Novel transient cytoplasmic rings stabilize assembling bacterial flagellar motors.

The process by which bacterial cells build their intricate flagellar motility apparatuses has long fascinated scientists. Our understanding of this process comes mainly from studies of purified flagella from two species, Escherichia coli and Salmonella enterica. Here, we used electron cryo-tomography (cryo-ET) to image the assembly of the flagellar motor in situ in diverse Proteobacteria: Hylemonella gracilis, Helicobacter pylori, Campylobacter jejuni, Pseudomonas aeruginosa, Pseudomonas fluorescens, and Shewanella oneidensis. Our results reveal the in situ structures of flagellar intermediates, beginning with the earliest flagellar type III secretion system core complex (fT3SScc) and MS-ring. In high-torque motors of Beta-, Gamma-, and Epsilon-proteobacteria, we discovered novel cytoplasmic rings that interact with the cytoplasmic torque ring formed by FliG. These rings, associated with the MS-ring, assemble very early and persist until the stators are recruited into their periplasmic ring; in their absence the stator ring does not assemble. By imaging mutants in Helicobacter pylori, we found that the fT3SScc proteins FliO and FliQ are required for the assembly of these novel cytoplasmic rings. Our results show that rather than a simple accretion of components, flagellar motor assembly is a dynamic process in which accessory components interact transiently to assist in building the complex nanomachine.

Manipulating chiral spin transport with ferroelectric polarization.

A magnon is a collective excitation of the spin structure in a magnetic insulator and can transmit spin angular momentum with negligible dissipation. This quantum of a spin wave has always been manipulated through magnetic dipoles (that is, by breaking time-reversal symmetry). Here we report the experimental observation of chiral spin transport in multiferroic BiFeO3 and its control by reversing the ferroelectric polarization (that is, by breaking spatial inversion symmetry). The ferroelectrically controlled magnons show up to 18% modulation at room temperature. The spin torque that the magnons in BiFeO3 carry can be used to efficiently switch the magnetization of adjacent magnets, with a spin-torque efficiency comparable to the spin Hall effect in heavy metals. Utilizing such controllable magnon generation and transmission in BiFeO3, an all-oxide, energy-scalable logic is demonstrated composed of spin-orbit injection, detection and magnetoelectric control. Our observations open a new chapter of multiferroic magnons and pave another path towards low-dissipation nanoelectronics.

In situ imaging of the bacterial flagellar motor disassembly and assembly processes.

The self-assembly of cellular macromolecular machines such as the bacterial flagellar motor requires the spatio-temporal synchronization of gene expression with proper protein localization and association of dozens of protein components. In Salmonella and Escherichia coli, a sequential, outward assembly mechanism has been proposed for the flagellar motor starting from the inner membrane, with the addition of each new component stabilizing the previous one. However, very little is known about flagellar disassembly. Here, using electron cryo-tomography and sub-tomogram averaging of intact Legionella pneumophila, Pseudomonas aeruginosa, and Shewanella oneidensis cells, we study flagellar motor disassembly and assembly in situ. We first show that motor disassembly results in stable outer membrane-embedded sub-complexes. These sub-complexes consist of the periplasmic embellished P- and L-rings, and bend the membrane inward while it remains apparently sealed. Additionally, we also observe various intermediates of the assembly process including an inner-membrane sub-complex consisting of the C-ring, MS-ring, and export apparatus. Finally, we show that the L-ring is responsible for reshaping the outer membrane, a crucial step in the flagellar assembly process.

Voltage-Controlled High-Bandwidth Terahertz Oscillators Based on Antiferromagnets.

Producing compact voltage-controlled frequency generators and sensors operating in the terahertz (THz) regime represents a major technological challenge. Here, we show that noncollinear antiferromagnets (NCAFMs) with kagome structure host gapless self-oscillations whose frequencies are tunable from 0 Hz to the THz regime via electrically induced spin-orbit torques (SOTs). The auto-oscillations' initiation, bandwidth, and amplitude are investigated by deriving an effective theory, which captures the reactive and dissipative SOTs. We find that the dynamics strongly depends on the ground state's chirality, with one chirality having gapped excitations, whereas the opposite chirality provides gapless self-oscillations. Our results reveal that NCAFMs offer unique THz functional components, which could play a significant role in filling the THz technology gap.

Ultra-sensitive voltage-controlled skyrmion-based spintronic diode.

We have designed a passive spintronic diode based on a single skyrmion stabilized in a magnetic tunnel junction and studied its dynamics induced by voltage-controlled magnetic anisotropy (VCMA) and Dzyaloshinskii-Moriya interaction (VDMI). We have demonstrated that the sensitivity (rectified output voltage over input microwave power) with realistic physical parameters and geometry can be larger than 10 kV W-1which is one order of magnitude larger than diodes employing a uniform ferromagnetic state. Our numerical and analytical results on the VCMA and VDMI-driven resonant excitation of skyrmions beyond the linear regime reveal a frequency dependence on the amplitude and no efficient parametric resonance. Skyrmions with a smaller radius produced higher sensitivities, demonstrating the efficient scalability of skyrmion-based spintronic diodes. These results pave the way for designing passive ultra-sensitive and energy efficient skyrmion-based microwave detectors.

Bacterial flagellar motor PL-ring disassembly subcomplexes are widespread and ancient.

The bacterial flagellum is an amazing nanomachine. Understanding how such complex structures arose is crucial to our understanding of cellular evolution. We and others recently reported that in several Gammaproteobacterial species, a relic subcomplex comprising the decorated P and L rings persists in the outer membrane after flagellum disassembly. Imaging nine additional species with cryo-electron tomography, here, we show that this subcomplex persists after flagellum disassembly in other phyla as well. Bioinformatic analyses fail to show evidence of any recent horizontal transfers of the P- and L-ring genes, suggesting that this subcomplex and its persistence is an ancient and conserved feature of the flagellar motor. We hypothesize that one function of the P and L rings is to seal the outer membrane after motor disassembly.

Periodontal bacterial species in hopeless dentitions with severe periodontitis: comparison of levels before extraction and 90 days after immediate implant placement.

The aim of this study was to evaluate changes in periodontal bacterial species during the transition from hopeless teeth to denture-supporting immediate implants. Biofilm and saliva samples were collected from 13 women and 7 men before the extraction of hopeless teeth with severe periodontitis (baseline) and 90 days after the placement of immediate implants that supported immediately loaded complete dentures (day 90). The levels of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, and Streptococcus oralis were analyzed by real-time polymerase chain reaction. Differences in the levels of bacterial species in the subgingival biofilm and saliva and between baseline and day 90 were evaluated by a 2-way analysis of variance followed by the Tukey test. There was a significant reduction in the levels of T forsythia from baseline to day 90 in saliva and subgingival biofilms (P < 0.05) and a tendency toward a reduction of the other bacterial species. The total bacterial load was higher in saliva than in subgingival biofilm at baseline and day 90 (P < 0.05), while the individual levels of all species were higher in the biofilm than in saliva at both times (P < 0.05). The results showed an overall reduction in the levels of pathogenic bacterial species, particularly T forsythia, during the transition from hopeless dentition to implant-supported dentures. The subgingival biofilm harbored considerable levels of pathogenic species, suggesting that implant placement immediately after extraction of teeth with severe periodontitis may induce changes that favor colonization by pathogenic microorganisms.

Effects and response of the Cerrado ground-layer to frost along the canopy cover gradient.

Frost effects on savanna plant communities have been considered as analogous to those from fire, both changing community structure and filtering species composition. However, while frost impacts have been well-studied for the woody component of savannas, it is still poorly explored for the ground-layer community. Here, we investigated effects of frost in the Cerrado along a gradient of tree cover, focusing on ground-layer plant species, near the southern limit of the Cerrado in Brazil. We aimed to elucidate if the pattern already described for the tree layer also extends to the ground layer in terms of mimicking the effects of fire on vegetation structure and composition. We assessed how damage severity differs across species and across the tree-cover gradient, and we examined the recovery process after frost in terms of richness and community structure along the canopy cover gradient. Frost caused immediate and widespread dieback of the perennial ground-layer, with greatest impact on community structure where tree cover was lowest. However, frost did not reduce the number of species, indicating community resilience to this natural disturbance. Although frost mimicked the effects of fire in some ways, in other ways it differed substantially from fire. Unlike fire, frost increases litter cover and decreases the proportion of bare soil, likely hindering crucial processes for recovery of plant populations, such as seed dispersal, seed germination and plant resprouting. This finding calls attention to the risk of misguided conclusions when the ground layer is neglected in ecological studies of tropical savannas and grasslands.

MiST 3.0: an updated microbial signal transduction database with an emphasis on chemosensory systems.

Bacteria and archaea employ dedicated signal transduction systems that modulate gene expression, second-messenger turnover, quorum sensing, biofilm formation, motility, host-pathogen and beneficial interactions. The updated MiST database provides a comprehensive classification of microbial signal transduction systems. This update is a result of a substantial scaling to accommodate constantly growing microbial genomic data. More than 125 000 genomes, 516 million genes and almost 100 million unique protein sequences are currently stored in the database. For each bacterial and archaeal genome, MiST 3.0 provides a complete signal transduction profile, thus facilitating theoretical and experimental studies on signal transduction and gene regulation. New software infrastructure and distributed pipeline implemented in MiST 3.0 enable regular genome updates based on the NCBI RefSeq database. A novel MiST feature is the integration of unique profile HMMs to link complex chemosensory systems with corresponding chemoreceptors in bacterial and archaeal genomes. The data can be explored online or via RESTful API (freely available at https://mistdb.com).

The chemosensory systems of Vibrio cholerae.

Vibrio cholerae, the causative agent of the acute diarrheal disease cholera, is able to thrive in diverse habitats such as natural water bodies and inside human hosts. To ensure their survival, these bacteria rely on chemosensory pathways to sense and respond to changing environmental conditions. These pathways constitute a highly sophisticated cellular control system in Bacteria and Archaea. Reflecting the complex life cycle of V. cholerae, this organism has three different chemosensory pathways that together contain over 50 proteins expressed under different environmental conditions. Only one of them is known to control motility, while the function of the other two remains to be discovered. Here, we provide an overview of the chemosensory systems in V. cholerae and the advances toward understanding their structure and function.

The distinct roles of water table depth and soil properties in controlling alternative woodland-grassland states in the Cerrado.

Open grassy vegetation and forests share riparian zones across the Neotropical savannas, characterizing alternative stable states. However, factors determining the occurrence and maintenance of each vegetation type are yet to be elucidated. To disentangle the role of environmental factors (soil properties and groundwater depth) constraining tree colonization of wet grasslands in the Cerrado, we assessed tree establishment during the early seedling and sapling stages and the influence of these factors on leaf gas exchange and leaf water potential of tree saplings. Three functionally distinct tree species were studied: (1) flood-tolerant species characteristic of gallery forests, (2) flood-intolerant species characteristic of seasonally dry savannas, and (3) generalist species found in both gallery forests and seasonally dry savannas. Savanna species was constrained by waterlogging, especially at the sapling stage, with restricted stomatal conductance and leaf water potential, resulting in low carbon assimilation, decreased plant size, and high mortality (above 80%). The gallery forest and the generalist species, however, were able to colonize the wet grasslands and survive, despite the low seedling emergence (below 30%) and sapling growth constrained by low gas exchange rates. Soil waterlogging is, therefore, an effective environmental filter that prevents savanna trees from expanding over wet grasslands. However, colonization by trees adapted to a shallow water table cannot be constrained by this or other soil properties, turning the wet grasslands dependent on natural disturbances to persist as an alternative state, sharing the waterlogged environments with the gallery forests in the Cerrado region.

Structure of the fission yeast actomyosin ring during constriction.

Cell division in many eukaryotes is driven by a ring containing actin and myosin. While much is known about the main proteins involved, the precise arrangement of actin filaments within the contractile machinery, and how force is transmitted to the membrane, remains unclear. Here we use cryosectioning and cryofocused ion beam milling to gain access to cryopreserved actomyosin rings in Schizosaccharomyces pombe for direct 3D imaging by electron cryotomography. Our results show that straight, overlapping actin filaments, running nearly parallel to each other and to the membrane, form a loose bundle of ∼150 nm in diameter that "saddles" the inward-bending membrane at the leading edge of the division septum. The filaments do not make direct contact with the membrane. Our analysis of the actin filaments reveals the variability in filament number, nearest-neighbor distances between filaments within the bundle, their distance from the membrane, and angular distribution with respect to the membrane.

Periodontal therapy on the oral health-related quality of life of obese and non-obese individuals.

Oral diseases such as periodontitis can have a more negative influence on the quality of life of obese than in normal-weight patients. The objective of the present study was to assess the impact of one-stage full-mouth disinfection (OSFMD) therapy on the oral health-related quality of life (OHRQL) of obese and non-obese individuals with periodontitis. Fifty-five obese and thirty-nine non-obese patients were evaluated. The questionnaires oral impacts on daily performance (OIDP) and oral health and quality of life (OHQoL) were given to all patients at baseline and 6 months after periodontal treatment by the OSFMD protocol. For statistical analysis, Chi-square, the two-factor repeated-measures ANOVA, and correlation tests were used. At baseline, mean global OHQoL and OIDP scores were similar for both groups (p > 0.05). At 6 months, OSFMD resulted in OHQoL and OIDP global scores improvements in both groups (p < 0.05), with no significant difference between groups. The most impaired activity at baseline was eating and cleaning teeth for both groups. Periodontal parameters were associated with worse values in the OHQoL and OIDP questionnaires only in obese patients. In conclusion, OSFMD yielded similar improvements in overall OHRQL in both obese and non-obese individuals. Periodontal parameters were associated with a worse quality of life in obese patients. Periodontal treatment can be an important component to improve the OHRQL of obese individuals, and clinicians should expect similar results as those obtained with non-obese patients.

Flammability thresholds or flammability gradients? Determinants of fire across savanna-forest transitions.

Vegetation-fire feedbacks are important for determining the distribution of forest and savanna. To understand how vegetation structure controls these feedbacks, we quantified flammability across gradients of tree density from grassland to forest in the Brazilian Cerrado. We experimentally burned 102 plots, for which we measured vegetation structure, fuels, microclimate, ignition success and fire behavior. Tree density had strong negative effects on ignition success, rate of spread, fire-line intensity and flame height. Declining grass biomass was the principal cause of this decline in flammability as tree density increased, but increasing fuel moisture contributed. Although the response of flammability to tree cover often is portrayed as an abrupt, largely invariant threshold, we found the response to be gradual, with considerable variability driven largely by temporal changes in atmospheric humidity. Even when accounting for humidity, flammability at intermediate tree densities cannot be predicted reliably. Fire spread in savanna-forest mosaics is not as deterministic as often assumed, but may appear so where vegetation boundaries are already sharp. Where transitions are diffuse, fire spread is difficult to predict, but should become increasingly predictable over multiple fire cycles, as boundaries are progressively sharpened until flammability appears to respond in a threshold-like manner.

Assigning chemoreceptors to chemosensory pathways in Pseudomonas aeruginosa.

In contrast to Escherichia coli, a model organism for chemotaxis that has 5 chemoreceptors and a single chemosensory pathway, Pseudomonas aeruginosa PAO1 has a much more complex chemosensory network, which consists of 26 chemoreceptors feeding into four chemosensory pathways. While several chemoreceptors were rigorously linked to specific pathways in a series of experimental studies, for most of them this information is not available. Thus, we addressed the problem computationally. Protein-protein interaction network prediction, coexpression data mining, and phylogenetic profiling all produced incomplete and uncertain assignments of chemoreceptors to pathways. However, comparative sequence analysis specifically targeting chemoreceptor regions involved in pathway interactions revealed conserved sequence patterns that enabled us to unambiguously link all 26 chemoreceptors to four pathways. Placing computational evidence in the context of experimental data allowed us to conclude that three chemosensory pathways in P. aeruginosa utilize one chemoreceptor per pathway, whereas the fourth pathway, which is the main system controlling chemotaxis, utilizes the other 23 chemoreceptors. Our results show that while only a very few amino acid positions in receptors, kinases, and adaptors determine their pathway specificity, assigning receptors to pathways computationally is possible. This requires substantial knowledge about interacting partners on a molecular level and focusing comparative sequence analysis on the pathway-specific regions. This general principle should be applicable to resolving many other receptor-pathway interactions.

In vivo structures of an intact type VI secretion system revealed by electron cryotomography.

The type VI secretion system (T6SS) is a versatile molecular weapon used by many bacteria against eukaryotic hosts or prokaryotic competitors. It consists of a cytoplasmic bacteriophage tail-like structure anchored in the bacterial cell envelope via a cytoplasmic baseplate and a periplasmic membrane complex. Rapid contraction of the sheath in the bacteriophage tail-like structure propels an inner tube/spike complex through the target cell envelope to deliver effectors. While structures of purified contracted sheath and purified membrane complex have been solved, because sheaths contract upon cell lysis and purification, no structure is available for the extended sheath. Structural information about the baseplate is also lacking. Here, we use electron cryotomography to directly visualize intact T6SS structures inside Myxococcus xanthus cells. Using sub-tomogram averaging, we resolve the structure of the extended sheath and membrane-associated components including the baseplate. Moreover, we identify novel extracellular bacteriophage tail fiber-like antennae. These results provide new structural insights into how the extended sheath prevents premature disassembly and how this sophisticated machine may recognize targets.

Chemotaxis cluster 1 proteins form cytoplasmic arrays in Vibrio cholerae and are stabilized by a double signaling domain receptor DosM.

Nearly all motile bacterial cells use a highly sensitive and adaptable sensory system to detect changes in nutrient concentrations in the environment and guide their movements toward attractants and away from repellents. The best-studied bacterial chemoreceptor arrays are membrane-bound. Many motile bacteria contain one or more additional, sometimes purely cytoplasmic, chemoreceptor systems. Vibrio cholerae contains three chemotaxis clusters (I, II, and III). Here, using electron cryotomography, we explore V. cholerae's cytoplasmic chemoreceptor array and establish that it is formed by proteins from cluster I. We further identify a chemoreceptor with an unusual domain architecture, DosM, which is essential for formation of the cytoplasmic arrays. DosM contains two signaling domains and spans the two-layered cytoplasmic arrays. Finally, we present evidence suggesting that this type of receptor is important for the structural stability of the cytoplasmic array.

Transovarial transmission of dengue 1 virus in Aedes aegypti larvae: real-time PCR analysis in a Brazilian city with high mosquito population density.

Transovarial transmission is among the reported factors able to influence environmental maintenance of dengue virus (DENV). Endemic areas with active transmission of dengue are suitable for studying transovarial transmission. Brazil is a country where dengue is endemic and where DENV-1 is the most common disease-related virus serotype. This study aimed to identify transovarial transmission of DENV-1 in Aedes aegypti larvae by reverse-transcriptase nested real-time polymerase chain reaction. Between March and October 2016, Culicidae larvae were collected using traps in 3 locations in Taubaté, São Paulo, Brazil, which has a high occurrence of dengue. The collected larvae were sacrificed in the 3rd or 4th larval stage, classified, and stored at -20 °C. The A. aegypti larvae samples (n = 910) were separated into 91 pools of 10 specimens each from which RNA was extracted, reverse transcribed into cDNA, and analyzed by nested qPCR. None of the pools tested positive for DENV-1. Due to the absence of detectable virus in the evaluated samples, we concluded that transovarial transmission may not be the primary mechanism for maintenance of DENV-1 in this particular environment.

Evolutionary Genomics Suggests That CheV Is an Additional Adaptor for Accommodating Specific Chemoreceptors within the Chemotaxis Signaling Complex.

Escherichia coli and Salmonella enterica are models for many experiments in molecular biology including chemotaxis, and most of the results obtained with one organism have been generalized to another. While most components of the chemotaxis pathway are strongly conserved between the two species, Salmonella genomes contain some chemoreceptors and an additional protein, CheV, that are not found in E. coli. The role of CheV was examined in distantly related species Bacillus subtilis and Helicobacter pylori, but its role in bacterial chemotaxis is still not well understood. We tested a hypothesis that in enterobacteria CheV functions as an additional adaptor linking the CheA kinase to certain types of chemoreceptors that cannot be effectively accommodated by the universal adaptor CheW. Phylogenetic profiling, genomic context and comparative protein sequence analyses suggested that CheV interacts with specific domains of CheA and chemoreceptors from an orthologous group exemplified by the Salmonella McpC protein. Structural consideration of the conservation patterns suggests that CheV and CheW share the same binding spot on the chemoreceptor structure, but have some affinity bias towards chemoreceptors from different orthologous groups. Finally, published experimental results and data newly obtained via comparative genomics support the idea that CheV functions as a "phosphate sink" possibly to off-set the over-stimulation of the kinase by certain types of chemoreceptors. Overall, our results strongly suggest that CheV is an additional adaptor for accommodating specific chemoreceptors within the chemotaxis signaling complex.

CDvist: a webserver for identification and visualization of conserved domains in protein sequences.

SUMMARY: Identification of domains in protein sequences allows their assigning to biological functions. Several webservers exist for identification of protein domains using similarity searches against various databases of protein domain models. However, none of them provides comprehensive domain coverage while allowing bulk querying and their visualization schemes can be improved. To address these issues, we developed CDvist (a comprehensive domain visualization tool), which combines the best available search algorithms and databases into a user-friendly framework. First, a given protein sequence is matched to domain models using high-specificity tools and only then unmatched segments are subjected to more sensitive algorithms resulting in a best possible comprehensive coverage. Bulk querying and rich visualization and download options provide improved functionality to domain architecture analysis. AVAILABILITY AND IMPLEMENTATION: Freely available on the web at http://cdvist.utk.edu CONTACT: oadebali@vols.utk.edu or ijouline@utk.edu.