Privileged frameworks from snake venom.

Venom as a form of chemical prey capture is a key innovation that has underpinned the explosive radiation of the advanced snakes (Caenophidia). Small venom proteins are often rich in disulfide bonds thus facilitating stable molecular scaffolds that present key functional residues on the protein surface. New toxin types are initially developed through the venom gland over-expression of normal body proteins, their subsequent gene duplication and diversification that leads to neofunctionalisation as random mutations modify their structure and function. This process has led to preferentially selected (privileged) cysteine-rich scaffolds that enable the snake to build arrays of toxins many of which may lead to therapeutic products and research tools. This review focuses on cysteine-rich small proteins and peptides found in snake venoms spanning natriuretic peptides to phospholipase enzymes, while highlighting their three-dimensional structures and biological functions as well as their potential as therapeutic agents or research tools.

Stable isotope and trace metal compositions of Australian prawns as a guide to authenticity and wholesomeness.

This research has explored the potential of stable isotope and trace metal profiles to distinguish Australian prawns from prawns imported from neighbouring Asian countries. Australian prawns were collected mostly from the Brisbane area. Strong differences in Australian vs. imported prawns were evident from both the isotope and trace element data, with the differences most likely occurring because imported prawns are typically reared in aquaculture facilities and frozen prior to sale in Australia. The aquaculture origins are characterised by comparatively; low δHVSMOW, δ(13)CVPDB values, low concentrations of arsenic, zinc and potassium, and high water contents (>80%). Relatively high arsenic and cadmium contents were found within Australian prawns, but the concentrations did not exceed local human health guidelines.

Accounting for the impact of conservation on human well-being.

Conservationists are increasingly engaging with the concept of human well-being to improve the design and evaluation of their interventions. Since the convening of the influential Sarkozy Commission in 2009, development researchers have been refining conceptualizations and frameworks to understand and measure human well-being and are starting to converge on a common understanding of how best to do this. In conservation, the term human well-being is in widespread use, but there is a need for guidance on operationalizing it to measure the impacts of conservation interventions on people. We present a framework for understanding human well-being, which could be particularly useful in conservation. The framework includes 3 conditions; meeting needs, pursuing goals, and experiencing a satisfactory quality of life. We outline some of the complexities involved in evaluating the well-being effects of conservation interventions, with the understanding that well-being varies between people and over time and with the priorities of the evaluator. Key challenges for research into the well-being impacts of conservation interventions include the need to build up a collection of case studies so as to draw out generalizable lessons; harness the potential of modern technology to support well-being research; and contextualize evaluations of conservation impacts on well-being spatially and temporally within the wider landscape of social change. Pathways through the smog of confusion around the term well-being exist, and existing frameworks such as the Well-being in Developing Countries approach can help conservationists negotiate the challenges of operationalizing the concept. Conservationists have the opportunity to benefit from the recent flurry of research in the development field so as to carry out more nuanced and locally relevant evaluations of the effects of their interventions on human well-being.

"Do it yourself" reference materials for δ13C determinations by isotope ratio mass spectrometry.

A method is described to prepare inexpensive organic reference materials for routine stable isotope work. The method blends isotope-enriched and isotope-depleted materials to match closely international measurement standards. Examples are presented showing how (13)C-enriched and (13)C-depleted glucose were blended to match the isotopic compositions of the international carbon isotope standards, NBS 19 and L-SVEC. Preparation of the blended standards was straightforward, and the prepared solutions proved easy to dispense into tin capsules and easy to combust in a conventional isotope analysis system. The blended materials allow a laboratory to perform a two-point calibration for isotopic composition on a daily basis, with the use of an international reference material recommended for routine quality assurance. The blending method can be applied widely to make isotope standards for other elements and compounds.

Ensuring the reliability of stable isotope ratio data--beyond the principle of identical treatment.

The need for inter-laboratory comparability is crucial to facilitate the globalisation of scientific networks and the development of international databases to support scientific and criminal investigations. This article considers what lessons can be learned from a series of inter-laboratory comparison exercises organised by the Forensic Isotope Ratio Mass Spectrometry (FIRMS) network in terms of reference materials (RMs), the management of data quality, and technical limitations. The results showed that within-laboratory precision (repeatability) was generally good but between-laboratory accuracy (reproducibility) called for improvements. This review considers how stable isotope laboratories can establish a system of quality control (QC) and quality assurance (QA), emphasising issues of repeatability and reproducibility. For results to be comparable between laboratories, measurements must be traceable to the international δ-scales and, because isotope ratio measurements are reported relative to standards, a key aspect is the correct selection, calibration, and use of international and in-house RMs. The authors identify four principles which promote good laboratory practice. The principle of identical treatment by which samples and RMs are processed in an identical manner and which incorporates three further principles; the principle of identical correction (by which necessary corrections are identified and evenly applied), the principle of identical scaling (by which data are shifted and stretched to the international δ-scales), and the principle of error detection by which QC and QA results are monitored and acted upon. To achieve both good repeatability and good reproducibility it is essential to obtain RMs with internationally agreed δ-values. These RMs will act as the basis for QC and can be used to calibrate further in-house QC RMs tailored to the activities of specific laboratories. In-house QA standards must also be developed to ensure that QC-based calibrations and corrections lead to accurate results for samples. The δ-values assigned to RMs must be recorded and reported with all data. Reference materials must be used to determine what corrections are necessary for measured data. Each analytical sequence of samples must include both QC and QA materials which are subject to identical treatment during measurement and data processing. Results for these materials must be plotted, monitored, and acted upon. Periodically international RMs should be analysed as an in-house proficiency test to demonstrate results are accurate.

Venom on ice: first insights into Antarctic octopus venoms.

The venom of Antarctic octopus remains completely unstudied. Here, a preliminary investigation was conducted into the properties of posterior salivary gland (PSG) extracts from four Antarctica eledonine (Incirrata; Octopodidae) species (Adelieledone polymorpha, Megaleledone setebos, Pareledone aequipapillae, and Pareledone turqueti) collected from the coast off George V's Land, Antarctica. Specimens were assayed for alkaline phosphatase (ALP), acetylcholinesterase (AChE), proteolytic, phospholipase A(2) (PLA(2)), and haemolytic activities. For comparison, stomach tissue from Cirroctopus sp. (Cirrata; Cirroctopodidae) was also assayed for ALP, AChE, proteolytic and haemolytic activities. Dietary and morphological data were collected from the literature to explore the ecological importance of venom, taking an adaptive evolutionary approach. Of the incirrate species, three showed activities in all assays, while P. turqueti did not exhibit any haemolytic activity. There was evidence for cold-adaptation of ALP in all incirrates, while proteolytic activity in all except P. turqueti. Cirroctopus sp. stomach tissue extract showed ALP, AChE and some proteolytic activity. It was concluded that the AChE activity seen in the PSG extracts was possibly due to a release of household proteins, and not one of the secreted salivary toxins. Although venom undoubtedly plays an important part in prey capture and processing by Antarctica eledonines, no obvious adaptations to differences in diet or morphology were apparent from the enzymatic and haemolytic assays. However, several morphological features including enlarged PSG, small buccal mass, and small beak suggest such adaptations are present. Future studies should be conducted on several levels: Venomic, providing more detailed information on the venom compositions as well as the venom components themselves; ecological, for example application of serological or genetic methods in identifying stomach contents; and behavioural, including observations on capture of different types of prey.

Population genetic study of the brain-derived neurotrophic factor (BDNF) gene.

Genetic variants in the brain-derived neurotrophic factor (BDNF) gene, predominantly the functional Val66Met polymorphism, have been associated with risk of bipolar disorder and other psychiatric disorders. However, not all studies support these findings, and overall the evidence for the association of BDNF with disease risk is weak. As differences in population genetic structure between patient samples could cause discrepant or spurious association results, we investigated this possibility by carrying out population genetic analyses of the BDNF genomic region. Substantial variation was detected in BDNF coding region single-nucleotide polymorphism (SNP) allele and haplotype frequencies between 58 global populations, with the derived Met allele of Val66Met ranging in frequency from 0 to 72% across populations. F(ST) analyses to assess diversity in the HapMap populations determined that the Val66Met F(ST) value was at the 99.8th percentile among all SNPs in the genome. As the BDNF population genetic differences may be due to local selection, we performed the long-range haplotype test for selection using 68 SNPs spanning the BDNF genomic region in 12 European-derived pedigrees. Evidence for positive selection was found for a high-frequency Val-carrying haplotype, with a relative extended haplotype homozygosity value above the 99 th percentile compared with HapMap data (P=4.6 x 10(-4)). In conclusion, we observed considerable BDNF allele and haplotype diversity among global populations and evidence for positive selection at the BDNF locus. These phenomena can have a profound impact on the detection of disease susceptibility genes and must be considered in gene association studies of BDNF.

Tentacles of venom: toxic protein convergence in the Kingdom Animalia.

The origin and evolution of venom in many animal orders remain controversial or almost entirely uninvestigated. Here we use cDNA studies of cephalopod posterior and anterior glands to reveal a single early origin of the associated secreted proteins. Protein types recovered were CAP (CRISP, Antigen 5 [Ag5] and Pathogenesis-related [PR-1]), chitinase, peptidase S1, PLA(2) (phospholipase A(2)), and six novel peptide types. CAP, chitinase, and PLA(2) were each recovered from a single species (Hapalochlaena maculosa, Octopus kaurna, and Sepia latimanus, respectively), while peptidase S1 transcripts were found in large numbers in all three posterior gland libraries. In addition, peptidase S1 transcripts were recovered from the anterior gland of H. maculata. We compare their molecular evolution to that of related proteins found in invertebrate and vertebrate venoms, revealing striking similarities in the types of proteins selected for toxic mutation and thus shedding light on what makes a protein amenable for use as a toxin.

The molecular basis of cross-reactivity in the Australian Snake Venom Detection Kit (SVDK).

The Snake Venom Detection Kit (SVDK) is of major medical importance in Australia, yet it has never been rigorously characterised in terms of its sensitivity and specificity, especially when it comes to reports of false-negative and false-positive results. This study investigates reactions and cross-reactions of five venoms the SVDK is directed against and a number of purified toxins. Snakes showing the closest evolutionary relationships demonstrated the lowest level of cross-reactivity between groups. This was, instead, far more evident between snakes that are extraordinarily evolutionary separated. These snakes: Pseudechis australis, Acanthophis antarcticus and Notechis scutatus, in fact displayed more false-positive results. Examination of individual toxin groups showed that phospholipase A(2)s (PLA(2)s) tends to react strongly and display considerable cross-reactivity across groups while the three-finger toxins (3FTx) reacted poorly in all but the Acanthophis well. The hook effect was evident for all venoms, particularly Oxyuranus scutellatus. The results of this study show considerable variation in toxin detection, with implications in further development of venom detection, both in Australia and other countries.

Positive natural selection in the human lineage.

Positive natural selection is the force that drives the increase in prevalence of advantageous traits, and it has played a central role in our development as a species. Until recently, the study of natural selection in humans has largely been restricted to comparing individual candidate genes to theoretical expectations. The advent of genome-wide sequence and polymorphism data brings fundamental new tools to the study of natural selection. It is now possible to identify new candidates for selection and to reevaluate previous claims by comparison with empirical distributions of DNA sequence variation across the human genome and among populations. The flood of data and analytical methods, however, raises many new challenges. Here, we review approaches to detect positive natural selection, describe results from recent analyses of genome-wide data, and discuss the prospects and challenges ahead as we expand our understanding of the role of natural selection in shaping the human genome.

Interprofessional postgraduate education in primary health care: is it making a difference?

This paper explores attitudes to, and perceptions of, the impact of interprofessional postgraduate education for primary health care professionals, based on a postal survey of 153 primary health care professionals undertaking postgraduate qualifications in New Zealand. The response rate was 75% (114/153 responses); comprising 79 doctors, 28 nurses, 7 other health professionals. As a result of their postgraduate education, 92% (104/113) reported improvement in their own practice; 68% (72/106) reported a positive influence on their workplace practice. Forty-eight percent (53/111) increased their understanding of their own professional role; 79% (77/98) increased their understanding of another professional groups' skills and competencies. Twenty-two percent (25/114) perceived increased career opportunities within a year; 56% (64/114) in the longer term. Only 12% (14/114) perceived future increases in income as a result of their study. Interprofessional postgraduate qualification study for primary health care professionals in New Zealand resulted in personal and professional benefit for individuals and their clinical practice, and increased understanding about their own and other health professionals' roles. The interprofessional nature of the education was seen as positive, contributing to a modest increase in collaboration between health professional groups. Barriers to furthering participation in interprofessional learning and increasing intersectorial collaboration in the workplace are identified and discussed.

Identification of Streptococcus spp. causing bovine mastitis by PCR-RFLP of 16S-23S ribosomal DNA.

Traditional microbiological methods for identification of Streptococcus spp. causing bovine mastitis have been demonstrated to be less than highly reliable. PCR-RFLP analysis of 16S-23S ribosomal DNA was used to characterise seven reference strains of streptococcal mastitis pathogens as well as four reference strains of other gram-positive, catalase-negative cocci of bovine origin to allow comparative identification of field isolates. RFLP analysis of PCR products, using a combination of two restriction endonucleases in single reactions (HaeIII and AluI, HaeIII and RsaI or AluI and RsaI) generated unique patterns for species of Streptococcus, Enterococcus and Lactococcus. One hundred field isolates of Streptococcus spp. collected from cows with clinical or subclinical mastitis were tested. Fifty-seven isolates, classified by conventional tests as S. uberis, were identified as 47 S. uberis and six S. parauberis by their unique RFLP patterns. The remaining four isolates had RFLP patterns distinct from the reference strains and two of these were identified as closely related to S. iniae and two to Aerococcus viridans by 16S rRNA sequencing. Conventional identification of 17 S. agalactiae and 26 S. dysgalactiae subsp. dysgalactiae was confirmed by RFLP. Use of a combination of restriction enzymes in a single tube enabled the rapid, accurate, cost effective and easily performed identification of all major streptococcal mastitis pathogens.

Presynaptic neuromuscular activity of venom from the brown-headed snake (Glyphodon tristis).

The brown-headed snake (Glyphodon tristis) inhabits the forest regions of Papua New Guinea, Torres Strait Islands, and far northern Queensland, Australia. Although bites by Glyphodon dunmalli have been reported, G. tristis was regarded as innocuous until 1989 when a healthy 20 year old man was bitten (Sutherland, S.K., Tibballs, J., 2001. Australian Animal Toxins, the Creatures, their Toxins and Care of the Poisoned Patient. University Press, Oxford). Treatment of envenomation by this species is empirical with no specific antivenom available. While no published studies on the venom of G. tristis are available, unpublished studies suggest neurotoxicity as being the main symptom of envenomation. In this study, the in vitro effects of G. tristis venom were examined using the chick biventer cervicis nerve muscle (CBCNM) preparation. Venom (10 microg/ml) inhibited indirect (0.2 ms, 0.1 Hz, supramaximal V) twitches of the CBCNM. This inhibition appeared to be presynaptic in origin as evidenced by the lack of effect of venom on responses to exogenous acetylcholine (1 mM), carbachol (20 microM) and KCl (40 mM) in the non-stimulated CBCNM. Prior addition (10 min) of polyvalent snake antivenom (5 U/ml; CSL Ltd) attenuated twitch inhibition. The venom (10-50 microg/ml) also appears to be myotoxic as indicated by a slowly developing contracture and inhibition of direct (2 ms, 0.1 Hz, supramaximal V, in the presence of tubocurarine 10 microM) twitches. Myotoxicity was confirmed by subsequent histological examination of tissues. This myotoxicity was prevented by the prior addition of polyvalent snake antivenom (30 U/ml). The phospholipase A inhibitor 4-BPB (1.8 mM) significantly attenuated the inhibition of indirect and direct twitches of the CBCNM preparation, indicating the involvement of a PLA2 component in the toxic action of the venom.

Eggs-only diet: its implications for the toxin profile changes and ecology of the marbled sea snake (Aipysurus eydouxii).

Studies so far have correlated the variation in the composition of snake venoms with the target prey population and snake's diet. Here we present the first example of an alternative evolutionary link between venom composition and dietary adaptation of snakes. We describe a dinucleotide deletion in the only three finger toxin gene expressed in the sea snake Aipysurus eydouxii (Marbled Sea Snake) venom and how it may have been the result of a significant change in dietary habits. The deletion leads to a frame shift and truncation with an accompanying loss of neurotoxicity. Due to the remarkable streamlining of sea snake venoms, a mutation of a single toxin can have dramatic effects on the whole venom, in this case likely explaining the 50- to 100-fold decrease in venom toxicity in comparison to that of other species in the same genus. This is a secondary result of the adaptation of A. eydouxii to a new dietary habit--feeding exclusively on fish eggs and, thus, the snake no longer using its venom for prey capture. This was parallel to greatly atrophied venom glands and loss of effective fangs. It is interesting to note that a potent venom was not maintained for use in defense, thus reinforcing that the primary use of snake venom is for prey capture.

Haploview: analysis and visualization of LD and haplotype maps.

UNLABELLED: Research over the last few years has revealed significant haplotype structure in the human genome. The characterization of these patterns, particularly in the context of medical genetic association studies, is becoming a routine research activity. Haploview is a software package that provides computation of linkage disequilibrium statistics and population haplotype patterns from primary genotype data in a visually appealing and interactive interface. AVAILABILITY: http://www.broad.mit.edu/mpg/haploview/ CONTACT: jcbarret@broad.mit.edu

The in vitro and in vivo pharmacological activity of Boiga dendrophila (mangrove catsnake) venom.

The great taxonomic and prey base diversity of colubrids (non-front-fanged snakes) suggests that their venoms may represent a 'literal gold mine' for scientists eager to find novel pharmacological probes. While pharmacological characterization is lacking for most of these venoms, this is even more so with regard to activity of colubrid venoms on the mammalian autonomic nervous system. This study characterizes the activity of venom from the colubrid, Boiga dendrophila using in vitro smooth muscle preparations and the anaesthetized rat. In the prostatic segment of the rat vas deferens, cumulative additions of venom (1-150 microg ml(-1)) induced concentration-dependent inhibition of electrically evoked (0.2 Hz, 0.3 ms, 70-100 V) twitches. The inhibitory effect of venom (100 microg ml(-1)) was attenuated by 8-phenyltheophylline (8-PT) (20 microM) and 8-cyclopentyl-1, 3-dipropylxanthine (20 microM) but not idazoxan (1 microM), or a combination of ranitidine (0.2 microM) and thioperamide (10 microM). The inhibitory effect of venom (100 microg ml(-1)) was augmented by dipyridamole (10 microM) but abolished by pretreatment with adenosine deaminase (7.5 units/100 microl) suggesting that it contains components with adenosine A(1) receptor activity, most likely adenosine. In isolated segments of guinea-pig ileum, venom (10-100 microg ml(-1)) caused concentration-dependent contractions which were inhibited by the muscarinic receptor antagonist atropine (0.1 microM) but not by the histamine receptor antagonist mepyramine (0.5 microM). In the anaesthetized rat, venom (5-7.5 mg kg(-1), i.v.) caused a hypotensive effect. Our data suggest that the venom contains components with purinergic and muscarinic receptor activity.

Assembling an arsenal: origin and evolution of the snake venom proteome inferred from phylogenetic analysis of toxin sequences.

We analyzed the origin and evolution of snake venom toxin families represented in both viperid and elapid snakes by means of phylogenetic analysis of the amino acid sequences of the toxins and related nonvenom proteins. Out of eight toxin families analyzed, five provided clear evidence of recruitment into the snake venom proteome before the diversification of the advanced snakes (Kunitz-type protease inhibitors, CRISP toxins, galactose-binding lectins, M12B peptidases, nerve growth factor toxins), and one was equivocal (cystatin toxins). In two others (phospholipase A(2) and natriuretic toxins), the nonmonophyly of venom toxins demonstrates that presence of these proteins in elapids and viperids results from independent recruitment events. The ANP/BNP natriuretic toxins are likely to be basal, whereas the CNP/BPP toxins are Viperidae only. Similarly, the lectins were recruited twice. In contrast to the basal recruitment of the galactose-binding lectins, the C-type lectins were shown to be Viperidae only, with the alpha-chains and beta-chains resulting from an early duplication event. These results provide strong additional evidence that venom evolved once, at the base of the advanced snake radiation, rather than multiple times in different lineages, with these toxins also present in the venoms of the "colubrid" snake families. Moreover, they provide a first insight into the composition of the earliest ophidian venoms and point the way toward a research program that could elucidate the functional context of the evolution of the snake venom proteome.

Using stable isotopes in mangrove fisheries research--a review and outlook.

Mangrove forests are important coastal wetlands in most of the tropics, but their importance to fisheries is poorly understood. Rationales for conserving these ecosystems are partly based on ideas that mangroves are important to food webs that support fisheries. Here we review use of stable isotopes to test the strength of the mangrove forest-fish connection, suggesting that a dual focus on both food web ecology and habitat use may be most profitable in future studies of this type.

Steady state models of stable isotopic distributions.

This study reviews common calculations and mathematical models used in stable isotopic studies. Some approximations are adopted to simplify the algebra for use in steady state models, with more rigorous mathematics outlined in an Appendix. Review shows that steady-state isotopic models are easy to construct, provide good approximations of system behavior, and are very helpful in evaluating isotopic cycling in many kinds of systems.

Molecular evolution and phylogeny of elapid snake venom three-finger toxins.

Animal venom components are of considerable interest to researchers across a wide variety of disciplines, including molecular biology, biochemistry, medicine, and evolutionary genetics. The three-finger family of snake venom peptides is a particularly interesting and biochemically complex group of venom peptides, because they are encoded by a large multigene family and display a diverse array of functional activities. In addition, understanding how this complex and highly varied multigene family evolved is an interesting question to researchers investigating the biochemical diversity of these peptides and their impact on human health. Therefore, the purpose of our study was to investigate the long-term evolutionary patterns exhibited by these snake venom toxins to understand the mechanisms by which they diversified into a large, biochemically diverse, multigene family. Our results show a much greater diversity of family members than was previously known, including a number of subfamilies that did not fall within any previously identified groups with characterized activities. In addition, we found that the long-term evolutionary processes that gave rise to the diversity of three-finger toxins are consistent with the birth-and-death model of multigene family evolution. It is anticipated that this "three-finger toxin toolkit" will prove to be useful in providing a clearer picture of the diversity of investigational ligands or potential therapeutics available within this important family.