Sequence analysis and distribution in Salmonella enterica serovars of IS3-like elements

The genome of Salmonella enterica serovar Enteritidis was shown to possess three IS3-like insertion elements, designated IS1230A, B and C, and each was cloned and their respective deoxynucleotide sequences determined. Mutations in elements IS1230A and B resulted in frameshifts in the open reading frames that encoded a putative transposase to be inactive. IS1230C was truncated at nucleotide 774 relative to IS1230B and therefore did not possess the 3' terminal inverted repeat. The three IS1230 derivatives were closely related to each other based on nucleotide sequence similarity. IS1230A was located adjacent to the sef operon encoding SEF14 fimbriae located at minute 97 of the genome of S. Enteritidis. IS1230B was located adjacent to the umuDC operon at minute 42.5 on the genome, itself located near to one terminus of an 815-kb genome inversion of S. Enteritidis relative to S. Typhimurium. IS1230C was located next to attB, the bacteriophage P22 attachment site, and proB, encoding gamma-glutamyl phosphate reductase. A truncated 3' remnant of IS1230, designated IS1230T, was identified in a clinical isolate of S. Typhimurium DT193 strain 2391. This element was located next to attB adjacent to which were bacteriophage P22-like sequences. Southern hybridisation of total genomic DNA from eighteen phage types of S. Enteritidis and eighteen definitive types of S. Typhimurium showed similar, if not identical, restriction fragment profiles in the respective serovars when probed with IS1230A.

Localization of calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1) in human gastrointestinal tract

Calcitonin gene-related peptide (CGRP) exerts its diverse effects on vasodilation, nociception, secretion, and motor function through a heterodimeric receptor comprising of calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1). Despite the importance of CLR.RAMP1 in human disease, little is known about its distribution in the human gastrointestinal (GI) tract, where it participates in inflammation and pain. In this study, we determined that CLR and RAMP1 mRNAs are expressed in normal human stomach, ileum and colon by RT-PCR. We next characterized antibodies that we generated to rat CLR and RAMP1 in transfected HEK cells. Having characterized these antibodies in vitro, we then localized CLR-, RAMP1-, CGRP- and intermedin-immunoreactivity (IMD-IR) in various human GI segments. In the stomach, nerve bundles in the myenteric plexus and nerve fibers throughout the circular and longitudinal muscle had prominent CLR-IR. In the proximal colon and ileum, CLR was found in nerve varicosities of the myenteric plexus and surrounding submucosal neurons. Interestingly, CGRP expressing fibers did not co-localize, but were in close proximity to CLR. However, CLR and RAMP1, the two subunits of a functional CGRP receptor were clearly localized in myenteric plexus, where they may form functional cell-surface receptors. IMD, another member of calcitonin peptide family was also found in close proximity to CLR, and like CGRP, did not co-localize with either CLR or RAMP1 receptors. Thus, CGRP and IMD appear to be released locally, where they can mediate their effect on their receptors regulating diverse functions such as inflammation, pain and motility.

Localization of calcitonin receptor-like receptor and receptor activity modifying protein 1 in enteric neurons, dorsal root ganglia, and the spinal cord of the rat

Calcitonin receptor-like receptor (CLR) and receptor activity modifying protein 1 (RAMP1) comprise a receptor for calcitonin gene related peptide (CGRP) and intermedin. Although CGRP is widely expressed in the nervous system, less is known about the localization of CLR and RAMP1. To localize these proteins, we raised antibodies to CLR and RAMP1. Antibodies specifically interacted with CLR and RAMP1 in HEK cells coexpressing rat CLR and RAMP1, determined by Western blotting and immunofluorescence. Fluorescent CGRP specifically bound to the surface of these cells and CGRP, CLR, and RAMP1 internalized into the same endosomes. CLR was prominently localized in nerve fibers of the myenteric and submucosal plexuses, muscularis externa and lamina propria of the gastrointestinal tract, and in the dorsal horn of the spinal cord of rats. CLR was detected at low levels in the soma of enteric, dorsal root ganglia (DRG), and spinal neurons. RAMP1 was also localized to enteric and DRG neurons and the dorsal horn. CLR and RAMP1 were detected in perivascular nerves and arterial smooth muscle. Nerve fibers containing CGRP and intermedin were closely associated with CLR fibers in the gastrointestinal tract and dorsal horn, and CGRP and CLR colocalized in DRG neurons. Thus, CLR and RAMP1 may mediate the effects of CGRP and intermedin in the nervous system. However, mRNA encoding RAMP2 and RAMP3 was also detected in the gastrointestinal tract, DRG, and dorsal horn, suggesting that CLR may associate with other RAMPs in these tissues to form a receptor for additional peptides such as adrenomedullin.

Phylogenetic relationships withinPelargonium sect. Peristera (Geraniaceae) inferred from nrDNA and cpDNA sequence comparisons

Phylogenetic analysis of nrDNA ITS and trnL (UAA) 5′ exon-trnF (GAA) chloroplast DNA sequences from 17 species ofPelargonium sect.Peristera, together with nine putative outgroups, suggests paraphyly for the section and a close relationship between the highly disjunct South African and Australian species of sect.Peristera. Representatives fromPelargonium sectt.Reniformia, Ligularia s. l. andIsopetalum (the St. Helena endemicP. cotyledonis) appear to be nested within thePeristera clade. The close relationship between the South African and AustralianPeristera is interpreted as being caused by long-range dispersal to Australia, probably as recent as the late Pliocene.

Localization and function of the Kv3.1b subunit in the rat medulla oblongata: focus on the nucleus tractus solitarii

The voltage-gated potassium channel subunit Kv3.1 confers fast firing characteristics to neurones. Kv3.1b subunit immunoreactivity (Kv3.1b-IR) was widespread throughout the medulla oblongata, with labelled neurones in the gracile, cuneate and spinal trigeminal nuclei. In the nucleus of the solitary tract (NTS), Kv3.1b-IR neurones were predominantly located close to the tractus solitarius (TS) and could be GABAergic or glutamatergic. Ultrastructurally, Kv3.1b-IR was detected in NTS terminals, some of which were vagal afferents. Whole-cell current-clamp recordings from neurones near the TS revealed electrophysiological characteristics consistent with the presence of Kv3.1b subunits: short duration action potentials (4.2 +/- 1.4 ms) and high firing frequencies (68.9 +/- 5.3 Hz), both sensitive to application of TEA (0.5 mm) and 4-aminopyridine (4-AP; 30 mum). Intracellular dialysis of an anti-Kv3.1b antibody mimicked and occluded the effects of TEA and 4-AP in NTS and dorsal column nuclei neurones, but not in dorsal vagal nucleus or cerebellar Purkinje cells (which express other Kv3 subunits, but not Kv3.1b). Voltage-clamp recordings from outside-out patches from NTS neurones revealed an outward K(+) current with the basic characteristics of that carried by Kv3 channels. In NTS neurones, electrical stimulation of the TS evoked EPSPs and IPSPs, and TEA and 4-AP increased the average amplitude and decreased the paired pulse ratio, consistent with a presynaptic site of action. Synaptic inputs evoked by stimulation of a region lacking Kv3.1b-IR neurones were not affected, correlating the presence of Kv3.1b in the TS with the pharmacological effects.

On the use of a time sequence of surface pressures in four-dimensional data assimilation

The possibility of using a time sequence of surface pressure observations in four-dimensional data assimilation is being investigated. It is shown that a linear multilevel quasi-geostrophic model can be updated successfully with surface data alone, provided the number of time levels are at least as many as the number of vertical levels. It is further demonstrated that current statistical analysis procedures are very inefficient to assimilate surface observations, and it is shown by numerical experiments that the vertical interpolation must be carried out using the structure of the most dominating baroclinic mode in order to obtain a satisfactory updating. Different possible ways towards finding a practical solution are being discussed.

Codon and mRNA sequence optimization of microdystrophin transgenes improves expression and physiological outcome in dystrophic mdx mice following AAV2/8 gene transfer

Duchenne muscular dystrophy is a fatal muscle-wasting disorder. Lack of dystrophin compromises the integrity of the sarcolemma and results in myofibers that are highly prone to contraction-induced injury. Recombinant adenoassociated virus (rAAV)-mediated dystrophin gene transfer strategies to muscle for the treatment of Duchenne muscular dystrophy (DMD) have been limited by the small cloning capacity of rAAV vectors and high titers necessary to achieve efficient systemic gene transfer. In this study, we assess the impact of codon optimization on microdystrophin (ΔAB/R3-R18/ΔCT) expression and function in the mdx mouse and compare the function of two different configurations of codon-optimized microdystrophin genes (ΔAB/R3-R18/ΔCT and ΔR4-R23/ΔCT) under the control of a muscle-restrictive promoter (Spc5-12). Codon optimization of microdystrophin significantly increases levels of microdystrophin mRNA and protein after intramuscular and systemic administration of plasmid DNA or rAAV2/8. Physiological assessment demonstrates that codon optimization of ΔAB/R3-R18/ΔCT results in significant improvement in specific force, but does not improve resistance to eccentric contractions compared with noncodon-optimized ΔAB/ R3-R18/ΔCT. However, codon-optimized microdystrophin ΔR4-R23/ΔCT completely restored specific force generation and provided substantial protection from contraction-induced injury. These results demonstrate that codon optimization of microdystrophin under the control of a muscle-specific promoter can significantly improve expression levels such that reduced titers of rAAV vectors will be required for efficient systemic administration.

Alternating sums of binomial coefficients with unit fraction arithmetic sequence coefficients

Expressions for finite sums involving the binomial coefficients with unit fraction coefficients whose denominators form an arithmetic sequence are determined.

Timing and localization of ionospheric signatures associated with substorm expansion phase onset

In this paper, we present case studies of the optical and magnetic signatures of the characteristics of the first minute of substorm expansion phase onset observed in the ionosphere. We find that for two isolated substorms, the onset of magnetic pulsations in the 24–96 s period wavelet band are colocated in time and space with the formation and development of small-scale optical undulations along the most equatorward preexisting auroral arc prior to auroral breakup. These undulations undergo an inverse spatial cascade into vortices prior to the release of the westward traveling surge. We also present a case study of a multiple activation substorm, whereby discrete onsets of ULF wave power above a predetermined quiet time threshold are shown to be associated with specific optical intensifications and brightenings. Moreover, in the multiple activation substorm event, we show that neither the formation of the small-scale undulations nor the formation of similar structures along a north–south aligned arc is sufficient to produce auroral breakup associated with expansion phase onset. It is only ∼10 min after these two disparate activation regions initiate that auroral breakup and the subsequent formation of a westward traveling surge occur. We discuss the implications of these results in terms of the triggering mechanisms likely to be occurring during these specific events.

Localization of deep water formation: role of atmospheric moisture transport and geometrical constraints on ocean circulation

A series of coupled atmosphere–ocean–ice aquaplanet experiments is described in which topological constraints on ocean circulation are introduced to study the role of ocean circulation on the mean climate of the coupled system. It is imagined that the earth is completely covered by an ocean of uniform depth except for the presence or absence of narrow barriers that extend from the bottom of the ocean to the sea surface. The following four configurations are described: Aqua (no land), Ridge (one barrier extends from pole to pole), Drake (one barrier extends from the North Pole to 35°S), and DDrake (two such barriers are set 90° apart and join at the North Pole, separating the ocean into a large basin and a small basin, connected to the south). On moving from Aqua to Ridge to Drake to DDrake, the energy transports in the equilibrium solutions become increasingly “realistic,” culminating in DDrake, which has an uncanny resemblance to the present climate. Remarkably, the zonal-average climates of Drake and DDrake are strikingly similar, exhibiting almost identical heat and freshwater transports, and meridional overturning circulations. However, Drake and DDrake differ dramatically in their regional climates. The small and large basins of DDrake exhibit distinctive Atlantic-like and Pacific-like characteristics, respectively: the small basin is warmer, saltier, and denser at the surface than the large basin, and is the main site of deep water formation with a deep overturning circulation and strong northward ocean heat transport. A sensitivity experiment with DDrake demonstrates that the salinity contrast between the two basins, and hence the localization of deep convection, results from a deficit of precipitation, rather than an excess of evaporation, over the small basin. It is argued that the width of the small basin relative to the zonal fetch of atmospheric precipitation is the key to understanding this salinity contrast. Finally, it is argued that many gross features of the present climate are consequences of two topological asymmetries that have profound effects on ocean circulation: a meridional asymmetry (circumpolar flow in the Southern Hemisphere; blocked flow in the Northern Hemisphere) and a zonal asymmetry (a small basin and a large basin).

PPARGC1A sequence variation and cardiovascular risk-factor levels: a study of the main genetic effects and gene x environment interactions in children from the European Youth Heart Study.

AIMS/HYPOTHESIS: The PPARGC1A gene coactivates multiple nuclear transcription factors involved in cellular energy metabolism and vascular stasis. In the present study, we genotyped 35 tagging polymorphisms to capture all common PPARGC1A nucleotide sequence variations and tested for association with metabolic and cardiovascular traits in 2,101 Danish and Estonian boys and girls from the European Youth Heart Study, a multicentre school-based cross-sectional cohort study. METHODS: Fasting plasma glucose concentrations, anthropometric variables and blood pressure were measured. Habitual physical activity and aerobic fitness were objectively assessed using uniaxial accelerometry and a maximal aerobic exercise stress test on a bicycle ergometer, respectively. RESULTS: In adjusted models, nominally significant associations were observed for BMI (rs10018239, p = 0.039), waist circumference (rs7656250, p = 0.012; rs8192678 [Gly482Ser], p = 0.015; rs3755863, p = 0.02; rs10018239, beta = -0.01 cm per minor allele copy, p = 0.043), systolic blood pressure (rs2970869, p = 0.018) and fasting glucose concentrations (rs11724368, p = 0.045). Stronger associations were observed for aerobic fitness (rs7656250, p = 0.005; rs13117172, p = 0.008) and fasting glucose concentrations (rs7657071, p = 0.002). None remained significant after correcting for the number of statistical comparisons. We proceeded by testing for gene x physical activity interactions for the polymorphisms that showed nominal evidence of association in the main effect models. None of these tests was statistically significant. CONCLUSIONS/INTERPRETATION: Variants at PPARGC1A may influence several metabolic traits in this European paediatric cohort. However, variation at PPARGC1A is unlikely to have a major impact on cardiovascular or metabolic health in these children.

Chaotic destruction of Anderson localization in a nonlinear lattice

We consider a scattering problem for a nonlinear disordered lattice layer governed by the discrete nonlinear Schrodinger equation. The linear state with exponentially small transparency, due to the Anderson localization, is followed for an increasing nonlinearity, until it is destroyed via a bifurcation. The critical nonlinearity is shown to decay with the lattice length as a power law. We demonstrate that in the chaotic regimes beyond the bifurcation the field is delocalized and this leads to a drastic increase of transparency. Copyright (C) EPLA, 2008

Draft genome sequence of pseudomonas corrugata, a phytopathogenic bacterium with potential industrial applications

Pseudomonas corrugata was first described as the causal agent of a tomato disease called 'pith necrosis' yet it is considered as a biological resource in various fields such as biocontrol of plant diseases and production of industrially promising microbial biopolymers (mcl-PHA). Here we report the first draft genome sequence of this species.

Sequence analysis of a CTX-M-1 IncI1 plasmid found in Salmonella enterica 4,5,12,i:-, Escherichia coli and Klebsiella pneumoniae on a single UK pig farm

OBJECTIVES: In 2009, CTX-M Enterobacteriaceae and Salmonella isolates were recovered from a UK pig farm, prompting studies into the dissemination of the resistance and to establish any relationships between the isolates. METHODS: PFGE was used to elucidate clonal relationships between isolates whilst plasmid profiling, restriction analysis, sequencing and PCR were used to characterize the CTX-M-harbouring plasmids. RESULTS: Escherichia coli, Klebsiella pneumoniae and Salmonella 4,5,12:i:- and Bovismorbificans resistant to cefotaxime (n = 65) were recovered and 63 were shown by PCR to harbour a group 1 CTX-M gene. The harbouring hosts were diverse, but the group 1 CTX-M plasmids were common. Three sequenced CTX-M plasmids from E. coli, K. pneumoniae and Salmonella enterica serotype 4,5,12:i:- were identical except for seven mutations and highly similar to IncI1 plasmid ColIb-P9. Two antimicrobial resistance regions were identified: one inserted upstream of yacABC harbouring ISCR2 transposases, sul2 and floR; and the other inserted within shfB of the pilV shufflon harbouring the ISEcp1 transposase followed by blaCTX-M-1. CONCLUSIONS: These data suggest that an ST108 IncI1 plasmid encoding a blaCTX-M-1 gene had disseminated across multiple genera on this farm, an example of horizontal gene transfer of the blaCTX-M-1 gene.