bluuebunny/crossref_metadata_2025_split huggingface dataset:
Loss of β2‐laminin alters calcium sensitivity and voltage‐gated calcium channel maturation of neurotransmission at the neuromuscular junction
Authors: Chand Kirat K. , Lee Kah Meng , Schenning Mitja P. , Lavidis Nickolas A. , Noakes Peter G. 
Published: November 2014

Key points
Neuromuscular junctions from β2‐laminin‐deficient mice exhibit lower levels of calcium sensitivity.
Loss of β2‐laminin leads to a failure in switching from N‐ to P/Q‐type voltage‐gated calcium channel (VGCC)‐mediated transmitter release that normally occurs with neuromuscular junction maturation.
The motor nerve terminals from β2‐laminin‐deficient mice fail to up‐regulate the expression of P/Q‐type VGCCs clusters and down‐regulate N‐type VGCCs clusters, as they mature.
There is decreased co‐localisation of presynaptic specialisations in β2‐laminin‐deficient neuromuscular junctions as a consequence of lesser P/Q‐type VGCC expression.
These findings support the idea that β2‐laminin is critical in the organisation and maintenance of active zones at the neuromuscular junction via its interaction with P/Q‐type VGCCs, which aid in stabilisation of the synapse.
Abstractβ2‐laminin is a key mediator in the differentiation and formation of the skeletal neuromuscular junction. Loss of β2‐laminin results in significant structural and functional aberrations such as decreased number of active zones and reduced spontaneous release of transmitter. In vitro β2‐laminin has been shown to bind directly to the pore forming subunit of P/Q‐type voltage‐gated calcium channels (VGCCs). Neurotransmission is initially mediated by N‐type VGCCs, but by postnatal day 18 switches to P/Q‐type VGCC dominance. The present study investigated the changes in neurotransmission during the switch from N‐ to P/Q‐type VGCC‐mediated transmitter release at β2‐laminin‐deficient junctions. Analysis of the relationship between quantal content and extracellular calcium concentrations demonstrated a decrease in the calcium sensitivity, but no change in calcium dependence at β2‐laminin‐deficient junctions. Electrophysiological studies on VGCC sub‐types involved in transmitter release indicate N‐type VGCCs remain the primary mediator of transmitter release at matured β2‐laminin‐deficient junctions. Immunohistochemical analyses displayed irregularly shaped and immature β2‐laminin‐deficient neuromuscular junctions when compared to matured wild‐type junctions. β2‐laminin‐deficient junctions also maintained the presence of N‐type VGCC clustering within the presynaptic membrane, which supported the functional findings of the present study. We conclude that β2‐laminin is a key regulator in development of the NMJ, with its loss resulting in reduced transmitter release due to decreased calcium sensitivity stemming from a failure to switch from N‐ to P/Q‐type VGCC‐mediated synaptic transmission.
Effects of spin‐lock field direction on the quantitative measurement of spin‐lattice relaxation time constant in the rotating frame (T1ρ) in a clinical MRI system
Authors: Yee Seonghwan , Gao Jia‐Hong 
Published: November 2014

Purpose:To investigate whether the direction of spin‐lock field, either parallel or antiparallel to the rotating magnetization, has any effect on the spin‐lock MRI signal and further on the quantitative measurement of T1ρ, in a clinical 3 T MRI system.Methods:The effects of inverted spin‐lock field direction were investigated by acquiring a series of spin‐lock MRI signals for an American College of Radiology MRI phantom, while the spin‐lock field direction was switched between the parallel and antiparallel directions. The acquisition was performed for different spin‐locking methods (i.e., for the single‐ and dual‐field spin‐locking methods) and for different levels of clinically feasible spin‐lock field strength, ranging from 100 to 500 Hz, while the spin‐lock duration was varied in the range from 0 to 100 ms.Results:When the spin‐lock field was inverted into the antiparallel direction, the rate of MRI signal decay was altered and the T1ρ value, when compared to the value for the parallel field, was clearly different. Different degrees of such direction‐dependency were observed for different spin‐lock field strengths. In addition, the dependency was much smaller when the parallel and the antiparallel fields are mixed together in the dual‐field method.Conclusions:The spin‐lock field direction could impact the MRI signal and further the T1ρ measurement in a clinical MRI system.
Pharmacokinetics, Microbial Response, and Pulmonary Outcomes of Multidose Intravenous Azithromycin in Preterm Infants at Risk for Ureaplasma Respiratory Colonization
Authors: Merchan L. Marcela , Hassan Hazem E. , Terrin Michael L. , Waites Ken B. , Kaufman David A. , Ambalavanan Namasivayam , Donohue Pamela , Dulkerian Susan J. , Schelonka Robert , Magder Laurence S. , Shukla Sagar , Eddington Natalie D. , Viscardi Rose M. 
Published: November 2014

ABSTRACT

The study objectives were to refine the population pharmacokinetics (PK) model, determine microbial clearance, and assess short-term pulmonary outcomes of multiple-dose azithromycin treatment in preterm infants at risk for
Ureaplasma
respiratory colonization. Fifteen subjects (7 of whom were
Ureaplasma
positive) received intravenous azithromycin at 20 mg/kg of body weight every 24 h for 3 doses. Azithromycin concentrations were determined in plasma samples obtained up to 168 h post-first dose by using a validated liquid chromatography-tandem mass spectrometry method. Respiratory samples were obtained predose and at three time points post-last dose for
Ureaplasma
culture, PCR, antibiotic susceptibility testing, and cytokine concentration determinations. Pharmacokinetic data from these 15 subjects as well as 25 additional subjects (who received either a single 10-mg/kg dose [
n
= 12] or a single 20-mg/kg dose [
n
= 13]) were analyzed by using a nonlinear mixed-effect population modeling (NONMEM) approach. Pulmonary outcomes were assessed at 36 weeks post-menstrual age and 6 months adjusted age. A 2-compartment model with all PK parameters allometrically scaled on body weight best described the azithromycin pharmacokinetics in preterm neonates. The population pharmacokinetics parameter estimates for clearance, central volume of distribution, intercompartmental clearance, and peripheral volume of distribution were 0.15 liters/h · kg
0.75
, 1.88 liters · kg, 1.79 liters/h · kg
0.75
, and 13 liters · kg, respectively. The estimated area under the concentration-time curve over 24 h (AUC
24
)/MIC
90
value was ∼4 h. All posttreatment cultures were negative, and there were no drug-related adverse events. One
Ureaplasma
-positive infant died at 4 months of age, but no survivors were hospitalized for respiratory etiologies during the first 6 months (adjusted age). Thus, a 3-day course of 20 mg/kg/day intravenous azithromycin shows preliminary efficacy in eradicating
Ureaplasma
spp. from the preterm respiratory tract.
Clostridium difficile Isolates with High Linezolid MICs Harbor the Multiresistance Gene
cfr
Authors: Marín Mercedes , Martín Adoración , Alcalá Luis , Cercenado Emilia , Iglesias Cristina , Reigadas Elena , Bouza Emilio 
Published: November 2014

ABSTRACT

We studied the molecular mechanisms of linezolid resistance in 9 isolates of toxigenic
Clostridium difficile
with high linezolid MICs. The activity of linezolid was determined against 891 clinical isolates of toxigenic
C. difficile
. The MIC
50
and MIC
90
of linezolid were 0.75 μg/ml and 1.5 μg/ml, respectively. Nine strains (1%) showed high linezolid MICs (6 μg/ml to 16 μg/ml) and also were resistant to clindamycin, erythromycin, and chloramphenicol. These strains were selected for molecular studies: sequencing of domain V of the
23 rRNA
gene, detection of the
cfr
methyltransferase gene, and sequencing of the ribosomal protein genes
rplC
and
rplD
. Molecular relatedness between strains was assessed using PCR ribotyping and MLVA (multilocus variable-number tandem-repeat analysis) typing. The strains belonged to ribotypes 001 (2/9), 017 (6/9), and 078 (1/9). MLVA showed that strains of ribotype 001 and 017 belonged to the same clonal complex in each ribotype. We did not detect mutations in the 23S rRNA gene. The
cfr
gene was detected in 7 of 9 strains. Sequencing of
cfr
amplicons revealed a similarity of 100% to a fragment of transposon Tn
6218
of
C. difficile
, which was annotated as a putative chloramphenicol/florfenicol resistance protein. We were unable to detect mechanisms of resistance to linezolid in the 2 strains belonging to ribotype 001. While the relevance of our results lies in the detection of the
cfr
gene as a possible mechanism of resistance to linezolid in
C. difficile
, our findings should be assessed by further investigations to characterize these possible
cfr
genes and their contribution to linezolid resistance.
In Vitro
Resistance Selection with Doravirine (MK-1439), a Novel Nonnucleoside Reverse Transcriptase Inhibitor with Distinct Mutation Development Pathways
Authors: Feng Meizhen , Wang Deping , Grobler Jay A. , Hazuda Daria J. , Miller Michael D. , Lai Ming-Tain 
Published: November 2014

ABSTRACT

Doravirine (DOR, formerly known as MK-1439) is a human immunodeficiency type 1 virus (HIV-1) nonnucleoside reverse transcriptase inhibitor (NNRTI) that is currently in phase 2b clinical trials.
In vitro
resistance selection of subtype B virus (MT4-green fluorescent protein [GFP] cells), as well as subtype A and C viruses (MT4-GFP/CCR5 cells) was conducted with DOR, rilpivirine (RPV), and efavirine (EFV) under low-multiplicity-of-infection conditions in a 96-well format. Resistance selection was performed with escalating concentrations of the NNRTIs ranging from the 95% effective concentration (1× EC
95
) to 1,000× EC
95
in the presence of 10% fetal bovine serum. In the resistance selection of subtype B virus with DOR, a V106A mutant virus led to two mutation pathways, followed by the emergence separately of either F227L or L234I. In the resistance selection of subtype A and C viruses, similar mutation development pathways were detected, in which a V106A or V106M mutant was also the starting virus in the pathways. Mutations that are commonly associated with RPV and EFV in clinical settings were also identified in subtype B viruses such as the E138K and K103N mutants, respectively, in this
in vitro
resistance selection study. The susceptibility of subtype B mutant viruses selected by DOR, RPV, and EFV to NNRTIs was evaluated. Results suggest that mutant viruses selected by DOR are susceptible to RPV and EFV and mutants selected by RPV and EFV are susceptible to DOR. When the replication capacity of the V106A mutant was compared with that of the wild-type (WT) virus, the mutant virus was 4-fold less fit than the WT virus.
Trends in Serotype Distribution and Antimicrobial Susceptibility in Salmonella enterica Isolates from Humans in Belgium, 2009 to 2013
Authors: Ceyssens Pieter-Jan , Mattheus Wesley , Vanhoof Raymond , Bertrand Sophie 
Published: November 2014

ABSTRACT

The Belgian National Reference Centre for
Salmonella
received 16,544 human isolates of
Salmonella enterica
between January 2009 and December 2013. Although 377 different serotypes were identified, the landscape is dominated by
S. enterica
serovars Typhimurium (55%) and Enteritidis (19%) in a ratio which is inverse to European Union averages. With outbreaks of
Salmonella
serotypes Ohio, Stanley, and Paratyphi B variant Java as prime examples, 20 serotypes displayed significant fluctuations in this 5-year period. Typhoid strains account for 1.2% of Belgian salmonellosis cases. Large-scale antibiotic susceptibility analyses (
n
= 4,561; panel of 12 antibiotics) showed declining resistance levels in
S
. Enteritis and Typhimurium isolates for 8 and 3 tested agents, respectively. Despite low overall resistance to ciprofloxacin (4.4%) and cefotaxime (1.6%), we identified clonal lineages of
Salmonella
serotypes Kentucky and Infantis displaying rising resistance against these clinically important drugs. Quinolone resistance is mainly mediated by serotype-specific mutations in GyrA residues Ser83 and Asp87 (92.2% not wild type), while an additional ParC_Ser80Ile mutation leads to ciprofloxacin resistance in 95.5%
S
. Kentucky isolates, which exceeds European averages. Plasmid-mediated quinolone resistance (PMQR) alleles
qnrA1
(
n
= 1),
qnrS
(
n
= 9),
qnrD1
(
n
= 4), and
qnrB
(
n
= 4) were found in only 3.0% of 533 isolates resistant to nalidixic acid. In cefotaxime-resistant isolates, we identified a broad range of Ambler class A and C β-lactamase genes (e.g.,
bla
SHV-12
,
bla
TEM-52
,
bla
CTX-M-14
, and
bla
CTX-M-15
) commonly associated with members of the family
Enterobacteriaceae
. In conclusion, resistance to fluoroquinolones and cefotaxime remains rare in human
S. enterica
, but clonal resistant serotypes arise, and continued (inter)national surveillance is mandatory to understand the origin and routes of dissemination thereof.
Disparities in Capreomycin Resistance Levels Associated with the
rrs
A1401G Mutation in Clinical Isolates of Mycobacterium tuberculosis
Authors: Reeves Analise Z. , Campbell Patricia J. , Willby Melisa J. , Posey James E. 
Published: November 2014

ABSTRACT

As the prevalence of multidrug-resistant and extensively drug-resistant tuberculosis strains continues to rise, so does the need to develop accurate and rapid molecular tests to complement time-consuming growth-based drug susceptibility testing. Performance of molecular methods relies on the association of specific mutations with phenotypic drug resistance and while considerable progress has been made for resistance detection of first-line antituberculosis drugs, rapid detection of resistance for second-line drugs lags behind. The
rrs
A1401G allele is considered a strong predictor of cross-resistance between the three second-line injectable drugs, capreomycin (CAP), kanamycin, and amikacin. However, discordance is often observed between the
rrs
A1401G mutation and CAP resistance, with up to 40% of
rrs
A1401G mutants being classified as CAP susceptible. We measured the MICs to CAP in 53 clinical isolates harboring the
rrs
A1401G mutation and found that the CAP MICs ranged from 8 μg/ml to 40 μg/ml. These results were drastically different from engineered A1401G mutants generated in isogenic
Mycobacterium tuberculosis
, which exclusively exhibited high-level CAP MICs of 40 μg/ml. These data support the results of prior studies, which suggest that the critical concentration of CAP (10 μg/ml) used to determine resistance by indirect agar proportion may be too high to detect all CAP-resistant strains and suggest that a larger percentage of resistant isolates could be identified by lowering the critical concentration. These data also suggest that differences in resistance levels among clinical isolates are possibly due to second site or compensatory mutations located elsewhere in the genome.
Neutralization of Mitochondrial Superoxide by Superoxide Dismutase 2 Promotes Bacterial Clearance and Regulates Phagocyte Numbers in Zebrafish
Authors: Peterman E. M. , Sullivan C. , Goody M. F. , Rodriguez-Nunez I. , Yoder J. A. , Kim C. H. 
Published: November 2014

ABSTRACT

Mitochondria are known primarily as the location of the electron transport chain and energy production in cells. More recently, mitochondria have been shown to be signaling centers for apoptosis and inflammation. Reactive oxygen species (ROS) generated as by-products of the electron transport chain within mitochondria significantly impact cellular signaling pathways. Because of the toxic nature of ROS, mitochondria possess an antioxidant enzyme, superoxide dismutase 2 (SOD2), to neutralize ROS. If mitochondrial antioxidant enzymes are overwhelmed during severe infections, mitochondrial dysfunction can occur and lead to multiorgan failure or death.
Pseudomonas aeruginosa
is an opportunistic pathogen that can infect immunocompromised patients. Infochemicals and exotoxins associated with
P. aeruginosa
are capable of causing mitochondrial dysfunction. In this work, we describe the roles of SOD2 and mitochondrial ROS regulation in the zebrafish innate immune response to
P. aeruginosa
infection.
sod2
is upregulated in mammalian macrophages and neutrophils in response to lipopolysaccharide
in vitro
, and
sod2
knockdown in zebrafish results in an increased bacterial burden. Further investigation revealed that phagocyte numbers are compromised in Sod2-deficient zebrafish. Addition of the mitochondrion-targeted ROS-scavenging chemical MitoTEMPO rescues neutrophil numbers and reduces the bacterial burden in Sod2-deficient zebrafish. Our work highlights the importance of mitochondrial ROS regulation by SOD2 in the context of innate immunity and supports the use of mitochondrion-targeted ROS scavengers as potential adjuvant therapies during severe infections.
Enterococcus faecalis 6-Phosphogluconolactonase Is Required for Both Commensal and Pathogenic Interactions with Manduca sexta
Authors: Holt Jonathan F. , Kiedrowski Megan R. , Frank Kristi L. , Du Jing , Guan Changhui , Broderick Nichole A. , Dunny Gary M. , Handelsman Jo 
Published: November 2014

ABSTRACT

Enterococcus faecalis
is a commensal and pathogen of humans and insects. In
Manduca sexta
,
E. faecalis
is an infrequent member of the commensal gut community, but its translocation to the hemocoel results in a commensal-to-pathogen switch. To investigate
E. faecalis
factors required for commensalism, we identified
E. faecalis
genes that are upregulated in the gut of
M. sexta
using recombinase-based
in vivo
expression technology (RIVET). The RIVET screen produced 113 clones, from which we identified 50 genes that are more highly expressed in the insect gut than in culture. The most frequently recovered gene was locus OG1RF_11582, which encodes a 6-phosphogluconolactonase that we designated
pglA
. A
pglA
deletion mutant was impaired in both pathogenesis and gut persistence in
M. sexta
and produced enhanced biofilms compared with the wild type in an
in vitro
polystyrene plate assay. Mutation of four other genes identified by RIVET did not affect persistence in caterpillar guts but led to impaired pathogenesis. This is the first identification of genetic determinants for
E. faecalis
commensal and pathogenic interactions with
M. sexta
. Bacterial factors identified in this model system may provide insight into colonization or persistence in other host-associated microbial communities and represent potential targets for interventions to prevent
E. faecalis
infections.
What Is a Host? Incorporating the Microbiota into the Damage-Response Framework
Authors: Casadevall Arturo , Pirofski Liise-anne 
Published: November 2014

ABSTRACT
Since proof of the germ theory of disease in the late 19th century, a major focus of the fields of microbiology and infectious diseases has been to seek differences between pathogenic and nonpathogenic microbes and the role that the host plays in microbial pathogenesis. Remarkably, despite the increasing recognition that host immunity plays a role in microbial pathogenesis, there has been little discussion about what constitutes a host. Historically, hosts have been viewed in the context of their fitness or immunological status and characterized by adjectives such as immune, immunocompetent, immunosuppressed, immunocompromised, or immunologically impaired. However, in recent years it has become apparent that the microbiota has profound effects on host homeostasis and susceptibility to microbial diseases in addition to its effects on host immunity. This raises the question of how to incorporate the microbiota into defining a host. This definitional problem is further complicated because neither host nor microbial properties are adequate to predict the outcome of host-microbe interaction because this outcome exhibits emergent properties. In this essay, we revisit the damage-response framework (DRF) of microbial pathogenesis and demonstrate how it can incorporate the rapidly accumulating information being generated by the microbiome revolution. We use the tenets of the DRF to put forth the following definition of a host: a host is an entity that houses an associated microbiome/microbiota and interacts with microbes such that the outcome results in damage, benefit, or indifference, thus resulting in the states of symbiosis, colonization, commensalism, latency, and disease.
Prolonged Growth of a Clinical Staphylococcus aureus Strain Selects for a Stable Small-Colony-Variant Cell Type
Authors: Bui Long M. G. , Hoffmann Peter , Turnidge John D. , Zilm Peter S. , Kidd Stephen P. 
Published: November 2014

ABSTRACT

An undetermined feature of
Staphylococcus aureus
pathogenesis is its persistence and then relapse of disease. This has been explained by its switch to alternative lifestyles, mainly as biofilm or small-colony variants (SCVs). Studying the native characteristics of SCVs has been problematic due to their reversion to the parental lifestyle. We have observed that for a number of
S. aureus
strains as they switch to an SCV lifestyle, there is the formation of an extracellular matrix. We focused our analysis on one strain, WCH-SK2. For bacterial survival in the host, the combination of low nutrients and the prolonged time frame forms a stress that selects for a specific cell type from the population. In this context, we used steady-state growth conditions with low nutrients and a controlled low growth rate for a prolonged time and with methylglyoxal. These conditions induced
S. aureus
WCH-SK2 into a stable SCV cell type; the cells did not revert after subculturing. Analysis revealed these cells possessed a metabolic and surface profile that was different from those of previously described SCVs or biofilm cells. The extracellular matrix was protein and extracellular DNA but not polysaccharide. The SCV cells induced expression of certain surface proteins (such as Ebh) and synthesis of lantibiotics while downregulating factors that stimulate the immune response (leucocidin, capsule, and carotenoid). Our data reveal cell heterogeneity within an
S. aureus
population and under conditions that resemble long-term survival in the host have identified a previously unnoticed
S. aureus
cell type with a distinctive metabolic and molecular profile.
An observational, longitudinal study on the home environment of people with chronic obstructive pulmonary disease: the research protocol of theHome Sweet Homestudy
Authors: Nakken Nienke , Janssen Daisy J A , van den Bogaart Esther H A , Vercoulen Jan H , Wouters Emiel F M , Spruit Martijn A 
Published: November 2014

IntroductionChronic obstructive pulmonary disease (COPD) represents an important public health challenge. Patients are confronted with limitations during activities of daily living (ADLs). Resident loved ones of patients with COPD may be uniquely positioned to witness these limitations. COPD may have an impact on not only the patients’ life, but also on the lives of the resident loved ones. Furthermore, COPD exacerbation-related hospital admissions often occur in patients with COPD. However, whether and to what extent these admissions influence resident loved ones’ burden and health status remains currently unknown. Therefore, the primary objectives of this study are to investigate the differences between patients with COPD and resident loved ones’ perceptions of patients’ health status and problematic ADLs and to study prospectively the effects of a COPD exacerbation on resident loved ones’ perceptions of patients’ health status and problematic ADLs.Methods and analysisAn observational, longitudinal study will be performed in 192 patients with COPD and their 192 resident loved ones. Primary outcomes are daily functioning, ADL, disease-specific health status, generic health status and dyspnoea. These will be assessed during home visits at baseline and after 12 months. Additional home visits will be performed when a COPD exacerbation-related hospital admission occurs during the 12-month follow-up period.Ethics and disseminationThis protocol was approved by the Medical Ethics Committee of the Catharina Hospital Eindhoven, the Netherlands (NL42721.060.12/M12-1280) and is registered in the Dutch Trial Register (NTR3941).