bluuebunny/crossref_metadata_2025_split huggingface dataset:
Requirement of the CroRS Two-Component System for Resistance to Cell Wall-Targeting Antimicrobials in Enterococcus faecium
Authors: Kellogg Stephanie L. , Little Jaime L. , Hoff Jessica S. , Kristich Christopher J. 
Published: February 2017

ABSTRACT

Enterococci are serious opportunistic pathogens that are resistant to many cell wall-targeting antibiotics. The CroRS two-component signaling system responds to antibiotic-mediated cell wall stress and is critical for resistance to cell wall-targeting antibiotics in
Enterococcus faecalis
. Here, we identify and characterize an orthologous two-component system found in
Enterococcus faecium
that is functionally equivalent to the CroRS system of
E. faecalis
. Deletion of
croRS
in
E. faecium
resulted in marked susceptibility to cell wall-targeting agents including cephalosporins and bacitracin, as well as moderate susceptibility to ampicillin and vancomycin. As in
E. faecalis
, exposure to bacitracin and vancomycin stimulates signaling through the CroRS system in
E. faecium
. Moreover, the CroRS system is critical in
E. faecium
for enhanced beta-lactam resistance mediated by overexpression of Pbp5. Expression of a Pbp5 variant that confers enhanced beta-lactam resistance cannot overcome the requirement for CroRS function. Thus, the CroRS system is a conserved signaling system that responds to cell wall stress to promote intrinsic resistance to important cell wall-targeting antibiotics in clinically relevant enterococci.
Mutations in
bla
KPC-3
That Confer Ceftazidime-Avibactam Resistance Encode Novel KPC-3 Variants That Function as Extended-Spectrum β-Lactamases
Authors: Haidar Ghady , Clancy Cornelius J. , Shields Ryan K. , Hao Binghua , Cheng Shaoji , Nguyen M. Hong 
Published: February 2017

ABSTRACT

We identified four
bla
KPC-3
mutations in ceftazidime-avibactam-resistant clinical
Klebsiella pneumoniae
isolates, corresponding to D179Y, T243M, D179Y/T243M, and EL165-166 KPC-3 variants. Using site-directed mutagenesis and transforming vectors into
Escherichia coli
, we conclusively demonstrated that mutant
bla
KPC-3
encoded enzymes that functioned as extended-spectrum β-lactamases; mutations directly conferred higher MICs of ceftazidime-avibactam and decreased the MICs of carbapenems and other β-lactams. Impact was strongest for the D179Y mutant, highlighting the importance of the KPC Ω-loop.
Enterobacter cloacae Complex Isolates Harboring
bla
NMC-A
or
bla
IMI
-Type Class A Carbapenemase Genes on Novel Chromosomal Integrative Elements and Plasmids
Authors: Boyd David A. , Mataseje Laura F. , Davidson Ross , Delport Johannes A. , Fuller Jeff , Hoang Linda , Lefebvre Brigitte , Levett Paul N. , Roscoe Diane L. , Willey Barbara M. , Mulvey Michael R. 
Published: February 2017

ABSTRACT

Carbapenem-resistant
Enterobacter cloacae
complex isolates submitted to a reference laboratory from 2010 to 2015 were screened by PCR for seven common carbapenemase gene groups, namely, KPC, NDM, OXA-48, VIM, IMP, GES, and NMC-A/IMI. Nineteen of the submitted isolates (1.7%) were found to harbor Ambler class A
bla
NMC-A
or
bla
IMI
-type carbapenemases. All 19 isolates were resistant to at least one carbapenem but susceptible to aminoglycosides, trimethoprim-sulfamethoxazole, tigecycline, and ciprofloxacin. Most isolates (17/19) gave positive results with the Carba-NP test for phenotypic carbapenemase detection. Isolates were genetically diverse by pulsed-field gel electrophoresis macrorestriction analysis, multilocus sequence typing, and
hsp60
gene analysis. The genes were found in various
Enterobacter cloacae
complex species; however,
bla
NMC-A
was highly associated with
Enterobacter ludwigii
. Whole-genome sequencing and bioinformatics analysis revealed that all NMC-A (
n
= 10), IMI-1 (
n
= 5), and IMI-9 (
n
= 2) producers harbored the carbapenemase gene on EludIMEX-1-like integrative mobile elements (EcloIMEXs) located in the identical chromosomal locus. Two novel genes,
bla
IMI-5
and
bla
IMI-6
, were harbored on different IncFII-type plasmids.
Enterobacter cloacae
complex isolates harboring
bla
NMC-A/IMI
-type carbapenemases are relatively rare in Canada. Though mostly found integrated into the chromosome, some variants are located on plasmids that may enhance their mobility potential.
Active Multienzyme Assemblies for Long-Chain Olefinic Hydrocarbon Biosynthesis
Authors: Christenson James K. , Jensen Matthew R. , Goblirsch Brandon R. , Mohamed Fatuma , Zhang Wei , Wilmot Carrie M. , Wackett Lawrence P. 
Published: February 2017

ABSTRACT

Bacteria from different phyla produce long-chain olefinic hydrocarbons derived from an OleA-catalyzed Claisen condensation of two fatty acyl coenzyme A (acyl-CoA) substrates, followed by reduction and oxygen elimination reactions catalyzed by the proteins OleB, OleC, and OleD. In this report, OleA, OleB, OleC, and OleD were individually purified as soluble proteins, and all were found to be essential for reconstituting hydrocarbon biosynthesis. Recombinant coexpression of tagged OleABCD proteins from
Xanthomonas campestris
in
Escherichia coli
and purification over His
6
and FLAG columns resulted in OleA separating, while OleBCD purified together, irrespective of which of the four Ole proteins were tagged. Hydrocarbon biosynthetic activity of copurified OleBCD assemblies could be reconstituted by adding separately purified OleA. Immunoblots of nondenaturing gels using anti-OleC reacted with
X. campestris
crude protein lysate indicated the presence of a large protein assembly containing OleC in the native host. Negative-stain electron microscopy of recombinant OleBCD revealed distinct large structures with diameters primarily between 24 and 40 nm. Assembling OleB, OleC, and OleD into a complex may be important to maintain stereochemical integrity of intermediates, facilitate the movement of hydrophobic metabolites between enzyme active sites, and protect the cell against the highly reactive β-lactone intermediate produced by the OleC-catalyzed reaction.

IMPORTANCE
Bacteria biosynthesize hydrophobic molecules to maintain a membrane, store carbon, and for antibiotics that help them survive in their niche. The hydrophobic compounds are often synthesized by a multidomain protein or by large multienzyme assemblies. The present study reports on the discovery that long-chain olefinic hydrocarbons made by bacteria from different phyla are produced by multienzyme assemblies in
X. campestris
. The OleBCD multienzyme assemblies are thought to compartmentalize and sequester olefin biosynthesis from the rest of the cell. This system provides additional insights into how bacteria control specific biosynthetic pathways.
Heterogeneity Confounds Establishment of “a” Model Microbial Strain
Authors: Keller Nancy P. 
Published: February 2017

ABSTRACT

Aspergillus fumigatus
is a ubiquitous environmental mold and the leading cause of diverse human diseases ranging from allergenic bronchopulmonary aspergillosis (ABPA) to invasive pulmonary aspergillosis (IPA). Experimental investigations of the biology and virulence of this opportunistic pathogen have historically used a few type strains; however, it is increasingly observed with this fungus that heterogeneity among isolates potentially confounds the use of these reference isolates. Illustrating this point, Kowalski et al. (mBio 7:e01515-16, 2016,
https://doi.org/10.1128/mBio.01515-16
) demonstrated that variation in 16 environmental and clinical isolates of
A. fumigatus
correlated virulence with fitness in low oxygen, whereas Fuller et al. (mBio 7:e01517-16, 2016,
https://doi.org/10.1128/mBio.01517-16
) showed wide variation in light responses at a physiological and protein functionality level in 15
A. fumigatus
isolates. In both studies, two commonly used type strains, Af293 and CEA10, displayed significant differences in physiological responses to abiotic stimuli and virulence in a murine model of IPA.
A Natural ChimericPseudomonasBacteriocin with Novel Pore-Forming Activity Parasitizes the Ferrichrome Transporter
Authors: Ghequire Maarten G. K. , Kemland Lieselore , Anoz-Carbonell Ernesto , Buchanan Susan K. , De Mot René 
Published: February 2017

ABSTRACTModular bacteriocins represent a major group of secreted protein toxins with a narrow spectrum of activity, involved in interference competition between Gram-negative bacteria. These antibacterial proteins include a domain for binding to the target cell and a toxin module at the carboxy terminus. Self-inhibition of producers is provided by coexpression of linked immunity genes that transiently inhibit the toxin’s activity through formation of bacteriocin-immunity complexes or by insertion in the inner membrane, depending on the type of toxin module. We demonstrate strain-specific inhibitory activity for PmnH, aPseudomonasbacteriocin with an unprecedented dual-toxin architecture, hosting both a colicin M domain, potentially interfering with peptidoglycan synthesis, and a novel colicin N-type domain, a pore-forming module distinct from the colicin Ia-type domain inPseudomonas aeruginosapyocin S5. A downstream-linked gene product confers PmnH immunity upon susceptible strains. This protein, ImnH, has a transmembrane topology similar to that ofPseudomonascolicin M-like and pore-forming immunity proteins, although homology with either of these is essentially absent. The enhanced killing activity of PmnH under iron-limited growth conditions reflects parasitism of the ferrichrome-type transporter for entry into target cells, a strategy shown here to be used as well by monodomain colicin M-like bacteriocins from pseudomonads. The integration of a second type of toxin module in a bacteriocin gene could offer a competitive advantage against bacteria displaying immunity against only one of both toxic activities.IMPORTANCEIn their continuous struggle for ecological space, bacteria face a huge load of contenders, including phylogenetically related strains that compete for the same niche. One important group of secreted antibacterial proteins assisting in eliminating these rivals are modular bacteriocins of Gram-negative bacteria, comprising a domain for docking onto the cell envelope of a target cell, a translocation domain enabling subsequent cellular entry, and a toxin module that kills target cells via enzymatic or pore-forming activity. We here demonstrate the antagonistic function of aPseudomonasbacteriocin with unique architecture that combines a putative enzymatic colicin M-like domain and a novel pore-forming toxin module. For target cell recognition and entry, this bacteriocin hybrid takes advantage of the ferrichrome transporter, also parasitized by enzymaticPseudomonasbacteriocins devoid of the pore-forming module. Bacteriocins with an expanded toxin potential may represent an inventive bacterial strategy to alleviate immunity in target cells.
Evolution and Epidemiology of Multidrug-Resistant
Klebsiella pneumoniae
in the United Kingdom and Ireland
Authors: Moradigaravand Danesh , Martin Veronique , Peacock Sharon J. , Parkhill Julian 
Published: February 2017

ABSTRACT

Klebsiella pneumoniae
is a human commensal and opportunistic pathogen that has become a leading causative agent of hospital-based infections over the past few decades. The emergence and global expansion of hypervirulent and multidrug-resistant (MDR) clones of
K. pneumoniae
have been increasingly reported in community-acquired and nosocomial infections. Despite this, the population genomics and epidemiology of MDR
K. pneumoniae
at the national level are still poorly understood. To obtain insights into these, we analyzed a systematic large-scale collection of invasive MDR
K. pneumoniae
isolates from hospitals across the United Kingdom and Ireland. Using whole-genome phylogenetic analysis, we placed these in the context of previously sequenced
K. pneumoniae
populations from geographically diverse countries and identified their virulence and drug resistance determinants. Our results demonstrate that United Kingdom and Ireland MDR isolates are a highly diverse population drawn from across the global phylogenetic tree of
K. pneumoniae
and represent multiple recent international introductions that are mainly from Europe but in some cases from more distant countries. In addition, we identified novel genetic determinants underlying resistance to beta-lactams, gentamicin, ciprofloxacin, and tetracyclines, indicating that both increased virulence and resistance have emerged independently multiple times throughout the population. Our data show that MDR
K. pneumoniae
isolates in the United Kingdom and Ireland have multiple distinct origins and appear to be part of a globally circulating
K. pneumoniae
population.

IMPORTANCE
Klebsiella pneumoniae
is a major human pathogen that has been implicated in infections in healthcare settings over the past few decades. Antimicrobial treatment of
K. pneumoniae
infections has become increasingly difficult as a consequence of the emergence and spread of strains that are resistant to multiple antimicrobials. To better understand the spread of resistant
K. pneumoniae
, we studied the genomes of a large-scale population of extensively antimicrobial-resistant
K. pneumoniae
in the United Kingdom and Ireland by utilizing the fine resolution that whole-genome sequencing of pathogen genomes provides. Our results indicate that the
K. pneumoniae
population is highly diverse and that, in some cases, resistant strains appear to have spread across the country over a few years. In addition, we found evidence that some strains have acquired antimicrobial resistance genes independently, presumably in response to antimicrobial treatment.
Identification of a
Pseudomonas aeruginosa
PAO1 DNA Methyltransferase, Its Targets, and Physiological Roles
Authors: Doberenz Sebastian , Eckweiler Denitsa , Reichert Olga , Jensen Vanessa , Bunk Boyke , Spröer Cathrin , Kordes Adrian , Frangipani Emanuela , Luong Khai , Korlach Jonas , Heeb Stephan , Overmann Jörg , Kaever Volkhard , Häussler Susanne 
Published: February 2017

ABSTRACT

DNA methylation is widespread among prokaryotes, and most DNA methylation reactions are catalyzed by adenine DNA methyltransferases, which are part of restriction-modification (R-M) systems. R-M systems are known for their role in the defense against foreign DNA; however, DNA methyltransferases also play functional roles in gene regulation. In this study, we used single-molecule real-time (SMRT) sequencing to uncover the genome-wide DNA methylation pattern in the opportunistic pathogen
Pseudomonas aeruginosa
PAO1. We identified a conserved sequence motif targeted by an adenine methyltransferase of a type I R-M system and quantified the presence of N
6
-methyladenine using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Changes in the PAO1 methylation status were dependent on growth conditions and affected
P. aeruginosa
pathogenicity in a
Galleria mellonella
infection model. Furthermore, we found that methylated motifs in promoter regions led to shifts in sense and antisense gene expression, emphasizing the role of enzymatic DNA methylation as an epigenetic control of phenotypic traits in
P. aeruginosa
. Since the DNA methylation enzymes are not encoded in the core genome, our findings illustrate how the acquisition of accessory genes can shape the global
P. aeruginosa
transcriptome and thus may facilitate adaptation to new and challenging habitats.

IMPORTANCE
With the introduction of advanced technologies, epigenetic regulation by DNA methyltransferases in bacteria has become a subject of intense studies. Here we identified an adenosine DNA methyltransferase in the opportunistic pathogen
Pseudomonas aeruginosa
PAO1, which is responsible for DNA methylation of a conserved sequence motif. The methylation level of all target sequences throughout the PAO1 genome was approximated to be in the range of 65 to 85% and was dependent on growth conditions. Inactivation of the methyltransferase revealed an attenuated-virulence phenotype in the
Galleria mellonella
infection model. Furthermore, differential expression of more than 90 genes was detected, including the small regulatory RNA
prrF1
, which contributes to a global iron-sparing response via the repression of a set of gene targets. Our finding of a methylation-dependent repression of the antisense transcript of the
prrF1
small regulatory RNA significantly expands our understanding of the regulatory mechanisms underlying active DNA methylation in bacteria.
Fault-controlled dolomitization in a rift basin
Authors: Hollis Cathy , Bastesen Eivind , Boyce Adrian , Corlett Hilary , Gawthorpe Robert , Hirani Jesal , Rotevatn Atle , Whitaker Fiona 
Published: February 2017

Abstract
There are numerous examples of fault-controlled, so-called hydrothermal dolomite (HTD), many of which host economic mineral deposits or hydrocarbons, but there remains a lack of consensus as to how they form. In particular, multiple phases of diagenetic overprinting can obscure geochemical fingerprints. Study of a Cenozoic succession with a relatively simple burial history here provides new insights into the development of differentially dolomitized beds. The Hammam Faraun fault (HFF) block within the Suez Rift, Egypt, hosts both massive and stratabound dolostone bodies. Non-fabric-selective massive dolostone is limited to the damage zone of the fault, while fabric-selective stratabound dolostone bodies penetrate nearly 2 km into the footwall. Oligo-Miocene seawater is interpreted to have been drawn down discrete faults into a deep aquifer and convected upwards along the HFF. Escape of fluids from the incipient HFF into the lower Thebes Formation led to differential, stratabound dolomitization. Once the HFF breached the surface, fluid circulation focused along the fault plane to form younger, massive dolostone bodies. This study provides a snapshot of dolomitization during the earliest phases of extension, unobscured by subsequent recrystallization and geochemical modification. Contrary to many models, stratabound dolomitization preceded non-stratabound dolomitization. Fluids were hydrothermal, but with little evidence for rapid cooling and brecciation common to many HTD bodies. These results suggest that many of the features used to interpret and predict the geometry of HTD in the subsurface form during later phases of structural deformation, perhaps overprinting less structurally complex dolomite bodies.
The mechanical, optical and thermoelectric properties of MCoF3(M = K and Rb) compounds
Authors: Mubarak A. A. 
Published: February 2017

This is an ab initio study instituted on the density functional theory (DFT) and the full-potential linearized augmented plane wave (FP-LAPW) calculations that are performed to analyze the mechanical, electronic, optical and thermoelectric properties of the cubic MCoF3compound (M = K and Rb). The studied compounds are found thermodynamically and mechanically stable. Moreover, these compounds are found to be elastically anisotropic and ductile. KCoF3and RbCoF3are classified as half-metallic and anti-ferromagnetic compounds. The optical properties are investigated from the dielectric function for the different energy ranges. The thermoelectric properties such as transport properties are determined as a function of temperature using BoltzTrape code in the range of 20–800 K. The present compounds are found to have p-type character. Also, the majority charge carriers are found to be electrons rather than hole. Useful mechanical, spintronic, optical and thermoelectric applications are predicted based upon the calculations.
Fixation target representation in prefrontal cortex during the antisaccade task
Authors: Zhou Xin , Constantinidis Christos 
Published: February 2017

Neurons that discharge strongly during the time period of fixation of a visual target and cease to discharge before saccade initiation have been described in the brain stem, superior colliculus, and cortical areas. In subcortical structures, fixation neurons play a reciprocal role with saccadic neurons during the generation of eye movements. Their role in the dorsolateral prefrontal cortex is less obvious, and it is not known if they are activated by fixation, inhibit saccade generation, or play a role in more complex functions such as the inhibition of inappropriate responses. We examined the properties of prefrontal fixation neurons in the context of an antisaccade task, which requires an eye movement directed away from a prepotent visual stimulus. We tested monkeys with variants of the task, allowing us to dissociate activity synchronized on the fixation offset, presentation of the visual stimulus, and saccadic onset. Fixation neuron activity latency was most strongly tied to the offset of the fixation point across task variants. It was not well predicted by the appearance of the visual stimulus, which is essential for planning of the correct eye movement and inhibiting inappropriate ones. Activity of fixation neurons was generally negatively correlated with that of saccade neurons; however, critical differences in timing make it unlikely that they provide precisely timed signals for the generation of eye movements. These results demonstrate the role of fixation neurons in the prefrontal cortex during tasks requiring timing of appropriate eye movement and inhibition of inappropriate actions. NEW & NOTEWORTHY Properties of neurons that discharge during eye fixation and go silent before saccade initiation have been described in subcortical structures involved in eye movement generation, but their role in the dorsolateral prefrontal cortex presents a puzzle. Our results demonstrate the role of fixation neurons in the prefrontal cortex during tasks requiring precise timing of appropriate eye movement and inhibition of inappropriate actions.
HSP70 Inhibition Synergistically Enhances the Effects of Magnetic Fluid Hyperthermia in Ovarian Cancer
Authors: Court Karem A. , Hatakeyama Hiroto , Wu Sherry Y. , Lingegowda Mangala S. , Rodríguez-Aguayo Cristian , López-Berestein Gabriel , Ju-Seog Lee , Rinaldi Carlos , Juan Eduardo J. , Sood Anil K. , Torres-Lugo Madeline 
Published: February 2017

Abstract
Hyperthermia has been investigated as a potential treatment for cancer. However, specificity in hyperthermia application remains a significant challenge. Magnetic fluid hyperthermia (MFH) may be an alternative to surpass such a challenge, but implications of MFH at the cellular level are not well understood. Therefore, the present work focused on the examination of gene expression after MFH treatment and using such information to identify target genes that when inhibited could produce an enhanced therapeutic outcome after MFH. Genomic analyzes were performed using ovarian cancer cells exposed to MFH for 30 minutes at 43°C, which revealed that heat shock protein (HSP) genes, including HSPA6, were upregulated. HSPA6 encodes the Hsp70, and its expression was confirmed by PCR in HeyA8 and A2780cp20 ovarian cancer cells. Two strategies were investigated to inhibit Hsp70-related genes, siRNA and Hsp70 protein function inhibition by 2-phenylethyenesulfonamide (PES). Both strategies resulted in decreased cell viability following exposure to MFH. Combination index was calculated for PES treatment reporting a synergistic effect. In vivo efficacy experiments with HSPA6 siRNA and MFH were performed using the A2780cp20 and HeyA8 ovarian cancer mouse models. A significantly reduction in tumor growth rate was observed with combination therapy. PES and MFH efficacy were also evaluated in the HeyA8 intraperitoneal tumor model, and resulted in robust antitumor effects. This work demonstrated that HSP70 inhibition combination with MFH generate a synergistic effect and could be a promising target to enhance MFH therapeutic outcomes in ovarian cancer. Mol Cancer Ther; 16(5); 966–76. ©2017 AACR.