Microfluidic chips for in vivo imaging of cellular responses to neural injury in Drosophila larvae

With powerful genetics and a translucent cuticle, the Drosophila larva is an ideal model system for live imaging studies of neuronal cell biology and function. Here, we present an easy-to-use approach for high resolution live imaging in Drosophila using microfluidic chips. Two different designs allow for non-invasive and chemical-free immobilization of 3(rd) instar larvae over short (up to 1 hour) and long (up to 10 hours) time periods. We utilized these 'larva chips' to characterize several sub-cellular responses to axotomy which occur over a range of time scales in intact, unanaesthetized animals. These include waves of calcium which are induced within seconds of axotomy, and the intracellular transport of vesicles whose rate and flux within axons changes dramatically within 3 hours of axotomy. Axonal transport halts throughout the entire distal stump, but increases in the proximal stump. These responses precede the degeneration of the distal stump and regenerative sprouting of the proximal stump, which is initiated after a 7 hour period of dormancy and is associated with a dramatic increase in F-actin dynamics. In addition to allowing for the study of axonal regeneration in vivo, the larva chips can be utilized for a wide variety of in vivo imaging applications in Drosophila.     

Genome sequence of Ensifer medicae strain WSM1115; an acid-tolerant Medicago-nodulating microsymbiont from Samothraki,Greece

Ensifer medicae strain WSM1115 forms effective nitrogen fixing symbioses with a range of annual Medicago species and is used in commercial inoculants in Australia. WSM1115 is an aerobic, motile, Gram-negative, non-spore-forming rod. It was isolated from a nodule recovered from the root of burr medic (Medicago polymorpha) collected on the Greek Island of Samothraki. WSM1115 has a broad host range for nodulation and N₂ fixation capacity within the genus Medicago, although this does not extend to all medic species. WSM1115 is considered saprophytically competent in moderately acid soils (pH(CaCl₂) 5.0), but it has failed to persist at field sites where soil salinity exceeded 10 ECe (dS/m). Here we describe the features of E. medicae strain WSM1115, together with genome sequence information and its annotation. The 6,861,065 bp high-quality-draft genome is arranged into 7 scaffolds of 28 contigs, contains 6,789 protein-coding genes and 83 RNA-only encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.     

Complete genome sequence of the orange-red pigmented, radioresistant Deinococcus proteolyticus type strain (MRP(T))

Deinococcus proteolyticus (ex Kobatake et al. 1973) Brook and Murray 1981 is one of currently 47 species in the genus Deinococcus within the family Deinococcaceae. Strain MRP(T) was isolated from feces of Lama glama and possesses extreme radiation resistance, a trait is shares with various other species of the genus Deinococcus, with D. proteolyticus being resistant up to 1.5 Mrad of gamma radiation. Strain MRP(T) is of further interest for its carotenoid pigment. The genome presented here is only the fifth completed genome sequence of a member of the genus Deinococcus (and the forth type strain) to be published, and will hopefully contribute to a better understanding of how members of this genus adapted to high gamma- or UV ionizing-radiation. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 2,886,836 bp long genome with its four large plasmids of lengths 97 kbp, 132 kbp, 196 kbp and 315 kbp harbors 2,741 protein-coding and 58 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Discrimination between Francisella tularensis and Francisella-Like Endosymbionts when Screening Ticks by PCR

The presence of Francisella-like endosymbionts in tick species known to transmit tularemia poses a potential diagnostic problem for laboratories that screen tick samples by PCR for Francisella tularensis. Tick samples initially considered positive for F. tularensis based on standard 16S rRNA gene PCR were found to be positive only for Francisella-like endosymbionts using a multitarget F. tularensis TaqMan assay (ISFtu2, tul4, and iglC) and 16S rRNA gene sequencing. Specificity of PCR-based diagnostics for F. tularensis should be carefully evaluated with appropriate specimen types prior to diagnostic use.     

Influence of inlet boundary conditions on the local haemodynamics of intracranial aneurysms

Haemodynamics is believed to play an important role in the initiation, growth and rupture of intracranial aneurysms. In this context, computational haemodynamics has been extensively used in an effort to establish correlations between flow variables and clinical outcome. It is common practice in the application of Dirichlet boundary conditions at domain inlets to specify transient velocities as either a flat (plug) profile or a spatially developed profile based on Womersley's analytical solution. This paper provides comparative haemodynamics measures for three typical cerebral aneurysms.

Three dimentional rotational angiography images of aneurysms at three common locations, viz. basilar artery tip, internal carotid artery and middle cerebral artery were obtained. The computational tools being developed in the European project @neurIST were used to reconstruct the fluid domains and solve the unsteady Navier–Stokes equations, using in turn Womersley and plug-flow inlet velocity profiles. The effects of these assumptions were analysed and compared in terms of relevant haemodynamic variables within the aneurismal sac. For the aneurysm at the basilar tip geometries with different extensions of the afferent vasculature were considered to study the plausibility of a fully-developed axial flow at the inlet boundaries.

The study shows that assumptions made on the velocity profile while specifying inlet boundary conditions have little influence on the local haemodynamics in the aneurysm, provided that a sufficient extension of the afferent vasculature is considered and that geometry is the primary determinant of the flow field within the aneurismal sac. For real geometries the Womersley profile is at best an unnecessary over-complication, and may even be worse than the plug profile in some anatomical locations (e.g. basilar confluence).     

Analysis of allelic variants in the catalase gene in patients with the skin depigmenting disorder vitiligo

Vitiligo is an acquired hypomelanotic skin disorder characterised by circumscribed depigmented macules resulting from the loss of functional melanocytes from the cutaneous epidermis. Conditions that might result in epidermal oxidative stress and consequently damage to pigment cells have been reported in the skin of vitiligo patients, including low catalase activity and increases in hydrogen peroxide levels. However, the cause of the decrease in catalase activity has not been equivocally determined. Several allelic variants in the catalase gene, a number of which have deleterious effects upon the expression or function of the enzyme, have been described and the aim of the present work was to assess the relevance of catalase gene variants in patients with vitiligo. Associations between ten separate allelic variants in the catalase gene and a predisposition to vitiligo were investigated in case-control studies with 166 English patients and 169 ethnically-matched controls using DNA sequencing and restriction fragment length polymorphism-polymerase chain reaction methods. Of the ten allelic variants analysed, only a C/T single nucleotide polymorphism in exon 9 of the catalase gene was associated with vitiligo. The C/T genotype was significantly over-represented in the vitiligo patient group compared with the control cohort. Of 166 vitiligo genotypes, 66 (39.8%) had the C/T variant compared to 45/169 (26.6%) control genotypes (P = 0.030). No evidence for an association between other allelic variants in the catalase gene and vitiligo susceptibility was found. The low catalase activity in vitiligo patient epidermis is more likely to result from environmental conditions such as inhibitory levels of hydrogen peroxide rather than allelic variations in the catalase gene which affect either expression or function of the enzyme.     

Dietary and environmental reconstruction with stable isotope analyses of herbivore tooth enamel from the Miocene locality of Fort Ternan, Kenya

Tooth enamel of nine Middle Miocene mammalian herbivores from Fort Ternan, Kenya, was analyzed for δ¹³C and δ¹⁸O. The δ¹⁸O values of the tooth enamel compared with pedogenic and diagenetic carbonate confirm the use of stable isotope analysis of fossil tooth enamel as a paleoenvironmental indicator. Furthermore, the δ¹⁸O of tooth enamel indicates differences in water sources between some of the mammals. The δ¹³C values of tooth enamel ranged from −8·6–−13·0‰ which is compatible with a pure C₃diet, though the possibility of a small C₄fraction in the diet of a few of the specimens sampled is not precluded. The carbon isotopic data do not support environmental reconstructions of a Serengeti-typed wooded grassland with a significant proportion of C₄grasses. This study does not preclude the presence of C₃grasses at Fort Ternan; it is possible that C₃grasses could have had a wider geographic range if atmospheric CO₂levels were higher than the present values.     

Olfactory and Behavioral Mechanisms Underlying Enhanced Mating Competitiveness Following Exposure to Ginger Root Oil and Orange Oil in Males of the Mediterranean Fruit Fly, Ceratitis capitata (Diptera: Tephritidae)

Males of the Mediterranean fruit fly, Ceratitis capitata, are strongly attracted to various plant odors, and previous work has demonstrated that male exposure to certain odors, including the scent of orange oil (OO) and ginger root oil (GRO), increases their mating success relative to non-exposed males. However, the mechanism(s) underlying this mating increase is not known. Here, we describe several experiments that further investigate the association between GRO- and OO-exposure and male signaling activity, pheromone attractiveness, and mating success in male medflies. Exposure to GRO or OO increased time spent pheromone calling but did not accelerate the rate of male sexual maturation. Using a wind tunnel, we compared female attraction to the pheromone of control, non-exposed males versus males previously exposed to OO or GRO. There was no evidence that GRO exposure enhanced the attractiveness of the male pheromone. The data for OO were inconclusive: females tended to spend more time on spheres emanating pheromone from OO-exposed males than on spheres emanating pheromone from non-exposed males, but the number of female landings did not differ between the two types of pheromone sources. Female choice tests confirmed that GRO- and OO-exposure boost male mating success relative to non-exposed males. Application of GRO directly to the abdomen reduced male mating success, whereas similar application of OO boosted male mating success. The potential role and mode of action of plant chemicals in the mating behavior of male medflies are evaluated in light of these findings.     

Spiders in the context of agricultural land abandonment in Greek Mountains: species responses,community structure and the need to preserve traditional agricultural landscapes

Agricultural land abandonment is a major conservation issue when it comes to remote Mediterranean mountainous ecosystems. Although its impact on taxa such as birds or butterflies is well known, knowledge remains poor for less studied invertebrate taxa such as spiders. We sampled ground spiders (Gnaphosidae, Liocranidae, Miturgidae and Corrinidae) in 20 randomly selected sites (1 km × 1 km; 15 pitfall traps) that well depicted a land abandonment gradient, in terms of four forest encroachment classes. Our results showed a negative effect of land abandonment on ground spider species richness and diversity (Simpson index), pinpointing that forested habitats with more than 75 % woody vegetation cover are relatively poor. We also provide evidence for the beneficial role of low intensity grazing (0.4–4 livestock units/km²) for ground spider abundance. Community analysis revealed four distinct clusters of co-occurring species, while Generalized Linear Models at cluster and species level showed the definitive role of forest encroachment, and secondarily of other environmental factors such as humidity, elevation and longitude, in regulating species distribution patterns. Conservation measures for ground spider diversity maintenance should focus on promoting traditional agricultural practices, including small-scale cultivation and mild livestock grazing in order to preserve open and semi-open rural mosaics.