<Emphasis Type="Italic">Drosophila</Emphasis> as a platform to predict the pathogenicity of novel aminoacyl-tRNA synthetase mutations in CMT

Charcot-Marie-Tooth disease (CMT) is the major form of inherited peripheral neuropathy in humans. CMT is clinically and genetically heterogeneous and four aminoacyl-tRNA synthetases have been implicated in disease etiology. Mutations in the YARS gene encoding a tyrosyl-tRNA synthetase (TyrRS) lead to Dominant Intermediate CMT type C (DI-CMTC). Three dominant YARS mutations were so far associated with DI-CMTC. To further expand the spectrum of CMT causing genetic defects in this tRNA synthetase, we performed DNA sequencing of YARS coding regions in a cohort of 181 patients with various types of peripheral neuropathy. We identified a novel K265N substitution that in contrast to all previously described mutations is located at the anticodon recognition domain of the enzyme. Further genetic analysis revealed that this variant represents a benign substitution. Using our recently developed DI-CMTC Drosophila model, we tested in vivo the pathogenicity of this new YARS variant. We demonstrated that the developmental and behavioral defects induced by all DI-CMTC causing mutations were not present upon ubiquitous or panneuronal TyrRS K265N expression. Thus, in line with our genetic studies, functional analysis confirmed that the K265N substitution does not induce toxicity signs in Drosophila. The consistency observed throughout this work underscores the robustness of our DI-CMTC animal model and identifies Drosophila as a valid read-out platform to ascertain the pathogenicity of novel mutations to be identified in the future.     

The ‘Hutchinsonian niche’ as an assemblage of demographic niches: implications for species geographic ranges

Hutchinson defined the ecological niche as a hypervolume shaped by the environmental conditions under which a species can ‘exist indefinitely’. Although several authors further discussed the need to adopt a demographic perspective of the ecological niche theory, very few have investigated the environmental requirements of different components of species’ life cycles (i.e. vital rates) in order to examine their internal niche structures. It therefore remains unclear how species’ demography, niches and distributions are interrelated. Using comprehensive demographic data for two well‐studied, short‐lived plants (Plantago coronopus, Clarkia xantiana), we show that the arrangement of species’ demographic niches reveals key features of their environmental niches and geographic distributions. In Plantago coronopus, opposing geographic trends in some individual vital rates, through different responses to environmental gradients (demographic compensation), stabilize population growth across the range. In Clarkia xantiana, a lack of demographic compensation underlies a gradient in population growth, which could translate in a directional geographic range shift. Overall, our results highlight that occurrence and performance niches cannot be assumed to be the same, and that studying their relationship is essential for a better understanding of species’ ecological niches. Finally, we argue for the value of considering the assemblage of species’ demographic niches when studying ecological systems, and predicting the dynamics of species geographical ranges.     

Search for unstable DNA in schizophrenia families with evidence for genetic anticipation.

Evidence for genetic anticipation has recently become an important subject of research in clinical psychiatric genetics. Renewed interest in anticipation was evoked by molecular genetic findings of a novel type of mutation termed "unstable DNA." The unstable DNA model can be construed as the "best fit" for schizophrenia twin and family epidemiological data. We have performed a large-scale Southern blot hybridization, asymmetrical PCR-based, and repeat expansion-detection screening for (CAG)n/(CTG)n and (CCG)n/(CGG)n expansions in eastern Canadian schizophrenia multiplex families demonstrating genetic anticipation. There were no differences in (CAG)n/(CTG)n and (CCG)n/(CGG)n pattern distribution either between affected and unaffected individuals or across generations. Our findings do not support the hypothesis that large (CAG)n/(CTG)n or (CCG)n/(CGG)n expansions are the major etiologic factor in schizophrenia. A separate set of experiments directed to the analysis of small (30-130 trinucleotides), Huntington disease-type expansions in individual genes is required in order to fully exclude the presence of (CAG)n/(CTG)n- or (CCG)n/(CGG)n-type unstable mutation.     

Fragments and dust after Holmium laser lithotripsy with or without “Moses technology”: How are they different?

Urinary stones can be readily disintegrated by Holmium:YAG laser (Holmium laser lithotripsy), resulting in a mixture of small stone dust particles, which will spontaneously evacuate with urine and larger residual fragments (RF) requiring mechanical retrieval. Differences between fragments and dust have not been well characterized. Also, it remains unknown how the recently introduced “Moses technology” may alter stone disintegration products. Three complementary analytical techniques have been used in this study to offer an in‐depth characterization of disintegration products after in vitro Holmium laser lithotripsy: stereoscopic microscopy, scanning electron microscopy and Fourier‐transform infrared spectroscopy. Dust was separated from fragments based on its floating ability in saline irrigation. Depending on initial crystalline constituents, stone dust either conserved attributes found in larger RFs or showed changes in crystalline organization. These included conversion of calcium oxalate dihydrate towards calcium oxalate monohydrate, changes in carbapatite spectra towards an amorphous phase, changes of magnesium ammonium phosphate towards a differing amorphous and crystalline phase and the appearance of hydroxyapatite on brushite fragments. Comparatively, “Moses technology” produced more pronounced changes. These findings provide new insights suggesting a photothermal effect occurring in Holmium laser lithotripsy. Figure: Appearance of hydroxyapatite hexagons on stone dust collected after Holmium laser lithotripsy of a brushite stone using “Moses technology.”      

High‐Throughput DNA sequencing of ancient wood

Reconstructing the colonization and demographic dynamics that gave rise to extant forests is essential to forecasts of forest responses to environmental changes. Classical approaches to map how population of trees changed through space and time largely rely on pollen distribution patterns, with only a limited number of studies exploiting DNA molecules preserved in wooden tree archaeological and subfossil remains. Here, we advance such analyses by applying high‐throughput (HTS) DNA sequencing to wood archaeological and subfossil material for the first time, using a comprehensive sample of 167 European white oak waterlogged remains spanning a large temporal (from 550 to 9,800 years) and geographical range across Europe. The successful characterization of the endogenous DNA and exogenous microbial DNA of 140 (~83%) samples helped the identification of environmental conditions favouring long‐term DNA preservation in wood remains, and started to unveil the first trends in the DNA decay process in wood material. Additionally, the maternally inherited chloroplast haplotypes of 21 samples from three periods of forest human‐induced use (Neolithic, Bronze Age and Middle Ages) were found to be consistent with those of modern populations growing in the same geographic areas. Our work paves the way for further studies aiming at using ancient DNA preserved in wood to reconstruct the micro‐evolutionary response of trees to climate change and human forest management.     

Therapeutic and pharmacokinetic characterizations of an anti-amyloidogenic bis-styrylbenzene derivative for Alzheimer’s disease treatment

Alzheimer’s disease drug discovery regarding exploration into the molecules and processes has focused on the intrinsic causes of the brain disorder correlated with the accumulation of amyloid-β. An anti-amyloidogenic bis-styrylbenzene derivative, KMS80013, showed excellent oral bioavailability (F = 46.2%), facilitated brain penetration (26%, iv) in mouse and target specific in vivo efficacy in acute AD mouse model attenuating the cognitive deficiency in Y-maze test. Acute toxicity (LD₅₀ >2000 mg/kg) and hERG channel inhibition (14% at 10 μM) results indicated safety of KMS80013.     

Cost-Effectiveness and Efficacy of spa,SCCmec, and PVL Genotyping of Methicillin-Resistant Staphylococcus aureus as Compared to Pulsed-Field Gel Electrophoresis

Pulsed-field gel electrophoresis (PFGE) is a valuable molecular typing assay used for methicillin-resistant Staphylococcus aureus (MRSA) surveillance and genotyping. However, there are several limitations associated with PFGE. In Alberta, Canada, the significant increase in the number of MRSA isolates submitted to the Provincial Laboratory for Public Health (ProvLab) for PFGE typing led to the need for an alternative genotyping method. In this study, we describe the transition from PFGE to Staphylococcus protein A (spa), Staphylococcal cassette chromosome (SCCmec), and Panton-Valentine leukocidin (PVL) typing. A total of 1915 clinical MRSA isolates collected from 2005 to 2009 were used to develop and validate an algorithm for assigning PFGE epidemic types using spa, SCCmec, and PVL typing and the resulting data was used to populate a new Alberta MRSA typing database. An additional 12620 clinical MRSA isolates collected from 2010 to 2012 as part of ongoing routine molecular testing at ProvLab were characterized using the new typing algorithm and the Alberta MRSA typing database. Switching to spa, SCCmec, and PVL from PFGE typing substantially reduced hands-on and turn-around times while maintaining historical PFGE epidemic type designations. This led to an approximate $77,000 reduction in costs from 2010 to 2012. PFGE typing is still required for a small subset of MRSA isolates that have spa types that are rare, novel, or associated with more than one PFGE epidemic type.     

3D Microstructural Architecture of Muscle Attachments in Extant and Fossil Vertebrates Revealed by Synchrotron Microtomography

Background

Firm attachments binding muscles to skeleton are crucial mechanical components of the vertebrate body. These attachments (entheses) are complex three-dimensional structures, containing distinctive arrangements of cells and fibre systems embedded in the bone, which can be modified during ontogeny. Until recently it has only been possible to obtain 2D surface and thin section images of entheses, leaving their 3D histology largely unstudied except by extrapolation from 2D data. Entheses are frequently preserved in fossil bones, but sectioning is inappropriate for rare or unique fossil material.

Methodology/Principal Findings

Here we present the first non-destructive 3D investigation, by propagation phase contrast synchrotron microtomography (PPC-SRµCT), of enthesis histology in extant and fossil vertebrates. We are able to identify entheses in the humerus of the salamander Desmognathus from the organization of bone-cell lacunae and extrinsic fibres. Statistical analysis of the lacunae differentiates types of attachments, and the orientation of the fibres, reflect the approximate alignment of the muscle. Similar histological structures, including ontogenetically related pattern changes, are perfectly preserved in two 380 million year old fossil vertebrates, the placoderm Compagopiscis croucheri and the sarcopterygian fish Eusthenopteron foordi.

Conclusions/Significance

We are able to determine the position of entheses in fossil vertebrates, the approximate orientation of the attached muscles, and aspects of their ontogenetic histories, from PPC-SRµCT data. Sub-micron microtomography thus provides a powerful tool for studying the structure, development, evolution and palaeobiology of muscle attachments.     

Gas exchange characteristics and nitrogen relations of two Mediterranean root hemiparasites:Bartsia trixago andParentucellia viscosa

Plant height, light-saturated rates of photosynthesis (A _max) and foliar nitrogen concentration (N ₁) were measured forBartsia trixago under field conditions in Mallorca. All three variables were postively correlated, and were also positively related to the abundance of nitrogen-fixing legumes in the associated vegetation (putative host species).A _max forB. trixago ranged from 7.7 to 18.8 mol m^-2 s^-1; similar rates were measured for a second hemiparasiteParentucellia viscosa, and both species were within the range of rates measured for six putative hosts (10.6–19.2 mol m^-2 s^-1). Fertilization of unattachedB. trixago plants with inorganic nitrogen (ammonium nitrate) elicited neither the growth nor the photosynthetic responses observed in plants considered to be parasitic on legumes and in receipt of an enriched organic nitrogen supply. Both hemiparasites had high diurnal leaf conductances (g _s) (469–2291 mmol m^-2 s^-1) and were at the upper end of the range of those measured in putative hosts (409–879 mmol m^-2 s^-1). In contrast with the latter, high nocturnal rates ofg _s were also recorded for the two hemiparasites (517–1862 mmol m^-2 s^-1). There was no clear relationship between eitherA _max orN ₁ and eitherg _s, transpiration (E) or water use efficiency (A _max/E) inB. trixago plants. The economics of water loss appear to be independent of both the supply of nitrogen from the host and autotrophic carbon fixation.