A technique to detect and to quantify fasciocutaneous blood vessels in small laboratory animals ex vivo

PURPOSE: A microangiographical technique is described, which allows visualization of small and capillary blood vessels and quantification of fasciocutaneous blood vessels by means of digital computer analysis in very small laboratory animals. MATERIALS AND METHODS: The left carotid artery of 20 nu/nu mice was cannulated (26 gauge) and a mixture of gelatin, bariumsulfate, and green ink was injected according to standardized protocol. Fasciocutaneous blood vessels were visualized by digital mammography and analyzed for vessel length and vessel surface area as standardized units [SU] by computer program. RESULTS: With the described microangiography method, fasciocutaneous blood vessels down to capillary size level can be clearly visualized. Regions of interest (ROIs) can be defined and the containing vascular network quantified. Comparable results may be obtained by calculating the microvascular area index (MAI) and the microvascular length index (MLI), related to the ROIs size. Identical ROIs showed a high reproducibility for measured [SU] < 0.01 +/- 0.0012%. CONCLUSION: Combining microsurgical techniques, pharmacological knowledge, and modern digital image technology, we were able to visualize small and capillary blood vessels even in small laboratory animals. By using our own computer analytical program, quantification of vessels was reliable, highly reproducible, and fast.

Effects of Novel Growth Hormone Secretagogues on Growth Hormone Secretion in Farm Animals

The requirement for growth hormone (GH) secretion by the anterior pituitary gland in beef calves is demonstrated by a complete lack of long bone-growth and muscle accretion after hypophysectomy (surgical removal of the pituitary gland). When the connecting link (hypophyseal stalk) to the basal region (hypothalamus) of the brain is surgically severed, long bone growth and body weight gain are greatly limited compared with sham-operated controls. This limited growth results from obliteration of episodic GH secretion and reduced basal blood concentration of the hormone compared with sham-operated controls. Thus, the hypophyseal stalk-transected (HST) calf provides an appropriate model to determine mechanisms by which hypothalamic neuropeptides from the brain regulate GH secretion, and thereby growth in the young calf. Neuropeptides have been isolated and characterized in bovine hypothalamus that stimulate GH secretion (GH-releasing hormone [GHRH]) or factor [GHRF] and inhibit GH secretion (GH release-inhibiting hormone [GHRIH] or somatostatin [SRIH]). A dose of .067 micrograms of GHRF per kilogram of body weight injected intravenously in HST calves abruptly increased plasma GH concentration to 55 nanograms per milliliter from the control period mean of 5 nanograms per milliliter. HST calves then were infused intravenously with .033 and .067 microgram somatostatin per kilogram of body weight, during which a pulse injection of .067 microgram of GHRF was administered. GH increase was limited to 9 and 5 micrograms per kilogram body weight during the .033- and .067 microgram SRIH infusions after GHRF; no GH rebound was observed after the SRIH was discontinued. GHRF from humans contains 40 to 44 amino acids. Rat hypothalamic GHRF analogs containing 29 to 32 amino acids elicited dose-dependent GH peak release in these HST calves. In 1977, Bowers and Monomy isolated novel GH releasing peptides consisting of only six amino acids; they caused GH release by isolated pituitary cells in culture and acute GH release when administered intravenously. We recently have utilized a novel nonpeptidyl GH secretagogue of low molecular weight in the pig to determine its mechanisms of action within the central nervous system.

The COLOSS BEEBOOK Volume I, Standard methods for Apis mellifera research: Introduction

The COLOSS BEEBOOK is a practical manual compiling standard methods in all fields of research on the western honey bee, Apis mellifera. The COLOSS network was founded in 2008 as a consequence of the heavy and frequent losses of managed honey bee colonies experienced in many regions of the world (Neumann and Carreck, 2010). As many of the world’s honey bee research teams began to address the problem, it soon became obvious that a lack of standardized research methods was seriously hindering scientists’ ability to harmonize and compare the data on colony losses obtained internationally. In its second year of activity, during a COLOSS meeting held in Bern, Switzerland, the idea of a manual of standardized honey bee research methods emerged. The manual, to be called the COLOSS BEEBOOK, was inspired by publications with similar purposes for fruit fly research (Lindsley and Grell, 1968; Ashburner 1989; Roberts, 1998; Greenspan, 2004).

The COLOSS BEEBOOK Volume II, Standard methods for Apis mellifera pest and pathogen research: Introduction

The COLOSS BEEBOOK is a practical manual compiling standard methods in all fields of research on the western honey bee, Apis mellifera. The COLOSS network was founded in 2008 as a consequence of the heavy and frequent losses of managed honey bee colonies experienced in many regions of the world (Neumann and Carreck, 2010). As many of the world’s honey bee research teams began to address the problem, it soon became obvious that a lack of standardized research methods was seriously hindering scientists’ ability to harmonize and compare the data on colony losses obtained internationally. In its second year of activity, during a COLOSS meeting held in Bern, Switzerland, the idea of a manual of standardized honey bee research methods emerged. The manual, to be called the COLOSS BEEBOOK, was inspired by publications with similar purposes for fruit fly research (Lindsley and Grell, 1968; Ashburner, 1989; Roberts, 1998; Greenspan, 2004).

Standard methods for Nosema research

Methods are described for working with Nosema apis and Nosema ceranae in the field and in the laboratory. For fieldwork, different sampling methods are described to determine colony level infections at a given point in time, but also for following the temporal infection dynamics. Suggestions are made for how to standardise field trials for evaluating treatments and disease impact. The laboratory methods described include different means for determining colony level and individual bee infection levels and methods for species determination, including light microscopy, electron microscopy, and molecular methods (PCR). Suggestions are made for how to standardise cage trials, and different inoculation methods for infecting bees are described, including control methods for spore viability. A cell culture system for in vitro rearing of Nosema spp. is described. Finally, how to conduct different types of experiments are described, including infectious dose, dose effects, course of infection and longevity tests

Minimising pain in farm animals: the 3S approach - 'Suppress, Substitute, Soothe'

Recently, the French National Institute for Agricultural Research appointed an expert committee to review the issue of pain in food-producing farm animals. To minimise pain, the authors developed a '3S' approach accounting for 'Suppress, Substitute and Soothe' by analogy with the '3Rs' approach of 'Reduction, Refinement and Replacement' applied in the context of animal experimentation. Thus, when addressing the matter of pain, the following steps and solutions could be assessed, in the light of their feasibility (technical constraints, logistics and regulations), acceptability (societal and financial aspects) and availability. The first solution is to suppress any source of pain that brings no obvious advantage to the animals or the producers, as well as sources of pain for which potential benefits are largely exceeded by the negative effects. For instance, tail docking of cattle has recently been eliminated. Genetic selection on the basis of resistance criteria (as e.g. for lameness in cattle and poultry) or reduction of undesirable traits (e.g. boar taint in pigs) may also reduce painful conditions or procedures. The second solution is to substitute a technique causing pain by another less-painful method. For example, if dehorning cattle is unavoidable, it is preferable to perform it at a very young age, cauterising the horn bud. Animal management and constraint systems should be designed to reduce the risk for injury and bruising. Lastly, in situations where pain is known to be present, because of animal management procedures such as dehorning or castration, or because of pathology, for example lameness, systemic or local pharmacological treatments should be used to soothe pain. These treatments should take into account the duration of pain, which, in the case of some management procedures or diseases, may persist for longer periods. The administration of pain medication may require the intervention of veterinarians, but exemptions exist where breeders are allowed to use local anaesthesia (e.g. castration and dehorning in Switzerland). Extension of such exemptions, national or European legislation on pain management, or the introduction of animal welfare codes by retailers into their meat products may help further developments. In addition, veterinarians and farmers should be given the necessary tools and information to take into account animal pain in their management decisions.

The COLOSS BEEBOOK Volume II: Standard methods for Apis mellifera pest and pathogen research

n recent years, declines of honey bee populations have received massive media attention worldwide, yet attempts to understand the causes have been hampered by a lack of standardisation of laboratory techniques. Published as a response to this, the COLOSS BEEBOOK is a unique collaborative venture involving 234 bee scientists from 34 countries, who have produced the definitive guide to how to carry out research on honey bees. It is hoped that these volumes will become the standards to be adopted by bee scientists worldwide. Volume II includes approximately 600 separate protocols dealing with the study of the pests and diseases of the honey bee, Apis mellifera. These cover epidemiology and surveying techniques, virus diseases, bacterial diseases such as European and American foulbrood, fungal and microsporidian diseases such as Nosema, mites such as Acarapis, Varroa and Tropilaelaps, and other pests such as the small hive beetle.

The COLOSS BEEBOOK Volume I: Standard Methods for Apis mellifera Research

In recent years, declines of honey bee populations have received massive media attention worldwide, yet attempts to understand the causes have been hampered by a lack of standardisation of laboratory techniques. Published as a response to this, the COLOSS BEEBOOK is a unique collaborative venture involving 234 bee scientists from 34 countries, who have produced the definitive guide to how to carry out research on honey bees. It is hoped that these volumes will become the standards to be adopted by bee scientists worldwide. Volume I includes approximately 1,100 separate protocols dealing with the study of the honey bee, Apis mellifera. These cover anatomy, behavioural studies, chemical ecology, breeding, genetics, instrumental insemination and queen rearing, pollination, molecular studies, statistics, toxicology and numerous other techniques

Conservation of the Cinereous Vulture Aegypius monachus in Spain (1966–2011): a bibliometric review of threats, research and adaptive management

Detecting and quantifying threats and researching and implementing management actions are key to improving the conservation status of endangered species. Bibliometric analysis can constitute a useful tool for the evaluation of such questions from a long-term perspective. Taking as a case study the Cinereous Vulture Aegypius monachus in Spain, we tested relationships between population dynamics, research efforts, existing threats and conservation milestones. The population growth of the species (from 206 pairs in 1976 to 2,068 in 2011) was parallelled by the increase in the total number of publications, the number of articles in SCI journals and the number of published works dealing with aspects of conservation, threats and management. These results are discussed in terms of cause-effect relationships taking into account that the influence of other non-mutually exclusive factors could also probably explain such associations. Similarly, we analysed the trend of the Cinereous Vulture breeding population with respect to different threats and indices of food availability, obtaining a positive correlation with the increase in big-game hunting bags in Spain. With respect to conservation milestones, we concluded that the current situation is positive in terms of the protection of the species and its habitat, with the situation in relation to food availability being unclear. Finally, we reviewed the main conservation actions that have been taken for the species in Spain and how these have been progressively modified based on new scientific and technical evidence, as an example of adaptive management applied to conservation.

Soil characteristics and Collembola and Acari abundance in control and warming plots at Abisco Research Station

Extreme weather events can have negative impacts on species survival and community structure when surpassing lethal thresholds. Extreme winter warming events in the Arctic rapidly melt snow and expose ecosystems to unseasonably warm air (2-10 °C for 2-14 days), but returning to cold winter climate exposes the ecosystem to lower temperatures by the loss of insulating snow. Soil animals, which play an integral part in soil processes, may be very susceptible to such events depending on the intensity of soil warming and low temperatures following these events. We simulated week-long extreme winter warming events - using infrared heating lamps, alone or with soil warming cables - for two consecutive years in a sub-Arctic dwarf shrub heathland. Minimum temperatures were lower and freeze-thaw cycles were 2-11 times more frequent in treatment plots compared with control plots. Following the second event, Acari populations decreased by 39%; primarily driven by declines of Prostigmata (69%) and the Mesostigmatic nymphs (74%). A community-weighted vertical stratification shift occurred from smaller soil dwelling (eu-edaphic) Collembola species dominance to larger litter dwelling (hemi-edaphic) species dominance in the canopy-with-soil warming plots compared with controls. The most susceptible groups to these winter warming events were the smallest individuals (Prostigmata and eu-edaphic Collembola). This was not apparent from abundance data at the Collembola taxon level, indicating that life forms and species traits play a major role in community assembly following extreme events. The observed shift in soil community can cascade down to the micro-flora affecting plant productivity and mineralization rates. Short-term extreme weather events have the potential to shift community composition through trait composition with potentially large consequences for ecosystem development.

The ARKdb: genome databases for farmed and other animals

The ARKdb genome databases provide comprehensive public repositories for genome mapping data from farmed species and other animals (http://www.thearkdb.org) providing a resource similar in function to that offered by GDB or MGD for human or mouse genome mapping data, respectively. Because we have attempted to build a generic mapping database, the system has wide utility, particularly for those species for which development of a specific resource would be prohibitive. The ARKdb genome database model has been implemented for 10 species to date. These are pig, chicken, sheep, cattle, horse, deer, tilapia, cat, turkey and salmon. Access to the ARKdb databases is effected via the World Wide Web using the ARKdb browser and Anubis map viewer. The information stored includes details of loci, maps, experimental methods and the source references. Links to other information sources such as PubMed and EMBL/GenBank are provided. Responsibility for data entry and curation is shared amongst scientists active in genome research in the species of interest. Mirror sites in the United States are maintained in addition to the central genome server at Roslin.

Overexpression of a Novel Arabidopsis Gene Related to Putative Zinc-Transporter Genes from Animals Can Lead to Enhanced Zinc Resistance and Accumulation

We describe the isolation of an Arabidopsis gene that is closely related to the animal ZnT genes (Zn transporter). The protein encoded by the ZAT (Zn transporter of Arabidopsis thaliana) gene has 398 amino acid residues and is predicted to have six membrane-spanning domains. To obtain evidence for the postulated function of the Arabidopsis gene, transgenic plants with the ZAT coding sequence under control of the cauliflower mosaic virus 35S promoter were analyzed. Plants obtained with ZAT in the sense orientation exhibited enhanced Zn resistance and strongly increased Zn content in the roots under high Zn exposure. Antisense mRNA-producing plants were viable, with a wild-type level of Zn resistance and content, like plants expressing a truncated coding sequence lacking the C-terminal cytoplasmic domain of the protein. The availability of ZAT can lead to a better understanding of the mechanism of Zn homeostasis and resistance in plants.

Polypeptide signaling for plant defensive genes exhibits analogies to defense signaling in animals.

The activation of plant defensive genes in leaves of tomato plants in response to herbivore damage or mechanical wounding is mediated by a mobile 18-amino acid polypeptide signal called systemin. Systemin is derived from a larger, 200-amino acid precursor called prosystemin, similar to polypeptide hormones and soluble growth factors in animals. Systemin activates a lipid-based signaling cascade, also analogous to signaling systems found in animals. In plants, linolenic acid is released from membranes and is converted to the oxylipins phytodienoic acid and jasmonic acid through the octadecanoid pathway. Plant oxylipins are structural analogs of animal prostaglandins which are derived from arachidonic acid in response to various signals, including polypeptide factors. Constitutive overexpression of the prosystemin gene in transgenic tomato plants resulted in the overproduction of prosystemin and the abnormal release of systemin, conferring a constitutive overproduction of several systemic wound-response proteins (SWRPs). The data indicate that systemin is a master signal for defense against attacking herbivores. The same defensive proteins induced by wounding are synthesized in response to oligosaccharide elicitors that are generated in leaf cells in response to pathogen attacks. Inhibitors of the octadecanoid pathway, and a mutation that interrupts this pathway, block the induction of SWRPs by wounding, systemin, and oligosaccharide elicitors, indicating that the octadecanoid pathway is essential for the activation of defense genes by all of these signals. The tomato mutant line that is functionally deficient in the octadecanoid pathway is highly susceptible to attacks by Manduca sexta larvae. The similarities between the defense signaling pathway in tomato leaves and those of the defense signaling pathways of macrophages and mast cells of animals suggests that both the plant and animal pathways may have evolved from a common ancestral origin.