes, ursanes, oleananes, lupanes, hopanes, etc.) and the sterol glycosides. This review aims to present in detail the biosynthesis patterns of the different aglycones from a common precursor and their glycosylation patterns to yield the functionally active glycoside. The review also presents recent advances in the pharmacological activities of these saponins, particularly as potent anti-neoplastic pharmacophores, antioxidants, or anti-viral/antibacterial agents. Since alternate production pedestals for these pharmacologically important triterpenes via cell and tissue cultures are an attractive option for their sustainable production, recent trends in the variety and scale of in vitro production of plant triterpenoids have also been discussed. [ABSTRACT FROM AUTHOR]

Amorphophallus konjac (Araceae) a perennial herb, it has high medicinal and industrial value. In this study, a simple and efficient system for direct somatic embryogenesis and plantlet regeneration of Amorphophallus konjac was developed. The floral axis was used as the experimental material. The primary callus, developed from the floral axis grown on Murashige and Skoog (MS) medium supplemented with different hormone combination at different concentrations. The highest rate of embryogenic callus formation was observed on the MS medium containing 9.04 µM 2, 4-dichlorophenoxyacetic acid (2, 4-D) and 5.37 µM naphthalene acetic acid (NAA). The maximum induction rate was 79.8%, and the embryogenic calli were able to subculture on a medium containing similar hormone combination for over 1 year. The calli were also placed on different media for regeneration and it produced complete plants with shoots and root systems simultaneously. The highest differentiation rate of the embryogenic calli grown on differentiation medium supplemented with 8.88 µM 6-benzylaminopurine (6-BA) and 5.37 µM NAA was 95.6%. Flow cytometry analysis showed no ploidy variation in all the regenerate plantlets.

The entomopathogenic nematode Heterorhabditis bacteriophora (Nematoda: Rhabditidae) is used in biological insect control. Their dauer juveniles (DJs) are free-living and developmentally arrested, invading host insects. They carry cells of their bacterial symbiont Photorhabdus spp. in the intestine. Once inside the insect´s hemolymph the DJs perceive a food signal, triggering them to exit the DJ stage and regurgitate the Photorhabdus cells into the insect's haemocoel, which kill the host and later provide essential nutrients for nematode reproduction. The exit from the DJ stage is called "recovery". For commercial pest control, nematodes are industrially produced in monoxenic liquid cultures. Artificial media are incubated with Photorhabdus before DJs are added. In absence of the insect's food signal, DJs depend on unknown bacterial food signals to trigger exit of the DJ stage. A synchronized and high DJ recovery determines the success of the industrial in vitro production and can significantly vary between nematode strains, inbred lines and mutants. In this study, fourteen bacterial strains from H. bacteriophora were isolated and identified as P. laumondii, P. kayaii and P. thracensis. Although the influence of bacterial supernatants on the DJ recovery of three inbred lines and two mutants differed significantly, the bacterial impact on recovery has a subordinate role whereas nematode factors have a superior influence. Recovery of inbred lines decreased with age of the DJs. One mutant (M31) had very high recovery in bacterial supernatant and spontaneous recovery in Ringer solution. Another mutant (M88) was recovery defective., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

The pig is an important livestock animal. Biotechnological interest in this species has increased due to its use, among others, in the generation of transgenic animals for use in biomedicine based on its greater physiological proximity to the human species than other large domestic animals. This development has paralleled an improvement in Assisted Reproduction Techniques (ART) used for this species. However, the ability to generate animals from embryos produced entirely in vitro is still limited and a wide margin for improvement remains. Here we review the procedures, additives, and devices used during pig in vitro fertilization (IVF), focusing on the main points of each step that have offered the best results in terms of increased efficiency of the system. The lack of standardized protocols and consensus on the parameters to be assessed makes it difficult to compare results across different studies, but some conclusions are drawn from the literature. We anticipate that new physiological protocols will advance the field of swine IVF, including induction of prefertilization ZP hardening with oviductal fluid, sperm preparation by swim-up method, increased viscosity through the addition of inert molecules or reproductive biofluids, and the incorporation of 3D devices. Here we also reflect on the need to expand the variables on which the efficiency of pig IVF is based, providing new parameters that should be considered to supply more objective and quantitative assessment of IVF additives and protocols. [ABSTRACT FROM AUTHOR]

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In vitro embryo production (IVP) in cattle has gained worldwide interest in recent years, but the efficiency of using IVP embryos for calf production is far from optimal. This review will examine the pregnancy retention rates of IVP embryos and explore causes for pregnancy failures. Based on work completed over the past 25 yr, only 27% of cattle receiving IVP embryos will produce a live calf. Approximately 60% of these pregnancies fail during the first 6 wk of gestation. When compared with embryos generated by superovulation, pregnancy rates are 10% to 40% lower for cattle carrying IVP embryos, exemplifying that IVP embryos are consistently less competent than in vivo-generated embryos. Several abnormalities have been observed in the morphology of IVP conceptuses. After transfer, IVP embryos are less likely to undergo conceptus elongation, have reduced embryonic disk diameter, and have compromised yolk sac development. Marginal binucleate cell development, cotyledon development, and placental vascularization have also been documented, and these abnormalities are associated with altered fetal growth trajectories. Additionally, in vitro culture conditions increase the risk of large offspring syndrome. Further work is needed to decipher how the embryo culture environment alters post-transfer embryo development and survival. The risk of these neonatal disorders has been reduced by the use of serum-free synthetic oviductal fluid media formations and culture in low oxygen tension. However, alterations are still evident in IVP oocyte and embryo transcript abundances, timing of embryonic cleavage events and blastulation, incidence of aneuploidy, and embryonic methylation status. The inclusion of oviductal and uterine-derived embryokines in culture media is being examined as one way to improve the competency of IVP embryos. To conclude, the evidence presented herein clearly shows that bovine IVP systems still must be refined to make it an economical technology in cattle production systems. However, the current shortcomings do not negate its current value for certain embryo production needs and for investigating early embryonic development in cattle. [ABSTRACT FROM AUTHOR]

This study assessed the lipid composition of oocytes from different follicle sizes and compared the expression of lipid-related genes and follicular fluid (FF) molecules between groups. We also investigated the functional consequences of differences on embryo development and blastocyst lipid deposits. Oocytes and FF were recovered from different follicle sizes. Oocytes from small (≤5 mm) and large (≥6mm) bovine follicles were used to produce Day 7 expanded blastocysts (Day7Ex) and blastocysts that only became expanded at Day 8 (Day8Ex) after insemination. Oocytes from >8 mm follicles had the highest lipid content. Few oocyte phospholipid variations were identified between groups. Very long chain fatty acid elongase 6 (ELOVL6) mRNA abundance was reduced in larger follicle-derived oocytes compared with the ≤2mm group. Increased levels of glucose, reactive oxygen species, glutathione and superoxide dismutase activity were also identified in FF from larger follicles. Large follicle-derived embryo development and lipid content of Day7Ex were greater than those derived from small follicles. Day8Ex had greater lipid deposition than Day7Ex. Oocytes and blastocysts exhibited follicle size-specific lipids. Large-follicle oocytes had increased lipid content and became Day7Ex with greater lipid deposition whereas delayed blastocoel expansion associated with a prolonged period of culture determined the lipid accumulation of Day8Ex. The FF microenvironment of large follicles seems to favour embryo development. Oocyte development is a highly complex process. Lipid composition of oocytes derived from different follicles sizes and their relationship with follicular fluid molecules, embryo development and blastocyst lipid deposit were identified. We suggest that maintaining the exposure of the oocyte to the follicular microenvironment allows oocytes to proceed with events that are necessary to reach maturity and the positive impact of lipid accumulation for development may be seen as a necessary step to the forthcoming intense proliferative activity of the growing embryo. [ABSTRACT FROM AUTHOR]

The efficiency of producing embryos using in vitro technologies in livestock species rarely exceeds the 30–40% threshold, indicating that the proportion of oocytes that fail to develop after in vitro fertilization and culture is considerably large. Considering that the intrinsic quality of the oocyte is one of the main factors affecting blastocyst yield, the precise identification of noninvasive cellular or molecular markers that predict oocyte competence is of major interest to research and practical applications. The aim of this review was to explore the current literature on different noninvasive markers associated with oocyte quality in the bovine model. Apart from some controversial findings, the presence of cycle-related structures in ovaries, a follicle size between 6 and 10 mm, large number of surrounding cumulus cells, slightly expanded investment without dark areas, large oocyte diameter (>120 microns), dark cytoplasm, and the presence of a round and smooth first polar body have been associated with better competence. In addition, the combination of oocyte and zygote selection via brilliant cresyl blue (BCB) test, spindle imaging, and the anti-Stokes Raman scattering microscopy together with studies decoding molecular cues in oocyte maturation have the potential to further optimize the identification of oocytes with better developmental competence for in-vitro-derived technologies in livestock species.

This study is performed to evaluate the effect of insulin in the porcine parthenogenetic embryo development. In porcine embryo culture, insulin is helpful factor in the process of embryo development. To identify this, insulin is used in pig embryos development. Therefore, this study was performed to investigate the effect of insulin on early embryonic development in pigs. For that, insulin positive or negative (0, 10 ug/mL) was supplemented in the porcine IVM media and then compared two groups divided by the cytoplasm of the black groups and white ring groups based on the distribution of lipid material of the cell cytoplasm in microscope. In maturation rates of porcine oocytes, significant higher black group rates were shown in the insulin positive groups compared with other groups (56.0±2.1 vs 46.2±0.3). In the embryo culture, black groups were showed the significant higher cleavage rates (82.1±0.8, 78.3±0.1 vs 63.2±0.3, 63.4±0.0), and blastocyst formation rates (15.5±3.6, 16.6±0.4 vs 11.7±1.3, 7.4±0.2) regardless of whether the addition of insulin. Also, black groups were showed higher cell number of blastocyst (33.2±2.5, 35.5±2.6 vs 31.2±2.1, 31.3±2.2). In conclusion, supplement of insulin producing black group in vitro maturation, it was effective in vitro maturation and embryonic development of pig embryos.

Although piglets have been delivered by embryo transfer (ET) with in vitro produced (IVP) embryos and blastocysts, a success rate has still remained lower level. Unlike mouse, human, and bovine, it is difficult to a production of piglets by in vitro fertilization (IVF) because of an inappropriate in vitro culture (IVC) system in pig. Therefore, the present study was conducted to investigate whether minimized exposure time in IVC can improve the pregnancy and delivery rates of piglets. Immediately after IVM, the oocytes were denuded and co-incubated with freshly ejaculated boar semen for 3.5 to 4 hours at 38.5 °C under 5% CO2 in air. To avoid long-term exposure to in vitro state, we emitted IVC step after IVF. After that the presumptive zygotes were transferred into both oviducts of the surrogate on the same day or 1 day after the onset of estrus. Pregnancy was diagnosed on day 28 after ET and then was checked regularly every month by ultrasound examination. The 3 out of 4 surrogates were determined as pregnant (75%) and a total of 5 piglets (2 females and 3 males) were delivered at 118.3 ± 2.5 days of pregnancy period. In conclusion, a short-term exposure time may be an important factor in the production of IVP-derived piglets. It can be apply to the in vitro production system of transgenic pig by IVF, cloning, and pronuclear microinjection methods.

The severely decreased platelet counts (10–30. 103 platelets/μL) frequently observed in patients undergoing chemotherapy, radiation treatment, or organ transplantation are associated with life-threatening increased bleeding risks. To circumvent these risks, platelet transfusion remains the treatment of choice, despite some limitations which include a limited shelf-life, storage-related deterioration, the development of alloantibodies in recipients and the transmission of infectious diseases. A sustained demand has evolved in recent years for controlled blood products, free of infectious, inflammatory, and immune risks. As a consequence, the challenge for blood centers in the near future will be to ensure an adequate supply of blood platelets, which calls for a reassessment of our transfusion models. To meet this challenge, many laboratories are now turning their research efforts toward the in vitro and customized production of blood platelets. In recent years, there has been a major enthusiasm for the cultured platelet production, as illustrated by the number of reviews that have appeared in recent years. The focus of the present review is to critically asses the arguments put forward in support of the culture of platelets for transfusion purposes. In light of this, we will recapitulate the main advances in this quickly evolving field, while noting the technical limitations to overcome to make cultured platelet a transfusional alternative.

Summary: Pluripotency-associated transcription factors (PATFs) modulate gene expression during early mammalian embryogenesis. Despite a strong understanding of PATFs during mouse embryogenesis, limited progress has been made in ruminants. This work aimed to describe the temporal expression of eight PATFs during both sheep and cattle preimplantation development. Transcript availability of PATFs was evaluated by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) in eggs, cleavage-stage embryos, morulae, and blastocysts. Transcripts of five genes were detected in all developmental stages of both species (KLF5, OCT4, RONIN, ZFP281, and ZFX). Furthermore, CMYC was detected in all cattle samples but was found from cleavage-stage onwards in sheep. In contrast, NR0B1 was detected in all sheep samples but was not detected in cattle morulae. GLIS1 displayed the most significant variation in temporal expression between species, as this PATF was only detected in cattle eggs and sheep cleavage-stage embryos and blastocysts. In silico analysis suggested that cattle and sheep PATFs share similar size, isometric point and molecular weight. A phenetic analysis showed two patterns of PATF clustering between cattle and sheep, among several mammalian species. In conclusion, the temporal expression of pluripotency-associated transcription factors differs between sheep and cattle, suggesting species-specific regulation during preimplantation development. [ABSTRACT FROM AUTHOR]

Flavonoids are small molecular secondary metabolites synthesized by plants with various biological activities such as anti-inflammatory, anti-carcinogenic, antioxidant, etc. Flavonoids occur naturally in fruits, vegetables, medicinal plants and beverages such as tea and wine. Biotechnology offers different in vitro systems which have been developed to exploit these active ingredients such as, in callus cultures, cell suspension cultures which are the emerging fields of biotechnology to investigate and enhance the production of these products. Along with this, hairy root culture and transformation techniques have been studied widely for flavonoid production. Nano-treatment is a novel way for improvement of flavonoids production. the present review focus on describing the flavonoids biochemistry, regulation of biosynthesis, as well as the Biotechnology and Nano-biotechnology tools for in vitro production of flavonoids. [ABSTRACT FROM AUTHOR]

The objective of this study was to find the optimal proportions of pregnancies from an in vitro-produced embryo transfer (IVP-ET) system and artificial insemination (AI) so that profitability is maximized over a range of prices for embryos and surplus dairy heifer calves. An existing stochastic, dynamic dairy model with genetic merits of 12 traits was adapted for scenarios where 0 to 100% of the eligible females in the herd were impregnated, in increments of 10%, using IVP-ET (ET0 to ET100, 11 scenarios). Oocytes were collected from the top donors selected for the trait lifetime net merit (NM$) and fertilized with sexed semen to produce IVP embryos. Due to their greater conception rates, first ranked were eligible heifer recipients based on lowest number of unsuccessful inseminations or embryo transfers, and then on age. Next, eligible cow recipients were ranked based on the greatest average estimated breeding values (EBV) of the traits cow conception rate and daughter pregnancy rate. Animals that were not recipients of IVP embryos received conventional semen through AI, except that the top 50% of heifers ranked for EBV of NM$ were inseminated with sexed semen for the first 2 AI. The economically optimal proportions of IVP-ET were determined using sensitivity analysis performed for 24 price sets involving 6 different selling prices of surplus dairy heifer calves at approximately 105 d of age and 4 different prices of IVP embryos. The model was run for 15 yr after the start of the IVP-ET program for each scenario. The mean ± standard error of true breeding values of NM$ of all cows in the herd in yr 15 was greater by $603 ± 2 per cow per year for ET100 when compared with ET0. The optimal proportion of IVP-ET ranged from ET100 (for surplus dairy heifer calves sold for ≥$300 along with an additional premium based on their EBV of NM$ and a ≤$100 embryo price) to as low as ET0 (surplus dairy heifer calves sold at $300 with a $200 embryo price). For the default assumptions, the profit/ cow in yr 15 was greater by $337, $215, $116, and $69 compared with ET0 when embryo prices were $50, $100, $150, and $200. The optimal use of IVP-ET was 100, 100, 62, and 36% of all breedings for these embryo prices, respectively. At the input price of $165 for an IVP embryo, the difference in the net present value of yr 15 profit between ET40 (optimal scenario) and ET0 was $33 per cow. In conclusion, some use of IVP-ET was profitable for a wide range of IVP-ET prices and values of surplus dairy heifer calves. [ABSTRACT FROM AUTHOR]

Higher plants undergo a variety of stresses and to combat those stresses they acclimatize themselves by producing diverse secondary metabolites. These secondary metabolites also have a wide range of industrial applications and hence they serve as candidates for commercialization. Owing to the constraints faced by natural plant extraction, plant cell/tissue culture has emerged as an alternative platform for the in vitro production of value added bioactive secondary metabolites. Implementation of several productivity enhancement strategies, including elicitation, can overcome the limitations faced by plant cell technology that hampers its extensive commercialization. Elicitation is a technique that involves exogenous addition of elicitors (abiotic or biotic) in the growth medium which consequently triggers stress response with concomitant enhancement in secondary metabolite production. Elicitor induced stress results in the activation of several defense-related genes or inactivation of non-defense-related genes, transient phosphorylation/dephosphorylation of proteins, expression of enzymes whose information can be used to ascertain the biosynthetic pathways of many secondary metabolites. Furthermore, integration of transcriptomics, proteomics and metabolomics with system biology can aid in discovery of novel genes, transcriptional factors and several biosynthetic pathways which in turn can serve as a valuable tool for metabolic engineering and gene manipulation for enhancing the yield and productivity of secondary metabolites. [ABSTRACT FROM AUTHOR]

• A strategy to improve the sustainability of the dairy sector is required. • Sex-sorted semen from dairy breed bulls reduces the number of male dairy calves. • Use of beef breed bulls will increase the beef merit of non-replacement calves. • Genetic gain (dairy and beef breeds) is accelerated by in vitro-produced embryos. • Reducing the number of male dairy calves could have transformative effect on the sustainability of milk and beef production. A unique aspect of seasonal-calving pasture-based systems of dairy production is the intense focus placed on achieving a concentrated herd-calving period in late winter and early spring. Hence, excellent reproductive performance is required during a short breeding period. A concentrated calving period also produces a problem in the form of a large number of male dairy calves being born at the same time; as these calves have little economic value due to poor beef merit, they present a potential welfare concern. A solution exists in the form of sex-sorted semen, but this is typically associated with poorer pregnancy per artificial insemination, and hence, the use of sex-sorted semen must be carefully considered. The logical strategy to use sex-sorted semen is to target the best genetic merit dams in the herd to generate replacement heifers, thereby accelerating herd genetic gain. On the other hand, if all dairy farmers adopt such a strategy, there will be a corresponding reduction in elite genetic merit male dairy calves being born, potentially reducing availability of the next generation of future bulls to be used for artificial insemination. Use of in vitro embryo production on elite dairy donors could avoid this problem by acting as a multiplier, potentially in tandem with Y-sorted semen to skew the offspring sex ratio towards more male calves. Use of sex-sorted semen on the best genetic merit dams can also facilitate a marked increase in the usage of beef semen on any dams that are deemed unsuitable for sex-sorted semen. The use of "beef on dairy" requires selection of beef bulls that generate offspring with traits that meet the key requirements of both the dairy farmer (e.g., gestation length and calving ease) and the beef farmer that must be motivated to purchase the calves (e.g., growth rate, age at slaughter, carcass value). Beef breed dams that have elite genetic merit for these traits could also be considered for in vitro embryo production, potentially in tandem with Y-sorted semen, to facilitate genetic gain for the growing "beef-on-dairy" market. It is possible to transfer a beef embryo (75–100% beef breed genetics) into dairy dams that are not required to generate replacements, but this is likely to remain a niche practice as there are many barriers to widespread adoption. Such combinations of assisted reproduction have the potential to improve the efficiency and sustainability metrics of seasonal-calving pasture-based dairy herds. [ABSTRACT FROM AUTHOR]

• Embryo transfer can improve genetic selection, fertility and postnatal phenotypes. • Overcoming inefficiencies in the system will make the practice more widespread. • One goal is to increase the number of oocytes from genetically elite females. • Another is to increase the efficiency of generating a live calf from each oocyte. • It may be possible to program development to optimize calf postnatal phenotype. The embryo produced by in vitro oocyte maturation, fertilization, and embryonic development is an important resource for genetic improvement and has the potential to improve female fertility and to be programmed to produce offspring with superior ability for health and production. The cultured embryo is also an important component of several realized and potential technologies such as gene editing, somatic cell nuclear cloning, stem cell technologies and gamete generation in vitro. Full realization of the opportunities afforded by the in vitro -produced embryo will require overcoming some technical obstacles to cost-effective implementation of an embryo transfer program. Among the research goals for improving the penetration of embryo transfer in the cattle industry are development of methods to increase the supply of oocytes from genetically elite females, enhance the proportion of oocytes that become transferrable embryos, improve the fraction of embryos that establish pregnancy after transfer, reduce pregnancy wastage after pregnancy diagnosis, and identify culture conditions to optimize postnatal phenotype. [ABSTRACT FROM AUTHOR]

• Overgrown offspring increase the incidence of dystocia and have low survival. • In vitro-produced offspring may experience large offspring syndrome. • Assisted reproductive technologies induce epimutations related with overgrowth. • Technological improvements are required to reduce the incidence of overgrowth. • Development of diagnostic tools for the early identification of overgrowth is crucial. Large/abnormal Offspring Syndrome (LOS/AOS) is a congenital overgrowth condition of cattle and sheep, characterized by macrosomia, abdominal wall defects, organomegaly, difficulty to stand and suckle at parturition. The condition was first described as an exclusive consequence of assisted reproductive technologies, such as in vitro production and somatic cell nuclear transfer (cloning). However, we recently reported the spontaneous occurrence of this syndrome in cattle. The etiology of LOS is unclear, although the syndrome is an epigenetic condition characterized by multi-locus loss-of-imprinting, global dysregulation of small and long RNAs, changes in DNA methylation, and altered chromosomal architecture. These molecular and epigenetic changes affect biological pathways implicated in organ size, cell proliferation, cell survival, resulting in the phenotypes which characterize LOS. The lack of accurate tools for the prediction and diagnosis of LOS and the prevention of dystocia resulting from fetal overgrowth is a major concern for the dairy and beef industries. Furthermore, death of the calf and/or dam during calving adds animal welfare issues and affects the net income of the industry. An early diagnosis of LOS/AOS during gestation is critical to facilitate the decision-making process on whether to allow the pregnancy to continue or not in order to prevent harm to the dam as well as to provide producers with the timely necessary information to prepare for a difficult birth. The present review summarizes the definition, traits, incidence, and molecular characteristics of LOS to provide information and serve as a guide for future investigations regarding the early identification of LOS during pregnancy in cattle. [ABSTRACT FROM AUTHOR]

In porcine embryo culture, one of reactive oxygen species (ROS) is harmful factors that are made during in vitro culture. To decrease the detrimental effect of ROS on embryo development, superoxide dismutase, catalase and glutathione peroxidase could be used in the embryo culture. Out of these antioxidants, 7,8-dihydroxyflavone (7,8-DHF) was reported its antioxidant effects to prevent the glutamine-triggered apoptosis. Therefore, this study was performed to investigate the most appropriate concentration of 7,8-DHF in porcine embryonic development. For that, 5 different concentration (0, 0.1, 0.5, 1, 2 uM) of 7,8-DHF was supplemented in the porcine IVM media and then maturation and blastocyst formation rates were compared among 5 groups. In maturation rates of porcine oocytes, significant higher maturation rates was shown in the 1.0 uM group compared with another 4 groups (83.3 ± 2.1 vs. 80.7 ± 1.4, 79.8 ± 1.4, 78.3 ± 1.2, 79.4 ± 1.6), respectively (P

Defective virus accumulations during baculovirus passages in insect cell culture are impediments to large scale baculovirus production. A genotypically defined virus inoculum comprises of stable genotypes was proposed for production of a Thailand isolated SeMNPV in Se-UCR1 insect cells. Targeted genotypes were from wild-type SeMNPV containing naturally mixed genotypes. Plaque assays, PCR screening and Xba I restriction analysis were employed for genotype purification, genotype selection and genome analysis, respectively. A selective marker was pif2 encoded per os infection factor which predominantly deleted, along with the adjacent pif1 , in defective viruses. A purified, genetically stable pif2+ (and pif1 +) genotype, namely SeTh pif2 +, was the first tryout. SeTh pif2 + occlusion bodies (OBs) possessed insecticidal activity but at lower level than the wild-type. When the SeTh pif2 + was co-infected with another purified, genetically stable pif1- (and pif2 -) genotype, SeTh pif2 -, at ratio of 3:1, respectively, mixed genotypes OBs had 2.8 times greater insecticidal activity than the SeTh pif2 + alone. Dilution of deleterious PIF1 of SeTh pif2 + by the pif1 deletion genotypes, SeTh pif2 -, was the key for this enhanced activity. A promising approach was described for SeMNPV production in vitro using the virus inoculum whose genotypes compositions were designed to mimic virus interactions in the wild-type, to generate per oral infective baculovirus. [ABSTRACT FROM AUTHOR]

Bananas and plantains ( Musa L. species) have medicinal applications against diseases such as hypoglycemia, hypertension, and neurological disorders, especially Alzheimer's disease. The demand for these plants is growing very fast, resulting in a relatively heavy load on Musaceae genetic resources. The study evaluated and compared the acetylcholinesterase (AChE) inhibition and antioxidant activity of field and in vitro plant materials of nine accessions of Musa spp. consisting of Tropical Musa banana (TMb: TMb 106, TMb 145, TMb 8, TMb 82, TMb 55) and Tropical Musa plantain (TMp: TMp 116, TMp 24, TMp 36) from the International Institute of Tropical Agriculture and Musa sapientum (MS) from the University of Ibadan Botanical garden, Nigeria. Musa accessions were estimated onto Murashige and Skoog (MS) medium supplemented with 0.18 mg/L indole acetic acid (IAA) and 4.5 mg/L benzyl amino purine (BAP). The in vitro grown accessions behaved differently with TMb 8 having the highest average shoot length of 5.03 ± 0.66 cm, and average number of leaves of 5.65 ± 0.38 cm at the end of 6 weeks. Leaf extracts provide more quantity of phenolics, flavonoids and higher 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity than the fruit extracts. The AChEI activity of the field plants ranged from 60.54 ± 0.54 to 98.46 ± 0.09 % and in vitro plants from 61.88 ± 0.11 to 76.60 ± 0.34 % at 200 µg/mL. Crude methanol extract (CME) of the in vitro plants showed higher DPPH antioxidant activity than the field plants with IC (extract concentration providing 50 % inhibition) values ranging from 9.57 ± 0.24 to 48.37 ± 0.62 µg/mL compared with CME of the field samples, which had IC ranging from 75.86 ± 1.76 to 162.20 ± 3.77 µg/mL. Plant tissue culture can be a reliable alternative cultivation method for mass propagation and conservation of Musa species for the production of antioxidant and acetylcholinesterase metabolites. [ABSTRACT FROM AUTHOR]

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Considering the advent of mesenchymal stem cells (MSCs) as a new source of somatic cells in embryo co-culture system, the current study was aimed to compare in vitro embryo development using embryonic MSCs monolayer with embryonic fibroblast cells (EFCs), oviductal epithelial cells (OECs), and cell-free culture system. The IVM/IVF presumptive sheep zygotes were randomly cultured in different culture conditions as follows: (1) SOFaaBSA medium for the whole culture period (SOF, n = 371), (2) SOFaaBSA medium for the first 3 days followed by co-culturing with MSCs for the next 5 days (SOF-MSCs, n = 120), (3) co-culturing with MSCs for the first 3 days followed by culture in SOFaaBSA medium for the next 5 days (MSCs-SOF, n = 133), (4) co-culturing with MSCs for the whole culture period (MSCs, n = 212), (5) SOFaaBSA medium for the first 3 days followed by co-culturing with EFCs for the next 5 days (SOF-EFCs, n = 132), (6) co-culturing with EFCs for the first 3 days followed by culture in SOFaaBSA medium for the next 5 days (EFCs-SOF, n = 165), (7) co-culturing with EFCs for the whole culture period (EFCs, n = 236), and (8) co-culturing with OECs for the whole culture period (OECs, n = 255). One-Way ANOVA by multiple pairwise comparisons using Tukey's test was performed. Co-culturing in MSCs group had no superiority over EFCs and OECs groups. Though, when co-culturing with MSCs and EFCs was limited to the first 3 days of culture, the embryo development indices were improved compared to the other co-cultured groups. Considering both the hatching rate and total cell number, the application of MSCs for the first 3 days of culture (MSCs-SOF) was superior to the other co-culture and SOF groups.

El objetivo del estudio fue evaluar el efecto de dos medios de cultivo en la tasa de maduración y fertilización de ovocitos de cabra. Los complejos cumulus ovocito (CCO´s) fueron obtenidos de ovarios de cabras post-mortem. En el experimento I (EXP I), 361 CCO´s fueron divididos al azar para su maduración en TCM-199 (n=171) y HECM-9 (n=190). Ambos medios fueron suplementados con LH y FSH (10µg/ml) y 50µg/ml de gentamicina. Los CCO´s fueron cultivados a 38.5 °C, 5% CO2 y a las 27 h post-maduración, fueron lavados en PBS y teñidos con Hoescht 33342 para evaluar la tasa de maduración. En el experimento II (EXP II) 667 CCO´s fueron distribuidos en TCM-199 (n=323) y HECM-9 (n=344), y 30 ovocitos de cada tratamiento fueron evaluados como en el EXP I, el resto fueron co-incubados con 2x104 espermatozoides caprinos en medio TBM + 0.03% de BSA para evaluar la tasa de fertilización a las 36 h después. En el EXP I, la tasa de maduración nuclear fue superior (p

This paper reviews the technical bases of in vitro embryo production in small ruminants with special attention to the results obtained by our group in Northeastern Brazil. The laparoscopic oocyte recovery in hormonally treated live animals indicates a promising future for the application of this technique to genetic improvement program. New molecular biology tools should provide information to improve the efficiency of in vitro maturation. Furthers efforts have to be made to improve the oocyte maturation and to standardize the semen-capacitating process.Este artigo revisa as bases técnicas da produção in vitro de embriões em pequenos ruminantes com especial atenção aos resultados obtidos por nosso grupo no Nordeste do Brasil. A colheita dos oócitos por laparoscopia em animais vivos e hormonalmente tratados aponta para um futuro promissor da aplicação dessa técnica em programas de melhoramento genético. Novas ferramentas de biologia molecular devem fornecer informações para melhorar a eficiência da maturação in vitro. Esforços posteriores devem ser realizados para melhorar os índices de maturação oocitária e para padronizar o processo de capacitação espermática.

In this study, the kinetic behavior of Sf9 and Sf21 cells used in the production of a baculovirus biopesticide to control the pest of corn Spodoptera frugiperda was analyzed. Kinetic variables such as maximum specific growth rate, cell productivity, mean rate of infection, as well as the mean rate of occlusion body production were determined during the infection of these cell-lines with the extracellular virus of the S. frugiperda nucleopolyhedrovirus (SfMNPV). The Sf9 cell-line resulted in better viral production results (5.0 x 10(8) OB/mL) than the Sf21 cell-line (2.5 x 10(8) OB/mL).Neste trabalho, analisou-se o comportamento cinético das células Sf9 e Sf21 utilizadas na produção de biopesticida para o controle de Spodoptera frugiperda. Variáveis cinéticas, como velocidade específica máxima de crescimento, produtividade em células, velocidade média de infecção e a velocidade média de produção de OB foram determinadas durante a infecção destas linhagens com o vírus extracelular do nucleopoliedrovirus de S. frugiperda. A linhagem Sf9 resultou em melhores resultados de produção do baculovírus (5 x 10(8) OB/mL), quando comparada à linhagem Sf21 (2,5 x 10(8) OB/mL) e outras linhagens da literatura.

Th1-type and Th2-type cytokine profiles and adhesion molecules in the serum of patients suffering from systemic lupus erythematosus and the cytokine production by peripheral blood mononuclear cells (PBMC) were stud- ied. Tumor necrosis factor-alpha (TNF-a), interferon- gamma (IFN-g), interleukin-1b (IL-1b), IL-4, IL-10, IL-13, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were measured using ELISA technique in the sera of 16 systemic lupus erythe- matosus patients without vasculitis (SLE), 30 SLE patients with vasculitis (LV), and in 18 healthy controls. The cytokines were also measured in the culture media of unstimulated, concanavalin-A (Con-A) and phorbol-12-myristate- 13-acetate (PMA) stimulated PBMC. TNF-a serum levels were significantly elevated in both SLE and LV patients and those of IL-1b in SLE patients. TNF-a was also significantly increased in SLE compared to LV patients. Serum levels of all three Th-2 cytokines were significantly elevated in both SLE and LV patients compared to healthy controls. Serum IFN-g and Th2 cytokine levels were significantly increased in patients with more active disease. Both ICAM- 1 and VCAM-1 were significantly increased in SLE patients and only VCAM-1 in LV patients. ICAM-1 showed a significant correlation with IL-1b, IFN-g, IL-4 and IL-10 in both patient groups. In the SLE group VCAM-1 correlated significantly only with ICAM-1, but in the LV group only with IL-1b and IFN-g. Compared to healthy controls, basal TNF- a and IL-4 production by unstimulated PBMC derived from SLE patients were significantly increased. Con-A-stimulated PBMC of both SLE groups produced significantly more IFN-g, IL-4 and IL-13 than Con-A-stimulated control cells. Con-A-stimulated cells derived from LV patients produced much more INF-g than cells from SLE patients. PMA strongly stimulated INFg, TNFa and IL-13 production by cells derived from both SLE groups but had no effect on IL- 4 production. In addition, it had little if any effect on the production of INFg and IL-13 by PBMC derived from healthy donors. These findings suggest that the altered pattern of cytokine production by PBMC may play an important role in the SLE pathophysiology, accounting for differences in the clinical expression of the disease. The differences in adhesion molecules production and their correlation with cytokines suggest ICAM-1 and VCAM-1 as useful markers in SLE patients stratification.

Recent advances in the field of animal assisted reproduction have allowed faster dissemination of animals with superior genetics. In cattle, the development of some biotechnologies such as the in vitro production (IVP) of embryos optimized the use of oocytes, which represents a great improvement in embryo production world-wide. Therefore, females with high net merit can be multiplied more efficiently to maximize the number of their offspring. The use of ultrasound-guided transvaginal follicle aspiration combined with IVP allowed wide commercial transfer of IVP-embryos with satisfactory pregnancy results. Yet, there are a number of unanswered areas including the level of repeatability and predictability in IVP systems. Thus, the aim of this study was to evaluate the repeatability of some variables related to IVP of bovine embryos from Nellore breed. The study was conducted from 2011 to 2012 using 42 Nellore cows housed at the Centro APTA Bovinos de Corte located in Sertãozinho-SP, Brazil. Donors were kept under the same management throughout the entire experimental period. All animals underwent a total of 25 transvaginal ultrasound-guided follicular aspiration sections. Oocytes were retrieved and handled similarly by Sexing Technologies using semen from 11 sires. Repeatability was evaluated in the following variables: total number of oocytes, embryos and viable oocytes, viable oocyte rate (number of viable oocytes divided by the total number of oocytes), and the embryo rate (total number of embryos divided by the number of viable oocytes). Statistical analyzes were performed with the PROC GLIMMIX (SAS Inst., Inc., Cary, NC). Resulting repeatability for the above mentioned variables were: total number of oocytes = 0.72; numbers of viable oocytes = 0.73; number of embryos = 0.65, rate of viable oocytes = 0.14 and embryo rate = 0.18. It was observed that variables having poisson distribution such as total oocytes, number of oocytes and viable embryos showed high repeatability over time. Interestingly, variables reported as percentages as for viable oocyte rate and embryo rate did not show high repeatability. As a result, donors who had high numbers of viable oocytes didn’t necessarily yield a consistent percentage of good quality embryos throughout the IVP sessions. In conclusion, among all studied variables, the total number of oocytes, number of viable oocytes and number of embryos showed high repeatability in Nellore cows.

The objectives of this study were (1) to evaluate in situ ruminal dry matter (DM), crude protein (CP) and starch degradation characteristics and in vitro gas production (GP) kinetics using a set of 20 different maize grain genotypes and (2) to predict the effective degradation (ED) of CP and starch from chemical and physical characteristics alone or in combination with in vitro GP measurements. Maize grains were characterised by different chemical and physical characteristics. Ruminal in situ degradation was measured in three lactating Jersey cows. Ground grains (sieve size: 2mm) were incubated in bags for 1, 2, 4, 8, 16, 24, 48 and 72 h. Bag residues were analysed for CP and starch content. Degradation kinetics was determined and the ED of DM, CP and starch calculated using a ruminal passage rate of 5%/h and 8%/h. The GP of the grains (sieve size: 1 mm) was recorded after 2, 4, 6, 8, 12, 24, 48 and 72 h incubation in buffered rumen fluid and fitted to an exponential equation to determine GP kinetics. Correlations and stepwise multiple linear regressions were evaluated for the prediction of ED calculated for a passage rate of 5%/h (ED5) for CP (EDCP5) and starch (EDST5). The in situ parameters and ED5 varied widely between genotypes with average values (±SD) of 64% ± 4.2, 62% ± 4.1 and 65% ± 5.2 for ED5 of DM, EDCP5 and EDST5 and were on average 10 percentage points lower for a passage rate of 8%/h. Degradation rates varied between 4.8%/h and 7.4%/h, 4.1%/h and 6.5%/h and 5.3%/h and 8.9%/h for DM, CP and starch, respectively. These rates were in the same range as GP rates (6.0-8.3%/h). The EDCP5 and EDST5 were related to CP concentration and could be evaluated in detail using CP fractions and specific amino acids. In vitro GP measurements and GP rates correlated well with EDCP5 and EDST5 and predicted EDCP5 and EDST5 in combination with the chemical characteristics of the samples. Equations can be used to obtain quick and cost effective information on ruminal degradation of CP and starch from maize grains. [ABSTRACT FROM AUTHOR]

The genus Astragalus L., Fabaceae (Leguminosae) is presented in Bulgarian flora by 31 species including local endemics, nine of them listed in the Red Data Book. The main biologically active compounds found in the Bulgarian Astragalus plants are saponins and flavonoids with a variety of effects: immunomodulatory, hepatoprotective, cytotoxic, antioxidant, antihypoxic and others. In the recent years volatiles with cytotoxic activity have also been determined. Saponins and flavonoids are still preferably produced by plants due to their complex structures. Variable quantities and qualities of the wild plant material, plants that need to grow several years before they are ready for harvesting and over-collecting of endangered Astragalus species are just a few of the problems connected with the production of these natural products. Therefore, cultured cells are possible alternative for production of high-value secondary metabolites. This review is focused on phytochemical, pharmacological and biotechnological studies of Bulgarian Astragalus plants and includes also results from our investigations. [ABSTRACT FROM AUTHOR]