Discovery of allosteric regulators with clinical potential to stabilize alpha-L-iduronidase in mucopolysaccharidosis type I.

Mucopolysaccharidosis type I (MPS I) is an inherited lysosomal disease caused by lowered activity of the enzyme alpha-L-iduronidase (IDUA). Current therapeutic options show limited efficacy and do not treat some important aspects of the disease. Therefore, it may be advantageous to identify strategies that could improve the efficacy of existing treatments. Pharmacological chaperones are small molecules that protect proteins from degradation, and their use in combination with enzyme replacement therapy (ERT) has been proposed as an alternative therapeutic strategy. Using the SEE-Tx® proprietary computational drug discovery platform, a new allosteric ligand binding cavity in IDUA was identified distal from the active site. Virtual high-throughput screening of approximately 5 million compounds using the SEE-Tx® docking platform identified a subset of small molecules that bound to the druggable cavity and functioned as novel allosteric chaperones of IDUA. Experimental validation by differential scanning fluorimetry showed an overall hit rate of 11.4%. Biophysical studies showed that one exemplary hit molecule GT-01803 bound to (Kd = 22 µM) and stabilized recombinant human IDUA (rhIDUA) in a dose-dependent manner. Co-administration of rhIDUA and GT-01803 increased IDUA activity in patient-derived fibroblasts. Preliminary in vivo studies have shown that GT-01803 improved the pharmacokinetic (PK) profile of rhIDUA, increasing plasma levels in a dose-dependent manner. Furthermore, GT-01803 also increased IDUA enzymatic activity in bone marrow tissue, which benefits least from standard ERT. Oral bioavailability of GT-01803 was found to be good (50%). Overall, the discovery and validation of a novel allosteric chaperone for rhIDUA presents a promising strategy to enhance the efficacy of existing treatments for MPS I. The compound's ability to increase rhIDUA activity in patient-derived fibroblasts and its good oral bioavailability underscore its potential as a potent adjunct to ERT, particularly for addressing aspects of the disease less responsive to standard treatment.

Cost Analysis of Magnetic Resonance Imaging and Computed Tomography in Cardiology: A Case Study of a University Hospital Complex in the Euro Region.

INTRODUCTION: In recent years, several hospitals have incorporated MRI equipment managed directly by their cardiology departments. The aim of our work is to determine the total cost per test of both CT and MRI in the setting of a Cardiology Department of a tertiary hospital. MATERIALS AND METHODS: The process followed for estimating the costs of CT and MRI tests consists of three phases: (1) Identification of the phases of the testing process; (2) Identification of the resources consumed in carrying out the tests; (3) Quantification and assessment of inputs. RESULTS: MRI involves higher personnel (EUR 66.03 vs. EUR 49.17) and equipment (EUR 89.98 vs. EUR 33.73) costs, while CT consumes higher expenditures in consumables (EUR 93.28 vs. EUR 22.95) and overheads (EUR 1.64 vs. EUR 1.55). The total cost of performing each test is higher in MRI (EUR 180.60 vs. EUR 177.73). CONCLUSIONS: We can conclude that the unit cost of each CT and MRI performed in that unit are EUR 177.73 and EUR 180.60, respectively, attributable to consumables in the case of CT and to amortization of equipment and staff time in the case of MRI.

Beta-Amyloid (Aß1-42) Increases the Expression of NKCC1 in the Mouse Hippocampus.

Alzheimer's disease (AD) is a neurodegenerative disorder with an increasing need for developing disease-modifying treatments as current therapies only provide marginal symptomatic relief. Recent evidence suggests the γ-aminobutyric acid (GABA) neurotransmitter system undergoes remodeling in AD, disrupting the excitatory/inhibitory (E/I) balance in the brain. Altered expression levels of K-Cl-2 (KCC2) and N-K-Cl-1 (NKCC1), which are cation-chloride cotransporters (CCCs), have been implicated in disrupting GABAergic activity by regulating GABAA receptor signaling polarity in several neurological disorders, but these have not yet been explored in AD. NKCC1 and KCC2 regulate intracellular chloride [Cl-]i by accumulating and extruding Cl-, respectively. Increased NKCC1 expression in mature neurons has been reported in these disease conditions, and bumetanide, an NKCC1 inhibitor, is suggested to show potential therapeutic benefits. This study used primary mouse hippocampal neurons to explore if KCC2 and NKCC1 expression levels are altered following beta-amyloid (Aß1-42) treatment and the potential neuroprotective effects of bumetanide. KCC2 and NKCC1 expression levels were also examined in 18-months-old male C57BL/6 mice following bilateral hippocampal Aß1-42 stereotaxic injection. No change in KCC2 and NKCC1 expression levels were observed in mouse hippocampal neurons treated with 1 nM Aß1-42, but NKCC1 expression increased 30-days post-Aß1-42-injection in the CA1 region of the mouse hippocampus. Primary mouse hippocampal cultures were treated with 1 nM Aß1-42 alone or with various concentrations of bumetanide (1 µM, 10 µM, 100 µM, 1 mM) to investigate the effect of the drug on cell viability. Aß1-42 produced 53.1 ± 1.4% cell death after 5 days, and the addition of bumetanide did not reduce this. However, the drug at all concentrations significantly reduced cell viability, suggesting bumetanide is highly neurotoxic. In summary, these results suggest that chronic exposure to Aß1-42 alters the balance of KCC2 and NKCC1 expression in a region-and layer-specific manner in mouse hippocampal tissue; therefore, this process most likely contributes to altered hippocampal E/I balance in this model. Furthermore, bumetanide induces hippocampal neurotoxicity, thus questioning its suitability for AD therapy. Further investigations are required to examine the effects of Aß1-42 on KCC2 and NKCC1 expression and whether targeting CCCs might offer a therapeutic approach for AD.

The Interplay Between Beta-Amyloid 1-42 (Aß1-42)-Induced Hippocampal Inflammatory Response, p-tau, Vascular Pathology, and Their Synergistic Contributions to Neuronal Death and Behavioral Deficits.

Alzheimer's disease (AD), the most common chronic neurodegenerative disorder, has complex neuropathology. The principal neuropathological hallmarks of the disease are the deposition of extracellular ß-amyloid (Aß) plaques and neurofibrillary tangles (NFTs) comprised of hyperphosphorylated tau (p-tau) protein. These changes occur with neuroinflammation, a compromised blood-brain barrier (BBB) integrity, and neuronal synaptic dysfunction, all of which ultimately lead to neuronal cell loss and cognitive deficits in AD. Aß1-42 was stereotaxically administered bilaterally into the CA1 region of the hippocampi of 18-month-old male C57BL/6 mice. This study aimed to characterize, utilizing immunohistochemistry and behavioral testing, the spatial and temporal effects of Aß1-42 on a broad set of parameters characteristic of AD: p-tau, neuroinflammation, vascular pathology, pyramidal cell survival, and behavior. Three days after Aß1-42 injection and before significant neuronal cell loss was detected, acute neuroinflammatory and vascular responses were observed. These responses included the up-regulation of glial fibrillary acidic protein (GFAP), cell adhesion molecule-1 (PECAM-1, also known as CD31), fibrinogen labeling, and an increased number of activated astrocytes and microglia in the CA1 region of the hippocampus. From day 7, there was significant pyramidal cell loss in the CA1 region of the hippocampus, and by 30 days, significant localized up-regulation of p-tau, GFAP, Iba-1, CD31, and alpha-smooth muscle actin (α-SMA) in the Aß1-42-injected mice compared with controls. These molecular changes in Aß1-42-injected mice were accompanied by cognitive deterioration, as demonstrated by long-term spatial memory impairment. This study is reporting a comprehensive examination of a complex set of parameters associated with intrahippocampal administration of Aß1-42 in mice, their spatiotemporal interactions and combined contribution to the disease progression. We show that a single Aß injection can reproduce aspects of the inflammatory, vascular, and p-tau induced pathology occurring in the AD human brain that lead to cognitive deficits.

Amyloid-beta1-42 induced glutamatergic receptor and transporter expression changes in the mouse hippocampus.

Alzheimer's disease (AD) is the leading type of dementia worldwide. With an increasing burden of an aging population coupled with the lack of any foreseeable cure, AD warrants the current intense research effort on the toxic effects of an increased concentration of beta-amyloid (Aß) in the brain. Glutamate is the main excitatory brain neurotransmitter and it plays an essential role in the function and health of neurons and neuronal excitability. While previous studies have shown alterations in expression of glutamatergic signaling components in AD, the underlying mechanisms of these changes are not well understood. This is the first comprehensive anatomical study to characterize the subregion- and cell layer-specific long-term effect of Aß1-42 on the expression of specific glutamate receptors and transporters in the mouse hippocampus, using immunohistochemistry with confocal microscopy. Outcomes are examined 30 days after Aß1-42 stereotactic injection in aged male C57BL/6 mice. We report significant decreases in density of the glutamate receptor subunit GluA1 and the vesicular glutamate transporter (VGluT) 1 in the conus ammonis 1 region of the hippocampus in the Aß1-42 injected mice compared with artificial cerebrospinal fluid injected and naïve controls, notably in the stratum oriens and stratum radiatum. GluA1 subunit density also decreased within the dentate gyrus dorsal stratum moleculare in Aß1-42 injected mice compared with artificial cerebrospinal fluid injected controls. These changes are consistent with findings previously reported in the human AD hippocampus. By contrast, glutamate receptor subunits GluA2, GluN1, GluN2A, and VGluT2 showed no changes in expression. These findings indicate that Aß1-42 induces brain region and layer specific expression changes of the glutamatergic receptors and transporters, suggesting complex and spatial vulnerability of this pathway during development of AD neuropathology. Read the Editorial Highlight for this article on page 7. Cover Image for this issue: https://doi.org/10.1111/jnc.14763.

Amyloid-Beta1-42 -Induced Increase in GABAergic Tonic Conductance in Mouse Hippocampal CA1 Pyramidal Cells.

Alzheimer's disease (AD) is a complex and chronic neurodegenerative disorder that involves a progressive and severe decline in cognition and memory. During the last few decades a considerable amount of research has been done in order to better understand tau-pathology, inflammatory activity and neuronal synapse loss in AD, all of them contributing to cognitive decline. Early hippocampal network dysfunction is one of the main factors associated with cognitive decline in AD. Much has been published about amyloid-beta1-42 (Aß1-42)-mediated excitotoxicity in AD. However, increasing evidence demonstrates that the remodeling of the inhibitory gamma-aminobutyric acid (GABAergic) system contributes to the excitatory/inhibitory (E/I) disruption in the AD hippocampus, but the underlying mechanisms are not well understood. In the present study, we show that hippocampal injection of Aß1-42 is sufficient to induce cognitive deficits 7 days post-injection. We demonstrate using in vitro whole-cell patch-clamping an increased inhibitory GABAergic tonic conductance mediated by extrasynaptic type A GABA receptors (GABAARs), recorded in the CA1 region of the mouse hippocampus following Aß1-42 micro injection. Such alterations in GABA neurotransmission and/or inhibitory GABAARs could have a significant impact on both hippocampal structure and function, causing E/I balance disruption and potentially contributing to cognitive deficits in AD.

Potential role of lava lizards as pollinators across the Galápagos Islands.

Lizards have been reported as important pollinators on several oceanic islands. Here we evaluate the potential role of Galápagos lava lizards (Microlophus spp.) as pollinators across their radiation. Over 3 years, we sampled pollen transport by 9 lava lizard species on the 10 islands where they are present, including 7 single-island endemics. Overall, only 25 of 296 individuals sampled (8.4%) transported pollen of 10 plant species, the most common being Prosopis juliflora, Exodeconus miersii, Sesuvium sp. and Cordia leucophlyctis. At least 8 of these plant species were native, and none were confirmed as introduced to the archipelago. Despite the low overall proportion of individuals carrying pollen, this was observed in 7 of the nine lizard species, and on 8 of the ten main islands (Española, Fernandina, Floreana, Isabela, Marchena, Pinta, Santa Cruz and Santiago), suggesting that this is a widespread interaction. The results reported here support the potential role of lava lizards as pollinators across their radiation, although they may represent a relatively modest contribution when compared with birds and insects. However, we cannot discard that lizards may be ecologically significant for particular plant species and ecosystems given the specific climatic condition and functional diversity of each island.

Small size does not restrain frugivory and seed dispersal across the evolutionary radiation of Galápagos lava lizards.

Frugivory in lizards is often assumed to be constrained by body size; only large individuals are considered capable of consuming fruits, with the potential of acting as seed dispersers. However, only one previous study has tested the correlation of frugivory with body and head size at an archipelago scale across closely related species. All nine lava lizards (Microlophus spp.) were studied on the eleven largest Galápagos islands from 2010 to 2016 to investigate whether frugivory is related to body and head size. We also tested whether fruit abundance influences fruit consumption and explored the effect of seed ingestion on seedling emergence time and percentage. Our results showed that across islands, lava lizards varied considerably in size (64-102 mm in mean snout-vent length) and level of frugivory (1-23%, i.e., percentage of droppings with seeds). However, level of frugivory was only weakly affected by size as fruit consumption was also common among small lizards. Lava lizards consumed fruits throughout the year and factors other than fruit abundance may be more important drivers of fruit selection (e.g., fruit size, energy content of pulp). From 2,530 droppings, 1,714 seeds of at least 61 plant species were identified, 76% of the species being native to the Galápagos. Most seeds (91%) showed no external structural damage. Seedling emergence time (44 versus 118 days) and percentage (20% versus 12%) were enhanced for lizard-ingested seeds compared to control (uningested) fruits. De-pulping by lizards (i.e., removal of pulp with potential germination inhibitors) might increase the chances that at least some seeds find suitable recruitment conditions. We concluded that lizards are important seed dispersers throughout the year and across the whole archipelago, regardless of body size.

Lanolin-Based Synthetic Membranes as Percutaneous Absorption Models for Transdermal Drug Delivery.

BACKGROUND: The major in vitro permeation studies are currently performed in Franz-type diffusion cells because of their simplicity, cost effectiveness and because the experimental conditions can be easily controlled. Apart from the skin, Franz-type diffusion cells can be used with synthetic membranes. Nevertheless, they do not emulate the nature of the lipidic matrix, which is responsible for the topical barrier function. OBJECTIVE: This paper offers two new approaches combining different synthetic membranes (Strat-M® and Nucleopore®) with lanolin, which provides lipidic components similar to the lipidic matrix. METHODS: The molecular structure of lanolin was studied in membranes by attenuated total reflectance infrared spectroscopy (ATR-IR). The water permeability and absorption of lidocaine, diclofenac sodium and betamethasone dipropionate were also studied and compared against free-lanolin membranes and skin. RESULTS: The results showed an increasing barrier function after lanolin application in both membranes, resulting in a decrease in water permeability. Observing the IR spectra, the lateral packaging of the lipid in the synthetic membranes seems to emulate the orthorhombic disposition from the stratum corneum. Moreover, the three substances applied to the lanolin-containing membranes have a similar absorption to that of the skin. CONCLUSIONS: In conclusion, combining synthetic membranes with lanolin may be a useful approach to mimic topical actives’ absorption.

The GABAergic system as a therapeutic target for Alzheimer's disease.

Glutamatergic and cholinergic dysfunction are well-attested features of Alzheimer's disease (AD), progressing with other pathological indices of the disorder and exacerbating neuronal and network dysfunction. However, relatively little attention has been paid to the inhibitory component of the excitatory/inhibitory (E/I) network, particularly dysfunction in the gamma-aminobutyric acid (GABA) signaling system. There is growing evidence in support of GABAergic remodeling in the AD brain, potentially beginning in early stages of disease pathogenesis, and this could thus be a valid molecular target for drug development and pharmacological therapies. Several GABAergic drugs have been tested for efficacy in attenuating or reversing various features and symptoms of AD, and this could represent a novel path by which we might address the growing need for more effective and benign therapies.

GABAA receptor subunit expression changes in the human Alzheimer's disease hippocampus, subiculum, entorhinal cortex and superior temporal gyrus.

Gamma-aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the central nervous system. GABA type A receptors (GABAA Rs) are severely affected in Alzheimer's disease (AD). However, the distribution and subunit composition of GABAA Rs in the AD brain are not well understood. This is the first comprehensive study to show brain region- and cell layer-specific alterations in the expression of the GABAA R subunits α1-3, α5, ß1-3 and γ2 in the human AD hippocampus, entorhinal cortex and superior temporal gyrus. In late-stage AD tissue samples using immunohistochemistry we found significant alteration of all investigated GABAA Rs subunits except for α3 and ß1 that were well preserved. The most prominent changes include an increase in GABAA R α1 expression associated with AD in all layers of the CA3 region, in the stratum (str.) granulare and hilus of the dentate gyrus. We found a significant increase in GABAA R α2 expression in the str. oriens of the CA1-3, str. radiatum of the CA2,3 and decrease in the str. pyramidale of the CA1 region in AD cases. In AD there was a significant increase in GABAA R α5 subunit expression in str. pyramidale, str. oriens of the CA1 region and decrease in the superior temporal gyrus. We also found a significant decrease in the GABAA R ß3 subunit immunoreactivity in the str. oriens of the CA2, str. granulare and str. moleculare of the dentate gyrus. In conclusion, these findings indicate that the expression of the GABAA R subunits shows brain region- and layer-specific alterations in AD, and these changes could significantly influence and alter GABAA R function in the disease. Cover Image for this issue: doi: 10.1111/jnc.14179.

Towards a Better Understanding of GABAergic Remodeling in Alzheimer's Disease.

γ-aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the vertebrate brain. In the past, there has been a major research drive focused on the dysfunction of the glutamatergic and cholinergic neurotransmitter systems in Alzheimer's disease (AD). However, there is now growing evidence in support of a GABAergic contribution to the pathogenesis of this neurodegenerative disease. Previous studies paint a complex, convoluted and often inconsistent picture of AD-associated GABAergic remodeling. Given the importance of the GABAergic system in neuronal function and homeostasis, in the maintenance of the excitatory/inhibitory balance, and in the processes of learning and memory, such changes in GABAergic function could be an important factor in both early and later stages of AD pathogenesis. Given the limited scope of currently available therapies in modifying the course of the disease, a better understanding of GABAergic remodeling in AD could open up innovative and novel therapeutic opportunities.

Flowers as a reservoir of yeast diversity: description of Wickerhamiella nectarea f.a. sp. nov., and Wickerhamiella natalensis f.a. sp. nov. from South African flowers and pollinators, and transfer of related Candida species to the genus Wickerhamiella as new combinations.

Flowers offer favourable microenvironments for yeast growth, and are increasingly recognised as a rich source of novel yeast species. Independent surveys of yeasts associated with flowers and pollinators in South Africa led to the discovery of 38 strains of two new species. Physiological profiles and analysis of the internal transcribed spacer and the D1/D2 domains of the large subunit rRNA gene showed that they represent two novel species that belong to the Wickerhamiella clade. We describe the species as Wickerhamiella nectarea f.a. sp. nov. (type strain EBDCdVSA11-1T, CBS 14162T, NRRL Y-63791T) and W. natalensis f.a. sp. nov. (type strain EBDCdVSA7-1T, CBS 14161T, NRRL Y-63790T). We extend the known range of flower-associated Wickerhamiella species to South Africa and discuss the ecology and phylogenetic relationships of the clade in relation to its host species and biogeography. Examination of growth characteristics supports that the Wickerhamiella clade exhibits a high degree of evolutionary lability, and that specialisation to different niches may occur rapidly. We review the current status of floral yeast biodiversity and nectar as a reservoir of species diversity, and the importance of pollinators and biogeography. In addition, 18 species formerly assigned to the genus Candida are reassigned formally to the genus Wickerhamiella.

Metschnikowia drakensbergensis sp. nov. and Metschnikowia caudata sp. nov., endemic yeasts associated with Protea flowers in South Africa.

In a taxonomic study of yeasts recovered from nectar of flowers and associated insects in South Africa, 11 strains were found to represent two novel species. Morphological and physiological characteristics and sequence analyses of the large-subunit rRNA gene D1/D2 region, as well as the actin, RNA polymerase II and elongation factor 2 genes, showed that the two novel species belonged to the genus Metschnikowia. Metschnikowia drakensbergensis sp. nov. (type strain EBD-CdVSA09-2(T) =CBS 13649(T) =NRRL Y-63721(T); MycoBank no. MB809688; allotype EBD-CdVSA10-2(A) =CBS13650(A) =NRRL Y-63720(A)) was recovered from nectar of Protea roupelliae and the beetle Heterochelus sp. This species belongs to the large-spored Metschnikowia clade and is closely related to Metschnikowia proteae, with which mating reactions and single-spored asci were observed. Metschnikowia caudata sp. nov. (type strain EBD-CdVSA08-1(T) =CBS 13651(T) =NRRL Y-63722(T); MycoBank no. MB809689; allotype EBD-CdVSA57-2(A) =CBS 13729(A) =NRRL Y-63723(A)) was isolated from nectar of Protea dracomontana, P. roupelliae and P. subvestita and a honeybee, and is a sister species to Candida hainanensis and Metschnikowia lopburiensis. Analyses of the four sequences demonstrated the existence of three separate phylotypes. Intraspecies matings led to the production of mature asci of unprecedented morphology, with a long, flexuous tail. A single ascospore was produced in all compatible crosses, regardless of sequence phylotype. The two species appear to be endemic to South Africa. The ecology and habitat specificity of these novel species are discussed in terms of host plant and insect host species.

Phylogenetic analysis of the angiosperm-floricolous insect-yeast association: have yeast and angiosperm lineages co-diversified?

Metschnikowia (Saccharomycetales, Metschnikowiaceae/Metschnikowia clade) is an ascomycetous yeast genus whose species are associated mostly with angiosperms and their insect pollinators over all continents. The wide distribution of the genus, its association with angiosperm flowers, and the fact that it includes some of the best-studied yeasts in terms of biogeography and ecology make Metschnikowia an excellent group to investigate a possible co-radiation with angiosperm lineages. We performed phylogenetic analyses implementing Bayesian inference and likelihood methods, using a concatenated matrix (≈2.6 Kbp) of nuclear DNA (ACT1, 1st and 2nd codon positions of EF2, Mcm7, and RPB2) sequences. We included 77 species representing approximately 90% of the species in the family. Bayesian and parsimony methods were used to perform ancestral character reconstructions within Metschnikowia in three key morphological characters. Patterns of evolution of yeast habitats and divergence times were explored in the Metschnikowia clade lineages with the purpose of inferring the time of origin of angiosperm-associated habitats within Metschnikowiaceae. This paper presents the first phylogenetic hypothesis to include nearly all known species in the family. The polyphyletic nature of Clavispora was confirmed and Metschnikowia species (and their anamorphs) were shown to form two groups: one that includes mostly floricolous, insect-associated species distributed in mostly tropical areas (the large-spored Metschnikowia clade and relatives) and another that comprises more heterogeneous species in terms of habitat and geographical distribution. Reconstruction of character evolution suggests that sexual characters (ascospore length, number of ascospores, and ascus formation) evolved multiple times within Metschnikowia. Complex and dynamic habitat transitions seem to have punctuated the course of evolution of the Metschnikowiaceae with repeated and independent origins of angiosperm-associated habitats. The origin of the family is placed in the Late Cretaceous (71.7 Ma) with most extant species arising from the Early Eocene. Therefore, the Metschnikowiaceae likely radiated long after the Mid-Cretaceous radiations of angiosperms and their diversification seems to be driven by repeated radiation on a pre-existing diverse resource.

Extensive pollen flow but few pollen donors and high reproductive variance in an extremely fragmented landscape.

Analysing pollen movement is a key to understanding the reproductive system of plant species and how it is influenced by the spatial distribution of potential mating partners in fragmented populations. Here we infer parameters related to levels of pollen movement and diversity of the effective pollen cloud for the wind-pollinated shrub Pistacia lentiscus across a highly disturbed landscape using microsatellite loci. Paternity analysis and the indirect KinDist and Mixed Effect Mating models were used to assess mating patterns, the pollen dispersal kernel, the effective number of males (N(ep)) and their relative individual fertility, as well as the existence of fine-scale spatial genetic structure in adult plants. All methods showed extensive pollen movement, with high rates of pollen flow from outside the study site (up to 73-93%), fat-tailed dispersal kernels and large average pollination distances (δ = 229-412 m). However, they also agreed in detecting very few pollen donors (N(ep) = 4.3-10.2) and a large variance in their reproductive success: 70% of males did not sire any offspring among the studied female plants and 5.5% of males were responsible for 50% of pollinations. Although we did not find reduced levels of genetic diversity, the adult population showed high levels of biparental inbreeding (14%) and strong spatial genetic structure (S(p) = 0.012), probably due to restricted seed dispersal and scarce safe sites for recruitment. Overall, limited seed dispersal and the scarcity of successful pollen donors can be contributing to generate local pedigrees and to increase inbreeding, the prelude of genetic impoverishment.

Metschnikowia proteae sp. nov., a nectarivorous insect-associated yeast species from Africa.

A collection of yeasts isolated from nectar of flowers of Protea caffra (Proteaceae) and associated scarab beetles (Atrichelaphinis tigrina, Cyrtothyrea marginalis, Trichostetha fascicularis and Heterochelus sp.) and drosophilid flies in South Africa, contained 28 isolates that could not be assigned to known species. Comparisons of the D1/D2 domains of the large subunit rRNA gene demonstrated the existence of three separate phylotypes with an affinity to the genus Metschnikowia and more specifically to the beetle-associated large-spored Metschnikowia clade. Twenty-six strains that had similar D1/D2 sequences were mixed in all pairwise combinations. They were found to mate and give rise to large asci typical of those in the clade. The name Metschnikowia proteae sp. nov. (type strain EBDT1Y1(T) = CBS 12522(T) = NRRL Y-48784(T); allotype strain EBDC2Y2 = CBS 12521 = NRRL Y-48785) is proposed to accommodate this novel species. The ecology of this novel yeast species is discussed in relation to its potential plant and insect host species. The additional two single strains isolated from Heterochelus sp. represent two novel undescribed species (Candida sp. 1 EBDM2Y3 and Candida sp. 2 EBDM8Y1). As these single strains are probably haploid mating types of Metschnikowia species, their description is deferred until the species are sufficiently well sampled to permit meaningful descriptions.

Psychometric properties of the Spence Child Anxiety Scale with adolescents from five European countries.

This study examined the factor structure, reliability, and validity of the original Spence Child Anxiety Scale (SCAS) and its four translations (German, Cypriot Greek, Swedish, Italian). A total of 2558 adolescents from five European countries (Germany = 495; Cyprus = 611; UK = 469; Sweden = 484; Italy = 499), ages 12-17 years, participated in the study. In addition to the SCAS, all participants completed the Strengths and Difficulties Questionnaire, a measure of general difficulties and positive attributes. The internal consistency and validity (convergent and discriminant) of the SCAS were excellent. Multiple group confirmatory factor analysis provided strong support for the generalizability of a 6-factor inter-correlated model across five European countries. Adolescents in the UK had significantly higher anxiety levels than adolescents in four other European countries. Our findings suggest that the SCAS is suitable for assessing anxiety disorder symptoms in adolescents in Germany, the UK, Cyprus, Sweden, and in Italy.

Adiponectin levels, cardiometabolic risk factors and markers of subclinical atherosclerosis in children.

BACKGROUND: Low levels of adiponectin have been associated with metabolic risk factors (RF) and cardiac disease. Minimal data is available about the relationship between adiponectin and subclinical atherosclerosis. OBJECTIVES: To determine the relationship of adiponectin to cardiometabolic RF, C-reactive protein (CRP), anthropometric parameters of obesity, and subclinical atherosclerosis in children. METHODS: Cross-sectional study in 103 children. We determined body mass index (BMI), waist circumference, percent fat mass, systolic and diastolic blood pressures, fasting lipid profile, glycemia and insulinemia, and CRP. Subclinical atherosclerosis was determined by carotid intima-media thickness (IMT) and flow-mediated dilation of the brachial artery (FMD). RESULTS: Mean age of the group was 12.4+/-1.9 years (47% girls; 20.4% prepubertal; 45 eutrophic, 23 overweight and 35 obese). Adiponectin levels were not statistically significantly different in eutrophic children versus obese+overweight: 17.7+/-5.6 and 15.9+/-5.3 microg/mL, respectively. Adiponectin levels in boys were no different from those in girls. Adiponectin correlated significantly with age, BMI, zBMI, waist circumference, systolic and diastolic blood pressures, HDL, insulinemia, and HOMA index. No statistically significant association with adiponectin was found for CRP, FMD or IMT. After adjusting by sex, pubertal status, and degree of obesity, the adiponectin levels associated significantly with HDL cholesterol and the HOMA index (r(2)=0.34, p<0.0001). CONCLUSIONS: Adiponectin levels were inversely correlated with anthropometric parameters of obesity and insulin resistance and directly correlated with HDL levels. However, no relationship with subclinical atherosclerosis was demonstrated in this study.