Mapping co-benefits for carbon storage and biodiversity to inform conservation policy and action.

Integrated high-resolution maps of carbon stocks and biodiversity that identify areas of potential co-benefits for climate change mitigation and biodiversity conservation can help facilitate the implementation of global climate and biodiversity commitments at local levels. However, the multi-dimensional nature of biodiversity presents a major challenge for understanding, mapping and communicating where and how biodiversity benefits coincide with climate benefits. A new integrated approach to biodiversity is therefore needed. Here, we (a) present a new high-resolution map of global above- and below-ground carbon stored in biomass and soil, (b) quantify biodiversity values using two complementary indices (BIp and BIr) representing proactive and reactive approaches to conservation, and (c) examine patterns of carbon-biodiversity overlap by identifying 'hotspots' (20% highest values for both aspects). Our indices integrate local diversity and ecosystem intactness, as well as regional ecosystem intactness across the broader area supporting a similar natural assemblage of species to the location of interest. The western Amazon Basin, Central Africa and Southeast Asia capture the last strongholds of highest local biodiversity and ecosystem intactness worldwide, while the last refuges for unique biological communities whose habitats have been greatly reduced are mostly found in the tropical Andes and central Sundaland. There is 38 and 5% overlap in carbon and biodiversity hotspots, for proactive and reactive conservation, respectively. Alarmingly, only around 12 and 21% of these proactive and reactive hotspot areas, respectively, are formally protected. This highlights that a coupled approach is urgently needed to help achieve both climate and biodiversity global targets. This would involve (1) restoring and conserving unprotected, degraded ecosystems, particularly in the Neotropics and Indomalaya, and (2) retaining the remaining strongholds of intactness. This article is part of the theme issue 'Climate change and ecosystems: threats, opportunities and solutions'.

The impact of hunting on tropical mammal and bird populations.

Hunting is a major driver of biodiversity loss, but a systematic large-scale estimate of hunting-induced defaunation is lacking. We synthesized 176 studies to quantify hunting-induced declines of mammal and bird populations across the tropics. Bird and mammal abundances declined by 58% (25 to 76%) and by 83% (72 to 90%) in hunted compared with unhunted areas. Bird and mammal populations were depleted within 7 and 40 kilometers from hunters' access points (roads and settlements). Additionally, hunting pressure was higher in areas with better accessibility to major towns where wild meat could be traded. Mammal population densities were lower outside protected areas, particularly because of commercial hunting. Strategies to sustainably manage wild meat hunting in both protected and unprotected tropical ecosystems are urgently needed to avoid further defaunation.

Circumpolar contaminant concentrations in polar bears (Ursus maritimus) and potential population-level effects.

Polar bears (Ursus maritimus) currently receive much attention in the context of global climate change. However, there are other stressors that might threaten the viability of polar bear populations as well, such as exposure to anthropogenic pollutants. Lipophilic organic compounds bio-accumulate and bio-magnify in the food chain, leading to high concentrations at the level of top-predators. In Arctic wildlife, including the polar bear, various adverse health effects have been related to internal concentrations of commercially used anthropogenic chemicals like PCB and DDT. The extent to which these individual health effects are associated to population-level effects is, however, unknown. In this study we assembled data on adipose tissue concentrations of ∑PCB, ∑DDT, dieldrin and ∑PBDE in individual polar bears from peer-reviewed scientific literature. Data were available for 14 out of the 19 subpopulations. We found that internal concentrations of these contaminants exceed threshold values for adverse individual health effects in several subpopulations. In an exploratory regression analysis we identified a clear negative correlation between polar bear population density and sub-population specific contaminant concentrations in adipose tissue. The results suggest that adverse health effects of contaminants in individual polar bears may scale up to population-level consequences. Our study highlights the need to consider contaminant exposure along with other threats in polar bear population viability analyses.

Metabolomics meets functional assays: coupling LC-MS and microfluidic cell-based receptor-ligand analyses.

INTRODUCTION: Metabolomics has become a valuable tool in many research areas. However, generating metabolomics-based biochemical profiles without any related bioactivity is only of indirect value in understanding a biological process. Therefore, metabolomics research could greatly benefit from tools that directly determine the bioactivity of the detected compounds. OBJECTIVE: We aimed to combine LC-MS metabolomics with a cell based receptor assay. This combination could increase the understanding of biological processes and may provide novel opportunities for functional metabolomics. METHODS: We developed a flow through biosensor with human cells expressing both the TRPV1, a calcium ion channel which responds to capsaicin, and the fluorescent intracellular calcium ion reporter, YC3.6. We have analysed three contrasting Capsicum varieties. Two were selected with contrasting degrees of spiciness for characterization by HPLC coupled to high mass resolution MS. Subsequently, the biosensor was then used to link individual pepper compounds with TRPV1 activity. RESULTS: Among the compounds in the crude pepper fruit extracts, we confirmed capsaicin and also identified both nordihydrocapsaicin and dihydrocapsaicin as true agonists of the TRPV1 receptor. Furthermore, the biosensor was able to detect receptor activity in extracts of both Capsicum fruits as well as a commercial product. Sensitivity of the biosensor to this commercial product was similar to the sensory threshold of a human sensory panel. CONCLUSION: Our results demonstrate that the TRPV1 biosensor is suitable for detecting bioactive metabolites. Novel opportunities may lie in the development of a continuous functional assay, where the biosensor is directly coupled to the LC-MS.

An allometric approach to quantify the extinction vulnerability of birds and mammals.

Methods to quantify the vulnerability of species to extinction are typically limited by the availability of species-specific input data pertaining to life-history characteristics and population dynamics. This lack of data hampers global biodiversity assessments and conservation planning. Here, we developed a new framework that systematically quantifies extinction risk based on allometric relationships between various wildlife demographic parameters and body size. These allometric relationships have a solid theoretical and ecological foundation. Extinction risk indicators included are (1) the probability of extinction, (2) the mean time to extinction, and (3) the critical patch size. We applied our framework to assess the global extinction vulnerability of terrestrial carnivorous and non-carnivorous birds and mammals. Irrespective of the indicator used, large-bodied species were found to be more vulnerable to extinction than their smaller counterparts. The patterns with body size were confirmed for all species groups by a comparison with IUCN data on the proportion of extant threatened species: the models correctly predicted a multimodal distribution with body size for carnivorous birds and a monotonic distribution for mammals and non-carnivorous birds. Carnivorous mammals were found to have higher extinction risks than non-carnivores, while birds were more prone to extinction than mammals. These results are explained by the allometric relationships, predicting the vulnerable species groups to have lower intrinsic population growth rates, smaller population sizes, lower carrying capacities, or larger dispersal distances, which, in turn, increase the importance of losses due to environmental stochastic effects and dispersal activities. Our study is the first to integrate population viability analysis and allometry into a novel, process-based framework that is able to quantify extinction risk of a large number of species without requiring data-intensive, species-specific information. The framework facilitates the estimation of extinction vulnerabilities of data-deficient species. It may be applied to forecast extinction vulnerability in response to a changing environment, by incorporating quantitative relationships between wildlife demographic parameters and environmental drivers like habitat alteration, climate change, or hunting.

Investigation of detection limits for diffuse optical tomography systems: I. Theory and experiment.

We present a statistical test using simulated photon migration data and a noise model derived from the hardware of a particular diffuse optical tomography system to predict its detection limits. Our method allows us to assess the spatial distribution of the detection sensitivity of arbitrary geometries and noise without requiring phantom measurements and reconstructions. We determine the minimal detectable lesion size at selected lesion positions and compare the predicted results with phantom measurements carried out in a cup geometry.

Effects of cysteine protease inhibitors on oviposition rate of the western flower thrips, Frankliniella occidentalis.

Proteolytic activity in whole insect extracts of the western flower thrips, Frankliniella occidentalis, was found to belong predominantly to the class of cysteine proteases. The pH optimum of the general proteolytic activity was determined to be 3.5, which is low when compared to other insects using cysteine proteases for protein digestion. The proteinaceous cysteine protease inhibitors chicken cystatin, potato cystatin and sea anemone equistatin inhibited in vitro more than 90% of the protease activity. To test in vivo the biological effect of such inhibitors on the oviposition rate of western flower thrips, recombinant potato cystatin and equistatin were fed to adult females. A gradual reduction in oviposition rate to about 45% of control was observed when reared on these PIs for a period of 5 days, with no increase in mortality. These results are discussed in the light of the application of protease inhibitors in transgenic plants to control this insect pest.

Characterization of recombinant mustard trypsin inhibitor 2 (MTI2) expressed in Pichia pastoris.

The mustard trypsin inhibitor MTI2 was expressed as secretory protein in the yeast Pichia pastoris. In order to evaluate the influence of the C-terminal amino acids of the precursor form on the inhibitor activity, the C-terminal precursor and the mature protein were both expressed. A third His-tagged construct was also designed to compare alternative purification procedures. Proteins were efficiently expressed at levels of 40-160 mg/l in shake flasks. Equilibrium dissociation constants demonstrated that the mature protein was a stronger inhibitor of bovine beta-trypsin compared to the precursor and His-tagged forms (0.01 nM vs. 0.58 nM and 0.71 nM, respectively). The recombinant proteins were active inhibitors of Spodoptera exigua gut proteases.

Identification of ErbB3-stimulated genes using modified representational difference analysis.

The epidermal growth factor receptor (EGFR) family of tyrosine kinases is involved in the growth of normal and tumour cells. The specific contribution of each of the four family members to these processes remains unclear. In the present study we have used a PCR-based subtractive approach to identify differences in messages induced in response to activation of ErbB3 and EGFR. The approach described is a modification of the representational difference analysis technique adapted for analysis of cDNA, which we have modified to permit identification of differential gene expression using as little as 20 microg of total RNA as the starting material. The mRNA obtained from EGF-stimulated NIH-3T3 cells expressing chimaeric EGFR-ErbB3 receptors provided the tester amplicons (small PCR-amplified fragments) which were subtracted against driver amplicons derived from unstimulated NIH-3T3 cells expressing the EGFR-ErbB3 chimaera or EGF-stimulated NIH-3T3 cells overexpressing the EGFR. A total of 22 different clones were isolated, 90% of which showed increased expression in the tester amplicons. Six of these, corresponding to known DNA sequences, were selected for further Northern blot analysis against total RNA prepared from the starting cell lines. Of these, the gene encoding the protein dlk (or a closely related protein, Pref-1) was identified as being regulated by ErbB3 but not by the EGFR. Other genes appeared to be elevated by both ErbB3 and EGFR, including those encoding c-jun, Ret finger protein (RFP), neuroleukin and amyloid protein precursor. One gene product, TIS11, was identified as being regulated by EGFR but not by ErbB3.

Eye disease in newly diagnosed leprosy patients in eastern Nepal.

To determine the magnitude of eye lesions in newly diagnosed leprosy patients we examined their eyes. The Eastern Leprosy Control Project was supported by The Netherlands Leprosy Relief Association; we used the regional clinic in Biratnagar and 5 mobile clinics in surrounding districts as our survey area. All patients who presented at the clinics over 10 weeks, diagnosed as having untreated leprosy were included. Of the 260 examined patients 97 (37.3%, 95% confidence interval 28.3-40.3%) had an eye lesion; 12/260 patients (4.6%, 95% confidence interval 2.0-7.2%) had sight-threatening lesions (lagophthalmos, iris involvement, corneal anaesthesia), directly related to leprosy; 46 (17.7%) patients were diagnosed as having some degree of cataract; 2 patients were aphakic; 3 patients (1.2%) were blind according to the WHO definition. In this series of new and untreated leprosy patients many eye lesions found are not relevant or leprosy related. There were 9 new patients with lagophthalmos, some too longstanding to treat with steroids. We found 3 patients with iris involvement. The figures we found for eye lesions, sight-threatening lesions and blindness are low when compared to other studies. The number of patients with any grade of cataract is high. The average total of leprosy patients who were blind can be compared with the average total who are blind in the general population.

Disabilities of hands, feet and eyes in newly diagnosed leprosy patients in eastern Nepal.

The objective of the study was to determine the magnitude of hand/feet/eye disabilities in newly diagnosed leprosy patients by examining all newly diagnosed leprosy patients who presented at the Eastern Leprosy Control Project (supported by The Netherlands Leprosy Relief Association), made up of a regional clinic in Biratnagar and 5 mobile clinics in surrounding districts. The study comprised of all new and previously untreated patients who presented at the clinics over a 10-week period who were diagnosed as leprosy sufferers. Of the 260 leprosy patients examined 12 (4.6%) had sight-threatening lesions (lagophthalmos, iris involvement, corneal anaesthesia); 3 patients were blind due to cataract; 96/260 patients (37.0%, 95% confidence interval 35.0-43.0%) had 1 or more disabilities of their hands and/or feet. The most frequently found disabilities were sensory loss of the hands and feet, claw hand and plantar ulcers. According to the WHO disability grading 60% had no disabilities, 19% had grade 1 and 21% had grade 2 disability. Disability assessment is very important not only to evaluate the effectiveness of the control programme but also for the patient, whose most important worry is the stigmatizing deformities leprosy patients suffer. The earlier detection of sensory loss might reduce these secondary deformities.

Paralysis of facial muscles in leprosy patients with lagophthalmos.

The objective of the study was to determine the pattern of involvement of facial muscles in lagophthalmos. Fifty-seven patients with lagophthalmos were examined to assess the degree of paralysis of facial muscles. Eighty-one percent of the patients with lagophthalmos had involvement of at least one other muscle group. In patients with lagophthalmos with a gap at mild closure of 5 mm or more, 27 of 30 (90%) had involvement of at least one other facial muscle. In lepromatous leprosy the pattern of involvement was symmetrical and "patchy," the right and left sides being affected equally. In tuberculoid leprosy, the ipsilateral muscles were more often involved, which is the pattern of involvement of a nerve trunk. The upper and lower facial muscles were affected in the same proportion. Hence, on clinical grounds, there is little support for the often postulated statement that the superficial course of the facial nerve above the zygomatic bone is decisive for exclusive paralysis of the zygomatic branch of the facial nerve.

A major substrate of maturation promoting factor identified as elongation factor 1 beta gamma delta in Xenopus laevis.

Protein synthesis is believed to be under control of the cell cycle during meiosis and mitosis. Any relationship between substrates for cdc2 kinase and components of the protein synthetic apparatus would therefore be of prime importance. During meiosis of Xenopus laevis oocytes one of the substrates for this kinase is a p47 protein, which is complexed to two other proteins, P36 and P30. Judged from partial amino acid sequence data on P47 and P30, the P30 and P47 proteins were reported to resemble the protein synthetic elongation factors (EF) 1 beta and 1 gamma from Artemia salina (Bellé, R., Derancourt, J., Poulhe, R., Capony, J.P., Ozon, R., and Mulner-Lorillon, O. (1989) FEBS Lett. 255, 101-104). This paper shows that the complex composed of P30, P47, and P36 from Xenopus is identical to the complex of EF-1 beta, EF-1 gamma, and EF-1 delta from Artemia according to two criteria. 1) Both stimulate elongation factor 1 alpha-mediated transfer RNA binding to ribosomes and exchange of guanine nucleotides on elongation factor 1 alpha to a comparable degree. 2) Each of the three subunits of the protein complex P30.P47.P36 from Xenopus shows a structural homology with one of the corresponding subunits of EF-1 beta gamma delta from Artemia. Presumably the phosphorylation of EF-1 gamma, which associates with tubulin at least in vitro, is important in processes following the onset of meiosis which is accompanied by a rise of protein synthesis.

Elongation factor EF-1 alpha from Artemia is specifically labeled at Lys-63 by the guanine nucleotide analog pyridoxal-5'-diphospho-5'-guanosine.

We have synthesized the guanine nucleotide analog pyridoxal-5'-diphospho-5'-guanosine. This compound specifically modifies a single lysine residue in elongation factor 1 alpha from Artemia, indicating that this residue is in close contact with the reactive part of the guanine nucleotide analog. This result is discussed in terms of the structure of the nucleotide-binding domain of the factor.

A conserved amino acid sequence around Arg-68 of Artemia elongation factor 1 alpha is involved in the binding of guanine nucleotides and aminoacyl transfer RNAs.

The rate of trypsin cleavage of elongation factor 1 alpha having bound GDP is low and increases on exchange of GDP for GTP. The cleavage occurs at a unique position of the protein chain, namely at arginine-68 of Artemia EF-1 alpha. This increase in trypsin sensitivity is enhanced further in the presence of charged or uncharged transfer RNA. The local unfolding of EF-alpha at residue 68 is discussed in terms of a model in which GTP hydrolysis controls the positioning of a short 3'-terminal section of transfer RNA near the centre of peptide bond synthesis.

Differences in specificity and catalytic efficiency between allozymes of esterase-4 from Drosophila mojavensis.

A more than 10-fold difference in the specificity and catalytic efficiency for 1-naphthyl esters was measured between two allozymes of esterase-4 from Drosophila mojavensis. This difference is mainly caused by a difference in the affinity for the 1-naphthyl esters. The amino acid compositions of the allozymes are not significantly different, which means that the difference in primary structure is small. Small differences in primary structure generally do not result in such a large increase in catalytic efficiency and such a large shift in substrate specificity as was found in the present study.