Evaluation of organic,conventional and intensive beef farm systems: health,management and animal production

The overall aim of the present study was to analyse and compare organic beef cattle farming in Spain with intensive and conventional systems. An on-farm study comparing farm management practices and animal health was carried out. The study also focussed on a slaughterhouse analysis by comparing impacts on the safety and quality of the cattle products. Twenty-four organic and 26 conventional farms were inspected, and farmers responded to a questionnaire that covered all basic data on their husbandry practices, farm management, veterinary treatments and reproductive performance during 2007. Furthermore, data on the hygiene and quality of 244, 2596 and 3021 carcasses of calves from organic, intensive and conventional farms, respectively, were retrieved from the official yearbook (2007) of a slaughterhouse. Differences found between organic and conventional farms across the farm analysis did not substantially reflect differences between both farm types in the predominant diseases that usually occur on beef cattle farms. However, calves reared organically presented fewer condemnations at slaughter compared with intensive and to a lesser extent with conventionally reared calves. Carcass performance also reflected differences between farm type and breed and was not necessarily better in organic farms.     

Expression of Helicobacter pylori TonB protein in transgenic Arabidopsis thaliana: toward production of vaccine antigens in plants

Background: The aim of this study was to produce a recombinant version of the highly antigenic Helicobacter pylori TonB (iron‐dependent siderophore transporter protein HP1341) in transgenic plants as a candidate oral vaccine antigen. Materials and Methods: Using Agrobacterium‐mediated gene transfer, we introduced three different constructs of the tonB gene into the genome of the model plant Arabidopsis thaliana. We investigated transgene insertion by PCR, produced TonB antibodies for analysis of the production of the recombinant protein in plants, verified the identity of the protein produced by mass spectrometry analysis, and analyzed the number of genetic inserts in the plants by Southern blotting. Results: Three different constructs of the expression cassette (full‐length tonB, tonB truncated in the 5′ end removing the codons for a transmembrane helix, and the latter construct with codons for the endoplasmic reticulum SEKDEL retention signal added to the 3′ end) were used to find the most effective way to express the TonB antigen. Production of TonB protein was detected in plants transformed with each of the constructs, confirmed by both Western blotting and mass spectrometry analysis. No considerable differences in protein expression from the three different constructs were observed. The protein concentration in the plants was at least 0.05% of the total soluble proteins. Conclusions: The Helicobacter pylori TonB protein can be produced in Arabidopsis thaliana plants in a form that is recognizable by rabbit anti‐TonB antiserum. These TonB‐expressing plants are highly suitable for animal studies of oral adminstration as a route for immunization against Helicobacter infections.     

Expression of antibodies and Fab fragments in transgenic potato plants: a case study for bulk production in crop plants

To use transgenic potato tubers (Solanum tuberosum cv. Désirée) for bulk production of recombinant antibodies, constructs were engineered for accumulating full-size IgGs and Fab fragments in the plant cell apoplast or endoplasmic reticulum (ER). An in-house transformation protocol was worked out for the efficient co-transformation of potato root explants. Accumulation levels in tubers of up to 0.5% of total soluble protein were found for antibodies targeted to the ER whereas five-fold lower accumulation levels were found for antibodies targeted for secretion. Additionally, different aspects important for the commercial exploitation of potato tubers as a heterologous production system were analysed. Tubers could be stored for up to 6 months without significant loss of antibody amount or activity. Minor variations in antibody accumulation levels were observed in tubers that originated from the same transformant. Most isolated IgGs and Fab fragments bound the antigen and had the correct molecular weight when compared with the hybridoma-derived standard. Processing to greenhouse or field trials, including in vitro propagation of a selected transformant, required only approximately 9 months from the start of transformation, a time frame in which hundreds of kilograms of transgenic potato tubers could easily be obtained. Small-scale purification of IgG was possible by using standard laboratory techniques. Thus, molecular farming in potato tubers can be a viable production system for economic production of clinically or industrially interesting macromolecules, such as antibodies.     

IBDV丝素蛋白/壳聚糖DNA微球疫苗的制备及免疫原性分析

为了制备传染性法氏囊病病毒(Infectious bursal disease virus,IBDV)DNA微球疫苗,并评价其免疫效果。以丝素蛋白(Silk fibroin,SF)/壳聚糖(Chitosan,CS)为壁材,IBDV的VP2/4/3 DNA疫苗为芯材,通过戊二醛和Na2SO4介导的乳化交联技术,制备出SF/CS复合微球疫苗,然后经肌肉注射14日龄非免疫鸡,2周后加强免疫一次,酶联免疫法(ELISA)定期监测鸡血清的IBDV抗体,以研究微球化疫苗的免疫原性。结果显示:戊二醛介导交联方法影响荷载DNA疫苗的活性,而Na2SO4介导的交联方法操作简单且不影响荷载DNA活性;建立了以壳聚糖浓度0.5%(pH 5.0)、丝素蛋白浓度0.6%,质粒DNA 500μg/mL溶解在2%Na2SO4溶液中的工作条件;SF-CS复合微球DNA疫苗荷载率89.14%,大小1.98μm,对DNaseⅠ的消化有保护作用。免疫后的抗IBDV血清ELISA抗体的检测显示,微球免疫组总体高于质粒疫苗免疫组(P0.05),而且SF/CS复合微球组免疫反应要略高于单纯CS包被的抗原组。研究表明,丝素蛋白/壳聚糖作为微球佐剂能提高IBDV DNA疫苗的临床免疫效果,有很好的应用前景。     

A practical validation approach for virus titer testing of avian infectious bursal disease live vaccine according to current regulatory guidelines

The method for virus titer determination of avian infectious bursal disease (IBD) live vaccine, developed long before regulatory validation guidelines is a cell culture based biological assay intended for use in vaccine release testing.The aim of our study was to perform a validation, based on fit-for-purpose principle, of an old 50% tissue culture infectious dose (TCID₅₀) method according to Guidelines of the International Cooperation on Harmonization of Technical Requirements for Registration of Veterinary Medicinal Products (VICH).This paper addresses challenges and discusses some key aspects that should be considered when validating biological methods. A different statistical approach and non-parametric statistics was introduced in validation protocol in order to derive useful information from experimental data. This approach is applicable for a wide range of methods.In conclusion, the previous virus titration method had showed to be precise, accurate, linear, robust and in accordance with current regulatory standards, which indicates that there is no need for additional re-development or upgrades of the method for its suitability for intended use.     

Transgenic tobacco plants producing caffeine: a potential new strategy for insect pest control

Caffeine (1,3,7–trimethylxanthine) is one of the most widely used plant secondary metabolites, primarily as a stimulant and an ingredient in drugs. In nature, caffeine is believed to function in chemical defense, acting as an antiherbivory and allelopathic agent, and therefore it might be employed to protect agriculturally important crop plants. In coffee plants, caffeine is synthesized from the precursor xanthosine in four steps, three N-methylations and removal of ribose. We had previously isolated genes encoding three distinct N-methyltransferases, and we demonstrated production of recombinant enzymes that yielded caffeine in in vitro reconstitution experiments. When these caffeine biosynthetic pathway genes were simultaneously expressed in tobacco plants (Nicotiana tabacum), caffeine was successfully produced up to 5 μg/g fresh weight in leaves. The leaves were unpalatable to tobacco cutworms (Spodoptera litura). This repellent action appeared to be more widely␣applicable to lepidopteran caterpillars as observed with small white (Pieris rapae) fed on Chinese cabbages that had been top-treated with caffeine. Our recent results suggest a novel approach to strengthen anti-herbivore traits by producing caffeine in crop plants.     

Transgenic tobacco plants as production platform for biologically active human interleukin 2 and its fusion with proteinase inhibitors

Transgenic plants offer a low‐cost approach for the production of pharmaceutically important and commercially valuable recombinant proteins. Our studies were focused on the plant‐based production of human interleukin 2 (hIL‐2) and its fusion with proteinase inhibitors, either SPI2 from Galleria mellonella or CMTI from Cucurbita maxima. Finally, five plant expression cassettes were obtained. Three of them contained the single cDNA encoding CMTI I, SPI2 and hIL‐2, respectively, while two of them contained the translational fusion, SPI2::hIL‐2 and CMTI::hIL‐2. In all cases, the transgenes were controlled by the RbcS1 promoter and terminator and the recombinant proteins were targeted to the endoplasmic reticulum. After tobacco transformation, five groups of transgenic plants were obtained and analysed. The level of recombinant proteins was estimated either by Western blot or by ELISA. The biological activity of plant‐produced hIL‐2 alone or in a fusion with SPI2 or CMTI was confirmed using the mammalian cells proliferation assay. The activities of proteinase inhibitors were confirmed in proteolysis assay using azocoll as a substrate. The usefulness of using proteinase inhibitor CMTI I in a fusion with hIL‐2 as a protective agent against trypsin digestion was demonstrated.     

Sociality and health: impacts of sociality on disease susceptibility and transmission in animal and human societies

This paper introduces a theme issue presenting the latest developments in research on the impacts of sociality on health and fitness. The articles that follow cover research on societies ranging from insects to humans. Variation in measures of fitness (i.e. survival and reproduction) has been linked to various aspects of sociality in humans and animals alike, and variability in individual health and condition has been recognized as a key mediator of these relationships. Viewed from a broad evolutionary perspective, the evolutionary transitions from a solitary lifestyle to group living have resulted in several new health-related costs and benefits of sociality. Social transmission of parasites within groups represents a major cost of group living, but some behavioural mechanisms, such as grooming, have evolved repeatedly to reduce this cost. Group living also has created novel costs in terms of altered susceptibility to infectious and non-infectious disease as a result of the unavoidable physiological consequences of social competition and integration, which are partly alleviated by social buffering in some vertebrates. Here, we define the relevant aspects of sociality, summarize their health-related costs and benefits, and discuss possible fitness measures in different study systems. Given the pervasive effects of social factors on health and fitness, we propose a synthesis of existing conceptual approaches in disease ecology, ecological immunology and behavioural neurosciences by adding sociality as a key factor, with the goal to generate a broader framework for organismal integration of health-related research.     

Infectious disease in animal metapopulations: the importance of environmental transmission

Motivated by an array of infectious diseases that threaten wildlife populations, a simple metapopulation model (subpopulations connected by animal movement) is developed, which allows for both movement-based and environmental transmission. The model demonstrates that for a range of plausible parameterizations of environmental transmission, increased movement rate of animals between discrete habitats can lead to a decrease in the overall proportion of sites that are occupied. This can limit the ability of the rescue effect to ensure locally extinct populations become recolonized and can drive metapopulations down in size so that extinction by mechanisms other than disease may become more likely. It further highlights that, in the context of environmental transmission, the environmental persistence time of pathogens and the probability of acquiring infection by environmental transmission can affect host metapopulations both qualitatively and quantitatively. Additional spillover sources of infection from alternate reservoir hosts are also included in the model and a synthesis of all three types of transmission, acting alone or in combination, is performed revealing that movement-based transmission is the only necessary condition for a decline in the proportion of occupied sites with increasing movement rate, but that the presence of other types of transmission can reverse this qualitative result. By including the previously neglected role of environmental transmission, this work contributes to the general discussion of when dispersal by wild animals is beneficial or detrimental to populations experiencing infectious disease.     

Efficacy of bacteriophage Lysed Pasteurella marker vaccine in laboratory animal models with a novel DIVA for haemorrhagic septicaemia

Objective

The present study aimed at evaluating the efficacy of an improved phage lysate marker vaccine for haemorrhagic septicaemia in mice and rabbit model and development of a DIVA ELISA based on iron restricted outer membrane protein (IROMP).

Open questions and recent advances in the control of a multi‐host infectious disease: animal tuberculosis

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Glycan-masking hemagglutinin antigens from stable CHO cell clones for H5N1 avian influenza vaccine development

Refocusing of B-cell responses can be achieved by preserving the overall fold of the antigen structure but selectively mutating the undesired antigenic sites with additional N-linked glycosylation motifs for glycan masking the vaccine antigen. We previously reported that glycan-masking recombinant H5 hemagglutinin (rH5HA) antigens on residues 83, 127, and 138 (g127 + g138 or g83 + g127 + 138 rH5HA) elicited broader neutralizing antibodies and protection against heterologous clades/subclades of high pathogenic avian influenza H5N1 viruses. In this study, we engineered the stably expressing Chinese hamster ovary (CHO) cell clones for producing the glycan-masking g127 + g138 and g83 + g127 + g138 rH5HA antigens. All of these glycan-masking rH5HA antigens produced in stable CHO cell clones were found to be mostly oligomeric structures. Only the immunization with the glycan-masking g127 + g138 but not g83 + g127 + g138 rH5HA antigens elicited more potent neutralizing antibody titers against four out of five heterologous clades/subclades of H5N1 viral strains. The increased neutralizing antibody titers against these heterologous viral strains were correlated with the increased amounts of stem-binding antibodies, only the glycan-masking g127 + g138 rH5HA antigens can translate into more protection against live viral challenges. The stable CHO cell line-produced glycan-masking g127 + g138 rH5HA can be used for H5N1 subunit vaccine development.     

Chemical evidence for covalent linkages of a semisynthetic glycoconjugate vaccine forHaemophilus influenzae type b disease

We have defined the nature of the covalent linkages in aHaemophilus influenzae type b oligosaccharide-CRM₁₉₇ conjugate vaccine, designated HbOC. The conjugate was acid hydrolyzed to release a novel amino-acid derivative,N-(2-hydroxyethyl)lysine (OHEt-Lys), identifiable with an amino-acid analyzer. This amino-acid derivative was formed by reduction of Schiff bases formed betweenH. influenzae type b oligosaccharides (HbO) and the lysyl -amino groups of CRM₁₉₇ (a non-toxic, cross-reactive variant of diphtheria toxin), followed by acid hydrolysis of HbOC. Quantification of OHEt-Lys per CRM₁₉₇ molecule allowed the determination of a covalency ratio, a useful parameter for evaluating the stoichiometry and consistency of HbOC preparations. Covalent association between HbO and CRM₁₉₇ was also demonstrated by the coincidence of immunoreactivity of gelelectrophoresed HbOC on a Western blot probed with anti-CRM₁₉₇ and anti-saccharide antisera.     

Evaluation of vaccines against enteric infections: a clinical and public health research agenda for developing countries

Enteric infections are a major cause of morbidity and mortality in developing countries. To date, vaccines have played a limited role in public health efforts to control enteric infections. Licensed vaccines exist for cholera and typhoid, but these vaccines are used primarily for travellers; and there are two internationally licensed vaccines for rotavirus, but they are mainly used in affluent countries. The reasons that enteric vaccines are little used in developing countries are multiple, and certainly include financial and political constraints. Also important is the need for more cogent evidence on the performance of enteric vaccines in developing country populations. A partial inventory of research questions would include: (i) does the vaccine perform well in the most relevant settings? (ii) does the vaccine perform well in all epidemiologically relevant age groups? (iii) is there adequate evidence of vaccine safety once the vaccines have been deployed in developing countries? (iv) how effective is the vaccine when given in conjunction with non-vaccine cointerventions? (v) what is the level of vaccine protection against all relevant outcomes? and (vi) what is the expected population level of vaccine protection, including both direct and herd vaccine protective effects? Provision of evidence addressing these questions will help expand the use of enteric vaccines in developing countries.     

Ethical models underpinning responses to threats to public health: a comparison of approaches to communicable disease control in Europe

Increases in international travel and migratory flows have enabled infectious diseases to emerge and spread more rapidly than ever before. Hence, it is increasingly easy for local infectious diseases to become global infectious diseases (GIDs). National governments must be able to react quickly and effectively to GIDs, whether naturally occurring or intentionally instigated by bioterrorism. According to the World Health Organisation, global partnerships are necessary to gather the most up-to-date information and to mobilize resources to tackle GIDs when necessary. Communicable disease control also depends upon national public health laws and policies. The containment of an infectious disease typically involves detection, notification, quarantine and isolation of actual or suspected cases; the protection and monitoring of those not infected; and possibly even treatment. Some measures are clearly contentious and raise conflicts between individual and societal interests. In Europe national policies against infectious diseases are very heterogeneous. Some countries have a more communitarian approach to public health ethics, in which the interests of individual and society are more closely intertwined and interdependent, while others take a more liberal approach and give priority to individual freedoms in communicable disease control. This paper provides an overview of the different policies around communicable disease control that exist across a select number of countries across Europe. It then proposes ethical arguments to be considered in the making of public health laws, mostly concerning their effectiveness for public health protection.     

Keeping the herds healthy and alert: implications of predator control for infectious disease

Predator control programmes are generally implemented in an attempt to increase prey population sizes. However, predator removal could prove harmful to prey populations that are regulated primarily by parasitic infections rather than by predation. We develop models for microparasitic and macroparasitic infection that specify the conditions where predator removal will (a) increase the incidence of parasitic infection, (b) reduce the number of healthy individuals in the prey population and (c) decrease the overall size of the prey population. In general, predator removal is more likely to be harmful when the parasite is highly virulent, macroparasites are highly aggregated in their prey, hosts are long‐lived and the predators select infected prey.