New Biotechniques and their Consequences for Farm Animal Welfare

Contents This paper considers (potentially) harmful consequences of new biotechnologies for farm animal welfare. The most important new biotechnologies that are currently used in farm animals breeding and husbandry include: multiple ovulation and embryo transfer (MOET) and in vitro embryo production (IVP). Cloning by nuclear transfer (NT) and transgenesis are still in development and mainly applied for experimental purposes with the prospect of a more widespread practical implemention in the future. Evidence is presented showing that generally accepted technologies such as MOET and IVP, relative to in vivo procedures, can result in a host of deleterious side-effects commonly known as the large offspring syndrome (LOS). Likewise, NT and transgenesis, which also typically include several in vitro reproductive manipulations, have clearly been associated with the occurrence of LOS symptoms. It is argued that transgenesis may constitute one additional set of factors that may negatively affect farm animal welfare: the expression of the transgene and the concomitant synthesis and release of a protein. NT might lead to incompletely reprogramming of the transferred genome. It is suggested that the introduction of new biotechnologies into farm animal husbandry should be accompanied by scientifically valid and systematic studies into the effects on animal welfare, with the help of a comprehensive welfare protocol.     

A brief overview of transgenic farm animals

Transgenesis offers new possibilities to rapidly modify the genome of living organisms. The application of transgenesis to farm animals faces many problems, more than those observed in the transgenesis of laboratory animals, as there are currently many different techniques available to obtain transgenic animals, which all have problems regarding low efficiency and high costs. When these techniques are applied to farm animals the problems concerning transgenesis are multiplied. Two main techniques, male pronuclear microinjection and sperm mediated gene transfer, utilised in farm animal transgenesis, are briefly presented. The improvement of these techniques and the employment of other biotechnologies such as cloning, could expand the uses of transgenic farm animals for human health.     

An efficient strategy for generation of transgenic mice by lentiviral transduction of male germline stem cells in vivo

Background: Male germline stem cells(MGSCs) are a subpopulation of germ cells in the testis tissue. MGSCs are capable of differentiation into spermatozoa and thus are perfect targets for genomic manipulation to generate transgenic animals.Method: The present study was to optimize a protocol of production of transgenic mice through transduction of MGSCs in vivo using lentiviral-based vectors. The recombinant lentiviral vectors with either EF-1 or CMV promoter to drive the expression of enhanced green fluorescent protein(e GFP) transgene were injected into seminiferous tubules or inter-tubular space of 7-day-old and 28-day-old mouse testes. At 5 or 6 wk post-surgery, these pre-founders were mated with wild-type C57BL/6J female mice(1.5 to 2.0-month-old).Results: Sixty-seven percent of F1 generation and 55.56 % of F2 offspring were positive for eG FP transgene under the control of EF-1 promoter via PCR analysis. The transgenic pups were generated in an injection site-and age-independent manner. The expression of transgene was displayed in the progeny derived from lentiviral vector containing CMV promoter to drive transgene, but it was silenced or undetectable in the offspring derived from lentiviral vector with transgene under EF-1 promoter. The methylation level of g DNA in the promoter region of transgene was much higher in the samples derived lentiviral vectors with EF-1 promoter than that with CMV promoter,suggesting e GFP transgene was suppressed by DNA methylation in vivo.Conclusion: This research reported here an effective strategy for generation of transgenic mice through transduction of MGSCs in vivo using lentivirus vectors with specific promoters, and the transgenic offspring were obtained in an injection site-and age-independent manner. This protocol could be applied to other animal species, leading to advancement of animal transgenesis in agricultural and biomedical fields.     

Perspectives for feed-efficient animal production

Modern animal breeding programs are largely based on biotechnological procedures, including AI and embryo transfer technology. Recent breakthroughs in reproductive technologies, such as somatic cell nuclear transfer and in vitro embryo production, and their combination with the emerging molecular genetic tools, will further advance progress and provide new opportunities for livestock breeding. This is urgently needed in light of the global challenges such as the ever-increasing human population, the limited resources of arable land, and the urgent environmental problems associated with farm animal production. Here, we focus on genomic breeding strategies and transgenic approaches for making farm animals more feed efficient. Based on studies in the mouse and rat model, we have identified a panel of genes that are critically involved in the regulation of feed uptake and that could contribute toward future breeding of farm animals with reduced environmental impact. We anticipate that genetically modified animals will play a significant role in shaping the future of feed-efficient and thus sustainable animal production, but will develop more slowly than the biomedical applications because of the complexity of the regulation of feed intake and metabolism.     

Use of Transgenic Animals to Improve Human Health and Animal Production

Contents Transgenic animals are more widely used for various purposes. Applications of animal transgenesis may be divided into three major categories: (i) to obtain information on gene function and regulation as well as on human diseases, (ii) to obtain high value products (recombinant pharmaceutical proteins and xeno-organs for humans) to be used for human therapy, and (iii) to improve animal products for human consumption. All these applications are directly or not related to human health. Animal transgenesis started in 1980. Important improvement of the methods has been made and are still being achieved to reduce cost as well as killing of animals and to improve the relevance of the models. This includes gene transfer and design of reliable vectors for transgene expression. This review describes the state of the art of animal transgenesis from a technical point of view. It also reports some of the applications in the medical field based on the use of transgenic animal models. The advance in the generation of pigs to be used as the source of organs for patients and in the preparation of pharmaceutical proteins from milk and other possible biological fluids from transgenic animals is described. The projects in course aiming at improving animal production by transgenesis are also depicted. Some the specific biosafety and bioethical problems raised by the different applications of transgenesis, including consumption of transgenic animal products are discussed.     

Characterization of transgenic livestock production

The objective of transgenic livestock improvement projects is to develop and bring to market superior breeding stock, as well as germplasm for the artificial insemination and embryo transfer industries. Livestock animal biotechnology programs hold the promise of achieving, in a single generation, improvements in commercially important livestock species previously possible only through long-term traditional selective breeding practices or by chance mutation. Transgenic farm animals harboring growth hormone or metabolically related structural genes have been created. Studies of these animals demonstrate the effects of inadequate regulation of transgene expression. Research continues to explore the intricacies of developmental regulation of such genes and phenotypic consequences of mammalian gene transfer. Ultimately, genetically engineered livestock will provide producers with the benefit of increased production efficiencies while the consumer will have healthier animal food products. Conceivably, products will be produced with lower levels of fat, cholesterol, feed additives and pharmaceutical residues from animals with altered carcass composition that will result in greater nutritional benefit for the consumer.     

Use of intracytoplasmic sperm injection (ICSI) to generate transgenic animals

Even though intracytoplasmic sperm injection (ICSI) has been widely used for the production of offspring in human infertility clinics and in reproductive research laboratories using mice, many researchers engaged in animal transgenesis still consider it somewhat cumbersome. The greatest advantage of ICSI-mediated transgenesis is that it allows introduction of very large DNA transgenes (e.g., yeast artificial chromosomes), with relatively high efficiency into the genomes of hosts, as compared to pronuclear injection. Recently, we have developed an active form of intracytoplasmic sperm injection-mediated transgenesis (ICSI-Tr) with fresh sperm utilizing transposons. The transgenic efficiencies rival all transgenic techniques except that of lentiviral methods.     

The complex relationship between welfare and reproduction in cattle

Using reproduction parameters as indicators for cattle welfare has limitations and, at best, these parameters should only be viewed as indirect indicators of welfare. On a farm level, measures such as fertility rates emphasize biological performance of the herd but fail to consider the welfare of individual animals. Even on an individual level, the relationship between reproductive effectiveness and animal welfare is complex. Good reproductive performance does not automatically signify good welfare, as domestication and targeted breeding programmes have led to prioritization of high productive and reproductive performance in most modern farm animal species. In this review, we synthesize literature regarding cattle husbandry, reproduction, welfare and their multidimensional relationships. We argue that practices such as artificial insemination or the use of sexed semen may provide potential welfare advantages as these practices reduce the risk of disease transmission and injury or enable selection of specific beneficial traits. Furthermore, they may offer a solution to current practices jeopardizing welfare, such as the management of surplus bull calves in the dairy industry. Conversely, the animals’ ability to perform natural behaviours such as oestrous expression, an aspect arguably contributing to welfare, is often limited on commercial farms; this limitation is particularly evident in housing systems such as tie stalls where movement is restricted. Moreover, common management practices such as oestrus manipulation may lead to negative attitudes in citizens who often regard naturalness as important element of animal welfare.     

The welfare implications of animal breeding and breeding technologies in commercial agriculture

The commercial applications in agriculture of new breeding technologies, as well as conventional breeding strategies, have the potential to influence animal welfare in both positive and negative ways. For example, the sexing of cattle semen might be used to reduce the number of unwanted male dairy calves provided that the technique had not been shown to produce adverse effects. On the other hand, inappropriate use of some breeding technologies may create new problems, or exacerbate welfare problems that may already have arisen within conventional livestock breeding.

It is the impact of any breeding technology or strategy that is important to welfare, whether it is the quality of life of the offspring that is compromised, or whether it is the application of the technology itself that causes pain, distress or lasting harm to the subject animal.

The aim of this paper is to provide clear and practical advice on the establishment of an appropriate framework within which developments in animal breeding and breeding technologies, and the outcome of such processes, may be considered, monitored and, where necessary, regulated. It builds on recommendations recently made by the FAWC to UK Government, and considers the wider public interest in these technologies.

Finally, the response from stakeholders is considered in attempting to ensure that such controls do not render production systems non-competitive in a global livestock industry.     

哺乳动物胚胎玻璃化冷冻研究进展

近40年来,利用冷冻保存技术将哺乳动物胚胎长期保存起来,建立"胚胎库"是保护物种资源和拯救濒危动物的有效手段,同时也是加快家畜品种改良、建立动物基因库和实施胚胎移植产业化的重要组成部分,也可以为克隆、转基因等现代生物技术提供丰富的试验材料,使胚胎的供给不受时间和空间的限制。胚胎冷冻保存技术在人类辅助生殖方面也具有广阔的应用前景。本文主要介绍了玻璃化冷冻过程中胚胎冷冻主体承载工具特点和应用,并展望未来哺乳动物胚胎冷冻发展的方向。     

Advances in farm animal transgenesis

The first transgenic livestock were produced in 1985 by microinjection of foreign DNA into zygotic pronuclei. This was the method of choice for more than 20 years, but more efficient protocols are now available, including somatic cell nuclear transfer and lentiviral transgenesis. Typical applications include carcass composition, lactational performance and wool production, as well as enhanced disease resistance and reduced environmental impact. Transgenic farm animal production for biomedical applications has found broad acceptance. In 2006 the European Medicines Agency (EMA) approved commercialization of the first recombinant pharmaceutical protein, antithrombin, produced in the mammary gland of transgenic goats. As the genome sequencing projects for various farm animal species are completed, it has become feasible to perform precise genetic modifications employing the emerging tools of lentiviral vectors, small interfering ribonucleic acids, meganucleases, zinc finger nucleases and transposons. We anticipate that genetic modification of farm animals will be instrumental in meeting global challenges in agricultural production and will open new horizons in biomedicine.     

Aspects of animal welfare in livestock production

The modern consumer is increasingly concerned about the welfare of farm animals which are kept in intensive systems on specialised farms where the health and well-being is almost completely dependent on the will, ability and care of the farmer. Further demands related to animal production are consumer health (quality and safety of food products), the protection of the environment and cheap food. The currently used husbandry systems are man made and emphasise automation which requires permanent critical observation of the welfare of the animals. Ethological indicators are equally important as health and performance to evaluate keeping systems. Future animal farming will be influenced by new technologies such as electronic animal identification and milking robots, and more important by biotechnology and genome analysis. Veterinary surgeons and farmers have to co-operate on the basis of scientifically sound animal welfare schemes which help to protect our farm animals in modern and intensive livestock production systems.     

Transgenic mouse offspring generated by ROSI

The production of transgenic animals is an important tool for experimental and applied biology. Over the years, many approaches for the production of transgenic animals have been tried, including pronuclear microinjection, sperm-mediated gene transfer, transfection of male germ cells, somatic cell nuclear transfer and the use of lentiviral vectors. In the present study, we developed a new transgene delivery approach, and we report for the first time the production of transgenic animals by co-injection of DNA and round spermatid nuclei into non-fertilized mouse oocytes (ROSI). The transgene used was a construct containing the human CMV immediate early promoter and the enhanced GFP gene. With this procedure, 12% of the live offspring we obtained carried the transgene. This efficiency of transgenic production by ROSI was similar to the efficiency by pronuclear injection or intracytoplasmic injection of male gamete nuclei (ICSI). However, ICSI required fewer embryos to produce the same number of transgenic animals. The expression of Egfp mRNA and fluorescence of EGFP were found in the majority of the organs examined in 4 transgenic lines generated by ROSI. Tissue morphology and transgene expression were not distinguishable between transgenic animals produced by ROSI or pronuclear injection. Furthermore, our results are of particular interest because they indicate that the transgene incorporation mediated by intracytoplasmic injection of male gamete nuclei is not an exclusive property of mature sperm cell nuclei with compact chromatin but it can be accomplished with immature sperm cell nuclei with decondensed chromatin as well. The present study also provides alternative procedures for transgene delivery into embryos or reconstituted oocytes.     

Establishing a laboratory animal model from a transgenic animal: RasH2 mice as a model for carcinogenicity studies in regulatory science

Transgenic animal models have been used in small numbers in gene function studies in vivo for a period of time, but more recently, the use of a single transgenic animal model has been approved as a second species, 6-month alternative (to the routine 2-year, 2-animal model) used in short-term carcinogenicity studies for generating regulatory application data of new drugs. This article addresses many of the issues associated with the creation and use of one of these transgenic models, the rasH2 mouse, for regulatory science. The discussion includes strategies for mass producing mice with the same stable phenotype, including constructing the transgene, choosing a founder mouse, and controlling both the transgene and background genes; strategies for developing the model for regulatory science, including measurements of carcinogen susceptibility, stability of a large-scale production system, and monitoring for uniform carcinogenicity responses; and finally, efficient use of the transgenic animal model on study. Approximately 20% of mouse carcinogenicity studies for new drug applications in the United States currently use transgenic models, typically the rasH2 mouse. The rasH2 mouse could contribute to animal welfare by reducing the numbers of animals used as well as reducing the cost of carcinogenicity studies. A better understanding of the advantages and disadvantages of the transgenic rasH2 mouse will result in greater and more efficient use of this animal model in the future.     

转基因山羊研究进展

近年来,动物转基因技术发展日淅完善,在牛、羊、猪等家畜上的应用日益增多.为全面了解山羊转基因技术的研究进展,论文简要回顾了国内外转基因山羊生产的里程碑事件,总结了生产转基因山羊的方法和原理,并对转基因山羊研究状况、应用前景及存在问题进行了较为系统的概括和分析,从而为借助现代生物技术丰富我国山羊育种手段,加快山羊新良种培...     

Assisted reproductive techniques for cattle breeding in developing countries: a critical appraisal of their value and limitations

Commercialization of animal biotechnologies, including those related to reproduction [also known as assisted reproductive techniques (ARTS)], is an increasing reality in developing countries, following the enormous flow of information around us and the increasing global commercial interests in areas where cattle production has its major assets. The present review discusses the achievements of various biotechnological tools for reproduction in cattle including semen handling for artificial insemination (AI), superovulation and embryo transfer (MOET), in vitro handling of oocytes and production of embryos, reproductive cloning and emerging technologies (sex selection, gene targeting and nuclear transfer for livestock transgenesis, genomics for marker-assisted selection, etc.). The application of these technologies for cattle breeding is critically discussed in relation to their impact in the improvement of the efficiency of dairy and beef production in developed and - particularly - in developing countries, which ultimately rule the possibilities of a competitive and sound production of food for human consumption. Despite the remarkable progress made and the punctual importance of some of the above-mentioned technologies, AI remains the most important assisted reproductive technology (ART) in developing countries. Any attempt to gain widespread of any other ART under the predominant economical conditions in developing countries ought to match the simplicity and the success of AI as a breeding tool.     

Production of transgenic livestock: promise fulfilled

The introduction of specific genes into the genome of farm animals and its stable incorporation into the germ line has been a major technological advance in agriculture. Transgenic technology provides a method to rapidly introduce "new" genes into cattle, swine, sheep, and goats without crossbreeding. It is a more extreme methodology, but in essence, not really different from crossbreeding or genetic selection in its result. Methods to produce transgenic animals have been available for more than 20 yr, yet recently lines of transgenic livestock have been developed that have the potential to improve animal agriculture and benefit producers and/or consumers. There are a number of methods that can be used to produce transgenic animals. However, the primary method to date has been the microinjection of genes into the pronuclei of zygotes. This method is one of an array of rapidly developing transgenic methodologies. Another method that has enjoyed recent success is that of nuclear transfer or "cloning." The use of this technique to produce transgenic livestock will profoundly affect the use of transgenic technology in livestock production. Cell-based, nuclear transfer or cloning strategies have several distinct advantages for use in the production of transgenic livestock that cannot be attained using pronuclear injection of DNA. Practical applications of transgenesis in livestock production include enhanced prolificacy and reproductive performance, increased feed utilization and growth rate, improved carcass composition, improved milk production and/or composition, and increased disease resistance. One practical application of transgenics in swine production is to improve milk production and/or composition. To address the problem of low milk production, transgenic swine over-expressing the milk protein bovine alpha-lactalbumin were developed and characterized. The outcomes assessed were milk composition, milk yield, and piglet growth. Our results indicate that transgenic overexpression of milk proteins may provide a means to improve swine lactation performance.     

Ethical stockmanship

The objective of this review is to consider the ethics of stockmanship, particularly from the perspective of the nature and extent of the duties of stockpeople to their farm animals. It will consider what science tells us about the impact of stockmanship on the animal, particularly the welfare of the farm animal. The effects of human-animal interactions on the stockperson will also be considered, since these interactions affect the work performance and job satisfaction of the stockperson and thus indirectly affect animal welfare. Animal ethics is broader than animal welfare and includes economic as well as philosophical, social, cultural and religious aspects. This paper is predicated on the view that farm animals can suffer, and that animal suffering is a key consideration in our moral obligations to animals. Housing and husbandry practices affect farm animal welfare and thus farmers and stockpeople have a responsibility to provide, at minimum, community-acceptable animal housing and husbandry standards for their animals. The farmer's or stockperson's attitudes and behaviour can directly affect the animal's welfare and thus they also have a responsibility to provide specific standards of stockmanship for these animals. However, research suggests that the behaviour of some stockpeople is not as correct as it might be. Such situations exemplify the inevitably unequal human - domestic animal relationship, and this inequality should be considered in analysing the boundary between right and wrong behaviour of humans. Thus ethical discussion, using science and other considerations and involving stockpeople, livestock industries, government and the general public, should be used to establish and assure acceptable stockperson competencies across the livestock industries. Training programs targeting the key attitudes and behaviour of stockpeople presently offer the livestock industries good opportunities to improve human-animal interactions.     

The importance of applied ethology for the execution of animal welfare legislation]

Applied ethology in general and farm animal ethology in particular have a great importance in connection with animal welfare regulations on a national and international level. They have through the legislation on animal welfare brought about important repercussions on housing of farm animals, wild animals and experimental animals in Switzerland. The animal welfare legislation has been a considerable boost to scientific research. The application of ethological knowledge in the legislation and in practice has indeed also its limits. The importance of applied ethology for animal welfare and for animal husbandry will still increase in future.     

The most important causes for quantitative and qualitative losses during breeding and turnover before slaughtering of farm animals

The present review article deals with the current knowledge on the most important reasons of quantitative and qualitative losses during breeding and turnover before slaughtering of farm animals. At present, the profitability of every economic activity of people is an indispensable necessity. As regards farm animals there are many factors, seemingly banal ones, that determine the final economical effect and satisfaction of such activity. Many essential errors are made by producers as a result of ignorance or not complying with the basic needs of their behaviour. The turnover only deepens these neglects. The most essential limitations of these quantitative and qualitative losses are the proper selection and joining of the animals as well as an exact knowledge and complying with their natural behaviour. Such activity decreases animal stress, improves a proper welfare and consequently increases economical effects considerably.