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To evaluate the skeletal and dentoalveolar effects after miniscrew assisted rapid palatal expansion (MARPE) and their correlation with the age of the patients. Settings and sample population: Sample comprised 19 patients with maxillary atresia and posteri

Objetivo: Analisar o perfil de utilização de medicamentos por hipertensos e/ou diabéticos de uma Estratégia de Saúde da Família do Sul de Mato Grosso. Métodos: Trata-se de uma pesquisa quantitativa e transversal com usuários cadastrados em uma Unidade Básica de Saúde. Os dados foram coletados em visitas domiciliares. Resultados: Os usuários consumiram uma média de 2,9 medicamentos. Não houve diferença estatística no consumo médio de medicamentos entre homens e mulheres. Os idosos utilizavam mais medicamentos. Os fármacos que atuam no sistema cardiovascular foram os mais consumidos. Conclusão: Os resultados apontam para a necessidade da promoção do uso racional de medicamentos, sobretudo na população idosa.

The clean development mechanism (CDM) was created with two main goals: help developed countries achieve the greenhouse gas emissions reduction targets set out in the Kyoto Protocol and provide sustainable development to developing countries who host the projects. Sustainable project management shares the same goals as the CDM project, which could be a valuable asset in the development of new projects. Much has been discussed about the effectiveness of the program in achieving those goals and even more in the post-2012 period when the CDM has experienced fewer new projects and lower certified emissions reduction (CER) prices compared to previous years. Based on Brazilian CDM project data, this works aims to analyze the economic aspect of sustainable management by observing the influence of the obtained CERs on the internal rates of return (IRRs) of a small hydro plant (SHP) according to the current CDM scenario, marked by CER prices reaching their lowest historical values. For this study, a spreadsheet was developed to simulate a project’s cash flows and thus determine its IRRs. The results show that using the current CER prices, carbon credits do not significantly affect SHP projects’ IRRs, with increases of less than 0.2% compared to a scenario with no credits. CDM projects may benefit more from optimizing the investment costs or increasing energy production. These results highlight a need to emphasize the sustainable benefits of the CDM program rather than only focusing on the economic return perspective.

To evaluate the stability of the transverse correction with a hybrid maxillary expansion appliance in the bone and tegumental piriformis opening in relation to bone age and maturation of the midpalatal suture (MPS).15 patients with a mean initial age of 14.9 years (SD=1.50), 7 (46.7%) were female and 8 (53.3%) were male, treated with a hybrid maxillary expander. Cone beam computed tomographic (CBCT) images were collected in three phases: T1 (orthodontic records), T2 (21.33 days (SD=10.68) after the end of expansion screw activation) and T3 after 9.13 months (DP=2.41) after the expansion screw was activated. In CBCT, measurements were performed in the nasal cavity considering the tegumental piriform opening (sagittal-axial sections) and bone (sagittal-axial-coronal sections) and the stage of MPS maturation (sagittal-axial sections). Repeated measures ANOVA was used for continuous variables and Friedman's ANOVA for the ordinal variable followed by Bonferroni's tests forThere were significant differences between T1 and T2 (The hybrid maxillary expansion appliance was effective in opening the midpalatal suture of all patients in the present study, without influence of the initial stage of MPS maturation and bone age. There was a relapse of the increase in the bone and tegumental piriform openings.

Contains fulltext : 229049.pdf (Publisher’s version ) (Open Access) The molecular organization of the membrane of the red blood cell controls cell morphology and function and is thereby a main determinant of red blood cell homeostasis in the circulation. The role of membrane organization is prominently reflected in red blood cell deformation and aggregation. However, there is little knowledge on whether they are controlled by the same membrane property and if so, to what extent. To address the potential interdependence of these two parameters, we measured deformation and aggregation in a variety of physiological as well as pathological conditions. As a first step, we correlated a number of deformability and aggregation parameters in red blood cells from healthy donors, which we obtained in the course of our studies on red blood cell homeostasis in health and disease. This analysis yielded some statistically significant correlations. Also, we found that most of these correlations were absent in misshapen red blood cells that have an inborn defect in the interaction between the membrane and the cytoskeleton. The observations suggest that deformability and aggregation share at least one common, membrane-related molecular mechanism. Together with data obtained after treatment with various agents known to affect membrane organization in vitro, our findings suggest that a phosphorylation-controlled interaction between the cytoskeleton and the integral membrane protein band 3 is part of the membrane-centered mechanism that plays a role in deformability as well as aggregation.

Microvesicle generation is an integral part of the aging process of red blood cells in vivo and in vitro. Extensive vesiculation impairs function and survival of red blood cells after transfusion, and microvesicles contribute to transfusion reactions. The triggers and mechanisms of microvesicle generation are largely unknown. In this study, we combined morphological, immunochemical, proteomic, lipidomic, and metabolomic analyses to obtain an integrated understanding of the mechanisms underlying microvesicle generation during the storage of red blood cell concentrates. Our data indicate that changes in membrane organization, triggered by altered protein conformation, constitute the main mechanism of vesiculation, and precede changes in lipid organization. The resulting selective accumulation of membrane components in microvesicles is accompanied by the recruitment of plasma proteins involved in inflammation and coagulation. Our data may serve as a basis for further dissection of the fundamental mechanisms of red blood cell aging and vesiculation, for identifying the cause-effect relationship between blood bank storage and transfusion complications, and for assessing the role of microvesicles in pathologies affecting red blood cells.

Extracellular vesicles (EVs) are emerging as important mediators of cell–cell communication as well as potential disease biomarkers and drug delivery vehicles. However, the mechanical properties of these vesicles are largely unknown, and processes leading to microvesicle-shedding from the plasma membrane are not well understood. Here an in depth atomic force microscopy force spectroscopy study of the mechanical properties of natural EVs is presented. It is found that several natural vesicles of different origin have a different composition of lipids and proteins, but similar mechanical properties. However, vesicles generated by red blood cells (RBC) at different temperatures/incubation times are different mechanically. Quantifying the lipid content of EVs reveals that their stiffness decreases with the increase in their protein/lipid ratio. Further, by maintaining RBC at “extreme” nonphysiological conditions, the cells are pushed to utilize different vesicle generation pathways. It is found that RBCs can generate protein-rich soft vesicles, possibly driven by protein aggregation, and low membrane–protein content stiff vesicles, likely driven by cytoskeleton-induced buckling. Since similar cortical cytoskeleton to that of the RBC exists on the membranes of most mammalian cells, our findings help advancing the understanding of the fundamental process of vesicle generation.

Contains fulltext : 225283.pdf (Publisher’s version ) (Open Access) Extracorporeal circulation is accompanied by changes in red blood cell morphology and structural integrity that affect cell function and survival, and thereby may contribute to the various side effects of heart-lung machine-assisted surgery. Our main objectives were to determine the effect of circulation of red blood cells in a stand-alone extracorporeal circuit on several parameters that are known to be affected by, as well as contribute to red blood cell aging. As a source of RBCs, we employed blood bank storage units of different ages. In order to assess the relevance of our in vitro observations for the characterization of extracorporal circulation technology, we compared these changes in those of patients undergoing extracorporeal circulation-assisted cardiac surgery. Our results show that circulation in a heart-lung machine is accompanied by changes in red blood cell volume, an increase in osmotic fragility, changes in deformability and aggregation behavior, and alterations in the exposure of phosphatidylserine and in microvesicle generation. RBCs from 1-week-old concentrates showed the highest similarities with the in vivo situation. These changes in key characteristics of the red blood cell aging process likely increase the susceptibility of red blood cells to the various mechanical, osmotic, and immunological stress conditions encountered during and after surgery in the patient's circulation, and thereby contribute to the side effects of surgery. Thus, aging-related parameters in red blood cell structure and function provide a foundation for the validation and improvement of extracorporeal circulation technology.

A subset of the red blood cells (RBCs) of patients with paroxysmal nocturnal hemoglobinuria (PNH) lacks GPI-anchored proteins. Some of these proteins, such as CD59, inhibit complement activation and protect against complement-mediated lysis. This pathology thus provides the possibility to explore the involvement of complement in red blood cell homeostasis and the role of GPI-anchored proteins in the generation of microvesicles (MVs) in vivo. Detailed analysis of morphology, volume, and density of red blood cells with various CD59 expression levels from patients with PNH did not provide indications for a major aberration of the red blood cell aging process in patients with PNH. However, our data indicate that the absence of GPI-anchored membrane proteins affects the composition of red blood cell-derived microvesicles, as well as the composition and concentration of platelet-derived vesicles. These data open the way toward a better understanding on the pathophysiological mechanism of PNH and thereby to the development of new treatment strategies.

Contains fulltext : 193256.pdf (Publisher’s version ) (Open Access) Red blood cells (RBCs) generate microvesicles to remove damaged cell constituents such as oxidized hemoglobin and damaged membrane constituents, and thereby prolong their lifespan. Damage to hemoglobin, in combination with altered phosphorylation of membrane proteins such as band 3, lead to a weakening of the binding between the lipid bilayer and the cytoskeleton, and thereby to membrane budding and microparticle shedding. Microvesicle generation is disturbed in patients with RBC-centered diseases, such as sickle cell disease, glucose 6-phosphate dehydrogenase deficiency, spherocytosis or malaria. A disturbance of the membrane-cytoskeleton interaction is likely to be the main underlying mechanism, as is supported by data obtained from RBCs stored in blood bank conditions. A detailed proteomic, lipidomic and immunogenic comparison of microvesicles derived from different sources is essential in the identification of the processes that trigger vesicle generation. The contribution of RBC-derived microvesicles to inflammation, thrombosis and autoimmune reactions emphasizes the need for a better understanding of the mechanisms and consequences of microvesicle generation.

During its 120 days sojourn in the circulation, the red blood cell (RBC) remodels its membrane. Acetylcholinesterase (AChE) is a glycosylphosphatidylinositol (GPI)-linked enzyme that may serve as a marker for membrane processes occurring this ageing-associated remodelling process.Expression and enzymatic activity of AChE were determined on RBCs of various ages, as obtained by separation based on volume and density (ageing in vivo), and on RBCs of various times of storage in blood bank conditions (ageing in vitro), as well as on RBC-derived vesicles.During ageing in vivo, the enzymatic activity of AChE decreases, but not the AChE protein concentration. In contrast, neither AChE activity nor concentration show a consistent, significant decrease during ageing in vitro. CD59, another GPI-linked protein that protects against complement-induced removal, also remains constant during storage. The cellular content of the integral membrane protein glycophorin A, however, decreases with storage time in the more dense RBC fractions. The latter are enriched in echinocytes and other misshapen cells during storage.Our findings suggest that, during RBC ageing, GPI-linked proteins and integral membrane proteins are differentially sorted. Also, the vesicles that are generated in vitro show a fast and extensive loss of AChE activity, but not of AChE expression. Thus, AChE characteristics may constitute sensitive biomarkers of RBC ageing in vivo, and a source of information on the structural and functional changes that GPI-linked proteins undergo during ageing in vivo and in vitro. This information may help to understand RBC homeostasis and the effects of transfusion, especially in immunologically compromised patients.

In this study, the nanostructures of pure ZnO and ZnO:xCe3+ were prepared using statistic design—factorial design 3(3−1)−3-level and mixed-level factorials and fractional with replicates in the central point, totaling 11 experiments. The experiments were performed using the microwave-assisted hydrothermal (MAH) method with Ce3+ concentration of 2 and 4 mol% at 60, 110, and 160 °C for 10, 20, and 30 min. X-ray diffraction (XRD), scanning electron microscopy (SEM), and optical diffuse reflectance were used to characterize the products. The fractional factorial design indicated the optimal design area, and the studies were continued by ridge analysis. The analysis of variance (ANOVA), the Pareto, and the model adjusted to the conditions proposed in this study due to the determination coefficient of 99.9%, variance (R 2), and response surface generated were satisfactory, thus having an optimization in the process of obtaining ZnO doped with Ce.

Contains fulltext : 168094.pdf (Publisher’s version ) (Open Access) During their passage through the circulation, red blood cells (RBCs) encounter severe physiological conditions consisting of mechanical stress, oxidative damage and fast changes in ionic and osmotic conditions. In order to survive for 120 days, RBCs adapt to their surroundings by subtle regulation of membrane organization and metabolism. RBC homeostasis depends on interactions between the integral membrane protein band 3 with other membrane and cytoskeletal proteins, and with key enzymes of various metabolic pathways. These interactions are regulated by the binding of deoxyhemoglobin to band 3, and by a signaling network revolving around Lyn kinase and Src family kinase-mediated phosphorylation of band 3. Here we show that manipulation of the interaction between the lipid bilayer and the cytoskeleton, using various pharmacological agents that interfere with protein-protein interactions and membrane lipid organization, has various effects on: (1) morphology, as shown by high resolution microscopy and quantitative image analysis; (2) organization of membrane proteins, as indicated by immunofluorescence confocal microscopy and quantitative as well as qualitative analysis of vesicle generation; (3) membrane lipid organization, as indicated by flow cytometric analysis of phosphatidylserine exposure; (4) deformability, as assessed in capillary-mimicking circumstances using a microfluidics system; (5) deformability as determined using a spleen-mimicking device; (6) metabolic activity as indicated by metabolomics. Our data show that there is a complex relationship between red cell morphology, membrane organization and deformability. Also, our data show that red blood cells have a relatively high resistance to disturbance of membrane organization in vitro, which may reflect their capacity to withstand mechanical, oxidative and osmotic stress in vivo.

The applicability of several Lattice-Boltzmann methods to wall-bounded turbulent flows is investigated. The various methods consist of the standard Bhatnagar–Gross–Krook (BGK) method with 19 (BGK19) and 27 (BGK27) discrete velocities, the multiple-relaxation-time (MRT) model with 19 discrete velocities and the cascaded Lattice-Boltzmann method (CLB). Based on the findings of turbulent channel flow it can be concluded that stability considerations, predicting the superiority of the advanced moment based schemes like the CLB and MRT method not necessarily hold for wall-bounded turbulent flows. Moreover, in some flow problems the simple BGK method with 19 discrete velocities delivers reasonable and stable results, where the other methods yield unphysical solutions.

Globin-coupled sensors are heme-binding signal transducers in Bacteria and Archaea in which an N-terminal globin controls the activity of a variable C-terminal domain. Here, we report that Bpe GReg, a globin-coupled diguanylate cyclase from the whooping cough pathogen Bordetella pertussis , synthesizes the second messenger bis-(3′–5′)-cyclic diguanosine monophosphate (c-di-GMP) upon oxygen binding. Expression of Bpe GReg in Salmonella typhimurium enhances biofilm formation, while knockout of the Bpe GReg gene of B. pertussis results in decreased biofilm formation. These results represent the first identification a signal ligand for any diguanylate cyclase and provide definitive experimental evidence that a globin-coupled sensor regulates c-di-GMP synthesis and biofilm formation. We propose that the synthesis of c-di-GMP by globin sensors is a widespread phenomenon in bacteria.

Francisella tularensis is a highly infectious bacterium with the potential to cause high fatality rates if infections are untreated. To aid in the development of rapid and accurate detection assays, we have sequenced and annotated the genomes of 18 F. tularensis and Francisella philomiragia strains.

The interaction of cellular proteins with the gap junction protein Connexin43 (Cx43) is thought to form a dynamic scaffolding complex that functions as a platform for the assembly of signaling, structural, and cytoskeletal proteins. A high stringency Scansite search of rat Cx43 identified the motif containing Ser373 (S373) as a 14-3-3 binding site. The S373 motif and the second best mode-1 motif, containing Ser244 (S244), are conserved in rat, mouse, human, chicken, and bovine, but not in Xenopus or zebrafish Cx43. Docking studies of a mouse/rat 14-3-3 homology model with the modeled phosphorylated S373 or S244 peptide ligands or their serine-to-alanine mutants, S373A or S244A, revealed that the pS373 motif facilitated a greater number of intermolecular contacts than the pS244 motif, thus supporting a stronger 14-3-3 binding interaction with the pS373 motif. The alanine substitution also reduced more than half the number of intermolecular contacts between 14-3-3 and the S373 motif, emphasizing the phosphorylation dependence of this interaction. Furthermore, the ability of the wild-type or the S244A GST-Cx43 C-terminal fusion protein, but not the S373A fusion protein, to interact with either 14-3-3 or 14-3-3zeta in GST pull-down experiments clearly demonstrated that the S373 motif mediates the direct interaction between Cx43 and 14-3-3 proteins. Blocking growth factor-induced Akt activation and presumably any Akt-mediated phosphorylation of the S373 motif in ROSE 199 cells did not prevent the down-regulation of Cx43-mediated cell-cell communication, suggesting that an Akt-mediated interaction with 14-3-3 was not involved in the disruption of Cx43 function.

The strategy for detecting oxygen, carbon monoxide, nitric oxide, and sulfides is predominantly through heme-based sensors utilizing either a globin domain or a PAS domain. Whereas PAS domains bind various cofactors, globins bind only heme. Globin-coupled sensors (GCSs) were first described as regulators of the aerotactic responses in Bacillus subtilis and Halobacterium salinarum. GCSs were also identified in diverse microorganisms that appear to have roles in regulating gene expression. Functional and evolutionary analyses of the GCSs, their protoglobin ancestor, and their relationship to the last universal common ancestor (LUCA) are discussed in the context of globin-based signal transduction.

Hemoglobins are ubiquitous in Eukarya and Bacteria but, until now, have not been found in Archaea . A phylogenetic analysis of the recently revealed microbial family of globin-coupled heme-based sensors suggests that these sensors descended from an ancient globin-only progenitor, or a protoglobin (Pgb). Here, we report the discovery and characterization of two Pgbs from the Archaea : Ap Pgb from the obligately aerobic hyperthermophile Aeropyrum pernix , and Ma Pgb from the strictly anaerobic methanogen Methanosarcina acetivorans . Both Ap Pgb and Ma Pgb bind molecular oxygen, nitric oxide, and carbon monoxide by means of a heme moiety that is coordinated to the protein through the F8 histidine (histidine 120). We postulate that these archaeal globins are the ancestors of contemporary hemoglobins.

Shigella flexneri causes shigellosis, severe and potentially life-threatening diarrhea, and accounts for 18% of shigellosis cases in the United States. Here, we present the 4.51-Mbp genome assembly of S. flexneri ATCC 12022, a quality control and reference strain, in 10 scaffolds.

The pleomorphic swarming bacilli of the genus Proteus are common human gut commensal organisms but also the causative agents of recurrent urinary tract infections and bacteremia. We sequenced and assembled the 3.99-Mbp genome of Proteus mirabilis ATCC 7002 (accession no. JOVJ00000000) and the 3.97-Mbp genome of Proteus vulgaris ATCC 49132 (accession no. JPIX00000000), both of which are commonly used reference strains.

Staphylococcus aureus is a major cause of bacterial infections in the United States, with high percentages of serious infections resistant to a variety of β-lactam antibiotics. Here, we present the scaffolded genome assembly into 16 contigs of S. aureus CDC73-57501 (ATCC 29247), a methicillin-resistant quality control strain.

Salmonella enterica constitutes a group of enteric pathogens with a broad host range, including humans, reptiles, and birds. S. enterica subsp. enterica is a common cause of inflammatory diarrhea in humans. We present the draft genome of S. enterica subsp. enterica serovar Enteritidis strain SEJ, including a 59-kbp plasmid.

Enterococcus faecalis is a nonmotile Gram-positive coccus, found both as a commensal organism in healthy humans and animals and as a causative agent of multiple diseases, in particular endocarditis. We sequenced the genome of E. faecalis ATCC 29212, a commonly used reference strain in laboratory studies, to complete “finished” annotated assembly (3 Mb).

Real-time tissue identification can benefit procedures such as stereotactic brain biopsy, functional neurosurgery and brain tumor excision. Optical scattering spectroscopy has been shown to be effective at discriminating cancer from noncancerous conditions in the colon, bladder and breast. The NASA Smart Probe extends the concept of ‘optical biopsy’ by using neural network techniques to combine the output from 3 microsensors contained within a cannula 2.7 mm in diameter (i.e. the diameter of a stereotactic brain biopsy needle). Experimental data from 5 rats show the clear differentiation between tissues such as brain, nerve, fat, artery and muscle that can be achieved with optical scattering spectroscopy alone. These data and previous findings with other modalities such as (1) analysis of the image from a fiberoptic neuroendoscope and (2) the output from a microstrain gauge suggest the Smart Probe multiple microsensor technique shows promise for real-time tissue identification in neurosurgical procedures.

Annotation of prokaryotic sequences can be separated into structural and functional annotation. Structural annotation is dependent on algorithmic interrogation of experimental evidence to discover the physical characteristics of a gene. This is done in an effort to construct accurate gene models, so understanding function or evolution of genes among organisms is not impeded. Functional annotation is dependent on sequence similarity to other known genes or proteins in an effort to assess the function of the gene. Combining structural and functional annotation across genomes in a comparative manner promotes higher levels of accurate annotation as well as an advanced understanding of genome evolution. As the availability of bacterial sequences increases and annotation methods improve, the value of comparative annotation will increase.

The flow in a real human nose is numerically simulated at steady inspiration and expiration. The analysis uses a Lattice Boltzmann method (LBM) which is particularly suited for flows in extremely intricate geometries. The nasal geometry is extracted from computer tomography (CT) data using a so-called reconstruction pipeline. Thus, for any nose the surface geometry can be defined and a numerical mesh can be generated. The focus of this investigation is on the analysis of the flow field at steady inspiration and expiration with respect to secondary flow structures. It is evidenced that strong vortical structures appear near the throat at inspiration forming a pair of counter-rotating vortices which disappear at expiration. Overall, at exhalation less vorticity is generated in the flow than at inhalation.

Renal hypoplasia results from an insufficient kidney volume caused, in part, by a deficient number of glomeruli. The purpose of this study was to apply tessellation analysis to determine whether glomerular point patterns differed between adult normal (WT) and mutant (Br) mice with heritable renal hypoplasia and to delineate a spatial distribution accounting for the observed patterns. Kidneys from adult WT and Br mice were collected, processed with routine light histology and representative transverse sections were photographed. Cortical area and perimeter were calculated from traced tissue contours and glomeruli were identified and digitized. Voronoi tessellations were constructed and average parameters for Voronoi polygon number, area, perimeter and edge counts as well as spatial metrics comprising nearest neighbor and centroidal distances were calculated and compared. Point distributions were simulated by randomizing glomerular coordinates from each section and plotting the new points utilizing uniform random, Gaussian random, or isotropic functions. Average nearest neighbor distances were generated for each specimen and ranked with respect to corresponding values generated from 1,000 iterations for each simulated set. Results showed that WT and Br were significantly different for each parameter suggesting that WT kidneys possessed more glomeruli, but these were less clustered compared to Br. Simulations suggested that WT and Br demonstrated similar, but not identical, underlying glomerular spatial distributions. Defective gene expression in Br is important for determining glomerular number and the defective pattern likely results from a heterochronic disturbance consisting of a truncated growth trajectory during embryonic kidney development.

Globin-coupled sensors (GCSs) are multidomain proteins, consisting of an N-terminal globin domain fused to a variety of C-terminal transmitter domains. Functional classification of GCSs is based on the transmitter domain(s) they possess, broadly falling under either aerotaxis or gene regulation. This chapter describes methods and strategies for cloning, expression, and purification of GCSs for spectroscopic analysis and determination of the minimum heme-binding domain, together with bioinformatic approaches for database searching and examination of domain architectures.

The flow field at inspiration and expiration in the upper human airways consisting of the trachea down to the sixth generation of the bronchial tree is numerically simulated. The three-dimensional steady flow at a hydraulic diameter-based Reynolds number Re(D)=1250 is computed via a lattice-Boltzmann method (LBM). The simulation is validated by the experimental data based on particle-image velocimetry (PIV) measurements. The good agreement between numerical and experimental results is evidenced by comparing velocity contours and distributions in a defined reference plane. The results show the LBM to be an accurate tool to numerically predict flow structures in the human lung. Using an automatic Cartesian grid generator, the overall process time from meshing to a steady-state solution is

The strategy for detecting oxygen, carbon monoxide, nitric oxide, and sulfides is predominantly through heme-based sensors utilizing either a globin domain or a PAS domain. Whereas PAS domains bind various cofactors, globins bind only heme. Globin-coupled sensors (GCSs) couple an N-terminal sensor globin domain to varied C-terminal signaling domains to effect aerotaxis and gene regulation. Having descended from an ancient protoglobin, GCSs are now ubiquitous and are encoded in the genome of several extremophiles (temperature, salt, and pH). We postulate that their role in regulating gene expression governs microbial processes critical to elemental recycling, bioremediation, and cellulose degradation. Functional and evolutionary analyses of the GCSs, their protoglobin ancestor, and their relationship to the Last Universal Common Ancestor (LUCA) are also discussed in the context of globin-based signal transduction.

Globin-coupled sensors (GCSs) are multidomain proteins, consisting of an N-terminal globin domain fused to a variety of C-terminal transmitter domains. Functional classification of GCSs is based on the transmitter domain(s) they possess, broadly falling under either aerotaxis or gene regulation. This chapter describes methods and strategies for cloning, expression, and purification of GCSs for spectroscopic analysis and determination of the minimum heme-binding domain, together with bioinformatic approaches for database searching and examination of domain architectures.

A major challenge in the field of shotgun metagenomics is the accurate identification of organisms present within a microbial community, based on classification of short sequence reads. Though existing microbial community profiling methods have attempted to rapidly classify the millions of reads output from modern sequencers, the combination of incomplete databases, similarity among otherwise divergent genomes, errors and biases in sequencing technologies, and the large volumes of sequencing data required for metagenome sequencing has led to unacceptably high false discovery rates (FDR). Here, we present the application of a novel, gene-independent and signature-based metagenomic taxonomic profiling method with significantly and consistently smaller FDR than any other available method. Our algorithm circumvents false positives using a series of non-redundant signature databases and examines Genomic Origins Through Taxonomic CHAllenge (GOTTCHA). GOTTCHA was tested and validated on 20 synthetic and mock datasets ranging in community composition and complexity, was applied successfully to data generated from spiked environmental and clinical samples, and robustly demonstrates superior performance compared with other available tools.

We report the complete genome sequence of the deep-sea γ-proteobacterium, Idiomarina loihiensis , isolated recently from a hydrothermal vent at 1,300-m depth on the Lōihi submarine volcano, Hawaii. The I. loihiensis genome comprises a single chromosome of 2,839,318 base pairs, encoding 2,640 proteins, four rRNA operons, and 56 tRNA genes. A comparison of I. loihiensis to the genomes of other γ-proteobacteria reveals abundance of amino acid transport and degradation enzymes, but a loss of sugar transport systems and certain enzymes of sugar metabolism. This finding suggests that I. loihiensis relies primarily on amino acid catabolism, rather than on sugar fermentation, for carbon and energy. Enzymes for biosynthesis of purines, pyrimidines, the majority of amino acids, and coenzymes are encoded in the genome, but biosynthetic pathways for Leu, Ile, Val, Thr, and Met are incomplete. Auxotrophy for Val and Thr was confirmed by in vivo experiments. The I. loihiensis genome contains a cluster of 32 genes encoding enzymes for exopolysaccharide and capsular polysaccharide synthesis. It also encodes diverse peptidases, a variety of peptide and amino acid uptake systems, and versatile signal transduction machinery. We propose that the source of amino acids for I. loihiensis growth are the proteinaceous particles present in the deep sea hydrothermal vent waters. I. loihiensis would colonize these particles by using the secreted exopolysaccharide, digest these proteins, and metabolize the resulting peptides and amino acids. In summary, the I. loihiensis genome reveals an integrated mechanism of metabolic adaptation to the constantly changing deep-sea hydrothermal ecosystem.

Real-time tissue identification can benefit procedures such as stereotactic brain biopsy, functional neurosurgery, and brain tumor excision. Optical scattering spectroscopy has been shown effective at discriminating cancer from non-cancerous conditions in the colon, bladder, and breast. The NASA Smart Probe extends the concept of "optical biopsy" by using neural network techniques to combine the output from three microsensors contained within a cannula 2.7 mm in diameter (i.e. the diameter of a stereotactic brain biopsy needle). Experimental data from five rats show the clear differentiation between tissues such as brain, nerve, fat, artery, and muscle that can be achieved with optical scattering spectroscopy alone. These data and previous findings with other modalities such as (1) analysis of the image from a fiberoptic neuroendoscope and (2) the output from a micro strain gauge suggest the Smart Probe multiple microsensor technique shows promise for real-time tissue identification in neurosurgical procedures.