Size-dependent influence of NOx on the growth rates of organic aerosol particles.

Atmospheric new-particle formation (NPF) affects climate by contributing to a large fraction of the cloud condensation nuclei (CCN). Highly oxygenated organic molecules (HOMs) drive the early particle growth and therefore substantially influence the survival of newly formed particles to CCN. Nitrogen oxide (NOx) is known to suppress the NPF driven by HOMs, but the underlying mechanism remains largely unclear. Here, we examine the response of particle growth to the changes of HOM formation caused by NOx. We show that NOx suppresses particle growth in general, but the suppression is rather nonuniform and size dependent, which can be quantitatively explained by the shifted HOM volatility after adding NOx. By illustrating how NOx affects the early growth of new particles, a critical step of CCN formation, our results help provide a refined assessment of the potential climatic effects caused by the diverse changes of NOx level in forest regions around the globe.

Positioning human body models for crashworthiness using model order reduction.

A new technique to accelerate the positioning of human body models (HBMs) by means of a dimensionality reduction of a database of precomputed simulations is presented. First, a set of important subspace deformation modes which are used to approximate the model's movements observed in the training simulations are computed. In the second step, a convex optimization problem is solved in order to obtain an optimal position of the human body model as described by the user. We apply the proposed method to a new reclined seating position of the Total Human Model for Safety (THUMS, v5).

How much do we know about the frequency of hybridisation and polyploidy in the Mediterranean region?

Natural hybridisation and polyploidy are currently recognised as drivers of biodiversity, despite early scepticism about their importance. The Mediterranean region is a biodiversity hotspot where geological and climatic events have created numerous opportunities for speciation through hybridisation and polyploidy. Still, our knowledge on the frequency of these mechanisms in the region is largely limited, despite both phenomena are frequently cited in studies of Mediterranean plants. We reviewed information available from biodiversity and cytogenetic databases to provide the first estimates of hybridisation and polyploidy frequency in the Mediterranean region. We also inspected the most comprehensive modern Mediterranean Flora (Flora iberica) to survey the frequency and taxonomic distribution of hybrids and polyploids in Iberian Peninsula. We found that <6% of Mediterranean plants were hybrids, although a higher frequency was estimated for the Iberian Peninsula (13%). Hybrids were concentrated in few families and in even fewer genera. The overall frequency of polyploidy (36.5%) was comparable with previous estimates in other regions; however our estimates increased when analysing the Iberian Peninsula (48.8%). A surprisingly high incidence of species harbouring two or more ploidy levels was also observed (21.7%). A review of the available literature also showed that the ecological factors driving emergence and establishment of new entities are still poorly studied in the Mediterranean flora, although geographic barriers seem to play a major role in polyploid complexes. Finally, this study reveals several gaps and limitations in our current knowledge about the frequency of hybridisation and polyploidy in the Mediterranean region. The obtained estimates might change in the future with the increasing number of studies; still, rather than setting the complete reality, we hope that this work triggers future studies on hybridisation and polyploidy in the Mediterranean region.

Structures of riboswitch RNA reaction states by mix-and-inject XFEL serial crystallography.

Riboswitches are structural RNA elements that are generally located in the 5' untranslated region of messenger RNA. During regulation of gene expression, ligand binding to the aptamer domain of a riboswitch triggers a signal to the downstream expression platform. A complete understanding of the structural basis of this mechanism requires the ability to study structural changes over time. Here we use femtosecond X-ray free electron laser (XFEL) pulses to obtain structural measurements from crystals so small that diffusion of a ligand can be timed to initiate a reaction before diffraction. We demonstrate this approach by determining four structures of the adenine riboswitch aptamer domain during the course of a reaction, involving two unbound apo structures, one ligand-bound intermediate, and the final ligand-bound conformation. These structures support a reaction mechanism model with at least four states and illustrate the structural basis of signal transmission. The three-way junction and the P1 switch helix of the two apo conformers are notably different from those in the ligand-bound conformation. Our time-resolved crystallographic measurements with a 10-second delay captured the structure of an intermediate with changes in the binding pocket that accommodate the ligand. With at least a 10-minute delay, the RNA molecules were fully converted to the ligand-bound state, in which the substantial conformational changes resulted in conversion of the space group. Such notable changes in crystallo highlight the important opportunities that micro- and nanocrystals may offer in these and similar time-resolved diffraction studies. Together, these results demonstrate the potential of 'mix-and-inject' time-resolved serial crystallography to study biochemically important interactions between biomacromolecules and ligands, including those that involve large conformational changes.

Past climate changes facilitated homoploid speciation in three mountain spiny fescues (Festuca, Poaceae).

Apart from the overwhelming cases of allopolyploidization, the impact of speciation through homoploid hybridization is becoming more relevant than previously thought. Much less is known, however, about the impact of climate changes as a driven factor of speciation. To investigate these issues, we selected Festuca picoeuropeana, an hypothetical natural hybrid between the diploid species F. eskia and F. gautieri that occurs in two different mountain ranges (Cantabrian Mountains and Pyrenees) separated by more than 400 km. To unravel the outcomes of this mode of speciation and the impact of climate during speciation we used a multidisciplinary approach combining genome size and chromosome counts, data from an extensive nuclear genotypic analysis, plastid sequences and ecological niche models (ENM). Our results show that the same homoploid hybrid was originated independently in the two mountain ranges, being currently isolated from both parents and producing viable seeds. Parental species had the opportunity to contact as early as 21000 years ago although niche divergence occurs nowadays as result of a climate-driven shift. A high degree of niche divergence was observed between the hybrid and its parents and no recent introgression or backcrossed hybrids were detected, supporting the current presence of reproductive isolation barriers between these species.

Facts and misfits in ultrasound therapy: steps to improve your treatment outcomes.

Ultrasound has been used as a therapeutic agent for decades. Unfortunately many treatments are ineffective because the wrong treatment parameters are used. In this paper I present much of my work over the past 20 years with the goal of improving ultrasound use. Seven steps to optimal ultrasound use are presented. These include: treatment size area; movement speed of the transducer; and the "stretching window".

Injuries restored to ROM using PSWD and mobilizations.

The purpose of this paper is to report the cases of 7 subjects who lacked full range of motion (ROM) in the elbow, and my treatment regimen of PSWD and joint mobilizations. 7 subjects presented with decreased elbow (ROM) due to extensive fractures and or dislocations from traumatic injuries. All subjects were post-surgical, 2 with internal fixation devices. Initial active extension for each patient was 17°, 23°, 28°, 25°, 45°, 30°, 26° respectively (range of 17-45 degrees). Treatment regimen consisted of PSWD to the anterior elbow for 20 min at 48 watts. Immediately after PSWD, mobilizations were administered to the elbow. Extension improved 19°, 21°, 25°, 23°, 20°, 30°, and 26°, respectively, after 6 or less treatments (range of 19-30 degrees). All but 1 patient returned to normal activities with functional ROM in all planes. Follow-up 4 weeks later indicated that all of the subjects maintained 85-100% of their extension. No negative effects were reported during the short-term follow-up. When precautions are taken, I propose PSWD (48 W) may be an appropriate adjunct to joint mobilizations to increase ROM in peripheral joints despite implanted metal.

Rapidly cooled horse spermatozoa: loss of viability is due to osmotic imbalance during thawing, not intracellular ice formation.

The cellular damage that spermatozoa encounter at rapid rates of cooling has often been attributed to the formation of intracellular ice. However, no direct evidence of intracellular ice has been presented. An alternative mechanism has been proposed by Morris (2006) that cell damage is a result of an osmotic imbalance encountered during thawing. This paper examines whether intracellular ice forms during rapid cooling or if an alternative mechanism is present. Horse spermatozoa were cooled at a range of cooling rates from 0.3 to 3,000 degrees C/min in the presence of a cryoprotectant. The ultrastructure of the samples was examined by Cryo Scanning Electron Microscopy (CryoSEM) and freeze substitution, to determine whether intracellular ice formed and to examine alternative mechanisms of cell injury during rapid cooling. No intracellular ice formation was detected at any cooling rate. Differential scanning Calorimetry (DSC) was employed to examine the amount of ice formed at different rate of cooling. It is concluded that cell damage to horse spermatozoa, at cooling rates of up to 3,000 degrees C/min, is not caused by intracellular ice formation. Spermatozoa that have been cooled at high rates are subjected to an osmotic shock when they are thawed.

Cryopreservation of horse semen under laboratory and field conditions using a Stirling Cycle freezer.

A Stirling Cycle freezer has been developed as an alternative to conventional liquid nitrogen controlled rate freezers. Horse semen samples were cooled in 0.25 ml straws and 15 ml bags in the Stirling Cycle freezer under laboratory conditions and as a portable device, powered from a car battery. For comparison, straws were frozen in a conventional liquid nitrogen controlled rate freezer. Upon thawing, motility and viability of samples frozen in the Stirling Cycle freezer were not significantly different when compared to samples frozen in the liquid nitrogen freezer. Unlike liquid nitrogen systems, the Stirling Cycle freezer does not pose a contamination risk, can be used in sterile conditions and has no need for a constant supply of cryogen. The freezer has potential for use in veterinary and genetic conservation applications.

Effectiveness of using a mat filled with a peroxygen disinfectant to minimize shoe sole contamination in a veterinary hospital.

OBJECTIVE: To determine the effectiveness of using a disinfectant mat filled with a peroxygen compound to prevent mechanical transmission of bacteria via contaminated footwear between the food animal ward and common breezeway of a veterinary teaching hospital. DESIGN: Observational study. SAMPLE POPULATION: Shoe soles of individuals entering and exiting from the ward. PROCEDURES: A mat filled with peroxygen disinfectant was placed at the entrance to the food animal ward, and participants wiped each shoe twice on the mat surface (n = 16) or walked on the mat surface but did not wipe their shoes (17) before entering and exiting from the ward. Swab specimens were collected from the shoe soles of participants before and after mat use and submitted for bacterial culture. RESULTS: For both study days, as participants entered the ward, median number of aerobic bacteria isolated from shoe swab specimens collected prior to use of the disinfectant mat was not significantly different from median number isolated after use of the disinfectant mat. However, as participants exited the ward, median number of aerobic bacteria isolated from shoe swab specimens collected prior to use of the disinfectant mat was significantly higher than median number isolated after use of the disinfectant mat. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that placing a mat filled with a peroxygen disinfectant at the exit from the food animal ward of a veterinary teaching hospital may help reduce mechanical transmission of bacteria on the footwear of individuals leaving the ward.

A thermodynamic framework for Mg2+ binding to RNA.

We present a model describing how Mg(2+) binds and stabilizes specific RNA structures. In this model, RNA stabilization arises from two energetically distinct modes of Mg(2+) binding: diffuse- and site-binding. Diffusely bound Mg(2+) are electrostatically attracted to the strong anionic field around the RNA and are accurately described by the Poisson-Boltzmann equation as an ensemble distributed according to the electrostatic potentials around the nucleic acid. Site-bound Mg(2+) are strongly attracted to specifically arranged electronegative ligands that desolvate the ion and the RNA binding site. Thus, site-binding is a competition between the strong coulombic attraction and the large cost of desolvating the ion and its binding pocket. By using this framework, we analyze three systems where a single site-bound Mg(2+) may be important for stability: the P5 helix and the P5b stem loop from the P4-P6 domain of the Tetrahymena thermophila group I intron and a 58-nt fragment of the Escherichia coli 23S ribosomal RNA. Diffusely bound Mg(2+) play a dominant role in stabilizing these RNA structures. These ions stabilize the folded structures, in part, by accumulating in regions of high negative electrostatic potential. These regions of Mg(2+) localization correspond to ions that are observed in the x-ray crystallographic and NMR structures of the RNA. In contrast, the contribution of site-binding to RNA stability is often quite small because of the large desolvation penalty. However, in special cases, site-binding of partially dehydrated Mg(2+) to locations with extraordinarily high electrostatic potential can also help stabilize folded RNA structures.

Translational repression of the Escherichia coli alpha operon mRNA: importance of an mRNA conformational switch and a ternary entrapment complex.

Ribosomal protein S4 represses synthesis of the four ribosomal proteins (including itself) in the Escherichia coli alpha operon by binding to a nested pseudoknot structure that spans the ribosome binding site. A model for the repression mechanism previously proposed two unusual features: (i) the mRNA switches between conformations that are "active" or "inactive" in translation, with S4 as an allosteric effector of the inactive form, and (ii) S4 holds the 30 S subunit in an unproductive complex on the mRNA ("entrapment"), in contrast to direct competition between repressor and ribosome binding ("displacement"). These two key points have been experimentally tested. First, it is found that the mRNA pseudoknot exists in an equilibrium between two conformers with different electrophoretic mobilities. S4 selectively binds to one form of the RNA, as predicted for an allosteric effector; binding of ribosomal 30 S subunits is nearly equal in the two forms. Second, we have used S4 labeled at a unique cysteine with either of two fluorophores to characterize its interactions with mRNA and 30 S subunits. Equilibrium experiments detect the formation of a specific ternary complex of S4, mRNA pseudoknot, and 30 S subunits. The existence of this ternary complex is unambiguous evidence for translational repression of the alpha operon by an entrapment mechanism.

Recognition of 16S rRNA by ribosomal protein S4 from Bacillus stearothermophilus.

Protein S4 is essential for bacterial small ribosomal subunit assembly and recognizes the 5' domain (approximately 500 nt) of small subunit rRNA. This study characterizes the thermodynamics of forming the S4-5' domain rRNA complex from a thermophile, Bacillus stearothermophilus, and points out unexpected differences from the homologous Escherichia coli complex. Upon incubation of the protein and RNA at temperatures between 35 and 50 degrees C under ribosome reconstitution conditions [350 mM KCl, 8 mM MgCl2, and 30 mM Tris (pH 7.5)], a complex with an association constant of > or = 10(9) M(-1) was observed, more than an order of magnitude tighter than previously found for the homologous E. coli complex under similar conditions. This high-affinity complex was shown to be stoichiometric, in equilibrium, and formed at rates on the order of magnitude expected for diffusion-controlled reactions ( approximately 10(7) M(-1) x s(-1)), though at low temperatures the complex became kinetically trapped. Heterologous binding experiments with E. coli S4 and 5' domain RNA suggest that it is the B. stearothermophilus S4, not the rRNA, that is activated by higher temperatures; the E. coli S4 is able to bind 5' domain rRNA equally well at 0 and 37 degrees C. Tight complex formation requires a low Mg ion concentration (1-2 mM) and is very sensitive to KCl concentration [- partial differential[log(K)]/partial differential(log[KCl]) = 9.3]. The protein has an unusually strong nonspecific binding affinity of 3-5 x 10(6) M(-1), detected as a binding of one or two additional proteins to the target 5' domain RNA or two to three proteins binding a noncognate 23S rRNA fragment of the approximately same size. This binding is not as sensitive to monovalent ion concentration [- partial differential[log(K)]/partial differential(log[KCl]) = 6.3] as specific binding and does not require Mg ion. These findings are consistent with S4 stabilizing a compact form of the rRNA 5' domain.

Thermal methods for the analysis of RNA folding pathways.

Once a model of the secondary structure of an RNA has been deduced, thermal melting analysis can be used to determine whether the model accounts for all intramolecular interactions of the RNA, or whether noncanonical and tertiary interactions make the structure more stable than predicted, or link parts of the structure in unexpected ways. It is also useful to determine the pH, salt, and temperature ranges under which the RNA adopts a stably folded structure, or to analyze unfolding pathways. This unit discusses sample preparation, instrumentation, and theoretical background. It also provide a sample analysis of tRNA unfolding.

Preterm premature rupture of membranes without labor is not associated with increased levels of matrix metalloproteinase-9 protein.

OBJECTIVES: Preterm premature rupture of membranes (PROM) accounts for 30-40% of all preterm births. The objectives of this study were to determine whether matrix metalloproteinase-9 (MMP-9) is increased in preterm PROM fetal membranes, whether labor or gestational age affects expression, and whether the increase is localized to the rupture site or is membrane-wide. METHODS: Fetal membranes were collected from 15 pregnancies complicated by preterm PROM and 26 control cases, which delivered at term or preterm without PROM. The preterm PROM cases represented both patients who labored and those who did not. Membrane samples at the rupture site and a remote site (approximately > 5 cm) were analyzed for MMP-9 protein and enzymatic activity by Western blot and gelatin zymography, respectively. RESULTS: MMP-9 levels in fetal membranes were similar at both the rupture and the remote sites. The highest levels of total MMP-9 protein were found in preterm PROM patients with labor (p < 0.05) and were increased four-fold over protein levels in non-laboring preterm PROM patients delivered by Cesarean section (p < 0.001). In preterm PROM patients without labor, levels of MMP-9 protein were similar to those of non-laboring patients at term and preterm. Zymography correlated with protein results in all membranes. CONCLUSIONS: Preterm PROM without labor is not associated with increased membrane levels of MMP-9 protein, suggesting that its local elevation does not play a role in early membrane rupture.

Levels of vaginal secretory leukocyte protease inhibitor are decreased in women with lower reproductive tract infections.

OBJECTIVE: Secretory leukocyte protease inhibitor contributes resistance to primary human immunodeficiency virus infection in the oral cavity. However, the levels of this inhibitor in the genital tract of women with sexually transmitted diseases or vaginitis are not well described. The objective was to determine vaginal inhibitor levels in women with symptomatic and asymptomatic genital infections. STUDY DESIGN: We tested 207 nonpregnant women for Neisseria gonorrhoeae, Trichomonas vaginalis, Chlamydia trachomatis, Candida species, and bacterial vaginosis by standard methods. A second group of symptom-free pregnant women (N = 231) was also studied. Secretory leukocyte protease inhibitor was measured by enzyme-linked immunosorbent assay, and results were compared by nonparametric methods. RESULTS: Vaginal levels of secretory leukocyte protease inhibitor in both groups were significantly lower in women with any sexually transmitted disease than in those without infection (P<.0001). Patients with bacterial vaginosis and those with bacterial vaginosis with yeast vaginitis also had decreased levels (P<.025). CONCLUSIONS: Levels of secretory leukocyte protease inhibitor in vaginal fluid are decreased in women with lower genital tract infection. This may represent a common mechanism of increasing susceptibility to infection with human immunodeficiency virus.

Structural preordering in the N-terminal region of ribosomal protein S4 revealed by heteronuclear NMR spectroscopy.

Protein S4, a component of the 30S subunit of the prokaryotic ribosome, is one of the first proteins to interact with rRNA in the process of ribosome assembly and is known to be involved in the regulation of this process. While the structure of the C-terminal 158 residues of Bacillus stearothermophilus S4 has been solved by both X-ray crystallography and NMR, that of the N-terminal 41 residues is unknown. Evidence suggests that the N-terminus is necessary both for the assembly of functional ribosomes and for full binding to 16S RNA, and so we present NMR data collected on the full-length protein (200 aa). Our data indicate that the addition of the N-terminal residues does not significantly change the structure of the C-terminal 158 residues. The data further indicate that the N-terminus is highly flexible in solution, without discernible secondary structure. Nevertheless, structure calculations based on nuclear Overhauser effect spectroscopic data combined with (15)N relaxation data revealed that two short segments in the N-terminus, S(12)RRL(15) and P(30)YPP(33), adopt transiently ordered states in solution. The major conformation of S(12)RRL(15) appears to orient the arginine side chains outward toward the solvent in a parallel fashion, while that of P(30)YPP(33) forms a nascent turn of a polyproline II helix. These segments contain residues that are highly conserved across many prokaryotic species, and thus they are reasonable candidates respectively for sites of interaction with RNA and other ribosomal proteins within the intact ribosome.

Case-controlled study of nursing home residents referred for treatment of vocally disruptive behavior.

The aim of this study was to identify factors associated with vocally disruptive behavior (VDB) in nursing home patients referred to aged care services for treatment, using a case-control methodology. Characteristics of the VDB, reasons for referral, perceived causal factors, and psychotropic use were noted. Twenty-five subjects and controls were examined with the Screaming Behavior Mapping Instrument, the Cornell Scale for Depression in Dementia, the Dementia Behavior Disturbance Scale, and measures of cognition, functional capacity, social activities, and emotional reactions of nursing staff. VDB was associated with other disturbed behaviors, depression, anxiety, severe dementia, functional impairment, communication difficulties, use of psychotropic medication, social isolation, and emotional distress in the nursing staff. Reasons for referral may relate more to the stress experienced by nursing home staff in managing VDB than to specific attributes of the VDB itself.