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

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

The case for considering the term 'mitochondrial vesicle' as a misnomer in publications about assisted reproductive technologies (ART) for amphibians.

The term 'mitochondrial vesicle' was first used in 2003 in a description of anuran sperm and persists to this day throughout the literature on assisted reproductive technologies (ART) for amphibians. In the present paper, we argue that the term is inappropriate because the widely accepted definition of a 'vesicle' refers to an integral structure with an enclosing lipid bilayer/membrane. Moreover, there are no electron micrographs that show a vesicular structure encapsulating mitochondria on amphibian sperm heads in the literature. In fact, in 1993, the mitochondria in the anuran sperm head had been described as positioned in 'mitochondrial collars' or 'mitochondrial sheaths' surrounded by the plasma membrane of the sperm head. On the other hand, mitochondrial-derived vesicles are defined as vesicles shed from mitochondria surfaces, potentially creating confusion. Therefore, our view is that the term 'mitochondrial vesicle' should be avoided in describing the positioning of mitochondria on sperm.

Could assisted reproductive techniques affect equine fetal membranes and neonatal outcome?

Embryo transfer (ET) and intracytoplasmic sperm injection (ICSI) are widely used in equine species, but their effects on fetal adnexa and neonates have not been investigated yet. The aim of this study was to retrospectively evaluate whether pregnancies obtained by ET or ICSI could be associated with the presence of macroscopic alterations of fetal membranes (FM) and umbilical cord (UC) and if the use of these techniques could influence neonatal outcome. Sixty-six light breed mares hospitalized at the Veterinary Teaching Hospital, University of Bologna, for attending delivery were included in the study. Mares were divided into Artificial Insemination (AI; 32/66 mares, 48 %), Embryo Transfer (ET; 12/66 mares, 18.2 %) and Intracytoplasmic Sperm Injection (ICSI; 22/66 mares, 33 %) groups. All the medical reports of mares and their foals were reviewed and data about mare, pregnancy, foaling, fetal membranes, umbilical cord and foal were recorded. The occurrence of dystocia resulted statistically different between AI group and ICSI group (p = 0.0066), and between AI group and ET group (p = 0.044). Macroscopic examination of FM revealed alterations in 30/66 mares (46 %): 8/32 in AI (25 %), 7/12 in ET (58 %) and 15/22 in ICSI (68 %) with significant lower incidence in AI compared to ET (p = 0.04) and ICSI (p = 0.002) groups. Alterations reported were chorionic villi hypoplasia, chorioallantois edema, allantois cysts, necrotic areas and greenish-grey concretions. Total length of UC resulted significantly shorter in ICSI group (49 ± 9 cm; p < 0.03) compared to AI (60 ± 17 cm) and ET (59 ± 15 cm). However, there were no differences in the incidence of foals' diseases at birth and in foals' survival among groups (p > 0.05). The results demonstrate that transfer of in vivo or in vitro produced embryos may lead to alterations of placental development, as observed in other species, without being associated with a higher incidence of neonatal morbidity and mortality. Further studies about trophoblast development, FM histological evaluation, and placental gene expression should be carried out to clarify the mechanisms underlying the placental alterations.

Reproduction technologies in pig farming: artificial insemination - literature review / Tecnologias de reprodução na suinocultura: inseminação artificial - revisão de literatura / Tecnologías de reproducción en la porcicultura: inseminación artificial - revisión de literatura

This summary addresses the use of reproduction technologies in swine farming, with an emphasis on artificial insemination (AI). Brazilian swine farming has been growing significantly and seeks new technologies to achieve high productive indices sustainably and competitively. Pigs present favorable characteristics such as high prolificacy, fertility, rapid growth, feed efficiency, and carcass yield, which has led to intensive development of the activity with advanced genetic selection. AI is widely employed to disseminate genetic material among different regions and farms. Several AI techniques are used in modern swine farming: intrauterine insemination (IUI) allows semen deposition in the uterine region, reducing costs; fixed-time insemination (FTAI) synchronizes estrus in various females, facilitating management and increasing efficiency; deep intrauterine insemination (DIUI) deposits semen in the uterine horns, obtaining better results; and cervical insemination (CI), a traditional technique widely used, although it may be more time-consuming and present higher reflux rates. The success of AI is related to knowledge of the reproductive cycle of sows, proper nutrition, and genetic and environmental factors. Semen quality is essential, requiring collection by trained professionals and evaluation of sperm motility and morphology. Although it is a consolidated technique, there are issues to be further explored to optimize its application, defining the exact moment for insemination, reducing reflux, and adopting effective protocols. AI is an essential tool for the growth of Brazilian swine farming, but it requires continuous studies to maximize its efficiency and results, considering the farm's production goal and the size of the enterprise to achieve high reproductive and productive indices.

Este resumo aborda o uso de tecnologias de reprodução na suinocultura, com ênfase na inseminação artificial (IA). A suinocultura brasileira vem crescendo significativamente e busca novas tecnologias para alcançar altos índices produtivos de maneira sustentável e competitiva. Os suínos apresentam características favoráveis, como alta prolificidade, fertilidade, rápido crescimento, eficiência alimentar e rendimento de carcaça, o que levou ao desenvolvimento intensivo da atividade com seleção genética avançada. A IA é amplamente empregada para disseminar material genético entre diferentes regiões e granjas. Diversas técnicas de IA são utilizadas na suinocultura moderna: a inseminação intrauterina (IAIU) permite a deposição do sêmen na região uterina, reduzindo custos; a inseminação em tempo fixo (IATF) sincroniza o estro em várias fêmeas, facilitando o manejo e aumentando a eficiência; a inseminação intrauterina profunda (IAUP) deposita o sêmen nos cornos uterinos, obtendo melhores resultados; e a inseminação cervical (IAC), técnica tradicional amplamente utilizada, embora possa ser mais demorada e apresentar maiores taxas de refluxo. O sucesso da IA estar relacionado ao conhecimento do ciclo reprodutivo das matrizes, à nutrição adequada e aos fatores genéticos e ambientais. A qualidade do sêmen é essencial, exigindo coleta por profissionais treinados e avaliação da motilidade e morfologia dos espermatozoides. Apesar de ser uma técnica consolidada, há questões a serem aprofundadas para otimizar sua aplicação, definindo o momento exato para a realização da inseminação, a redução do refluxo e adoção de protocolos eficazes. A IA é uma ferramenta essencial para o crescimento da suinocultura brasileira, mas requer estudos contínuos para maximizar sua eficiência e resultados, considerando o objetivo produtivo da granja e o tamanho do empreendimento para alcançar altos índices reprodutivos e produtivos.

Este resumen aborda el uso de tecnologías de reproducción en la producción porcina, con énfasis en la inseminación artificial (IA). La producción porcina brasileña ha crecido significativamente y busca nuevas tecnologías para alcanzar altos índices de productividad de manera sostenible y competitiva. Los cerdos presentan características favorables, como alta prolificidad, fertilidad, rápido crecimiento, eficiencia alimentaria y rendimiento de la canal, lo que ha llevado al desarrollo intensivo de la actividad con selección genética avanzada. La IA se utiliza ampliamente para difundir material genético entre diferentes regiones y granjas. Diversas técnicas de IA son utilizadas en la producción porcina moderna: la inseminación intrauterina (IAIU) permite la deposición del semen en la región uterina, reduciendo costos; la inseminación a tiempo fijo (IATF) sincroniza el estro en varias hembras, facilitando el manejo y aumentando la eficiencia; la inseminación intrauterina profunda (IAUP) deposita el semen en los cuernos uterinos, obteniendo mejores resultados; y la inseminación cervical (IAC), técnica tradicional ampliamente utilizada, aunque puede ser más demorada y presentar mayores tasas de reflujo. El éxito de la IA está relacionado con el conocimiento del ciclo reproductivo de las hembras, la nutrición adecuada y los factores genéticos y ambientales. La calidad del semen es esencial, requiriendo la recolección por profesionales capacitados y la evaluación de la motilidad y morfología de los espermatozoides. A pesar de ser una técnica consolidada, hay aspectos que deben ser profundizados para optimizar su aplicación, como la definición precisa del momento de la inseminación, la reducción del reflujo y la adopción de protocolos eficaces. La IA es una herramienta esencial para el crecimiento de la producción porcina brasileña, pero requiere estudios continuos para maximizar su eficiencia y resultados, considerando el objetivo productivo de la granja y el tamaño del emprendimiento para alcanzar altos índices reproductivos y productivos.

The oocyte: the key player in the success of assisted reproduction technologies.

The ovulation of a mature oocyte at metaphase II of meiosis, with optimal potential to undergo fertilisation by a sperm cell, complete meiosis and sustain the switch to mitotic division, and support early embryo development, involves a protracted and disrupted/delayed series of processes. Many of these are targeted for exploitation in vivo , or recapitulation in vitro , by the livestock industry. Reproductive technologies, including AI, multiple ovulation embryo transfer, ovum pick-up, in vitro embryo production, and oestrus and ovulation synchronisation, offer practitioners and producers the opportunity to produce offspring from genetically valuable dams in much greater numbers than they would normally have in their lifetime, while in vitro oocyte and follicle culture are important platforms for researchers to interrogate the physiological mechanisms driving fertility. The majority of these technologies target the ovarian follicle and the oocyte within; thus, the quality and capability of the recovered oocyte determine the success of the reproductive intervention. Molecular and microscopical technologies have grown exponentially, providing powerful platforms to interrogate the molecular mechanisms which are integral to or affected by ART. The development of the bovine oocyte from its differentiation in the ovary to ovulation is described in the light of its relevance to key aspects of individual interventions, while highlighting the historical timeline.

Assisted reproductive technologies (ARTs) in Mithun (Bos frontalis): What progress has been made so far? An overview.

Mithun, a unique bovine species, endemic to parts of North East India and plays an important role in the socioeconomic, cultural and religious fabrics of the local tribal population. To date, Mithuns are reared in a traditional free-range system by communities and increased deforestation, agricultural commercialization, disease outbreaks and indiscriminate slaughtering of elite Mithun for table purposes have significantly decreased its habitat and the elite Mithun population. Greater genetic gain is achieved with the implementation and effective use of assisted reproductive technologies (ARTs); however, presently it is limited to organized Mithun farms. At a slow pace, Mithun farmers are adopting semi-intensive rearing systems and interest in the use of ARTs is gradually escalating in Mithun husbandry. This article reviews the current status of ARTs such as semen collection and cryopreservation, estrus synchronization and timed artificial insemination (TAI), multiple ovulation and embryo transfer and in vitro embryo production and future perspectives in Mithun. Mithun semen collection and cryopreservation have been standardized, and estrus synchronization and TAI are suitable technologies that can be easily implemented under field conditions in near future. The establishment of an open nucleus-breeding system under community participatory mode along with the introduction of the ARTs is an alternative to the traditional breeding system for rapid genetic improvement of Mithun. Finally, the review considers the potential benefits of ARTs in Mithun and future research should include the use of these ARTs which will provide additional opportunities for improved breeding regimens in Mithun.

Gestational and health outcomes of dairy cows conceived by assisted reproductive technologies compared to artificial insemination.

Herd gestation and health management are key aspects of effective dairy farm operations and animal welfare improvement. Unfortunately, very little is known about the developmental divergences induced by assisted reproduction technologies (ART) and their consequences once the animal is mature. Indeed, the gestational and health outcomes of this subset of the Holstein population is yet to be characterized. In this study, the intergenerational impacts of ART conception were assessed by looking at the gestation and health outcomes of a large cohort of cows (n = 284,813) for which the conception methods were known. Our results showed that cows conceived by multiple ovulation embryo transfer (MOET) and in vitro fertilisation (IVF) displayed longer gestations: +0.37 ± 0.079 and +0.65 ± 0.21 day compared to cows conceived by artificial insemination (AI). Surprisingly, animals conceived by all methods experienced a similar 1-day decline in average gestation length from 2012 to 2019. Cows conceived by IVF were not more likely to experience stillbirths but were affected by common diseases such as ovarian cysts, mastitis, and uterine diseases in different proportions compared to cows conceived by other methods. This study provides new and unique information on ART animals regarding perinatal mortality and general health outcomes.

Emerging arguments for reproductive technologies in wildlife and their implications for assisted reproduction and conservation of threatened marsupials.

Assisted reproductive technologies (ARTs) have significant potential to make a meaningful contribution to the conservation of threatened wildlife. This is true of Australia's iconic, and endangered koala (Phascolarctos cinereus). If developed, ARTs could offer a solution to manage genetic diversity and costs in breeding programs and may provide frozen repositories for either insurance or the practical production of genetically resilient koalas for release and on-ground recovery. Holding back the wider use of ARTs for koalas and other wildlife is a lack of funding to close the remaining knowledge gaps in the marsupial reproductive sciences and develop the reproductive tools needed. This lack of funding is arguably driven by a poor understanding of the potential contribution ARTs could make to threatened species management. We present a review of our cross-disciplinary and accessible strategy to draw much needed public attention and funding for the development of ARTs in wildlife, using emerging cost and genetic modelling arguments and the koala as a case study.

Effects of activation and assisted reproduction techniques on the composition, structure, and properties of the sauger (Sander Canadensis) spermatozoa plasma membrane.

The sperm plasma membrane is a multifunctional organelle essential to fertilization. However, assisted reproduction techniques often negatively affect this structure, resulting in reduced fertility. These reductions have been attributed to plasma membrane damage in a wide array of species, including fish. Considerable research has been conducted on the fish sperm membrane, but few have examined the effect of cryopreservation and other assisted reproduction techniques (ARTs) on not only membrane composition, but also specific characteristics (e.g., fluidity) and organization (e.g., lipid rafts). Herein, we determined the effects of three ARTs (testicular harvest, strip spawning, and cryopreservation) on the sperm plasma membrane, using Sauger (Sander canadensis) sperm as a model. To this end, a combination of fluorescent dyes (e.g., merocyanine 540, filipin III, cholera toxin subunit ß), liquid chromatography - mass spectroscopy (LC-MS) analysis of membrane lipids, and membrane ultracentrifugation coupled with plate assays and immunofluorescence were used to describe and compare sperm fluidity, membrane composition, as well as lipid raft composition and distribution among sperm types. Stripped sperm became more fluid following motility activation (40% increase in highly fluid cells characterized by a 2 × increase in fluorescence) and contained lipid rafts restricted to the midpiece. Testicular harvest yielded sperm with characteristics similar to stripped sperm. By contrast, cryopreservation impacted every aspect of membrane physiology. Two cell populations, one highly fluid and the other rigid, resulted from the freeze-thaw process. Cryopreservation reduced lipid raft cholesterol content by 44% and flotilin-2 (a lipid raft marker) was partially displaced owing to a decrease in buoyancy. Unlike stripped and testicular sperm, LC-MS analysis revealed increases in oxidative damage markers, membrane destabilization, and apoptotic signaling in cryopreserved sperm. Ultrastructural analysis also revealed widespread physical damage to the membrane following freeze-thaw. Sperm motility, however, was unrelated to any measure of membrane physiology used in this study. Our results demonstrate that ARTs have the potential to substantially affect the sperm plasma membrane, but not always detrimentally. These results provide multiple potential biomarkers of sperm quality as well as insight into sources of sub-fertility resulting from use of ARTs.

Current status of assisted reproductive technologies in buffaloes.

Buffaloes are raised by small farm holders primarily as source of draft power owing to its resistance to hot climate, disease, and stress conditions. Over the years, transformation of these animals from draft to dairy was deliberately carried out through genetic improvement program leading to the development of buffalo-based enterprises. Buffalo production is now getting more attention and interest from buffalo raisers due to its socioeconomic impact as well as its contribution to propelling the livestock industry in many developing countries. Reproduction of buffaloes, however, is confronted with huge challenge and concern as being generally less efficient to reproduce compared with cattle due to both intrinsic and extrinsic factors such as poor estrus manifestation, silent heat, marked seasonal infertility, postpartum anestrus, long calving interval, delayed puberty, inherently low number of primordial follicles in their ovaries, high incidence of atresia, and apoptosis. Assisted reproductive technologies (ARTs) are major interventions for the efficient utilization of follicle reserve in buffaloes. The present review focuses on estrus and ovulation synchronization for fixed time artificial insemination, in vitro embryo production, intracytoplasmic sperm injection, cryopreservation of oocytes and embryos, somatic cell nuclear transfer, the factors affecting utilization in various ARTs, and future perspectives in buffaloes.

Common goals, different stages: the state of the ARTs for reptile and amphibian conservation.

Amphibians and reptiles are highly threatened vertebrate taxa with large numbers of species threatened with extinction. With so many species at risk, conservation requires the efficient and cost-effective application of all the tools available so that as many species as possible are assisted. Biobanking of genetic material in genetic resource banks (GRBs) in combination with assisted reproductive technologies (ARTs) to retrieve live animals from stored materials are two powerful, complementary tools in the conservation toolbox for arresting and reversing biodiversity decline for both amphibians and reptiles. However, the degree of development of the ARTs and cryopreservation technologies differ markedly between these two groups. These differences are explained in part by different perceptions of the taxa, but also to differing reproductive anatomy and biology between the amphibians and reptiles. Artificial fertilisation with cryopreserved sperm is becoming a more widely developed and utilised technology for amphibians. However, in contrast, artificial insemination with production of live progeny has been reported in few reptiles, and while sperm have been successfully cryopreserved, there are still no reports of the production of live offspring generated from cryopreserved sperm. In both amphibians and reptiles, a focus on sperm cryopreservation and artificial fertilisation or artificial insemination has been at the expense of the development and application of more advanced technologies such as cryopreservation of the female germline and embryonic genome, or the use of sophisticated stem cell/primordial germ cell cryopreservation and transplantation approaches. This review accompanies the publication of ten papers on amphibians and twelve papers on reptiles reporting advances in ARTs and biobanking for the herpetological taxa.

Reptile assisted reproductive technologies: can ART help conserve 300 million years of evolution by preserving extant reptile biodiversity?

Biodiversity loss is the greatest environmental problem threatening ecosystem, animal, and human health. Anthropogenic induced changes to climate, habitat, disease, species distributions, poaching, and unsustainable trade have accelerated extinction rates in all vertebrates, including reptiles. Preventing reptile extinctions will require humans to acknowledge these losses and develop ex situ and in situ plans to preserve them. Assisted reproductive technologies (ART) are management tools used to protect numerous vertebrate taxa; however, progress in developing ART for reptiles has lagged. Creating functional and sustainable reptile ART will strengthen our conservation capacity by capturing genetic material from select individuals to overcome natural or manmade boundaries. Utilising short-term gamete storage and genome resource banking, in conjunction with timed artificial insemination (AI) or ex ovo incubation, could lead to profound advances in reptile conservation, mitigating the loss of reptile biodiversity. In this article, we review ART reptile research completed since the 1970s. Topics include AI, hormonal control of reproduction, gamete collection, gamete storage, and genome resource banking. Additionally, we review the potential application of advanced reproductive methodologies, including in vitro /ex ovo fertilisation, intracytoplasmic sperm injection, cloning (somatic cell nuclear transfer), and genetic editing.

Potential applications of assisted reproductive technologies (ART) in reindeer (Rangifer tarandus).

Interest in the use of assisted reproductive technology in reindeer husbandry has gradually increased during the last decades. This article reviews Western and Russian literature on reindeer semen collection, semen cryopreservation and artificial insemination. In addition, literature on the synchronisation of the stage of reindeer oestrous cycle among females, recovery of in vivo embryos, embryo transfer, the production of in vitro-produced embryos and pregnancy diagnosis is reviewed.

Controlled spermatozoa-oocyte interaction improves embryo quality in sheep.

The current protocols of in vitro fertilization and culture in sheep rely on paradigms established more than 25 years ago, where Metaphase II oocytes are co-incubated with capacitated spermatozoa overnight. While this approach maximizes the number of fertilized oocytes, on the other side it exposes them to high concentration of reactive oxygen species (ROS) generated by active and degenerating spermatozoa, and positively correlates with polyspermy. Here we set up to precisely define the time frame during which spermatozoa effectively penetrates and fertilizes the oocyte, in order to drastically reduce spermatozoa-oocyte interaction. To do that, in vitro matured sheep oocytes co-incubated with spermatozoa in IVF medium were sampled every 30 min (start of incubation time 0) to verify the presence of a fertilizing spermatozoon. Having defined the fertilization time frame (4 h, data from 105 oocytes), we next compared the standard IVF procedures overnight (about 16 h spermatozoa/oocyte exposure, group o/nIVF) with a short one (4 h, group shIVF). A lower polyspermic fertilization (> 2PN) was detected in shIVF (6.5%) compared to o/nIVF (17.8%), P < 0.05. The o/nIVF group resulted in a significantly lower 2-cell stage embryos, than shIVF [34.6% (81/234) vs 50.6% (122/241) respectively, P < 0.001]. Likewise, the development to blastocyst stage confirmed a better quality [29% (70/241) vs 23.5% (55/234), shIVF vs o/nIVF respectively] and an increased Total Cell Number (TCN) in shIVF embryos, compared with o/n ones. The data on ROS have confirmed that its generation is IVF time-dependent, with high levels in the o/nIVF group. Overall, the data suggest that a shorter oocyte-spermatozoa incubation results in an improved embryo production and a better embryo quality, very likely as a consequence of a shorter exposure to the free oxygen radicals and the ensuing oxidative stress imposed by overnight culture.

Invited review: Use of assisted reproduction techniques to accelerate genetic gain and increase value of beef production in dairy herds.

The contribution of the calf enterprise to the profit of the dairy farm is generally considered small, with beef bull selection on dairy farms often not considered a high priority. However, this is likely to change in the future as the rapid rate of expansion of the dairy herd in some countries is set to plateau and improvements in dairy herd fertility combine to reduce the proportion of dairy breed calves required on dairy farms. This presents the opportunity to increase the proportion of beef breed calves born, increasing both the value of calf sales and the marketability of the calves. Beef embryos could become a new breeding tool for dairies as producers need to reassess their breeding policy as a consequence of welfare concerns and poor calf prices. Assisted reproductive technologies can contribute to accelerated genetic gain by allowing an increased number of offspring to be produced from genetically elite dams. There are the following 3 general classes of donor females of interest to an integrated dairy-beef system: (1) elite dairy dams, from which oocytes are recovered from live females using ovum pick-up and fertilized in vitro with semen from elite dairy bulls; (2) elite beef dams, where the oocytes are recovered from live females using ovum pick-up and fertilized with semen from elite beef bulls; and (3) commercial beef dams (≥50% beef genetics), where ovaries are collected from the abattoir postslaughter, and oocytes are fertilized with semen from elite beef bulls that are suitable for use on dairy cows (resulting embryo with ≥75% beef genetics). The expected benefits of these collective developments include accelerated genetic gain for milk and beef production in addition to transformation of the dairy herd calf crop to a combination of good genetic merit dairy female calves and premium-quality beef calves. The aim of this review is to describe how these technologies can be harnessed to intensively select for genetic improvement in both dairy breed and beef breed bulls suitable for use in the dairy herd.

Multi-locus imprinting disturbances of Beckwith-Wiedemann and Large offspring syndrome/Abnormal offspring syndrome: A brief review.

In vitro fertilization and somatic cell nuclear transfer are assisted reproduction technologies commonly used in humans and cattle, respectively. Despite advances in these technologies, molecular failures can occur, increasing the chance of the onset of imprinting disorders in the offspring. Large offspring syndrome/abnormal offspring syndrome (LOS/AOS) has been described in cattle and has features such as hypergrowth, malformation of organs, and skeletal and placental defects. In humans, Beckwith-Wiedemann syndrome (BWS) has phenotypic characteristics similar to those found in LOS/AOS. In both syndromes, disruption of genomic imprinting associated with loss of parental-specific expression and parental-specific epigenetic marks is involved in the molecular etiology. Changes in the imprinting pattern of these genes lead to loss of imprinting (LOI) due to gain or loss of methylation, inducing the emergence of these syndromes. Several studies have reported locus-specific alterations in these syndromes, such as hypomethylation in imprinting control region 2 (KvDMR1) in BWS and LOS/AOS. These LOI events can occur at multiple imprinted loci in the same affected individual, which are called multi-locus methylation defect (MLMD) events. Although the bovine species has been proposed as a developmental model for human imprinting disorders, there is little information on bovine imprinted genes in the literature, even the correlation of epimutation data with clinical characteristics. In this study, we performed a systematic review of all the multi-locus LOI events described in human BWS and LOS/AOS, in order to determine in which imprinted genes the largest changes in the pattern of DNA methylation and expression occur, helping to fill gaps for a better understanding of the etiology of both syndromes.

Perturbations of the hepatic proteome behind the onset of metabolic disorders in mouse offspring developed following embryo manipulation.

Assisted Reproductive Technologies (ART) allowed the births of >8 million babies worldwide. Even if ART children are healthy at birth, several studies reported that ART may cause changes in foetal programming, leading to an increased predisposition to metabolic disorders in adulthood. Previous studies on mouse model showed obesity, glucose intolerance, and hepatic lipid accumulation in ART offspring. A cumulative effect of the different components of ART protocol has been previously described, for example, in the occurrence of epigenetic defects. Here, we investigated whether there is a cumulative effect of embryo transfer (ET), in vitro culture (IVC) and blastomere biopsy (BB) in the onset of metabolic disorders in mouse offspring vs those naturally conceived (Control - CTR). To this aim, proteomic analysis was performed on the livers from adult mouse offspring developed following ET, IVC and BB vs CTR. We observed deregulated expression of proteins involved in lipid, carbohydrate, energy metabolisms and cellular processes in ART offspring. Moreover, we found increased body weight in all ART offspring while i) insulin resistance in BB male, ii) females glucose intolerance and high level of triglycerides and cholesterol in BB females and iii) low levels of interleukin-6 in BB, IVC and ET males. In conclusion, our study suggests that the use of various embryo manipulations influences the metabolic health of adult offspring, resulting in an increased predisposition to hepatic diseases and metabolic syndrome in a sex-specific manner.

The ART of bringing extinction to a freeze - History and future of species conservation, exemplified by rhinos.

The ongoing mass extinction of animal species at an unprecedented rate is largely caused by human activities. Progressive habitat destruction and fragmentation is resulting in accelerated loss of biodiversity on a global scale. Over decades, captive breeding programs of non-domestic species were characterized by efforts to optimize species-specific husbandry, to increase studbook-based animal exchange, and to improve enclosure designs. To counter the ongoing dramatic loss of biodiversity, new approaches are warranted. Recently, new ideas, particularly the application of assisted reproduction technologies (ART), have been incorporated into classical zoo breeding programs. These technologies include semen and oocyte collection, artificial insemination, and in-vitro embryo generation. More futuristic ideas of advanced ART (aART) implement recent advances in biotechnology and stem-cell related approaches such as cloning, inner cell mass transfer (ICM), and the stem-cell-associated techniques (SCAT) for the generation of gametes and ultimately embryos of highly endangered species, such as the northern white rhinoceros (Ceratotherium simum cottoni) of which only two female individuals are left. Both, ART and aART greatly depend on and benefit from the rapidly evolving cryopreservation techniques and biobanking not only of genetic, but also of viable cellular materials suitable for the generation of induced pluripotent stem cells (iPSC). The availability of cryopreserved materials bridges gaps in time and space, thereby optimizing the available genetic variability and enhancing the chance to restore viable populations.

Development of assisted reproductive technologies for Mus spretus†.

The genus Mus consists of many species with high genetic diversity. However, only one species, Mus musculus (the laboratory mouse), is common in biomedical research. The unavailability of assisted reproductive technologies (ARTs) for other Mus species might be a major reason for their limited use in laboratories. Here, we devised ARTs for Mus spretus (the Algerian mouse), a commonly used wild-derived Mus species. We found that in vitro production of M. spretus embryos was difficult because of low efficacies of superovulation with equine chorionic gonadotropin or anti-inhibin serum (AIS) (5-8 oocytes per female) and a low fertilization rate following in vitro fertilization (IVF; 15.2%). The primary cause of this was the hardening of the zona pellucida but not the sperm's fertilizing ability, as revealed by reciprocal IVF with laboratory mice. The largest number of embryos (16 per female) were obtained when females were injected with AIS followed by human chorionic gonadotropin and estradiol injections 24 h later, and then by natural mating. These in vivo-derived 2-cell embryos could be vitrified/warmed with a high survival rate (94%) using an ethylene glycol-based solution. Importantly, more than 60% of such embryos developed into healthy offspring following interspecific embryo transfer into (C57BL/6 × C3H) F1 female mice. Thus, we have devised practical ARTs for Mus spretus mice, enabling efficient production of embryos and animals, with safe laboratory preservation of their strains. In addition, we have demonstrated that interspecific embryo transfer is possible in murine rodents.