Use of a commercial tissue dissociation system to detect Salmonella-contaminated poultry products.

Successful detection of bacterial pathogens in food can be challenging due to the physical and compositional complexity of the matrix. Different mechanical/physical and chemical methods have been developed to separate microorganisms from food matrices to facilitate detection. The present study benchmarked a commercial tissue digestion system that applies both chemical and physical methods to separate microorganisms from tissues against stomaching, a standard process currently utilized by commercial and regulatory food safety laboratories. The impacts of the treatments on the physical properties of the food matrix were characterized along with the compatibility of the methods with downstream microbiological and molecular detection assays. The results indicate the tissue digestion system can significantly reduce the average particle size of the chicken sample relative to processing via a stomacher (P < 0.001) without adversely affecting either real-time PCR (qPCR) or plate counting assays, which are typically used to detect Salmonella. Furthermore, inoculated chicken treated with the GentleMACS resulted in a significant increase (P < 0.003) in the qPCR's detection capabilities relative to stomached controls. Cohen kappa (κ) coefficient and McNemar's test indicate the plating assays and PCR results agree with measurements obtained via the 3 M Molecular Detection System as defined in the MLG standard (κ > 0.62; P > 0.08). Collectively, the results demonstrate that the technique enables detection of pathogens in meat at lower levels of contamination using current industry standard technologies.

Fe-Doped g-C3N4/Bi2MoO6 Heterostructured Composition with Improved Visible Photocatalytic Activity for Rhodamine B Degradation.

The binary heterostructured semiconducting visible light photocatalyst of the iron-doped graphitic carbon nitride/bismuth molybdate (Fe-g-C3N4/Bi2MoO6) composite was prepared by coupling with Fe-doped g-C3N4 and Bi2MoO6 particles. In the present study, a comparison of structural characteristics, optical properties, and photocatalytic degradation efficiency and activity between Fe-doped g-C3N4 particles, Bi2MoO6 particles, and Fe-g-C3N4/Bi2MoO6 composite was investigated. The results of X-ray diffraction (XRD) examination indicate that the hydrothermal Bi2MoO6 particles have a single orthorhombic phase and Fourier transform infrared (FTIR) spectroscopy analysis confirms the formation of Fe-doped g-C3N4. The optical bandgaps of the Fe-doped g-C3N4 and Bi2MoO6 particles are 2.74 and 2.73 eV, respectively, as estimated from the Taut plots obtained from UV-Vis diffuse reflectance spectroscopy (DRS) spectra. This characteristic indicates that the two semiconductor materials are suitable for absorbing visible light. The transmission electron microscopy (TEM) micrograph reveals the formation of the heterojunction Fe-g-C3N4/Bi2MoO6 composite. The results of photocatalytic degradation revealed that the developed Fe-g-C3N4/Bi2MoO6 composite photocatalyst exhibited significantly better photodegradation performance than the other two single semiconductor photocatalysts. This property can be attributed to the heterostructured nanostructure, which could effectively prevent the recombination of photogenerated carriers (electron-hole pairs) and enhance photocatalytic activity. Furthermore, cycling test showed that the Fe-g-C3N4/Bi2MoO6 heterostructured photocatalyst exhibited good reproducibility and stability for organic dye photodegradation.

Aurora B functions at the apical surface after specialized cytokinesis during morphogenesis in C. elegans.

Although cytokinesis has been intensely studied, the way it is executed during development is not well understood, despite a long-standing appreciation that various aspects of cytokinesis vary across cell and tissue types. To address this, we investigated cytokinesis during the invariant Caenorhabditis elegans embryonic divisions and found several parameters that are altered at different stages in a reproducible manner. During early divisions, furrow ingression asymmetry and midbody inheritance is consistent, suggesting specific regulation of these events. During morphogenesis, we found several unexpected alterations to cytokinesis, including apical midbody migration in polarizing epithelial cells of the gut, pharynx and sensory neurons. Aurora B kinase, which is essential for several aspects of cytokinesis, remains apically localized in each of these tissues after internalization of midbody ring components. Aurora B inactivation disrupts cytokinesis and causes defects in apical structures, even if inactivated post-mitotically. Therefore, we demonstrate that cytokinesis is implemented in a specialized way during epithelial polarization and that Aurora B has a role in the formation of the apical surface.

Missing Piece in Colloidal Stability─Morphological Factor of Hydrophobic Nanoparticles.

Hydrophobic nanoparticles (NPs) in water were considered unstable because they lack the repulsive electrostatic interaction and steric effect to prevent aggregation. In this study, porous hydrophobic NPs of two star-shaped giant molecules, POSS-(R)8, were found to be stable in water and able to retain their kinetic stability in a wide range of temperatures, pH values, and ionic strengths. Unlike the solid hydrophobic NPs that aggregate even with the negative zeta potential (ζ) induced by surface-structured hydrogen-bonded (SHB) water, the porous morphology of POSS-(R)8 NPs reduces the entropically driven hydrophobic effect to prevent aggregation. With the porous morphology, the hydrophobic NPs are stable without the hydrophilic or charged surface functional groups and demonstrate good encapsulation capability. The morphological factor of colloids is thus one of the missing pieces in the theory of colloidal stability that extends our understanding of colloidal science.

The complex multicellular morphology of the food spoilage bacteria Brochothrix thermosphacta strains isolated from ground chicken.

Herein we describe a highly structured, filamentous growth phenotype displayed by an isolate of the food spoilage microorganism Brochothrix thermosphacta. The growth morphology of this B. thermosphacta strain (strain BII) was dependent on environmental factors such as the growth media, incubation temperatures, and the inoculum concentration. Inoculation of cultures in highly dilute suspensions resulted in the formation of isolated, tight aggregates resembling fungal growth in liquid media. This same strain also formed stable, mesh-like structures in 6-well tissue culture plates under specific growth conditions. The complex growth phenotype does not appear to be unique to strain BII but was common among B. thermosphacta strains isolated from chicken. Light and electron micrographs showed that the filaments of multiple BII cells can organize into complex, tertiary structures resembling multistranded cables. Time-lapse microscopy was employed to monitor the development of such aggregates over 18 h and revealed growth originating from short filaments into compact ball-like clusters that appeared fuzzy due to protruding filaments or cables. This report is the first to document this complex filamentous growth phenotype in a wild-type bacterial isolate of B. thermosphacta.

WSe2 Homojunction Devices: Electrostatically Configurable as Diodes, MOSFETs, and Tunnel FETs for Reconfigurable Computing.

In this paper, electrostatically configurable 2D tungsten diselenide (WSe2 ) electronic devices are demonstrated. Utilizing a novel triple-gate design, a WSe2 device is able to operate as a tunneling field-effect transistor (TFET), a metal-oxide-semiconductor field-effect transistor (MOSFET) as well as a diode, by electrostatically tuning the channel doping to the desired profile. The implementation of scaled gate dielectric and gate electrode spacing enables higher band-to-band tunneling transmission with the best observed subthreshold swing (SS) among all reported homojunction TFETs on 2D materials. Self-consistent full-band atomistic quantum transport simulations quantitatively agree with electrical measurements of both the MOSFET and TFET and suggest that scaling gate oxide below 3 nm is necessary to achieve sub-60 mV dec-1 SS, while further improvement can be obtained by optimizing the spacers. Diode operation is also demonstrated with the best ideality factor of 1.5, owing to the enhanced electrostatic control compared to previous reports. This research sheds light on the potential of utilizing electrostatic doping scheme for low-power electronics and opens a path toward novel designs of field programmable mixed analog/digital circuitry for reconfigurable computing.

Effects of Arginine Deimination and Citrulline Side-Chain Length on Peptide Secondary Structure Formation.

Post-translational modifications expand the chemical functionality of peptides and proteins beyond that originating from the encoded amino acids, but studies on the structural effects of these modifications have been limited. Arginine undergoes deimination to give citrulline (Cit), converting the positively charged guanidinium moiety into a neutral urea group. Herein, we report the effect of Arg deimination on secondary structure formation. To understand the reason for the number of methylene units in Cit, the effect of Cit side-chain length on secondary structure formation was also studied. Ala-based peptides and ß-hairpin peptides were used to study α-helix and ß-sheet formation, respectively. Peptides containing Cit analogues were prepared by an orthogonal protecting group strategy coupled with solid-phase carbamylation. The CD data for the Ala-based peptides were analyzed by using modified Lifson-Roig theory, showing that the helix propensity of Arg decreased upon deimination and that either shortening or lengthening Cit also decreased the helix propensity. The ß-hairpin peptides were analyzed by NMR methods, showing minimal change in strand formation energetics upon Arg deimination. Altering the Cit side-chain length did not affect strand formation energetics either. These results should be useful for the preparation of urea-bearing systems and the design of peptides incorporating urea-bearing residues with varying side-chain length.

Implementation of normalized retention time (iRT) for bottom-up proteomic analysis of the aminoglycoside phosphotransferase enzyme facilitating method distribution.

Improvements in mass spectrometry technology to include instrument duty cycle, resolution, and sensitivity suggest mass spectrometry as a highly competitive alternative to conventional microbiological proteomic techniques. Targeted mass spectral analysis, sans prior empirical measurements, has begun to solely use the enormous amount of available genomic information for assay development. An in silico tryptic digestion of a suspected antibiotic-resistant enzyme using only its genomic information for assay development was achieved. Both MRM and full-scan MS2 independent data acquisitions were obtained for an antibiotic-resistance microbe not previously measured using mass spectrometry. In addition, computation methods to determine highest responding peptides in positive ion mode liquid chromatography-mass spectrometry (LC-MS) were evaluated. Employment of the relative retention time (iRT) concept using a homemade peptide standard set revealed facile method transfer between two fundamental different mass spectral platforms: an ultra-high-pressure liquid chromatography triple quadrupole-mass spectrometer (UHPLC-MS) and nano-liquid chromatography parallel reaction monitoring (nano-LC-PRM) hybrid quadrupole orbitrap Q-exactive mass spectrometer supporting easy dissemination and rapid method implementation between laboratories. Graphical Abstract.

Rapid detection and quantification of aminoglycoside phosphorylation products using direct-infusion high-resolution and ultra-high-performance liquid chromatography/mass spectrometry.

RATIONALE: Worldwide efforts are underway to determine the extent of antimicrobial resistance (AMR). In 2015, the World Health Organization (WHO) founded the Global Antimicrobial Surveillance System (GLASS) focusing on surveillance and dissemination of data. In addition, the WHO advocates method development focused on rapid determination and close to real-time monitoring of antibiotic usage and its effectiveness. Rapid determination of aminoglycoside modification by O-phosphorylation, the most prevalent mechanism of clinical resistance, was performed using direct flow and liquid chromatography/mass spectrometry (LC/MS). METHODS: A strain of Escherichia coli carrying a plasmid encoding an aminoglycoside modification enzyme (O-phosphotransferase) was incubated with kanamycin, an aminoglycoside. The antibiotic and its modified form were observed using direct flow and LC/MS. Direct flow high-resolution mass spectrometry (HRMS) using a Thermo Fisher Q-Exactive hybrid quadrupole-orbitrap mass spectrometer was employed for quantitative analysis and structural elucidation. Liquid chromatography coupled with the AB Sciex QTRAP 6500+ was also used for quantitative analysis. RESULTS: Detection of phosphorylated kanamycin was achieved in less than 4 h of incubation. Calibration curves for modified kanamycin from 2.5-250 and 10-200 µg mL-1  µg mL-1 were obtained for LC/MS and direct injection high-resolution experiments, respectively. The high-resolution measurements were employed for conformation and structural elucidation of the novel precursor and product ion biomarkers with high mass accuracy (≤7 ppm). These results confirm previous in vitro O-phosphotransferase metabolite measurements. CONCLUSIONS: A new analytical method capable of determination and quantification of the most common form of aminoglycoside resistance (via phosphorylation) was developed requiring short incubation times for a positive confirmation 100-fold lower than the minimum inhibitory concentration (MIC). High-resolution data simultaneously revealed quantitative abilities and provided numerous novel product ions confirming placement of the phosphate group on kanamycin.

Detection of acetyltransferase modification of kanamycin, an aminoglycoside antibiotic, in bacteria using ultrahigh-performance liquid chromatography tandem mass spectrometry.

RATIONALE: The occurrence of antibiotic-resistant bacteria is a worldwide issue that has the potential, if not addressed, to eliminate classes of antibiotics that have extended life expectancy in the last century. An approach to confront this threat is the development of technologies that greatly accelerate the detection of antibiotic resistance to minimize unnecessary treatment involving antibiotics. Development of an analytical method for rapid detection of aminoglycoside resistance using liquid chromatography/mass spectrometry (LC/MS) has not been reported in the literature and is described here. METHODS: A strain of Escherichia coli carrying a plasmid encoding an aminoglycoside-modifide enzyme (N-acetyltransferase) was incubated with kanamycin, an aminoglycoside. The antibiotic and its modified form were observed using LC/MS. An ABSciex QTrap 6500+ was used for kinetic and quantitative analysis and high-resolution structural elucidation was performed using a Thermo Fisher Q-Exactive hybrid quadrupole-orbitrap mass spectrometer. RESULTS: Detection of kanamycin modification was achieved in less than an hour of incubation. Calibration curves for both modified and unmodified kanamycin from 0.5 to 50 µg mL-1 were obtained. Generation and depletion of modified and unmodified kanamycin as a function of time were performed. High-resolution mass spectrometry was employed for confirmation and structural elucidation of the novel precursor and product ion biomarkers with high mass accuracy (≤7 ppm). CONCLUSIONS: A newly developed analytical method is able to determine bacterial resistance to aminoglycosides (via acetylation of kanamycin), qualitatively and quantitatively, within 30 minutes and 6 hours of incubation with kanamycin, respectively. High-resolution data support the placement of an acetyl group on kanamycin confirming aminoglycoside resistance and its mechanism. Quantification was achieved for both forms of the antibiotic 50- to 100-fold lower than the minimum inhibitory concentration for the resistant bacteria and can be used to replace conventional antimicrobial susceptibility tests.

Serogroup-level resolution of the "Super-7" Shiga toxin-producing Escherichia coli using nanopore single-molecule DNA sequencing.

DNA sequencing and other DNA-based methods are now broadly used for detection and identification of bacterial foodborne pathogens. For the identification of foodborne bacterial pathogens, taxonomic assignments must be made to the species or even subspecies level. Long-read DNA sequencing provides finer taxonomic resolution than short-read sequencing. Here, we demonstrate the potential of long-read shotgun sequencing obtained from the Oxford Nanopore Technologies (ONT) MinION single-molecule sequencer, in combination with the Basic Local Alignment Search Tool (BLAST) with custom sequence databases, for foodborne pathogen identification. A library of mixed DNA from strains of the "Super-7" Shiga toxin-producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, O145, and O157[:H7]) was sequenced using the ONT MinION resulting in 44,245 long-read sequences. The ONT MinION sequences were compared to a custom database composed of the E. coli O-antigen gene clusters. A vast majority of the sequence reads were from outside of the O-antigen cluster and did not align to any sequences in the O-antigen database. However, 58 sequences (0.13% of the total sequence reads) did align to a specific Super-7 O-antigen gene cluster, with each O-antigen cluster aligning to at least four sequence reads. BLAST analysis against a custom whole-genome database revealed that 5096 (11.5%) of the MinION sequence reads aligned to one and only one sequence in the database, of which 99.6% aligned to a sequence from a "Super-7" STEC. These results demonstrate the ability of the method to resolve STEC to the serogroup level and the potential general utility of the MinION for the detection and typing of foodborne pathogens.

Genome amplification and promoter mutation expand the range of csgD-dependent biofilm responses in an STEC population.

Expression of the major biofilm components of E. coli, curli fimbriae and cellulose, requires the CsgD transcription factor. A complex regulatory network allows environmental control of csgD transcription and biofilm formation. However, most clinical serotype O157 : H7 strains contain prophage insertions in the csgD regulator, mlrA, or mutations in other regulators that restrict csgD expression. These barriers can be circumvented by certain compensating mutations that restore higher csgD expression. One mechanism is via csgD promoter mutations that switch sigma factor utilization. Biofilm-forming variants utilizing RpoD rather than RpoS have been identified in glycerol freezer stocks of the non-biofilm-forming food-borne outbreak strain, ATCC 43894. In this study we used whole genome sequencing and RNA-seq to study genotypic and transcriptomic differences between those strains. In addition to defining the consequences of the csgD promoter switch and identifying new csgD-controlled genes, we discovered a region of genome amplification in our laboratory stock of 43894 (designated 43894OW) that contributed to the regulation of csgD-dependent properties.

Genome sequencing and comparative genomics provides insights on the evolutionary dynamics and pathogenic potential of different H-serotypes of Shiga toxin-producing Escherichia coli O104.

BACKGROUND: Various H-serotypes of the Shiga toxin-producing Escherichia coli (STEC) O104, including H4, H7, H21, and H¯, have been associated with sporadic cases of illness and have caused food-borne outbreaks globally. In the U.S., STEC O104:H21 caused an outbreak associated with milk in 1994. However, there is little known on the evolutionary origins of STEC O104 strains, and how genotypic diversity contributes to pathogenic potential of various O104 H-antigen serotypes isolated from different ecological niches and/or geographical regions. RESULTS: Two STEC O104:H21 (milk outbreak strain) and O104:H7 (cattle isolate) strains were shot-gun sequenced, and the genomes were closed. The intimin (eae) gene, involved in the attaching-effacing phenotype of diarrheagenic E. coli, was not found in either strain. Examining various O104 genome sequences, we found that two "complete" left and right end portions of the locus of enterocyte effacement (LEE) pathogenicity island were present in 13 O104 strains; however, the central portion of LEE was missing, where the eae gene is located. In O104:H4 strains, the missing central portion of the LEE locus was replaced by a pathogenicity island carrying the aidA (adhesin involved in diffuse adherence) gene and antibiotic resistance genes commonly carried on plasmids. Enteroaggregative E. coli-specific virulence genes and European outbreak O104:H4-specific stx2-encoding Escherichia P13374 or Escherichia TL-2011c bacteriophages were missing in some of the O104:H4 genome sequences available from public databases. Most of the genomic variations in the strains examined were due to the presence of different mobile genetic elements, including prophages and genomic island regions. The presence of plasmids carrying virulence-associated genes may play a role in the pathogenic potential of O104 strains. CONCLUSIONS: The two strains sequenced in this study (O104:H21 and O104:H7) are genetically more similar to each other than to the O104:H4 strains that caused an outbreak in Germany in 2011 and strains found in Central Africa. A hypothesis on strain evolution and pathogenic potential of various H-serotypes of E. coli O104 strains is proposed.

The Influence of Phase Separation on Bioactivity of Spray Pyrolyzed Bioactive Glass.

In this study, bioactive glass (BG) particles were synthesized directly using spray pyrolysis (SP). Since the bioactivity of glass particles is well correlated with their chemical composition, how to obtain homogenous bioactive glass becomes an important issue. For SP, the main reason for chemical inhomogeneity was considered to be caused by the difference in the precipitation speed of each precursor. So, two Si-containing precursors of BG, namely tetraethyl orthosilicate (TEOS) and silicon acetate (SiA), have been applied to prepare BG particles. The bioglasses were characterized by X-ray diffraction, scanning electron microscopy, transmission electron microscopy and energy dispersive spectroscopy to examine their phase composition, and surface structures, inner morphologies and chemical compositions. It was observed that, under the calcination temperature of 700 degrees C, TEOS-derived powder contained Si-rich nanoparticles and Si-deficit submicron particles as inhomogeneity, whereas the SiA-derived powder was homogenous. The reason of inhomogeneity is that TEOS dissolves in "volatile" ethanol more readily than in water via the SP mechanism of "gas-to-particle-conversion" to form Si-rich nanoparticles. The presence of Si-rich nanoparticles causes Si-deficit "wollastonite submicron particles" to form, which impairs the bioactivity. Finally, BG particle formation mechanisms from different precursors have been proposed.

The near-quantitative sampling of genomic DNA from various food-borne Eubacteria.

BACKGROUND: The disruption of the bacterial cell wall plays an important part in achieving quantitative extraction of DNA from Eubacteria essential for accurate analyses of genetic material recovered from environmental samples. RESULTS: In this work we have tested a dozen commercial bacterial genomic DNA extraction methodologies on an average of 7.70 × 10(6) (±9.05%), 4.77 × 10(8) (±31.0%), and 5.93 × 10(8) (±4.69%) colony forming units (CFU) associated with 3 cultures (n = 3) each of Brochothrix thermosphacta (Bt; Gram-positive), Shigella sonnei (Ss; Gram-negative), and Escherichia coli O79 (Ec; Gram-negative). We have utilized real-time PCR (qPCR) quantification with two specific sets of primers associated with the 16S rRNA "gene" to determine the number of copies CFU(-1) by comparing the unknown target DNA qPCR results with standards for each primer set. Based upon statistical analyses of our results, we determined that the Agencourt Genfind v2, High Pure PCR Template Prep Kit, and Omnilyse methods consistently provided the best yield of genomic DNA ranging from 141 to 934, 8 to 21, and 16 to 27 16S rDNA copies CFU(-1) for Bt, Ss, and Ec. If one assumes 6-7 copies of the 16S rRNA gene per genome, between 1 and 3 genomes per actively dividing cell and ≥ 100 cells CFU(-1) for Bt (found to be a reasonable assumption using an optical method expounded upon herein) or between 1 and 2 cells CFU(-1) for either Ss or Ec, then the Omnilyse procedure provided nearly quantitative extraction of genomic DNA from these isolates (934 ± 19.9 copies CFU(-1) for Bt; 20.8 ± 2.68 copies CFU(-1) for Ss; 26.9 ± 3.39 copies CFU(-1) for Ec). The Agencourt, High Pure, and Omnilyse technologies were subsequently assessed using 5 additional Gram-positive and 10 Gram-negative foodborne isolates (n = 3) using a set of "universal" 16S rDNA primers. CONCLUSION: Overall, the most notable DNA extraction method was found to be the Omnilyse procedure which is a "bead blender" technology involving high frequency agitation in the presence of zirconium silicate beads.

Detection and Isolation of Campylobacter spp. from Raw Meat.

This article presents a rapid yet robust protocol for isolating Campylobacter spp. from raw meats, specifically focusing on Campylobacter jejuni and Campylobacter coli. The protocol builds upon established methods, ensuring compatibility with the prevailing techniques employed by regulatory bodies such as the Food and Drug Administration (FDA) and the U.S. Department of Agriculture (USDA) in the USA, as well as the International Organization for Standardization (ISO) in Europe. Central to this protocol is collecting a rinsate, which is concentrated and resuspended in Bolton Broth media containing horse blood. This medium has been proven to facilitate the recovery of stressed Campylobacter cells and reduce the required enrichment duration by 50%. The enriched samples are then transferred onto nitrocellulose membranes on brucella plates. To improve the sensitivity and specificity of the method, 0.45 µm and 0.65 µm pore-size filter membranes were evaluated. Data revealed a 29-fold increase in cell recovery with the 0.65 µm pore-size filter compared to the 0.45 µm pore-size without impacting specificity. The highly motile characteristics of Campylobacter allow cells to actively move through the membrane filters towards the agar medium, which enables effective isolation of pure Campylobacter colonies. The protocol incorporates multiplex quantitative real-time polymerase chain reaction (mqPCR) assay to identify the isolates at the species level. This molecular technique offers a reliable and efficient means of species identification. Investigations conducted over the past twelve years involving retail meats have demonstrated the ability of this method to enhance recovery of Campylobacter from naturally contaminated meat samples compared to current reference methods. Furthermore, this protocol boasts reduced preparation and processing time. As a result, it presents a promising alternative for the efficient recovery of Campylobacter from meat. Moreover, this procedure can be seamlessly integrated with DNA-based methods, facilitating rapid screening of positive samples alongside comprehensive whole-genome sequencing analysis.

Synthesis and Characterization of Sol-Gelled Barium Zirconate as Novel MTA Radiopacifiers.

Barium zirconate (BaZrO3, BZO), which exhibits superior mechanical, thermal, and chemical stability, has been widely used in many applications. In dentistry, BZO is used as a radiopacifier in mineral trioxide aggregates (MTAs) for endodontic filling applications. In the present study, BZO was prepared using the sol-gel process, followed by calcination at 700-1000 °C. The calcined BZO powders were investigated using X-ray diffraction and scanning electron microscopy. Thereafter, MTA-like cements with the addition of calcined BZO powder were evaluated to determine the optimal composition based on radiopacity, diametral tensile strength (DTS), and setting times. The experimental results showed that calcined BZO exhibited a majority BZO phase with minor zirconia crystals. The crystallinity, the percentage, and the average crystalline size of BZO increased with the increasing calcination temperature. The optimal MTA-like cement was obtained by adding 20% of the 700 °C-calcined BZO powder. The initial and final setting times were 25 and 32 min, respectively. They were significantly shorter than those (70 and 56 min, respectively) prepared with commercial BZO powder. It exhibited a radiopacity of 3.60 ± 0.22 mmAl and a DTS of 3.02 ± 0.18 MPa. After 28 days of simulated oral environment storage, the radiopacity and DTS decreased to 3.36 ± 0.53 mmAl and 2.84 ± 0.27 MPa, respectively. This suggests that 700 °C-calcined BZO powder has potential as a novel radiopacifier for MTAs.

Effects of Calcination Temperature on the Synthesis of One-Pot Sol-Gelled Barium Titanate Powder and Its Performance as an Endodontic Radiopacifier.

Barium titanate (BaTiO3, BTO), conventionally used for dielectric and ferroelectric applications, has been assessed for biomedical applications, such as its utilization as a radiopacifier in mineral trioxide aggregates (MTA) for endodontic treatment. In the present study, BTO powders were prepared using the sol-gel process, followed by calcination at 400-1100 °C. The X-ray diffraction technique was then used to examine the as-prepared powders to elucidate the effect of calcination on the phase composition and crystalline size of BTO. Calcined BTO powders were then used as radiopacifiers for MTA. MTA-like cements were investigated to determine the optimal calcination temperature based on the radiopacity and diametral tensile strength (DTS). The experimental results showed that the formation of BTO phase was observed after calcination at temperatures of 600 °C and above. The calcined powders were a mixture of BaTiO3 phase with residual BaCO3 and/or Ba2TiO4 phases. The performance of MTA-like cements with BTO addition increased with increasing calcination temperature up to 1000 °C. The radiopacity, however, decreased after 7 days of simulated oral environmental storage, whereas an increase in DTS was observed. Optimal MTA-like cement was obtained by adding 40 wt.% 1000 °C-calcined BTO powder, with its resulting radiopacity and DTS at 4.83 ± 0.61 mmAl and 2.86 ± 0.33 MPa, respectively. After 7 days, the radiopacity decreased slightly to 4.69 ± 0.51 mmAl, accompanied by an increase in DTS to 3.13 ± 0.70 MPa. The optimal cement was biocompatible and verified using MG 63 and L929 cell lines, which exhibited cell viability higher than 95%.

Phage insertion in mlrA and variations in rpoS limit curli expression and biofilm formation in Escherichia coli serotype O157: H7.

Biofilm formation in Escherichia coli is a tightly controlled process requiring the expression of adhesive curli fibres and certain polysaccharides such as cellulose. The transcriptional regulator CsgD is central to biofilm formation, controlling the expression of the curli structural and export proteins and the diguanylate cyclase adrA, which indirectly activates cellulose production. CsgD itself is highly regulated by two sigma factors (RpoS and RpoD), multiple DNA-binding proteins, small regulatory RNAs and several GGDEF/EAL proteins acting through c-di-GMP. One such transcription factor MlrA binds the csgD promoter to enhance the RpoS-dependent transcription of csgD. Bacteriophage, often carrying the stx1 gene, utilize an insertion site in the proximal mlrA coding region of E. coli serotype O157 : H7 strains, and the loss of mlrA function would be expected to be the major factor contributing to poor curli and biofilm expression in that serotype. Using a bank of 55 strains of serotype O157 : H7, we investigated the consequences of bacteriophage insertion. Although curli/biofilm expression was restored in many of the prophage-bearing strains by a wild-type copy of mlrA on a multi-copy plasmid, more than half of the strains showed only partial or no complementation. Moreover, the two strains carrying an intact mlrA were found to be deficient in biofilm formation. However, RpoS mutations that attenuated or inactivated RpoS-dependent functions such as biofilm formation were found in >70 % of the strains, including the two strains with an intact mlrA. We conclude that bacteriophage interruption of mlrA and RpoS mutations provide major obstacles limiting curli expression and biofilm formation in most serotype O157 : H7 strains.

The influence of leader's spiritual values of servant leadership on employee motivational autonomy and eudaemonic well-being.

This research examined the role of leader's spiritual values in terms of the "servant leadership" in the process of promoting employee's autonomous motivation and eudaemonic well-being. Sample consists of 265 Chinese supervisor-subordinate dyads recruited from a variety of industries in Taiwan. Spiritual values perceived by the subordinates, as well as the discrepancy between leader-subordinate perceptions, but not the leader's self-perceptions of spiritual values, were found to contribute significantly beyond transactional leadership in predicting subordinate motivational autonomy and eudaemonic well-being, and subordinate autonomous motivations fully mediates the relationship between spiritual values and eudaemonic well-being.