bluuebunny/crossref_metadata_2025_split huggingface dataset:
A New ABC Half-Transporter in Leishmania major Is Involved in Resistance to Antimony
Authors: Manzano J. I. , García-Hernández R. , Castanys S. , Gamarro F. 
Published: May 2013

ABSTRACT

The characterization of ABCI4, a new intracellular ATP-binding cassette (ABC) half-transporter in
Leishmania major
, is described. We show that ABCI4 is involved in heavy metal export, thereby conferring resistance to Pentostam, to Sb(III), and to As(III) and Cd(II). Parasites overexpressing ABCI4 showed a lower mitochondrial toxic effect of antimony by decreasing reactive oxygen species production and maintained higher values of both the mitochondrial electrochemical potential and total ATP levels with respect to controls. The ABCI4 half-transporter forms homodimers as determined by a coimmunoprecipitation assay. A combination of subcellular localization studies under a confocal microscope and a surface biotinylation assay using parasites expressing green fluorescent protein- and FLAG-tagged ABCI4 suggests that the transporter presents a dual localization in both mitochondria and the plasma membrane. Parasites overexpressing ABCI4 present an increased replication in mouse peritoneal macrophages. We have determined that porphyrins are substrates for ABCI4. Consequently, the overexpression of ABCI4 confers resistance to some toxic porphyrins, such as zinc-protoporphyrin, due to the lower accumulation resulting from a significant efflux, as determined using the fluorescent zinc-mesoporphyrin, a validated heme analog. In addition, ABCI4 has a significant ability to efflux thiol after Sb(III) incubation, thus meaning that ABCI4 could be considered to be a potential thiol-X-pump that is able to recognize metal-conjugated thiols. In summary, we have shown that this new ABC transporter is involved in drug sensitivity to antimony and other compounds by efflux as conjugated thiol complexes.
Compensation of the Metabolic Costs of Antibiotic Resistance by Physiological Adaptation in Escherichia coli
Authors: Händel Nadine , Schuurmans J. Merijn , Brul Stanley , ter Kuile Benno H. 
Published: May 2013

ABSTRACT

Antibiotic resistance is often associated with metabolic costs. To investigate the metabolic consequences of antibiotic resistance, the genomic and transcriptomic profiles of an amoxicillin-resistant
Escherichia coli
strain and the wild type it was derived from were compared. A total of 125 amino acid substitutions and 7 mutations that were located <1,000 bp upstream of differentially expressed genes were found in resistant cells. However, broad induction and suppression of genes were observed when comparing the expression profiles of resistant and wild-type cells. Expression of genes involved in cell wall maintenance, DNA metabolic processes, cellular stress response, and respiration was most affected in resistant cells regardless of the absence or presence of amoxicillin. The SOS response was downregulated in resistant cells. The physiological effect of the acquisition of amoxicillin resistance in cells grown in chemostat cultures consisted of an initial increase in glucose consumption that was followed by an adaptation process. Furthermore, no difference in maintenance energy was observed between resistant and sensitive cells. In accordance with the transcriptomic profile, exposure of resistant cells to amoxicillin resulted in reduced salt and pH tolerance. Taken together, the results demonstrate that the acquisition of antibiotic resistance in
E. coli
is accompanied by specifically reorganized metabolic networks in order to circumvent metabolic costs. The overall effect of the acquisition of resistance consists not so much of an extra energy requirement, but more a reduced ecological range.
Single-Dose Pharmacodynamics of Amphotericin B against Aspergillus Species in an
In Vitro
Pharmacokinetic/Pharmacodynamic Model
Authors: Al-Saigh Rafal , Siopi Maria , Siafakas Nikolaos , Velegraki Aristea , Zerva Loukia , Meletiadis Joseph 
Published: May 2013

ABSTRACT

Conventional MIC testing of amphotericin B results in narrow MIC ranges challenging the detection of resistant strains. In order to discern amphotericin B pharmacodynamics, the
in vitro
activity of amphotericin B was studied against
Aspergillus
isolates with the same MICs by using a new
in vitro
pharmacokinetic/pharmacodynamic (PK/PD) model that simulates amphotericin B human plasma levels. Clinical isolates of
Aspergillus fumigatus
,
A. terreus
, and
A. flavus
with the same Clinical and Laboratory Standards Institute modal MICs of 1 mg/liter were exposed to amphotericin B concentrations following the plasma concentration-time profile after single-bolus administration with
C
max
values of 0.6, 1.2, 2.4, and 4.8 mg/liter. Fungal growth was monitored for up to 72 h based on galactomannan production. Complete growth inhibition was observed only against
A. fumigatus
with amphotericin B with a
C
max
of ≥2.4 mg/liter. At the lower
C
max
values 0.6 and 1.2 mg/liter, significant growth delays of 34 and 52 h were observed, respectively (
P
< 0.001). For
A. flavus
, there was no complete inhibition but a progressive growth delay of 1 to 50 h at an amphotericin B
C
max
of 0.6 to 4.8 mg/liter (
P
< 0.001). For
A. terreus
, the growth delay was modest (up to 8 h) at all
C
max
s (
P
< 0.05). The
C
max
(95% confidence interval) associated with 50% activity for
A. fumigatus
was 0.60 (0.49 to 0.72) mg/liter, which was significantly lower than for
A. flavus
3.06 (2.46 to 3.80) mg/liter and for
A. terreus
7.90 (5.20 to 12.29) mg/liter (
P
< 0.001). A differential
in vitro
activity of amphotericin B was found among
Aspergillus
species despite the same MIC in the order
A. fumigatus
>
A. flavus
>
A. terreus
in the
in vitro
PK/PD model, possibly reflecting the different concentration- and time-dependent inhibitory/killing activities amphotericin B exerted against these species.
FeoB-Mediated Uptake of Iron by Francisella tularensis
Authors: Thomas-Charles Cindy A. , Zheng Huaixin , Palmer Lance E. , Mena Patricio , Thanassi David G. , Furie Martha B. 
Published: May 2013

ABSTRACT

Francisella tularensis
, the bacterial cause of tularemia, infects the liver and replicates in hepatocytes
in vivo
and
in vitro
. However, the factors that govern adaptation of
F. tularensis
to the intrahepatocytic niche have not been identified. Using cDNA microarrays, we determined the transcriptional profile of the live vaccine strain (LVS) of
F. tularensis
grown in the FL83B murine hepatocytic cell line compared to that of
F. tularensis
cultured in broth. The
fslC
gene of the
fsl
operon was the most highly upregulated. Deletion of
fslC
eliminated the ability of the LVS to produce siderophore, which is involved in uptake of ferric iron, but it did not impair its growth in hepatocytes, A549 epithelial cells, or macrophages. Therefore, we sought an alternative means by which
F. tularensis
might obtain iron. Deletion of
feoB
, which encodes a putative ferrous iron transporter, retarded replication of the LVS in iron-restricted media, reduced its growth in hepatocytic and epithelial cells, and impaired its acquisition of iron. Survival of mice infected intradermally with a lethal dose of the LVS was slightly improved by deletion of
fslC
but was not altered by loss of
feoB
. However, the Δ
feoB
mutant showed diminished ability to colonize the lungs, liver, and spleen of mice that received sublethal inocula. Thus, FeoB represents a previously unidentified mechanism for uptake of iron by
F. tularensis
. Moreover, failure to produce a mutant strain lacking both
feoB
and
fslC
suggests that FeoB and the proteins of the
fsl
operon are the only major means by which
F. tularensis
acquires iron.
Escherichia coli Heat-Stable Toxin b Impairs Intestinal Epithelial Barrier Function by Altering Tight Junction Proteins
Authors: Ngendahayo Mukiza Clément , Dubreuil J. Daniel 
Published: May 2013

ABSTRACT

Escherichia coli
heat-stable toxin b (STb) causes diarrhea in animals. STb binds to sulfatide, its receptor, and is then internalized. In the cytoplasm, through a cascade of events, STb triggers the opening of ion channels, allowing ion secretion and water loss and leading to diarrhea. Tight junctions (TJs) are well known for controlling paracellular traffic of ions and water by forming a physical intercellular barrier in epithelial cells, and some bacterial toxins are known to affect adversely TJs. The present study aimed at determining the effect of STb on TJs. T84 cells were treated for 24 h with purified STb and a nontoxic STb mutant (D30V). Transepithelial resistance (TER), paracellular flux marker, and confocal microscopy were used to analyze the effect of STb on TJs. Purified STb caused a significant reduction of TER parallel to an increase in paracellular permeability compared to the results seen in untreated cells or mutant D30V. The increased paracellular permeability was associated with a marked alteration of F-actin stress fibers. F-actin filament dissolution and condensation were accompanied by redistribution and/or fragmentation of ZO-1, claudin-1, and occludin. These changes were also observed following treatment of T84 cells with an 8-amino-acid peptide found in the STb sequence corresponding to a consensus sequence of
Vibrio cholerae
Zot toxin. These effects were not observed with a scrambled peptide or mutant D30V. Our findings indicate that STb induces epithelial barrier dysfunction through changes in TJ proteins that could contribute to diarrhea.
Differential Virulence and Disease Progression following Mycobacterium tuberculosis Complex Infection of the Common Marmoset (Callithrix jacchus)
Authors: Via Laura E. , Weiner Danielle M. , Schimel Daniel , Lin Philana Ling , Dayao Emmanuel , Tankersley Sarah L. , Cai Ying , Coleman M. Teresa , Tomko Jaime , Paripati Praveen , Orandle Marlene , Kastenmayer Robin J. , Tartakovsky Michael , Rosenthal Alexander , Portevin Damien , Eum Seok Yong , Lahouar Saher , Gagneux Sebastien , Young Douglas B. , Flynn JoAnne L. , Barry Clifton E. 
Published: May 2013

ABSTRACTExisting small-animal models of tuberculosis (TB) rarely develop cavitary disease, limiting their value for assessing the biology and dynamics of this highly important feature of human disease. To develop a smaller primate model with pathology similar to that seen in humans, we experimentally infected the common marmoset (Callithrix jacchus) with diverse strains ofMycobacterium tuberculosisof various pathogenic potentials. These included recent isolates of the modern Beijing lineage, the Euro-American X lineage, andM. africanum. All three strains produced fulminant disease in this animal with a spectrum of progression rates and clinical sequelae that could be monitored in real time using 2-deoxy-2-[18F]fluoro-d-glucose (FDG) positron emission tomography (PET)/computed tomography (CT). Lesion pathology at sacrifice revealed the entire spectrum of lesions observed in human TB patients. The three strains produced different rates of progression to disease, various extents of extrapulmonary dissemination, and various degrees of cavitation. The majority of live births in this species are twins, and comparison of results from siblings with different infecting strains allowed us to establish that the infection was highly reproducible and that the differential virulence of strains was not simply host variation. Quantitative assessment of disease burden by FDG-PET/CT provided an accurate reflection of the pathology findings at necropsy. These results suggest that the marmoset offers an attractive small-animal model of human disease that recapitulates both the complex pathology and spectrum of disease observed in humans infected with variousM. tuberculosisstrain clades.
Human Embryonic Stem Cell Lines Model Experimental Human Cytomegalovirus Latency
Authors: Penkert Rhiannon R. , Kalejta Robert F. 
Published: May 2013

ABSTRACT

Herpesviruses are highly successful pathogens that persist for the lifetime of their hosts primarily because of their ability to establish and maintain latent infections from which the virus is capable of productively reactivating. Human cytomegalovirus (HCMV), a betaherpesvirus, establishes latency in CD34
+
hematopoietic progenitor cells during natural infections in the body. Experimental infection of CD34
+
cells
ex vivo
has demonstrated that expression of the viral gene products that drive productive infection is silenced by an intrinsic immune defense mediated by Daxx and histone deacetylases through heterochromatinization of the viral genome during the establishment of latency. Additional mechanistic details about the establishment, let alone maintenance and reactivation, of HCMV latency remain scarce. This is partly due to the technical challenges of CD34
+
cell culture, most notably, the difficulty in preventing spontaneous differentiation that drives reactivation and renders them permissive for productive infection. Here we demonstrate that HCMV can establish, maintain, and reactivate
in vitro
from experimental latency in cultures of human embryonic stem cells (ESCs), for which spurious differentiation can be prevented or controlled. Furthermore, we show that known molecular aspects of HCMV latency are faithfully recapitulated in these cells. In total, we present ESCs as a novel, tractable model for studies of HCMV latency.

IMPORTANCE
Human cytomegalovirus (HCMV) is a significant human pathogen that is known for causing birth defects, blindness in AIDS patients, and organ transplant rejection. The ability of HCMV to cause disease is dependent upon its capacity to establish and maintain latent infections. Very few of the molecular mechanisms of latency have been elucidated, due in part to the lack of a tractable cell culture model. Here we present embryonic stem cells (ESCs) as a model for HCMV latency, one in which genome maintenance and reactivation could be closely monitored. HCMV establishes latency in ESCs in the same fashion as it does in CD34
+
cells, the currently favored
in vitro
model. Hence, ESCs represent a novel model with unique properties, such as the ability to be genetically manipulated and cultured indefinitely in an undifferentiated state, that will facilitate the mechanistic examination of certain aspects of HCMV latency that have proven technically challenging in other model systems.
Metagenomic
De Novo
Assembly of an Aquatic Representative of the Verrucomicrobial Class
Spartobacteria
Authors: Herlemann Daniel P. R. , Lundin Daniel , Labrenz Matthias , Jürgens Klaus , Zheng Zongli , Aspeborg Henrik , Andersson Anders F. 
Published: May 2013

ABSTRACT

The verrucomicrobial subdivision 2 class
Spartobacteria
is one of the most abundant bacterial lineages in soil and has recently also been found to be ubiquitous in aquatic environments. A 16S rRNA gene study from samples spanning the entire salinity range of the Baltic Sea indicated that, in the pelagic brackish water, a phylotype of the
Spartobacteria
is one of the dominating bacteria during summer. Phylogenetic analyses of related 16S rRNA genes indicate that a purely aquatic lineage within the
Spartobacteria
exists. Since no aquatic representative from the
Spartobacteria
has been cultured or sequenced, the metabolic capacity and ecological role of this lineage are yet unknown. In this study, we reconstructed the genome and metabolic potential of the abundant Baltic Sea
Spartobacteria
phylotype by metagenomics. Binning of genome fragments by nucleotide composition and a self-organizing map recovered the near-complete genome of the organism, the gene content of which suggests an aerobic heterotrophic metabolism. Notably, we found 23 glycoside hydrolases that likely allow the use of a variety of carbohydrates, like cellulose, mannan, xylan, chitin, and starch, as carbon sources. In addition, a complete pathway for sulfate utilization was found, indicating catabolic processing of sulfated polysaccharides, commonly found in aquatic phytoplankton. The high frequency of glycoside hydrolase genes implies an important role of this organism in the aquatic carbon cycle. Spatiotemporal data of the phylotype’s distribution within the Baltic Sea indicate a connection to
Cyanobacteria
that may be the main source of the polysaccharide substrates.

IMPORTANCE
The ecosystem roles of many phylogenetic lineages are not yet well understood. One such lineage is the class
Spartobacteria
within the
Verrucomicrobia
that, despite being abundant in soil and aquatic systems, is relatively poorly studied. Here we circumvented the difficulties of growing aquatic
Verrucomicrobia
by applying shotgun metagenomic sequencing on a water sample from the Baltic Sea. By using a method based on sequence signatures, we were able to
in silico
isolate genome fragments belonging to a phylotype of the
Spartobacteria
. The genome, which represents the first aquatic representative of this clade, encodes a diversity of glycoside hydrolases that likely allow degradation of various complex carbohydrates. Since the phylotype cooccurs with
Cyanobacteria
, these may be the primary producers of the carbohydrate substrates. The phylotype, which is highly abundant in the Baltic Sea during summer, may thus play an important role in the carbon cycle of this ecosystem.
Identification of Promiscuous KIF20A Long Peptides Bearing Both CD4+ and CD8+ T-cell Epitopes: KIF20A-Specific CD4+ T-cell Immunity in Patients with Malignant Tumor
Authors: Tomita Yusuke , Yuno Akira , Tsukamoto Hirotake , Senju Satoru , Kuroda Yasuhiro , Hirayama Masatoshi , Irie Atsushi , Kawahara Kenta , Yatsuda Junji , Hamada Akinobu , Jono Hirofumi , Yoshida Koji , Tsunoda Takuya , Kohrogi Hirotsugu , Yoshitake Yoshihiro , Nakamura Yusuke , Shinohara Masanori , Nishimura Yasuharu 
Published: May 2013

AbstractPurpose: To identify long peptides (LP) derived from a novel tumor-associated antigen (TAA), kinesin family member 20A (KIF20A), which induce tumor-specific T-helper type 1 (TH1) cells and CTLs.Experimental Design: We combined information from a recently developed computer algorithm predicting HLA class II–binding peptides with KIF20A-derived CTL-epitope sequences presented by HLA-A2 (A*02:01) or HLA-A24 (A*24:02) to select candidate promiscuous TH1-cell epitopes containing CTL epitopes. Peripheral blood mononuclear cells (PBMC) derived from healthy donors or patients with head-and-neck malignant tumor (HNMT) were used to study the immunogenicity of KIF20A-LPs, and the in vitro cross-priming potential of KIF20A-LPs bearing CTL epitopes. We used HLA-A24 transgenic mice to address whether vaccination with KIF20A-LP induces efficient cross-priming of CTLs in vivo. The TH1-cell response to KIF20A-LPs in HNMT patients receiving immunotherapy with TAA-derived CTL-epitope peptides was analyzed using IFN-γ enzyme-linked immunospot assays.Results: We identified promiscuous KIF20A-LPs bearing naturally processed epitopes recognized by CD4+ T cells and CTLs. KIF20A-specific CTLs were induced by vaccination with a KIF20A-LP in vivo. KIF20A expression was detected in 55% of HNMT by immunohistochemistry, and significant frequencies of KIF20A-specific TH1 cell responses were detected after short-term in vitro stimulation of PBMCs with KIF20A-LPs in 50% of HNMT patients, but not in healthy donors. Furthermore, these responses were associated with KIF20A expression in HNMT tissues.Conclusions: These are the first results showing the presence of KIF20A-specific TH1 cell responses in HNMT patients and underline the possible utility of KIF20A-LPs for propagation of TH1 cells and CTLs. Clin Cancer Res; 19(16); 4508–20. ©2013 AACR.
Risk Factors for Cerebral Embolization After Carotid Artery Stenting With Embolic Protection
Authors: Bijuklic Klaudija , Wandler Andreas , Varnakov Yuriy , Tuebler Thilo , Schofer Joachim 
Published: May 2013

Background—
Meta-analyses of randomized trials have shown an increased risk of periprocedural stroke after carotid artery stenting (CAS) compared with carotid endarterectomy, which may differ in specific patient subgroups. Knowledge of risk factors for cerebral embolic lesions during CAS may impact treatment decisions for the individual patient, but these factors have not been extensively studied. We aimed to identify factors predictive for cerebral ischemic lesions during embolic protected CAS.

Methods and Results—
Preprocedural and postprocedural diffusion-weighted magnetic resonance imaging was performed for evaluation of new cerebral ischemic lesions in 728 (86.9%) of 837 consecutive patients undergoing CAS with cerebral embolic protection. Multivariable logistic regression analyses were performed to identify factors predictive for embolic lesions. New ischemic lesions were found in 32.8% of patients. Age, hypertension, lesion length, lesion eccentricity, and aortic arch type III were significantly associated with new ischemic lesions; calcified lesions were negatively associated. In 25% of these patients embolic lesions were also found in the contralateral hemisphere. Predictive factors for contralateral lesions were age, >50% stenosis of the contralateral internal carotid artery, and an aortic arch type II, with a trend for aortic arch type III.

Conclusions—
Age, hypertension, lesion morphology, and aortic arch type were predictive for procedural-related cerebral embolic lesions during embolic protected CAS. Age, significant contralateral carotid stenosis, and complex aortic arch type were predictive for bilateral ischemic lesions. The clinical implications of ischemic lesions are not yet fully understood.
Thyroxin Treatment Protects Against White Matter Injury in The Immature Brain via Brain-Derived Neurotrophic Factor
Authors: Hung Pi-Lien , Huang Chao-Ching , Huang Hsiu-Mei , Tu Dom-Gene , Chang Ying-Chao 
Published: May 2013

Background and Purpose—
Low level of thyroid hormone is a strong independent risk factor for white matter (WM) injury, a major cause of cerebral palsy, in preterm infants. Thyroxin upregulates brain-derived neurotrophic factor during development. We hypothesized that thyroxin protected against preoligodendrocyte apoptosis and WM injury in the immature brain via upregulation of brain-derived neurotrophic factor.

Methods—
Postpartum (P) day-7 male rat pups were exposed to hypoxic ischemia (HI) and intraperitoneally injected with thyroxin (T4; 0.2 mg/kg or 1 mg/kg) or normal saline immediately after HI at P9 and P11. WM damage was analyzed for myelin formation, axonal injury, astrogliosis, and preoligodendrocyte apoptosis. Neurotrophic factor expression was assessed by real-time polymerase chain reaction and immunohistochemistry. Neuromotor functions were measured using open-field locomotion (P11 and P21), inclined plane climbing (P11), and beam walking (P21). Intracerebroventricular injection of TrkB-Fc or systemic administration of 7,8-dihydroxyflavone was performed.

Results—
On P11, the HI group had significantly lower blood T4 levels than the controls. The HI group showed ventriculomegaly and marked reduction of myelin basic protein immunoreactivities in the WM. T4 (1 mg/kg) treatment after HI markedly attenuated axonal injury, astrocytosis, and microgliosis, and increased preoligodendrocyte survival. In addition, T4 treatment significantly increased myelination and selectively upregulated brain-derived neurotrophic factor expression in the WM, and improved neuromotor deficits after HI. The protective effect of T4 on WM myelination and neuromotor performance after HI was significantly attenuated by TrkB-Fc. Systemic 7,8-dihydroxyflavone treatment ameliorated hypomyelination after HI injury.

Conclusions—
T4 protects against WM injury at both pathological and functional levels via upregulation of brain-derived neurotrophic factor–TrkB signaling in the immature brain.