The genome of deep-sea vent chemolithoautotroph Thiomicrospira crunogena XCL-2

Presented here is the complete genome sequence of Thiomicrospira crunogena XCL-2, representative of ubiquitous chemolithoautotrophic sulfur-oxidizing bacteria isolated from deep-sea hydrothermal vents. This gammaproteobacterium has a single chromosome (2,427,734 base pairs), and its genome illustrates many of the adaptations that have enabled it to thrive at vents globally. It has 14 methyl-accepting chemotaxis protein genes, including four that may assist in positioning it in the redoxcline. A relative abundance of coding sequences (CDSs) encoding regulatory proteins likely control the expression of genes encoding carboxysomes, multiple dissolved inorganic nitrogen and phosphate transporters, as well as a phosphonate operon, which provide this species with a variety of options for acquiring these substrates from the environment. Thiom. crunogena XCL-2 is unusual among obligate sulfur-oxidizing bacteria in relying on the Sox system for the oxidation of reduced sulfur compounds. The genome has characteristics consistent with an obligately chemolithoautotrophic lifestyle, including few transporters predicted to have organic allocrits, and Calvin-Benson-Bassham cycle CDSs scattered throughout the genome.     

Functional molecular mass of Escherichia coli K92 polysialyltransferase as determined by radiation target analysis

The polysialyltransferase of Escherichia coli K92 catalyzes the transfer of sialic acid from CMP-sialic acid to a growing chain of polysialic acid at the nonreducing end. The enzyme encoded by the neuS gene is membrane-associated and has been suggested to be organized within a complex of several proteins encoded by the K92 gene cluster. Attempts to prepare a soluble active NeuS enzyme have been unsuccessful. Recent results suggest that de novo synthesis of polysialic acid requires coexpression of four genes from the cluster: neuS, neuE, kpsC, and kpsS. However, elongation of preexisting polysialic acid chains only requires expression of neuS. The molecular organization of the catalytic unit of bacterial polysialyltransferases has not been described. We used radiation inactivation to measure the size of the minimum functional unit catalyzing the polysialyltransferase chain extension and de novo reactions. Membranes harboring NeuS in the presence and absence of other products of the K92 gene cluster were exposed to high-energy electrons. The rate of loss of polysialyltransferase activity reveals the mass of the molecules essential for catalytic activity. We observed that the transfer of neuNAc from CMP-neuNAc to a polysialic acid acceptor is catalyzed by a complex with a target size larger than that of monomeric NeuS. The target size of the unit catalyzing the extension of existing polysialic acid chains does not differ significantly from the size of the unit catalyzing transfer of sialic acid to the endogenous acceptor. Parallel samples of membranes containing NeuS and a green fluorescent protein (GFP) chimera were compared by target analysis. The target size of this structural unit was estimated by analysis of the rate of decay of the GFP-NeuS chimera band migrating in the immunoblots. The target size of the structural unit is larger than expected for a monomer. The results of these experiments show that while the target size of the catalytic activity for K92 polysialyltransferase is larger than a monomer of NeuS, a large complex is not required for catalysis.     

Seawater DMS in a perturbed coastal ecosystem

Intra- and interspecific niche overlap for two mayfly species with similar life cycle timing, Rhithrogena semicolorata and Ecdyonurus sp. gr. venosus, were investigated. The nymphs were classified into 5 classes according to size and spatial overlaps are measured along a substratum roughness gradient. Substratum roughness selection was investigated by defining utilisation curves, optimum and tolerance values of the nymphs in relation to larval growth. Differences between species and size classes within each species were observed. Ecdyonurus sp. gr. venosus dominated on rough substrates, whereas R. semicolorata was most abundant on smooth substrates. An intermediate value of total interspecific substratum roughness overlap (0.49) was found. Higher intraspecific than interspecific overlap values suggested a spatial niche segregation between the two species. The results suggested that the spatial niches measured, were closer to the `fundamental niches' than could be expected if competition was acting on the two studied populations.     

Nuclear magnetic resonance-based metabolomics of OCT-embedded frozen kidney samples in mouse and man following standardized pre-analytics

Introduction

Pre-analytical processing significantly affects tissue metabolomes. Since most frozen kidney samples are stored after embedding, standardization of cryoprotective medium removal before metabolomics is essential.

Objectives

We used rodent and human kidney samples to develop an easy and robust pre-analytical procedure compatible with ¹H-nuclear magnetic resonance (NMR)-based metabolomics.

Methods

In mice, renal ischemia was induced for 30 min, followed by 48-h reperfusion (I/R, n?=?6). Right kidneys were transversally cut in two fragments, and snap-frozen in liquid nitrogen (LN2) or in Optimal Cutting Temperature ^® (OCT) fixative. In man, double kidney biopsies were simultaneously obtained before transplantation (n?=?15), and snap-frozen in LN2 or OCT.

Results

¹H-NMR spectrum of pure OCT highlighted two major peaks, i.e. from 3.4 to 4.2 ppm (47.2%) and from 1.2 to 2.2 ppm (42.5%). ¹H-NMR spectra of mouse OCT kidneys were biased at 3.7. By contrast, ¹H-NMR analyses of mouse OCT kidneys iteratively rinsed in saline significantly discriminated sham versus I/R groups, with Q² at 0.695 (to be compared with Q² at 0.866 for LN2 sham vs. I/R kidneys). Discriminant metabolites were analogous in both OCT and LN2 kidneys, with a correlation coefficient of 0.83. In man, iteratively rinsing OCT kidneys in saline eliminated the spectral 3.7-peak, thereby making metabolomes of OCT kidneys interpretable and similar to LN2 samples, with a correlation coefficient of 0.73.

Conclusion

NMR metabolomics using OCT-frozen kidney samples is valuable in mouse and man, following standardized OCT removal. This may help use residual biobanked human tissues to better understand renal pathophysiology.

     

Super‐resolution structure of DNA significantly differs in buccal cells of controls and Alzheimer's patients

The advent of super‐resolution microscopy allowed for new insights into cellular and physiological processes of normal and diseased cells. In this study, we report for the first time on the super‐resolved DNA structure of buccal cells from patients with Alzheimer's disease (AD) versus age‐ and gender‐matched healthy, non‐caregiver controls. In this super‐resolution study cohort of 74 participants, buccal cells were collected and their spatial DNA organization in the nucleus examined by 3D Structured Illumination Microscopy (3D‐SIM). Quantitation of the super‐resolution DNA structure revealed that the nuclear super‐resolution DNA structure of individuals with AD significantly differs from that of their controls (p < 0.05) with an overall increase in the measured DNA‐free/poor spaces. This represents a significant increase in the interchromatin compartment. We also find that the DNA structure of AD significantly differs in mild, moderate, and severe disease with respect to the DNA‐containing and DNA‐free/poor spaces. We conclude that whole genome remodeling is a feature of buccal cells in AD.     

A decade of colonization: the spread of the Asian tiger mosquito in Pennsylvania and implications for disease risk

In recent decades, the Asian tiger mosquito expanded its geographic range throughout the northeastern United States, including Pennsylvania. The establishment of Aedes albopictus in novel areas raises significant public health concerns, since this species is a highly competent vector of several arboviruses, including chikungunya, West Nile, and dengue. In this study, we used geographic information systems (GIS) to examine a decade of colonization by Ae. albopictus throughout Pennsylvania between 2001 and 2010. We examined the spatial and temporal distribution of Ae. albopictus using spatial statistical analysis and examined the risk of dengue virus transmission using a model that captures the probability of transmission. Our findings show that since 2001, the Ae. albopictus population in Pennsylvania has increased, becoming established and expanding in range throughout much of the state. Since 2010, imported cases of dengue fever have been recorded in Pennsylvania. Imported cases of dengue, in combination with summer temperatures conducive for virus transmission, raise the risk of local disease transmission.     

Sex hormone-binding globulin and thrombin generation in women using hormonal contraception

Objectives: We investigated the impact of serum sex hormone-binding globulin (SHBG) on thrombin generation (TG) in women according to hormonal contraception.

Patients and methods: A cross-sectional study of SHBG and TG measured via calibrated automated thrombography was conducted in 150 healthy women, including 75 users of combined oral contraceptives (COC), 22 users of progestin-only contraceptives (POC) and 53 nonusers.

Results: COC but not POC-users had significantly higher SHBG levels compared with nonusers. In hormonal contraceptive users, SHBG was positively associated with both activated protein C (APC) resistance and baseline TG, and protein S and prothrombin were important mediators.

Conclusion: These data provide further evidence that SHBG may be used as a biomarker in assessing prothrombotic profile of hormonal contraception.     

Identical consensus sequence and conserved genomic polymorphism of hepatitis E virus during controlled interspecies transmission

High-throughput sequencing of bile and feces from two pigs experimentally infected with human hepatitis E virus (HEV) of genotype 3f revealed the same full-length consensus sequence as in the human sample. Twenty-nine percent of polymorphic sites found in HEV from the human sample were conserved throughout the infection of the heterologous host. The interspecies transmission of HEV quasispecies is the result of a genomic negative-selection pressure on random mutations which can be deleterious to the viral population. HEV intrahost nucleotide diversity was found to be in the lower range of other human RNA viruses but correlated with values found for zoonotic viruses. HEV transmission between humans and pigs does not seem to be modulated by host-specific mutations, suggesting that adaptation is mainly regulated by ecological drivers.     

Non‐invasive fecal hormone analysis and behavioral observations for monitoring stress responses in captive western lowland gorillas (Gorilla gorilla gorilla)

Non‐invasive techniques for monitoring the stress response in captive western lowland gorillas (Gorilla gorilla gorilla) were investigated. Fecal samples for cortisol measurement and concurrent behavioral data were collected from six individuals in a socially housed gorilla group (one adult male, three adult females and their three offspring) over a 7‐month period. Despite inter‐individual variation in the dynamics of fecal cortisol concentrations over time, several major secretory peaks coincided across individuals. High cortisol concentrations in feces were correlated with induced stressors or behavioral observations indicating high social tension, with a 1–2 day lag period. Entry progression order of the gorillas into a den complex and a supplant‐based dominance index were suitable indicators of overall dominance hierarchies, and fluctuations over time reflected periods of instability. Diurnal variation in fecal cortisol was not apparent when comparing afternoon and morning samples, however the sample collection interval was relatively short (3–5 hr). These results demonstrate the feasibility of monitoring stress responses based on the dynamics of both fecal cortisol excretion and behavior. This non‐invasive approach may be used for gauging responses to changes in husbandry, environment and group structure of captive gorillas. Zoo Biol 0:1–15, 2005. © 2005 Wiley‐Liss, Inc.     

p27 controls autophagic vesicle trafficking in glucose-deprived cells via the regulation of ATAT1-mediated microtubule acetylation

The cyclin-dependent kinase inhibitor p27^Kip1 (p27) has been involved in promoting autophagy and survival in conditions of metabolic stress. While the signaling cascade upstream of p27 leading to its cytoplasmic localization and autophagy induction has been extensively studied, how p27 stimulates the autophagic process remains unclear. Here, we investigated the mechanism by which p27 promotes autophagy upon glucose deprivation. Mouse embryo fibroblasts (MEFs) lacking p27 exhibit a decreased autophagy flux compared to wild-type cells and this is correlated with an abnormal distribution of autophagosomes. Indeed, while autophagosomes are mainly located in the perinuclear area in wild-type cells, they are distributed throughout the cytoplasm in p27-null MEFs. Autophagosome trafficking towards the perinuclear area, where most lysosomes reside, is critical for autophagosome–lysosome fusion and cargo degradation. Vesicle trafficking is mediated by motor proteins, themselves recruited preferentially to acetylated microtubules, and autophagy flux is directly correlated to microtubule acetylation levels. p27^−/− MEFs exhibit a marked reduction in microtubule acetylation levels and restoring microtubule acetylation in these cells, either by re-expressing p27 or with deacetylase inhibitors, restores perinuclear positioning of autophagosomes and autophagy flux. Finally, we find that p27 promotes microtubule acetylation by binding to and stabilizing α-tubulin acetyltransferase (ATAT1) in glucose-deprived cells. ATAT1 knockdown results in random distribution of autophagosomes in p27^+/+ MEFs and impaired autophagy flux, similar to that observed in p27^−/− cells. Overall, in response to glucose starvation, p27 promotes autophagy by facilitating autophagosome trafficking along microtubule tracks by maintaining elevated microtubule acetylation via an ATAT1-dependent mechanism.Subject terms: Tumour-suppressor proteins, Macroautophagy